Aryl hydrocarbon receptor（AhR）activation

91-20-3

UFWIBTONFRDIAS-UHFFFAOYSA-N

Naphthalene

Albocarbon Dezodorator Moth flakes naftaleno, puro Naphtalene NAPHTHALENE SCALES Naphthalin Naphthene Napthalene NSC 37565 Tar camphor UN 2304 White tar

DTXSID8020913

83-32-9

CWRYPZZKDGJXCA-UHFFFAOYSA-N

Acenaphthene

Acenaphthylene, 1,2-dihydro- 1,2-Dihydroacenaphthylene 1,8-Ethylenenaphthalene acenafteno Acenaphtene Acenaphthen Naphthyleneethylene NSC 7657 peri-Ethylenenaphthalene

DTXSID3021774

208-96-8

HXGDTGSAIMULJN-UHFFFAOYSA-N

Acenaphthylene

acenaftileno acenaphthyene Acenaphthylen Acenaphtylene Cyclopenta[de]naphthalene NSC 59821

DTXSID3023845

86-73-7

NIHNNTQXNPWCJQ-UHFFFAOYSA-N

Fluorene

9H-Fluorene 2,2'-Methylenebiphenyl Diphenylenemethane Fluoren fluoreno Methane, diphenylene- NSC 6787 o-Biphenylenemethane

DTXSID8024105

85-01-8

YNPNZTXNASCQKK-UHFFFAOYSA-N

Phenanthrene

[3]Helicene fenantreno, puro NSC 26256 Phenanthren Ravatite

DTXSID6024254

120-12-7

MWPLVEDNUUSJAV-UHFFFAOYSA-N

Anthracene

Anthracen Anthracin antraceno, puro Green Oil NSC 7958 Paranaphthalene Tetra Olive N2G UN 1136

DTXSID0023878

206-44-0

GVEPBJHOBDJJJI-UHFFFAOYSA-N

Fluoranthene

ClusterCarbon 1,2-(1,8-Naphthylene)benzene Benzene, 1,2-(1,8-naphthalenediyl)- Benzo[jk]fluorene fluoranteno Fluoranthen NSC 6803

DTXSID3024104

129-00-0

BBEAQIROQSPTKN-UHFFFAOYSA-N

Pyrene

Benzo (d,e,f) phenanthrene Benzo[def]phenanthrene NSC 17534 NSC 66449 β-Pyrene

DTXSID3024289

218-01-9

WDECIBYCCFPHNR-UHFFFAOYSA-N

Chrysene

[4]Phenacene 1,2-Benzophenanthrene 1,2-Benzphenanthrene Benzo(a)phenanthrene Benzo[a]phenanthrene Chrysen criseno NSC 6175

DTXSID0022432

205-99-2

FTOVXSOBNPWTSH-UHFFFAOYSA-N

Benzo(b)fluoranthene

Benz[e]acephenanthrylene 2,3-Benzfluoranthene 2,3-Benzofluoranthrene 3,4-Benz(e)acephenanthrylene 3,4-Benz[e]acephenanthrylene 3,4-Benzfluoranthene 3,4-Benzofluoranthene Benz(e)acephenanthrylen benz(e)acephenanthrylene Benzo(b)flluoranthene benzo(e)acefenantrileno benzo(e)acephenanthrylene Benzo(e)fluoranthene Benzo[b]fluoranthene Benzo[e]fluoranthene NSC 89265

DTXSID0023907

207-08-9

HAXBIWFMXWRORI-UHFFFAOYSA-N

Benzo(k)fluoranthene

11,12-Benzofluoranthene 2,3,1',8'-Binaphthylene 8,9-Benzfluoranthene 8,9-Benzofluoranthene benzo(k)fluoranteno Benzo(k)fluoranthen Dibenzo[b,jk]fluorene

DTXSID0023909

50-32-8

FMMWHPNWAFZXNH-UHFFFAOYSA-N

Benzo(a)pyrene

BaP Benzo[a]pyrene 3,4-Benz[a]pyrene 3,4-Benzopyrene 3,4-Benzpyrene 6,7-Benzopyrene BENZ(A)PYREN Benz(a)pyrene Benz[a]pyrene Benzo[d,e,f]chrysene Benzo[def]chrysen Benzo[def]chrysene benzo[def]criseno NSC 21914 Benzo(d,e,f)chrysene 3,4-Benzo(a)pyrene 3,4-Benz(a)pyrene EINECS 200-028-5 RCRA waste number U022 3,4-Benzopirene 3,4-Benzpyren UNII-3417WMA06D

DTXSID2020139

56-55-3

DXBHBZVCASKNBY-UHFFFAOYSA-N

Benz(a)anthracene

Benz[a]anthracene 1,2-Benz[a]anthracene 1,2-Benzanthracene 1,2-Benzanthrene 1,2-Benzoanthracene 2,3-Benzophenanthrene 2,3-Benzphenanthrene BENZ(A)ANTHRACEN Benz[a]anthracen Benzanthracene Benzanthrene Benzo(a)anthracene Benzo[a]anthracene benzo[a]antraceno Benzo[b]phenanthrene Benzoanthracene Naphthanthracene NSC 30970 Tetraphene

DTXSID5023902

193-39-5

SXQBHARYMNFBPS-UHFFFAOYSA-N

Indeno(1,2,3-cd)pyrene

Indeno[1,2,3-cd]pyrene 1,10-(1,2-Phenylene)pyrene 1,10-(o-Phenylene)pyrene Indeno(1,2,3-c,d)pyrene indeno[1,2,3-cd]pireno Indeno[1,2,3-cd]pyren

DTXSID8024153

53-70-3

LHRCREOYAASXPZ-UHFFFAOYSA-N

Dibenz(a,h)anthracene

Dibenz[a,h]anthracene 1,2:5,6-Benzanthracene 1,2:5,6-Dibenz[a]anthracene 1,2:5,6-Dibenzanthracen 1,2:5,6-Dibenzanthracene 1,2:5,6-Dibenzoanthracene DIBENZ(A,H)ANTHRACEN Dibenz[a,h]anthracen Dibenzo(a,h)anthracene Dibenzo[a,h]anthracene dibenzo[a,h]antraceno NSC 22433

DTXSID9020409

191-24-2

GYFAGKUZYNFMBN-UHFFFAOYSA-N

Benzo(g,h,i)perylene

Benzo[ghi]perylene 1,12-Benzoperylene 1,12-Benzperylene BENZO(GHI)PERYLEN Benzo(ghi)perylene benzo[ghi]perileno Benzo[ghi]perylen NSC 89275

DTXSID5023908

60-35-5

DLFVBJFMPXGRIB-UHFFFAOYSA-N

Acetamide

Acetamid acetamida Acetic acid amide Acetimidic acid Ethanamide Ethanimidic acid Methanecarboxamide NSC 25945

DTXSID7020005

103-90-2

RZVAJINKPMORJF-UHFFFAOYSA-N

Acetaminophen

4-Acetamidophenol APAP Paracetamol 4-hydroxyacetanilide Acetamide, N-(4-hydroxyphenyl)- 4-(Acetylamino)phenol 4-(N-Acetylamino)phenol 4-Acetaminophenol 4'-Hydroxyacetanilide Abensanil Acetagesic Acetalgin ACETAMIDE, N-(4-HYDROXYPHENYL) Acetaminofen Acetanilide, 4'-hydroxy- ACETANILIDE, 4-HYDROXY- Algotropyl Alvedon Anaflon Apamide Banesin Ben-u-ron Bickie-mol Biocetamol Cetadol Citramon P Claratal Clixodyne Dafalgan Daphalgan Dial-a-gesic Disprol Doliprane Dolprone Dymadon Efferalgan Endophy Febrilex Febrilix Febro-Gesic Febrolin Fepanil Finimal Gattaphen T Gelocatil Gutte Enteric Homoolan Jin Gang Lestemp Liquagesic Lonarid Lyteca Syrup Minoset Momentum N-(4-Hydroxyphenyl)acetamide N-Acetyl-4-aminophenol N-Acetyl-4-hydroxyaniline N-Acetyl-p-aminophenol Napafen Naprinol Nobedon NSC 109028 NSC 3991 Ortensan p-(Acetylamino)phenol p-Aceaminophenol Pacemol p-Acetamidophenol p-Acetoaminophen P-ACETYLAMINOPHENOL Paldesic panadeine Panadol Panadol Actifast Panadol Extend Panaleve Panasorb Panodil Paracetamol DC Paracetamole Parageniol Paramol Paraspen Parelan Pasolind N Perfalgan Phenaphen Phendon p-Hydroxyacetanilide Prodafalgan Puerxitong Pyrinazine Resfenol Resprin Rhodapop NCR Salzone Tabalgin Tachipirina Tempanal Tralgon Tylenol TylolHot Valadol Valgesic Vermidon Vick Pyrena

DTXSID2020006

968-81-0

VGZSUPCWNCWDAN-UHFFFAOYSA-N

Acetohexamide

Benzenesulfonamide, 4-acetyl-N-[(cyclohexylamino)carbonyl]- 1-(p-Acetylbenzenesulfonyl)-3-cyclohexylurea 1-[(p-Acetylphenyl)sulfonyl]-3-cyclohexylurea Acetohexamid acetohexamida Dimelin Dimelor Dymelor Gamadiabet Hypoglicil Metaglucina Minoral N-(p-Acetylphenylsulfonyl)-N'-cyclohexylurea Ordimel Tsiklamid Urea, 1-[(p-acetylphenyl)sulfonyl]-3-cyclohexyl-

DTXSID7020007

67-66-3

HEDRZPFGACZZDS-UHFFFAOYSA-N

Chloroform

Trichloromethane Methane, trichloro- CARBON TRICHLORIDE Chloroforme cloroformo Formyl trichloride Methane trichloride Methane,trichloro- NSC 77361 Trichloroform UN 1888

DTXSID1020306

110-00-9

YLQBMQCUIZJEEH-UHFFFAOYSA-N

Furan

Divinylene oxide furanne Furfuran Oxacyclopentadiene Tetrole UN 2389

DTXSID6020646

7429-90-5

XAGFODPZIPBFFR-UHFFFAOYSA-N

AZDRQVAHHNSJOQ-UHFFFAOYSA-N

Aluminum

Aisin Metal Fiber Al 050P-H24 ALC Fine Alcan XI 1391 Almi-Paste SSP 303AR Aloxal 3010 Alpaste 00-0506 Alpaste 0100M Alpaste 0100MA Alpaste 0100M-C Alpaste 0200M Alpaste 0200T Alpaste 0230M Alpaste 0230T Alpaste 0241M Alpaste 0300M Alpaste 0500M Alpaste 0539X Alpaste 0620MS Alpaste 0625TS Alpaste 0638-70C Alpaste 0700M Alpaste 0780M Alpaste 0900M Alpaste 100M Alpaste 100MS Alpaste 100MSR Alpaste 1100M Alpaste 1100MA Alpaste 1100N Alpaste 1100NA Alpaste 1109MA Alpaste 1109MC Alpaste 1200M Alpaste 1200T Alpaste 1260MS Alpaste 1500MA Alpaste 1700NL Alpaste 1810YL Alpaste 1830YL Alpaste 1900M Alpaste 1900XS Alpaste 1950M Alpaste 1950N Alpaste 210N Alpaste 2172EA Alpaste 2173 Alpaste 240T Alpaste 241M Alpaste 417 Alpaste 46-046 Alpaste 4-621 Alpaste 4919 Alpaste 50-63 Alpaste 50-635 Alpaste 51-148B Alpaste 51-231 Alpaste 5205N Alpaste 5207N Alpaste 52-509 Alpaste 52-568 Alpaste 5301N Alpaste 5302N Alpaste 53-119 Alpaste 5422NS Alpaste 54-452 Alpaste 54-497 Alpaste 54-542 Alpaste 55-516 Alpaste 55-519 Alpaste 55-574 Alpaste 5620NS Alpaste 5630NS Alpaste 5640NS Alpaste 56-501 Alpaste 5650NS Alpaste 5653NS Alpaste 5654NS Alpaste 5680N Alpaste 5680NS Alpaste 60-600 Alpaste 60-760 Alpaste 60-768 Alpaste 62-356 Alpaste 6340NS Alpaste 6370NS Alpaste 6390NS Alpaste 640NS Alpaste 65-388 Alpaste 66NLB Alpaste 710N Alpaste 7130N Alpaste 7160N Alpaste 7160NS Alpaste 725N Alpaste 740NS Alpaste 7430NS Alpaste 7580NS Alpaste 7620NS Alpaste 7640NS Alpaste 7670M Alpaste 7670NS Alpaste 7675NS Alpaste 7679NS Alpaste 7680N Alpaste 7680NS Alpaste 76840NS Alpaste 7730N Alpaste 7770N Alpaste 7830N Alpaste 8004 Alpaste 8080N Alpaste 8260NAR Alpaste 891K Alpaste 91-0562 Alpaste 92-0592 Alpaste 93-0595 Alpaste 93-0647 Alpaste 94-2315 Alpaste 95-0570 Alpaste 96-0635 Alpaste 96-2104 Alpaste 97-0510 Alpaste 97-0534 Alpaste AW 520B Alpaste AW 612 Alpaste AW 9800 Alpaste F 795 Alpaste FM 7680K Alpaste FX 440 Alpaste FX 910 Alpaste FZ 0534 Alpaste FZU 40C Alpaste G Alpaste HR 8801 Alpaste HS 2 Alpaste J Alpaste K 9800 Alpaste MC 666 Alpaste MC 707 Alpaste MF 20 Alpaste MG 01 Alpaste MG 1000 Alpaste MG 1300 Alpaste MG 500 Alpaste MG 600 Alpaste MH 6601 Alpaste MH 8801 Alpaste MH 9901 Alpaste MR 7000 Alpaste MR 9000 Alpaste MS 630 Alpaste N 1700NL Alpaste NS 7670 Alpaste O 100N Alpaste O 2130 Alpaste O 300M Alpaste P 0100 Alpaste P 1950 Alpaste S Alpaste SAP 110 Alpaste SAP 414P Alpaste SAP 550N Alpaste SCR 5070 Alpaste TCR 2020 Alpaste TCR 2060 Alpaste TCR 2070 Alpaste TCR 3010 Alpaste TCR 3030 Alpaste TCR 3040 Alpaste TCR 3130 Alpaste TD 200T Alpaste UF 500 Alpaste WB 0230 Alpaste WD 500 Alpaste WJP-U 75C Alpaste WX 0630 Alpaste WX 7830 Alpaste WXA 7640 Alpaste WXM 0630 Alpaste WXM 0650 Alpaste WXM 0660 Alpaste WXM 1415 Alpaste WXM 1440 Alpaste WXM 5422 Alpaste WXM 760b Alpaste WXM 7640 Alpaste WXM 7675 Alpaste WXM-T 60B Alpaste WXM-U 75 Alpaste WXM-U 75C Altop X Aluchrome Ultrafin Super Alumat 1600 Alumet H 30 aluminio Aluminium Aluminium Flake Aluminum 27 Aluminum atom Aluminum element Aluminum Flake PCF 7620 Aluminum granules ALUMINUM METAL/GRANULE ALUMINUM PASTE ALUMINUM PIGMENT ALUMINUM TURNINGS Alumi-paste 640NS Alumipaste 91-0562 Alumipaste 98-1822T Alumipaste AW 620 Alumipaste CR 300 Alumipaste GX 180A Alumipaste GX 201A Alumipaste HR 7000 Alumipaste HR 850 Alumipaste MG 11 Alumipaste MH 8801 Aquamet NPW 2900 Aquapaste 205-5 Aquasilver LPW Astroflake 40 Astroflake Black N 020 Astroflake Black N 070 Astroflake LG 40 Astroflake LG 70 Astroflake Silver N 040 Astroshine NJ 1600 Astroshine T 8990 Atomizalumi VA 200 C.I. PIGMENT METAL 1 Chromal IV Chromal X Decomet 1001/10 Decomet 2018/10 Decomet High Gloss Al 1002/10 Ecka AS 081 Eckart 9155 Eterna Brite 301-1 Eterna Brite 601-1 Eterna Brite 651-1 Eterna Brite EBP 251PA Eterna Brite Primier 251PA Ferro FX 53-038 Friend Color F 500GR-W Friend Color F 500WT Friend Color F 700RE-W Friend Color F 701RE-W Hi Print 60T High Print 60T Hisparkle HS 2 Hydro Paste 8726 Hydrolac WHH 2153 Hydrolan 3560 Hydrolux Reflexal 100 Hydroshine WS 1001 JISA 51010P Kryal Z Lansford 243 LE Sheet 800 Leafing Alpaste LG-H Silver 25 Lunar Al-V 95 Metallux 161 Metallux 2154 Metallux 2192 Metalure Metalure 55350 Metalure L 55350 Metalure L 59510 Metalure W 2001 Metapor Metasheen 1800 Metasheen HR 0800 Metasheen KM 100 Metasheen KM 1000 Metasheen Slurry 1807 Metasheen Slurry 1811 Metasheen Slurry KM 100 Metax G Metax S Mirror Glow 1000 Mirror Glow 600 Mirrorsheen Noral Aluminium Noral Ink Grade Aluminium Obron 10890 Offset FM 4500 Puratronic Reflexal 145 Reynolds 400 Reynolds 4-301 Reynolds 4-591 Reynolds 667 SAP 260PW-HS SAP-FM 4010 SBC 516-20Z Scotchcal 7755SE Serumekku Setanium 50MIS-H8 Siberline ET 2025 Siberline ST 21030E1 Silvar A Silver VT 522 Silverline SSP 353 Silvex 793-20C Sparkle Silver 3141ST Sparkle Silver 3500 Sparkle Silver 3641 Sparkle Silver 5000AR Sparkle Silver 516AR Sparkle Silver 5242AR Sparkle Silver 5245AR Sparkle Silver 5271AR Sparkle Silver 5500 Sparkle Silver 5745 Sparkle Silver 7000AR Sparkle Silver 7005AR Sparkle Silver 7500 Sparkle Silver 960-25E1 Sparkle Silver E 1745AR Sparkle Silver L 1526AR Sparkle Silver Premier 751 Sparkle Silver SS 3130 Sparkle Silver SS 5242AR Sparkle Silver SS 5588 Sparkle Silver SSP 132AR Special PCR 507 Splendal 6001BG Spota Mobil 801 SSP 760-20C Stapa Aloxal PM 2010 Stapa Aloxal PM 3010 Stapa Aloxal PM 4010 Stapa Hydrolac BG 8n.1 Stapa Hydrolac BGH Chromal X Stapa Hydrolac PM Chromal VIII Stapa Hydrolac W 60NL Stapa Hydrolac WH 16 Stapa Hydrolac WH 66NL Stapa Hydrolux 2192 Stapa Hydrolux 8154 Stapa IL Hydrolan 2192-55900G Stapa Metallic R 607 Stapa Metallux 1050 Stapa Metallux 211 Stapa Metallux 212 Stapa Metallux 2196 Stapa Metallux 274 Stapa Mobilux 181 Stapa Offset 3000 Stapa PV 10 Stapa VP 46432G Starbrite 2100 Super Fine 18000 Super Fine 22000 Supramex 2022 Toyo Aluminum 02-0005 Toyo Aluminum 93-3040 Transmet K 102HE Tufflake 3645 Tufflake 5843 UN 1396 US Aluminum 809 Valimet H 2 Valimet H 3 White Silver 7080N White Silver 7130N

DTXSID3040273

7440-43-9

BDOSMKKIYDKNTQ-UHFFFAOYSA-N

Cadmium

Cadimium CADMIUM BLUE CADMIUM, IN PLATTEN, STANGEN, BROCKEN,KOERNER

DTXSID1023940

7439-97-6

QSHDDOUJBYECFT-UHFFFAOYSA-N

Mercury

Liquid silver Mercure MERCURIC METAL TRIPLE DISTILLED mercurio Mercury element Quecksilber Quicksilver UN 2024 UN 2809

DTXSID1024172

7440-61-1

JFALSRSLKYAFGM-UHFFFAOYSA-N

Uranium

Uranium, isotope of mass 238 238U Element UN 2979 (DOT) Uranium I

DTXSID1042522

7440-38-2

RQNWIZPPADIBDY-UHFFFAOYSA-N

Arsenic

As Arsenic black ARSENIC METAL arsenico Grey arsenic UN 1558

DTXSID4023886

7440-22-4

BQCADISMDOOEFD-UHFFFAOYSA-N

Silver

Ag Nanopaste NPS-J 90 Ag Sphere 2 Ag-C-GS Algaedyn Arctic Silver 3 Argentum Astroflake 5 Carey Lea silver Colloidal silver Dotite XA 208 Du Pont 4943 ECM 100AF4810 Enlight 600 Enlight silver plate 600 Epinall Finesphere SVND 102 Fordel DC FP 5369-502 Jelcon SH 1 Jungindai Takasago 300 KS (metal) LCP 1-19SFS Metz 3000-1 Nanomelt AGC-A Nanomelt Ag-XA 301 Nanomelt Ag-XF 301 Nanomelt Ag-XF 301H Nanopaste NPS-J 90 Perfect Silver Puff Silver X 1200 RT 1710S-C1 SD (metal) Shell Silver Silbest E 20 Silbest F 20 Silbest J 18 Silbest TC 12 Silbest TC 20E Silbest TC 25A Silbest TCG 1 Silbest TCG 7 Silcoat AgC 103 Silcoat AgC 2011 Silcoat AgC 209 Silcoat AgC 2190 Silcoat AgC 222 Silcoat AgC 2411 Silcoat AgC 74T Silcoat AgC-A Silcoat AgC-AO Silcoat AgC-B Silcoat AgC-BO Silcoat AgC-D Silcoat AgC-G Silcoat AgC-GS Silcoat AgC-L Silcoat AgC-O Silcoat GS Silcoat RF 200 Silflake 135 Silsphere 514 Silver atom Silver element Silver Flake 1 Silver Flake 25 Silver Flake 52 Silver Flake 7A SILVER FLAKES Silver metal Silvest TCG 11N Technic 299 Technic 450 Techno Alpha 175

DTXSID4024305

7439-96-5

PWHULOQIROXLJO-UHFFFAOYSA-N

Manganese

Colloidal manganese Cutaval Manganese element Manganese fulleride Manganese metal alloy Manganese-55 manganeso

DTXSID2024169

7440-02-0

PXHVJJICTQNCMI-UHFFFAOYSA-N

Nickel

Carbonyl 255 Carbonyl Ni 123 Carbonyl Ni 283 Carbonyl Nickel 123 Carbonyl Nickel 283 Carbonyl Nickel 287 Cerac N 2003 CNS 10 Micron Exmet 4 Ni X-4/0 Fibrex P Incofoam Nickel element NICKEL ROUND ANODES Nicrobraz LM:BNi 2 Ni-Flake 95 Novamet 123 Novamet 4SP Novamet 4SP10 Novamet 525 Novamet CNS 400 Novamet HCA 1 Novamet NI 255 Raney nickel Raney nickel 2800 UN 1325 UN 2881

DTXSID2020925

7440-66-6

HCHKCACWOHOZIP-UHFFFAOYSA-N

Zinc

Zn Asarco L 15 C.I. Pigment Black 16 Merrillite NC-Zinc Rheinzink Stapa TE Zinc AT UF (metal) UN 1436 Zinc dust Zinc Dust 3 Zinc Dust 500 mesh Zinc Dust LS 2 Zinc Dust MCS Zinc Flakes GTT ZINC METAL ZINC MOSSY ZINC STRIP ZINC, MOSSY Zincsalt GTT

DTXSID7035012

CHEBI:16991

CHEBI deoxyribonucleic acid

GO:0005739

GO mitochondrion

MP:0003674

MP oxidative stress

GO:0006954

GO inflammatory response

WIKI increased

WIKI functional change

Naphthalene

2026-07-03T11:28:12

Acenaphthene

2026-07-09T06:10:36

Acenaphthylene

2026-07-09T06:14:20

Fluorene

2026-07-09T06:14:57

Phenanthrene

2016-11-29T18:42:22

Anthracene

2026-07-03T11:29:36

Fluoranthene

2026-07-09T06:28:08

Pyrene

2026-07-03T11:30:22

Chrysene

2026-07-03T11:31:50

Benzo(b)fluoranthene

2026-07-03T11:22:50

Benzo(k)fluoranthene

2016-11-29T18:42:08

Benzo(a)pyrene

2020-03-20T20:17:42

Benz(a)anthracene

2026-07-03T11:31:06

Indeno(1,2,3-cd)pyrene

2026-07-09T07:32:19

Dibenz(a,h)anthracene

2026-07-09T07:32:49

Benzo(g,h,i)perylene

2026-07-09T07:33:24

Acetaminophen

2016-11-29T18:42:26

Chloroform

2016-11-29T18:42:27

furan

2020-05-01T14:35:22

Platinum

2022-02-04T14:36:54

Aluminum

2022-02-04T14:42:11

Cadmium

2017-10-25T08:33:12

Mercury

2016-11-29T18:42:19

Uranium

2021-08-05T14:28:50

Arsenic

2021-04-27T00:15:21

Silver

2022-02-03T11:20:11

Manganese

2022-02-04T14:47:23

Nickel

2022-02-04T14:47:59

Zinc

2022-02-04T15:05:00

nanoparticles

2016-12-21T09:40:06

Ionizing Radiation Ionizing radiation can vary in energy, dose, charge, and in the spatial distributions of energy transferred to other matter (linear energy transfer per unit length or LET) (ICRU 1970). At the same dose, low and high LET both generate energy deposition events, including many higher energy events (Goodhead and Nikjoo 1989). However, they differ in the spatial distribution and upper range of intensity of energy deposited. Lower LET such as gamma rays sparsely deposit many individual excitations or small clusters of excitations of low energy (Goodhead 1988). In contrast, high LET such as alpha particles have fewer tracks but readily transfer their energy to matter and therefore deposit their energy over a much smaller area (Goodhead 1994). Consequently, alpha and other high LET particles penetrate less deeply into tissue, interactions are densely focused on a narrow track, and individual energy depositions can be large (Goodhead 1988). These different energy deposition patterns can lead to differences in radiation effects including the pattern of DNA damage.

Exposure to ionizing radiation can come from natural and industrial sources. Space and terrestrial radiation includes a range of LET particles, while diagnostic radiation methods such as X-ray imaging, mammography and CT scans use low LET X-rays. Radiation therapy can use an external beam to direct radiation on a focused tissue area, or deposit solid or liquid radioactive materials in the body that release (mostly gamma) radiation internally. External radiotherapy typically uses X-rays but is moving towards higher LET charged particles such as protons and heavy ions (Durante, Orecchia et al. 2017).

2019-05-03T12:36:36

2019-05-07T12:12:13

Estrogen

2019-05-08T11:40:27

Nanoparticles and Micrometer Particles

2022-02-04T13:43:43

WikiUser_26

ApacheUser rodents

9606

NCBI Homo sapiens

WCS_9606

common toxicological species human

10090

NCBI mouse

10116

NCBI rat

WCS_7227

common ecological species Drosophila melanogaster

6239

NCBI Caenorhabditis elegans

10090

NCBI Mus musculus

10116

NCBI Rattus norvegicus

Aryl hydrocarbon receptor（AhR）activation

AhR activation

Molecular

AOPWiki 2026-07-09T03:41:46

2026-07-09T03:41:46

Increase, Oxidative Stress

Molecular

Oxidative stress is defined as an imbalance in the production of reactive oxygen species (ROS) and antioxidant defenses. High levels of oxidizing free radicals can be very damaging to cells and molecules within the cell.  As a result, the cell has important defense mechanisms to protect itself from ROS. For example, Nrf2 is a transcription factor and master regulator of the oxidative stress response. During periods of oxidative stress, Nrf2-dependent changes in gene expression are important in regaining cellular homeostasis (Nguyen, et al., 2009) and can be used as indicators of the presence of oxidative stress in the cell.  In addition to the directly damaging actions of ROS, cellular oxidative stress also changes cellular activities on a molecular level. Redox sensitive proteins have altered physiology in the presence and absence of ROS, which is caused by the oxidation of sulfhydryls to disulfides on neighboring amino acids (Antelmann & Helmann 2011). Importantly Keap1, the negative regulator of Nrf2, is regulated in this manner (Itoh, et al. 2010).  ROS also undermine the mitochondrial defense system from oxidative damage. The antioxidant systems consist of superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase, as well as antioxidants such as α-tocopherol and ubiquinol, or antioxidant vitamins and minerals including vitamin E, C, carotene, lutein, zeaxanthin, selenium, and zinc (Fletcher, 2010). The enzymes, vitamins and minerals catalyze the conversion of ROS to non-toxic molecules such as water and O2. However, these antioxidant systems are not perfect and endogenous metabolic processes and/or exogenous oxidative influences can trigger cumulative oxidative injuries to the mitochondria, causing a decline in their functionality and efficiency, which further promotes cellular oxidative stress (Balasubramanian, 2000; Ganea & Harding, 2006; Guo et al., 2013; Karimi et al., 2017).   However, an emerging viewpoint suggests that ROS-induced modifications may not be as detrimental as previously thought, but rather contribute to signaling processes (Foyer et al., 2017).    Sources of ROS Production  Direct Sources: Direct sources involve the deposition of energy onto water molecules, breaking them into active radical species. When ionizing radiation hits water, it breaks it into hydrogen (H*) and hydroxyl (OH*) radicals by destroying its bonds. The hydrogen will create hydroxyperoxyl free radicals (HO2*) if oxygen is available, which can then react with another of itself to form hydrogen peroxide (H2O2) and more O2 (Elgazzar and Kazem, 2015). Antioxidant mechanisms are also affected by radiation, with catalase (CAT) and peroxidase (POD) levels rising as a result of exposure (Seen et al. 2018; Ahmad et al. 2021).   Indirect Sources: An indirect source of ROS is the mitochondria, which is one of the primary producers in eukaryotic cells (Powers et al., 2008).  As much as 2% of the electrons that should be going through the electron transport chain in the mitochondria escape, allowing them an opportunity to interact with surrounding structures. Electron-oxygen reactions result in free radical production, including the formation of hydrogen peroxide (H2O2) (Zhao et al., 2019). The electron transport chain, which also creates ROS, is activated by free adenosine diphosphate (ADP), O2, and inorganic phosphate (Pi) (Hargreaves et al. 2020; Raimondi et al. 2020; Vargas-Mendoza et al. 2021). The first and third complexes of the transport chain are the most relevant to mammalian ROS production (Raimondi et al., 2020). The mitochondria has its own set of DNA and it is a prime target of oxidative damage (Guo et al., 2013). ROS is also produced through nicotinamide adenine dinucleotide phosphate oxidase (Nox) stimulation, an event commenced by angiotensin II, a product/effector of the renin-angiotensin system (Nguyen Dinh Cat et al. 2013; Forrester et al. 2018). Other ROS producers include xanthine oxidase, immune cells (macrophage, neutrophils, monocytes, and eosinophils), phospholipase A2 (PLA2), monoamine oxidase (MAO), and carbon-based nanomaterials (Powers et al. 2008; Jacobsen et al. 2008; Vargas-Mendoza et al. 2021).

Oxidative Stress: Direct measurement of ROS is difficult because ROS are unstable. The presence of ROS can be assayed indirectly by measurement of cellular antioxidants, or by ROS-dependent cellular damage. Listed below are common methods for detecting the KE, however there may be other comparable methods that are not listed  <ul> <li>Detection of ROS by chemiluminescence (https://www.sciencedirect.com/science/article/abs/pii/S0165993606001683) </li> <li>Detection of ROS by chemiluminescence is also described in OECD TG 495 to assess phototoxic potential. </li> <li>Glutathione (GSH) depletion. GSH can be measured by assaying the ratio of reduced to oxidized glutathione (GSH:GSSG) using a commercially available kit (e.g., http://www.abcam.com/gshgssg-ratio-detection-assay-kit-fluorometric-green- ab138881.html). </li> <li>TBARS. Oxidative damage to lipids can be measured by assaying for lipid peroxidation using TBARS (thiobarbituric acid reactive substances) using a commercially available kit. </li> <li>8-oxo-dG. Oxidative damage to nucleic acids can be assayed by measuring 8-oxo-dG adducts (for which there are a number of ELISA based commercially available kits),or HPLC, described in Chepelev et al. (Chepelev, et al. 2015). </li> </ul>    Molecular Biology: Nrf2. Nrf2’s transcriptional activity is controlled post-translationally by oxidation of Keap1. Assay for Nrf2 activity include:  <ul> <li>Immunohistochemistry for increases in Nrf2 protein levels and translocation into the nucleus Western blot for increased Nrf2 protein levels </li> <li>Western blot of cytoplasmic and nuclear fractions to observe translocation of Nrf2 protein from the cytoplasm to the nucleus qPCR of Nrf2 target genes (e.g., Nqo1, Hmox-1, Gcl, Gst, Prx, TrxR, Srxn), or by commercially available pathway-based qPCR array (e.g., oxidative stress array from SABiosciences) </li> <li>Whole transcriptome profiling by microarray or RNA-seq followed by pathway analysis (in IPA, DAVID, metacore, etc.) for enrichment of the Nrf2 oxidative stress response pathway (e.g., Jackson et al. 2014) </li> <li>OECD TG422D describes an ARE-Nrf2 Luciferase test method </li> </ul> In general, there are a variety of commercially available colorimetric or fluorescent kits for detecting Nrf2 activation. <table border="1"> <tbody> <tr> <td> Assay Type & Measured Content  </td> <td> Description  </td> <td> Dose Range Studied  </td> <td> Assay Characteristics (Length/Ease of use/Accuracy)  </td> </tr> <tr> <td> ROS  Formation in the Mitochondria assay (Shaki et al., 2012)  </td> <td> “The mitochondrial ROS measurement was performed flow cytometry using DCFH-DA. Briefly, isolated kidney mitochondria were incubated with UA (0, 50, 100 and 200 µM) in respiration buffer containing (0.32 mM sucrose, 10mM Tris, 20 mM Mops, 50 µM EGTA, 0.5 mM MgCl2, 0.1 mM KH2PO4 and 5 mM sodium succinate) [32]. In the interval times of 5, 30 and 60 min following the UA addition, a sample was taken and DCFH-DA was added (final concentration, 10 µM) to mitochondria and was then incubated for 10 min.Uranyl acetate-induced ROS generation in isolated kidney mitochondria were determined through the flow cytometry (Partec, Deutschland) equipped with a 488-nm argon ion laser and supplied with the Flomax software and the signals were obtained using a 530-nm bandpass filter (FL-1 channel). Each determination is based on the mean fluorescence intensity of 15,000 counts.”    </td> <td> 0, 50,100 and 200 µM of Uranyl Acetate    </td> <td>  Long/ Easy High accuracy    </td> </tr> <tr> <td> Mitochondrial Antioxidant Content Assay Measuring GSH content (Shaki et al., 2012)    </td> <td> “GSH content was determined using DTNB as the indicator and spectrophotometer method for the isolated mitochondria. The mitochondrial fractions (0.5 mg protein/ml) were incubated with various concentrations of uranyl acetate for 1 h at 30 °C and then 0.1 ml of mitochondrial fractions was added into 0.1 mol/l of phosphate buffers and 0.04% DTNB in a total volume of 3.0 ml (pH 7.4). The developed yellow color was read at 412 nm on a spectrophotometer (UV-1601 PC, Shimadzu, Japan). GSH content was expressed as µg/mg protein.”  </td> <td> 0, 50,  100, or  200 µM  Uranyl Acetate  </td> <td>   </td> </tr> <tr> <td> H2O2 Production Assay Measuring H2O2 Production in isolated mitochondria (Heyno et al., 2008)    </td> <td> “Effect of CdCl2 and antimycin A (AA) on H2O2 production in isolated mitochondria from potato. H2O2 production was measured as scopoletin oxidation. Mitochondria were incubated for 30 min in the measuring buffer  (see the Materials and Methods) containing 0.5 mM succinate as an electron donor and 0.2 µM mesoxalonitrile 3‐chlorophenylhydrazone (CCCP) as an uncoupler, 10 U horseradish peroxidase and 5 µM scopoletin.”   </td> <td> 0, 10, 30  µM Cd2+     2 µM antimycin A  </td> <td>   </td> </tr> <tr> <td> Flow Cytometry ROS & Cell Viability (Kruiderig et al., 1997)    </td> <td> “For determination of ROS, samples taken at the indicated time points were directly transferred to FACScan tubes. Dih123 (10 mM, final concentration) was added and cells were incubated at 37°C in a humidified atmosphere (95% air/5% CO2) for 10 min. At t 5 9, propidium iodide (10 mM, final concentration) was added, and cells were analyzed by flow cytometry at 60 ml/min. Nonfluorescent Dih123 is cleaved by ROS to fluorescent R123 and detected by the FL1 detector as described above for Dc (Van de Water 1995)”“For determination of ROS, samples taken at the indicated time points were directly transferred to FACScan tubes. Dih123 (10 mM, final concentration) was added and cells were incubated at 37°C in a humidified atmosphere (95% air/5% CO2) for 10 min. At t 5 9, propidium iodide (10 mM, final concentration) was added, and cells were analyzed by flow cytometry at 60 ml/min. Nonfluorescent Dih123 is cleaved by ROS to fluorescent R123 and detected by the FL1 detector as described above for Dc (Van de Water 1995)”  </td> <td>   </td> <td>           Strong/easy medium  </td> </tr> <tr> <td> DCFH-DA  Assay Detection of hydrogen peroxide production (Yuan et al.,  2016)  </td> <td> Intracellular ROS production was measured using DCFH-DA as a probe. Hydrogen peroxide oxidizes DCFH to DCF. The probe is hydrolyzed intracellularly to DCFH carboxylate anion. No direct reaction with H2O2 to form fluorescent production.    </td> <td> 0-400  µM  </td> <td> Long/ Easy High accuracy  </td> </tr> <tr> <td> H2-DCF-DAAssay Detection of superoxide production (Thiebault etal., 2007)    </td> <td> This dye is a stable nonpolar compound which diffuses readily into the cells and yields H2-DCF. Intracellular OH or ONOO- react with H2-DCF when cells contain peroxides, to form the highly fluorescent compound DCF, which effluxes the cell. Fluorescence intensity of DCF is measured using a fluorescence spectrophotometer.  </td> <td> 0–600  µM  </td> <td> Long/ Easy High accuracy  </td> </tr> <tr> <td> CM-H2DCFDA  Assay (Eruslanov  & Kusmartsev, 2009)  </td> <td> The dye (CM-H2DCFDA) diffuses into the cell and is cleaved by esterases, the thiol reactive chlormethyl group reacts with intracellular glutathione which can be detected using flow cytometry.  </td> <td>   </td> <td> Long/Easy/ High Accuracy  </td> </tr> </tbody> </table>   <table border="1"> <tbody> <tr> <td> Method of Measurement   </td> <td> References   </td> <td> Description   </td> <td colspan="2"> OECD-Approved Assay  </td> </tr> <tr> <td> Chemiluminescence   </td> <td> (Lu, C. et al., 2006;   Griendling, K. K., et al., 2016)  </td> <td> ROS can induce electron transitions in molecules, leading to electronically excited products. When the electrons transition back to ground state, chemiluminescence is emitted and can be measured. Reagents such as luminol and lucigenin are commonly used to amplify the signal.   </td> <td colspan="2"> No    </td> </tr> <tr> <td> Spectrophotometry   </td> <td> (Griendling, K. K., et al., 2016)  </td> <td> NO has a short half-life. However, if it has been reduced to nitrite (NO2-), stable azocompounds can be formed via the Griess Reaction, and further measured by spectrophotometry.   </td> <td colspan="2"> No  </td> </tr> <tr> <td> Direct or Spin Trapping-Based electron paramagnetic resonance (EPR) Spectroscopy   </td> <td> (Griendling, K. K., et al., 2016)  </td> <td> The unpaired electrons (free radicals) found in ROS can be detected with EPR and is known as electron paramagnetic resonance. A variety of spin traps can be used.   </td> <td colspan="2"> No  </td> </tr> <tr> <td> Nitroblue Tetrazolium Assay   </td> <td> (Griendling, K. K., et al., 2016)  </td> <td> The Nitroblue Tetrazolium assay is used to measure O2.− levels. O2.− reduces nitroblue tetrazolium (a yellow dye) to formazan (a blue dye), and can be measured at 620 nm.   </td> <td colspan="2"> No  </td> </tr> <tr> <td> Fluorescence analysis of dihydroethidium (DHE) or Hydrocyans   </td> <td> (Griendling, K. K., et al., 2016)  </td> <td> Fluorescence analysis of DHE is used to measure O2.− levels.  O2.− is reduced to O2 as DHE is oxidized to 2-hydroxyethidium, and this reaction can be measured by fluorescence. Similarly, hydrocyans can be oxidized by any ROS, and measured via fluorescence.   </td> <td colspan="2"> No  </td> </tr> <tr> <td> Amplex Red Assay   </td> <td> (Griendling, K. K., et al., 2016)  </td> <td> Fluorescence analysis to measure extramitochondrial or extracellular H2O2 levels. In the presence of horseradish peroxidase and H2O2, Amplex Red is oxidized to resorufin, a fluorescent molecule measurable by plate reader.   </td> <td colspan="2"> No  </td> </tr> <tr> <td> Dichlorodihydrofluorescein Diacetate (DCFH-DA)   </td> <td> (Griendling, K. K., et al., 2016)  </td> <td> An indirect fluorescence analysis to measure intracellular H2O2 levels.  H2O2 interacts with peroxidase or heme proteins, which further react with DCFH, oxidizing it to dichlorofluorescein (DCF), a fluorescent product.   </td> <td colspan="2"> No  </td> </tr> <tr> <td> HyPer Probe   </td> <td> (Griendling, K. K., et al., 2016)  </td> <td> Fluorescent measurement of intracellular H2O2 levels. HyPer is a genetically encoded fluorescent sensor that can be used for in vivo and in situ imaging.   </td> <td colspan="2"> No  </td> </tr> <tr> <td> Cytochrome c Reduction Assay   </td> <td> (Griendling, K. K., et al., 2016)  </td> <td> The cytochrome c reduction assay is used to measure O2.− levels. O O2.− is reduced to O2 as ferricytochrome c is oxidized to ferrocytochrome c, and this reaction can be measured by an absorbance increase at 550 nm.   </td> <td colspan="2"> No  </td> </tr> <tr> <td> Proton-electron double-resonance imaging (PEDRI)   </td> <td> (Griendling, K. K., et al., 2016)  </td> <td> The redox state of tissue is detected through nuclear magnetic resonance/magnetic resonance imaging, with the use of a nitroxide spin probe or biradical molecule.   </td> <td colspan="2"> No              </td> </tr> <tr> <td> Glutathione (GSH) depletion   </td> <td> (Biesemann, N. et al., 2018)   </td> <td> A downstream target of the Nrf2 pathway is involved in GSH synthesis. As an indication of oxidation status, GSH can be measured by assaying the ratio of reduced to oxidized glutathione (GSH:GSSG) using a commercially available kit (e.g., <a href="http://www.abcam.com/gshgssg-ratio-detection-assay-kit-fluorometric-green-ab138881.html" rel="noreferrer noopener" target="_blank">http://www.abcam.com/gshgssg-ratio-detection-assay-kit-fluorometric-green-ab138881.html</a>).    </td> <td colspan="2"> No  </td> </tr> <tr> <td> Thiobarbituric acid reactive substances (TBARS)   </td> <td> (Griendling, K. K., et al., 2016)  </td> <td> Oxidative damage to lipids can be measured by assaying for lipid peroxidation with TBARS using a commercially available kit.    </td> <td colspan="2"> No  </td> </tr> <tr> <td> Protein oxidation (carbonylation)  </td> <td> (Azimzadeh et al., 2017; Azimzadeh et al., 2015; Ping et al., 2020)  </td> <td> Can be determined with ELISA or a commercial assay kit. Protein oxidation can indicate the level of oxidative stress.  </td> <td colspan="2"> No  </td> </tr> <tr> <td> Seahorse XFp Analyzer  </td> <td> Leung et al. 2018  </td> <td> The Seahorse XFp Analyzer provides information on mitochondrial function, oxidative stress, and metabolic dysfunction of viable cells by measuring respiration (oxygen consumption rate; OCR) and extracellular pH (extracellular acidification rate; ECAR).  </td> <td> No  </td> </tr> </tbody> </table>   Molecular Biology: Nrf2. Nrf2’s transcriptional activity is controlled post-translationally by oxidation of Keap1. Assays for Nrf2 activity include:   <table border="1"> <tbody> <tr> <td> Method of Measurement   </td> <td> References   </td> <td> Description   </td> <td> OECD-Approved Assay  </td> </tr> <tr> <td> Immunohistochemistry   </td> <td> (Amsen, D., de Visser, K. E., and Town, T., 2009)  </td> <td> Immunohistochemistry for increases in Nrf2 protein levels and translocation into the nucleus    </td> <td> No  </td> </tr> <tr> <td> qPCR   </td> <td> (Forlenza et al., 2012)  </td> <td> qPCR of Nrf2 target genes (e.g., Nqo1, Hmox-1, Gcl, Gst, Prx, TrxR, Srxn), or by commercially available pathway-based qPCR array (e.g., oxidative stress array from SABiosciences)   </td> <td> No  </td> </tr> <tr> <td> Whole transcriptome profiling via microarray or via RNA-seq followed by a pathway analysis  </td> <td> (Jackson, A. F. et al., 2014)  </td> <td> Whole transcriptome profiling by microarray or RNA-seq followed by pathway analysis (in IPA, DAVID, metacore, etc.) for enrichment of the Nrf2 oxidative stress response pathway  </td> <td> No  </td> </tr> </tbody> </table>

Taxonomic applicability: Occurrence of oxidative stress is not species specific.   Life stage applicability: Occurrence of oxidative stress is not life stage specific.  Sex applicability: Occurrence of oxidative stress is not sex specific.  Evidence for perturbation by prototypic stressor: There is evidence of the increase of oxidative stress following perturbation from a variety of stressors including exposure to ionizing radiation and altered gravity (Bai et al., 2020; Ungvari et al., 2013; Zhang et al., 2009).

High Mixed

High

All life stages

High High

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(2013), “Angiotensin II, NADPH oxidase, and redox signaling in the vasculature”, Antioxidants & redox signaling, Vol. 19/10, Mary Ann Liebert, Larchmont, <a href="https://doi.org/10.1089/ars.2012.4641" rel="noreferrer noopener" target="_blank">https://doi.org/10.1089/ars.2012.4641</a>  Ping, Z. et al. (2020), “Oxidative Stress in Radiation-Induced Cardiotoxicity”, Oxidative Medicine and Cellular Longevity, Vol. 2020, Hindawi, <a href="https://doi.org/10.1155/2020/3579143" rel="noreferrer noopener" target="_blank">https://doi.org/10.1155/2020/3579143</a>  Powers, S.K. and M.J. Jackson. (2008), “Exercise-Induced Oxidative Stress: Cellular Mechanisms and Impact on Muscle Force Production”, Physiological Reviews, Vol. 88/4, American Physiological Society, Rockville, <a href="https://doi.org/10.1152/physrev.00031.2007" rel="noreferrer noopener" target="_blank">https://doi.org/10.1152/physrev.00031.2007</a>  Raimondi, V., F. Ciccarese and V. Ciminale. (2020), “Oncogenic pathways and the electron transport chain: a dangeROS liason”, British Journal of Cancer, Vol. 122/2, Nature Portfolio, London, <a href="https://doi.org/10.1038/s41416-019-0651-y" rel="noreferrer noopener" target="_blank">https://doi.org/10.1038/s41416-019-0651-y</a>  Seen, S. and L. Tong. (2018), “Dry eye disease and oxidative stress”, Acta Ophthalmologica, Vol. 96/4, John Wiley & Sons, Inc., Hoboken, <a href="https://doi.org/10.1111/aos.13526" rel="noreferrer noopener" target="_blank">https://doi.org/10.1111/aos.13526</a>  Ungvari, Z. et al. (2013), “Ionizing Radiation Promotes the Acquisition of a Senescence-Associated Secretory Phenotype and Impairs Angiogenic Capacity in Cerebromicrovascular Endothelial Cells: Role of Increased DNA Damage and Decreased DNA Repair Capacity in Microvascular Radiosensitivity”, The Journals of Gerontology Series A: Biological Sciences and Medical Sciences, Vol. 68/12, Oxford University Press, Oxford, <a href="https://doi.org/10.1093/gerona/glt057." rel="noreferrer noopener" target="_blank">https://doi.org/10.1093/gerona/glt057.</a>     Vargas-Mendoza, N. et al. (2021), “Oxidative Stress, Mitochondrial Function and Adaptation to Exercise: New Perspectives in Nutrition”, Life, Vol. 11/11, Multidisciplinary Digital Publishing Institute, Basel, <a href="https://doi.org/10.3390/life11111269" rel="noreferrer noopener" target="_blank">https://doi.org/10.3390/life11111269</a>  Wang, H. et al. (2019), “Radiation-induced heart disease: a review of classification, mechanism and prevention”, International Journal of Biological Sciences, Vol. 15/10, Ivyspring International Publisher, Sydney, <a href="https://doi.org/10.7150/ijbs.35460" rel="noreferrer noopener" target="_blank">https://doi.org/10.7150/ijbs.35460</a>   Zhang, R. et al. (2009), “Blockade of AT1 receptor partially restores vasoreactivity, NOS expression, and superoxide levels in cerebral and carotid arteries of hindlimb unweighting rats”, Journal of applied physiology, Vol. 106/1, American Physiological Society, Rockville, <a href="https://doi.org/10.1152/japplphysiol.01278.2007" rel="noreferrer noopener" target="_blank">https://doi.org/10.1152/japplphysiol.01278.2007</a>.  Zhao, R. Z. et al. (2019), “Mitochondrial electron transport chain, ROS generation and uncoupling”, International journal of molecular medicine, Vol. 44/1, Spandidos Publishing Ltd., Athens, <a href="https://doi.org/10.3892/ijmm.2019.4188" rel="noreferrer noopener" target="_blank">https://doi.org/10.3892/ijmm.2019.4188</a>  AOPWiki 2017-05-30T13:58:17

2026-02-11T07:05:27

Increase, DNA damage

Increase, DNA Damage

Molecular

DNA nucleotide damage, single, and double strand breaks occur in the course of cellular operations such as DNA repair and replication and can be induced directly and in neighboring “bystander” cells by internal or external stressors like reactive oxygen species, chemicals, and radiation. Ionizing radiation and RONS such as hydroxyl radicals or peroxide can create a range of lesions (a change in molecular structure) in the base of the nucleotide, with guanine particularly vulnerable because of its low redox potential (David, O'Shea et al. 2007). The same stressors can also break the sugar (deoxyribose)-phosphate backbone creating a single strand break. Simultaneous proximal breaks in both strands of DNA form double strand breaks, which are considered to be more destructive and mutagenic than lesions or single strand breaks. Double strand breaks can generate chromosomal abnormalities including changes in chromosomal number, breaks and gaps, translocations, inversions, and deletions (Yang, Craise et al. 1992; Haag, Hsu et al. 1996; Ponnaiya, Cornforth et al. 1997; Yang, Georgy et al. 1997; Unger, Wienberg et al. 2010; Behjati, Gundem et al. 2016; Morishita, Muramatsu et al. 2016). However, DNA lesions and single strand breaks can also be destructive and mutagenic. Lesions can lead to point mutations (David, O'Shea et al. 2007) or single strand breaks (Regulus, Duroux et al. 2007). Lesions and single strand breaks can also promote the formation of double strand breaks: replication fork collapse and double strand breaks sometimes occur during mitosis when the replisome encounters an unrepaired single strand break (Kuzminov 2001), and clustered lesions and closely opposed single strand breaks can also form double strand breaks (Chaudhry and Weinfeld 1997; Vispe and Satoh 2000; Shiraishi, Shikazono et al. 2017). Complex damage consists of any combination of closely opposed DNA lesions, abasic sites, crosslinks, single, or double strand breaks in proximity. While classically induced by ionizing radiation, there is also evidence that it can be induced by oxidative activity (Sharma, Collins et al. 2016) or even by a single oxidizing particle (Ravanat, Breton et al. 2014). Complex damage is more difficult to repair (Kuhne, Rothkamm et al. 2000; Stenerlow, Hoglund et al. 2000; Pinto, Prise et al. 2005; Rydberg, Cooper et al. 2005). DNA damage and resulting repair activity can trigger a halt in the cell cycle, cell death (apoptosis), and cause permanent changes to DNA including deletions, translocations, and sequence changes. DNA damage is also associated with an increase in genomic instability - the new appearance of DNA damage including double strand breaks, mutations, and chromosomal damage following repair of initial damage in affected cells or in clonal descendants or neighbors of DNA damaged cells. The mechanism behind this long term DNA damage is not clear, but telomere erosion appears to play a major role (Murnane 2012; Sishc, Nelson et al. 2015). Genomic instability is more common and longer lasting following complex damage (Ponnaiya, Cornforth et al. 1997), and is influenced by multiple factors including variants in DNA repair genes (Ponnaiya, Cornforth et al. 1997; Yu, Okayasu et al. 2001; Yin, Menendez et al. 2012), RONS (Dayal, Martin et al. 2008), estrogen (Kutanzi and Kovalchuk 2013), caspases (Liu, He et al. 2015), and telomeres (Sishc, Nelson et al. 2015).

DNA damage can be studied in isolated DNA, fixed cells, or living cells. Types of damage that can be detected include single and double strand breaks, nucleotide damage, complex damage, and chromosomal or telomere damage. The OECD test guideline for DNA synthesis Test No. 486 (OECD 1997) detects nucleotide excision repair, so it will reflect the formation of bulky DNA adducts but not the majority of oxidative damage to nucleotides, which is typically repaired via the Base Excision Repair pathway. The OECD test guideline alkaline comet assay Test No. 489 (OECD 2016) detects single and double strand breaks, including those arising from repair as well as some (alkali sensitive) nucleotide lesions including some lesions from oxidative damage. OECD tests for chromosomal damage and micronuclei Test No. 473, 475, 483, and 487 measure longer term effects of DNA damage but these tests require the damaged cell to subsequently undergo replication (OECD 2016; OECD 2016; OECD 2016; OECD 2016).  They can therefore reflect a wider range of sources of DNA damage including changes in mitosis. Finally, tests for mutations reveal past DNA damage that resulted in a heritable change, and these are described in the key event ‘Increase in Mutation’. Many other (non-test guideline) techniques have been used to examine specific forms of DNA damage (Table 1). Double strand breaks are commonly reported because of the significant risk attributed to breaks and the relative ease of detecting and quantifying them. Historically, single and double strand breaks were measured using gel electrophoresis, but are now commonly visualized microscopically using fluorescent or other labeled probes for double and single strand break repair such as H2AX and XRCC2.  Base lesions can also be detected using labeled probes for base excision repair enzymes, or by chemical methods such as mass spectroscopy. Refinements on these methods can be used to characterize complex or clustered damage, in which various forms of damage occur in close proximity on a DNA molecule (Lorat, Timm et al. 2016; Nikitaki, Nikolov et al. 2016). Certain challenges are common to all methods of detecting DNA damage. In the time required to initiate the detection method, some DNA may already be repaired, leading to undercounting of damage. On the other hand, apoptotic DSBs may be incorrectly included in a measurement of direct (non-apoptotic) induction of DSB damage unless controlled. All methods have difficulty distinguishing individual components of clustered lesions, and microscopic methods may undercount disparate breaks that are processed together in repair centers (Barnard, Bouffler et al. 2013). Methods that use isolated DNA (gel electrophoresis, analytical chemistry) are vulnerable to artifacts and must ensure that the DNA sample is protected from oxidative damage during extraction (Pernot, Hall et al. 2012; Barnard, Bouffler et al. 2013; Ravanat, Breton et al. 2014). Table 1. Common methods of detecting DNA damage <table border="1" cellpadding="0" cellspacing="0"> <tbody> <tr> <td style="height:22px; width:127px"> Target </td> <td style="height:22px; width:167px"> Name </td> <td style="height:22px; width:133px"> Method </td> <td style="height:22px; width:211px"> Strengths/Weaknesses </td> </tr> <tr> <td style="height:22px; width:127px"> Nucleotide damage </td> <td style="height:22px; width:167px"> Single cell gel electrophoresis (comet assay) with restriction enzymes (Collins 2004) </td> <td style="height:22px; width:133px"> Gel electrophoresis   </td> <td style="height:22px; width:211px"> A variant of the comet assay in which restriction enzymes allow the identification of different types of nucleotide damage. The comet assay is more sensitive than PFGE, detecting damage from 0.1 Gy ionizing radiation (Pernot, Hall et al. 2012). A reproducible high-throughput application of the assay is available (Ge, Prasongtanakij et al. 2014; Sykora, Witt et al. 2018), and the test requires only a small (single cell) sample. Requires destruction of the cell. </td> </tr> <tr> <td style="height:22px; width:127px"> Nucleotide damage </td> <td style="height:22px; width:167px"> Labeled probes including Biotrin OxyDNA and anti- 8-oxoguanine-DNA glycosylase (OGG1) for oxidative damage and AP endonuclease (APE1) for Base Excision Repair of less bulky lesions such as oxidative damage. </td> <td style="height:22px; width:133px"> Microscopy, FACS </td> <td style="height:22px; width:211px"> Most useful with FACS or other measures of average or relative intensity, as locations and numbers of damaged nucleotides can be difficult to distinguish using fluorescence microscopy. (Ogawa, Kobayashi et al. 2003; Nikitaki, Nikolov et al. 2016). </td> </tr> <tr> <td style="height:22px; width:127px"> Nucleotide damage </td> <td style="height:22px; width:167px"> High performance liquid chromatography (HPLC), tandem mass spectrometry (MS/MS) </td> <td style="height:22px; width:133px"> Analytical chemistry </td> <td style="height:22px; width:211px"> Capable of quantifying low levels of specific nucleotide lesions (Madugundu, Cadet et al. 2014; Ravanat, Breton et al. 2014). Requires destruction of the cell. </td> </tr> <tr> <td style="height:22px; width:127px"> Nucleotide damage </td> <td style="height:22px; width:167px"> Unscheduled DNA synthesis test OECD Test Guideline 486 (OECD 1997) </td> <td style="height:22px; width:133px"> Autoradiography </td> <td style="height:22px; width:211px"> Measures DNA damage that is repaired using Nucleotide Excision Repair - mostly bulky adducts (OECD (Organisation for Economic Co-operation and Development) 2016). </td> </tr> <tr> <td style="height:22px; width:127px"> Non-specific DNA strand breaks </td> <td style="height:22px; width:167px"> Single cell gel electrophoresis (comet assay), alkali conditions OECD Test Guideline 489 (OECD 2016) </td> <td style="height:22px; width:133px"> Gel electrophoresis </td> <td style="height:22px; width:211px"> When used in alkali conditions, the comet assay reveals single and double strand breaks and alkali-sensitive nucleotide lesions. See single cell gel electrophoresis (comet assay) with restriction enzymes above for further comments.     </td> </tr> <tr> <td style="height:22px; width:127px"> Single strand breaks </td> <td style="height:22px; width:167px"> Labeled probe pXRCC1 (Lorat, Brunner et al. 2015) </td> <td style="height:22px; width:133px"> Microscopy </td> <td style="height:22px; width:211px"> Fluorescent probes can label single strand breaks in cells, while immunogold labeling is able to distinguish multiple single strand breaks in clusters (Lorat, Timm et al. 2016; Nikitaki, Nikolov et al. 2016). </td> </tr> <tr> <td style="height:22px; width:127px"> Double strand breaks </td> <td style="height:22px; width:167px"> Single cell gel electrophoresis (comet assay), neutral conditions </td> <td style="height:22px; width:133px"> Gel electrophoresis </td> <td style="height:22px; width:211px"> Neutral conditions help minimize the release of single strand breaks coiled DNA and alkali lesions, allowing the measurement of double strand breaks. Since single strand breaks can still appear, assay is not very sensitive or specific to double strand breaks (Pernot, Hall et al. 2012). See single cell gel electrophoresis (comet assay) with restriction enzymes above for further comments. </td> </tr> <tr> <td style="height:22px; width:127px"> Double strand breaks </td> <td style="height:22px; width:167px"> Pulsed field gel electrophoresis (PFGE) </td> <td style="height:22px; width:133px"> Gel electrophoresis </td> <td style="height:22px; width:211px"> Permits the quantitative measurement of double strand breaks, and can be combined with immunoblotting to detect DNA-associated proteins (Lobrich, Rydberg et al. 1995; Kawashima, Yamaguchi et al. 2017). Considered less sensitive than comet assay, but detected damage from 0.25 Gy ionizing radiation (Gradzka and Iwanenko 2005). Requires destruction of the cell. </td> </tr> <tr> <td style="height:22px; width:127px"> Double strand breaks </td> <td style="height:22px; width:167px"> Labeled probes including phosphorylated H2AX, 53BP1, Ku70, ATM (Lorat, Brunner et al. 2015) </td> <td style="height:22px; width:133px"> Microscopy </td> <td style="height:22px; width:211px"> Fluorescent probes can label individual double breaks in cells allowing for quantification, with immunogold labeling resolving breaks in clusters (Lorat, Timm et al. 2016; Nikitaki, Nikolov et al. 2016). Sensitive: detects damage from 0.001 Gy ionizing radiation (Rothkamm and Lobrich 2003; Ojima, Ban et al. 2008). </td> </tr> <tr> <td style="height:22px; width:127px"> Chromosomal damage </td> <td style="height:22px; width:167px"> Chromosomal aberrations and micronuclei OECD Test Guidelines 473, 475, 483, and 487 (OECD 2016; OECD 2016; OECD 2016; OECD 2016) </td> <td style="height:22px; width:133px"> Microscopy </td> <td style="height:22px; width:211px"> Detects major DNA damage resulting from large breaks and rearrangements, or mitotic failures. Damage does not appear until DNA undergoes mitosis, so slower and limited to damage in replicating cells. Insensitive tosmall deletions and substitutions. </td> </tr> </tbody> </table>

CL:0000255

CL eukaryotic cell

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"Novel gene rearrangements in transformed breast cells identified by high-resolution breakpoint analysis of chromosomal aberrations." Endocrine-related cancer 17(1): 87-98.</a> <a name="_ENREF_45">Vispe, S. and M. S. Satoh (2000). "DNA repair patch-mediated double strand DNA break formation in human cells." The Journal of biological chemistry 275(35): 27386-27392.</a> <a name="_ENREF_46">Yang, T.-H., L. M. Craise, et al. (1992). "Chromosomal changes in cultured human epithelial cells transformed by low- and high-LET radiation." Adv Space Res 12(2-3): 127-136.</a> <a name="_ENREF_47">Yang, T. C., K. A. Georgy, et al. (1997). "Initiation of oncogenic transformation in human mammary epithelial cells by charged particles." Radiat Oncol Investig 5(3): 134-138.</a> <a name="_ENREF_48">Yin, Z., D. Menendez, et al. (2012). "RAP80 is critical in maintaining genomic stability and suppressing tumor development." Cancer research 72(19): 5080-5090.</a> <a name="_ENREF_49">Yu, Y., R. Okayasu, et al. (2001). "Elevated breast cancer risk in irradiated BALB/c mice associates with unique functional polymorphism of the Prkdc (DNA-dependent protein kinase catalytic subunit) gene." Cancer Res 61(5): 1820-1824.</a> AOPWiki 2016-11-29T18:41:30

2019-05-08T12:28:46

Increase, Cell cycle arrest

Cell cycle arrest

Cellular

AOPWiki 2026-07-09T04:05:55

2026-07-09T04:05:55

Impact on sirtuins-related aging signaling pathway gene expression

Molecular

AOPWiki 2026-07-09T03:52:59

2026-07-09T03:52:59

Increase, Myocardial and vascular structural remodeling

Myocardial and vascular structural remodeling

Tissue

AOPWiki 2026-07-09T05:00:47

2026-07-09T05:00:47

Increase, Cardiovascular aging

Cardiovascular aging

Individual

AOPWiki 2026-07-09T05:08:16

2026-07-09T05:08:16

Increase, Telomere attrition

Molecular

AOPWiki 2026-07-09T06:05:15

2026-07-09T06:05:15

Increase, Mitochondrial dysfunction

Cellular

Mitochondrial dysfunction is a consequence of inhibition of the respiratory chain leading to oxidative stress. Mitochondria can be found in all cells and are considered the most important cellular consumers of oxygen. Furthermore, mitochondria possess numerous redox enzymes capable of transferring single electrons to oxygen, generating the superoxide (O2-). Some mitochondrial enzymes that are involved in reactive oxygen species (ROS) generation include the electron-transport chain (ETC) complexes I, II and III; pyruvate dehydrogenase (PDH) and glycerol-3-phosphate dehydrogenase (GPDH). The transfer of electrons to oxygen, generating superoxide, happens mainly when these redox carriers are charged enough with electrons and the potential energy for transfer is elevated, like in the case of high mitochondrial membrane potential. In contrast, ROS generation is decreased if there are not enough electrons and the potential energy for the transfer is not sufficient (reviewed in Lin and Beal, 2006). Cells are also able to detoxify the generated ROS due to an extensive antioxidant defence system that includes superoxide dismutases, glutathione peroxidases, catalase, thioredoxins, and peroxiredoxins in various cell organelles (reviewed in Lin and Beal, 2006). It is worth mentioning that, as in the case of ROS generation, antioxidant defences are also closely related to the redox and energetic status of mitochondria. If mitochondria are structurally and functionally healthy, an antioxidant defence mechanism balances ROS generation, and there is not much available ROS production. However, in case of mitochondrial damage, the antioxidant defence capacity drops and ROS generation takes over. Once this happens, a vicious cycle starts and ROS can further damage mitochondria, leading to more free-radical generation and further loss of antioxidant capacity. During mitochondrial dysfunction the availability of ATP also decreases, which is considered necessary for repair mechanisms after ROS generation. A number of proteins bound to the mitochondria or endoplasmic reticulum (ER), especially in the mitochondria-associated ER membrane (MAM), are playing an important role of communicators between these two organelles (reviewed Mei et al., 2013). ER stress induces mitochondrial dysfunction through regulation of Ca2+ signaling and ROS production (reviewed Mei et al., 2013). Prolonged ER stress leads to release of Ca2+ at the MAM and increased Ca2+ uptake into the mitochondrial matrix, which induces Ca2+-dependent mitochondrial outer membrane permeabilization and apoptosis. At the same, ROS are produced by proteins in the ER oxidoreductin 1 (ERO1) family. ER stress activates ERO1 and leads to excessive production of ROS, which, in turn, inactivates SERCA and activates inositol-1,4,5- trisphosphate receptors (IP3R) via oxidation, resulting in elevated levels of cytosolic Ca2+, increased mitochondrial uptake of Ca2+, and ultimately mitochondrial dysfunction. Just as ER stress can lead to mitochondrial dysfunction, mitochondrial dysfunction also induces ER Stress (reviewed Mei et al., 2013). For example, nitric oxide disrupts the mitochondrial respiratory chain and causes changes in mitochondrial Ca2+ flux which induce ER stress. Increased Ca2+ flux triggers loss of mitochondrial membrane potential (MMP), opening of mitochondrial permeability transition pore (mPTP), release of cytochrome c and apoptosis inducing factor (AIF), decreasing ATP synthesis and rendering the cells more vulnerable to both apoptosis and necrosis (Wang and Qin, 2010). Metal-induced Mitochondrial Dysfunction Mitochondria are an important site of Ca2+ regulation and storage, taking up Ca2+ ions electrophoretically from the cytosol through a Ca2+ uniporter, which can then accumulate in the mitochondria (Roos et al., 2012; Orrenius et al., 2015). Similarities between calcium and metals, such as cadmium and lead, makes the entrance and accumulation of these metals into the mitochondria via calcium metals possible by mode of molecular mimicry (Mathews et al., 2013; Adiele et al., 2012). The outer mitochondrial membrane also contains the divalent metal transporter (DMT1), which allows for mitochondrial uptake of divalent metals such as Fe and Mn. When cells are under heavy metal-induced stress, DMT has been shown to be overexpressed in the mitochondrial membrane, making the mitochondria targets of metal toxicity and accumulation. Heavy metal exposure in aerobic organisms increases ROS formation through redox cycling, where metals with different valence states (Fe, Cu, Cr, etc.) directly produce ROS as they are reduced by cellular antioxidants and then react with oxygen (Shaki et al., 2012; Shaki et al., 2013; Pourahmad et al., 2006; Santos et al., 2007). The production of highly reactive hydroxyl radicals under mitochondrial oxidative stress and in the presence of transition metals occurs via the Fenton reaction or Haber-Weiss reaction (Hancock et al., 2001; Valko et al., 2005; Adam-Vizi et al., 2010). Metals and ROS are capable of damaging mitochondrial DNA as well as mechanisms of DNA repair and proliferation arrest (Valko et al., 2005). Metals and ROS have the potential to directly damage mitochondrial membranes and structure by binding to and oxidizing membrane lipids and proteins. This structural damage can collapse the MMP and lead to the opening of the MPTP (Orrenius et al., 2015; Roos et al., 2012; Pourahmad et al., 2006). Uranium and mercury, for example, have both been shown to directly inhibit the mitochondrial electron transport chain and interfere with ATP production (Shaki et al., 2012; Roos et al., 2012). Furthermore, as previously mentioned, metals have been shown to inhibit ROS-detoxifying enzymes. By binding to these enzymes, metals can inhibit their antioxidant functions, and cause an accumulation of ROS and increased synthesis of more antioxidant enzymes in order to combat the oxidative stress (Blajszczak and Bonini, 2017). Summing up: Mitochondria play a pivotal role in cell survival and cell death because they are regulators of both energy metabolism and apoptotic/necrotic pathways (Fiskum, 2000; Wieloch, 2001; Friberg and Wieloch, 2002). The production of ATP via oxidative phosphorylation is a vital mitochondrial function (Kann and Kovács, 2007; Nunnari and Suomalainen, 2012). The ATP is continuously required for signalling processes (e.g. Ca2+ signalling), maintenance of ionic gradients across membranes, and biosynthetic processes (e.g. protein synthesis, heme synthesis or lipid and phospholipid metabolism) (Kang and Pervaiz, 2012), and (Green, 1998; McBride et al., 2006). Inhibition of mitochondrial respiration contributes to various cellular stress responses, such as deregulation of cellular Ca2+ homeostasis (Graier et al., 2007) and ROS production (Nunnari and Suomalainen, 2012; reviewed Mei et al., 2013).). It is well established in the existing literature that mitochondrial dysfunction may result in: (a) an increased ROS production and a decreased ATP level, (b) the loss of mitochondrial protein import and protein biosynthesis, (c) the reduced activities of enzymes of the mitochondrial respiratory chain and the Krebs cycle, (d) the loss of the mitochondrial membrane potential, (e) the loss of mitochondrial motility, causing a failure to re-localize to the sites with increased energy demands (f) the destruction of the mitochondrial network, and (g) increased mitochondrial Ca2+ uptake, causing Ca2+ overload (reviewed in Lin and Beal, 2006; Graier et al., 2007), (h) the rupture of the mitochondrial inner and outer membranes, leading to (i) the release of mitochondrial pro-death factors, including cytochrome c (Cyt. c), apoptosis-inducing factor, or endonuclease G (Braun, 2012; Martin, 2011; Correia et al., 2012; Cozzolino et al., 2013), which eventually leads to apoptotic, necrotic or autophagic cell death (Wang and Qin, 2010). Due to their structural and functional complexity, mitochondria present multiple targets for various compounds.

Mitochondrial dysfunction can be detected using isolated mitochondria, intact cells or cells in culture as well as in vivo studies. Such assessment can be performed with a large range of methods (revised by Brand and Nicholls, 2011) for which some important examples are given. All approaches to assess mitochondrial dysfunction fall into two main categories: the first assesses the consequences of a loss-of-function, i.e. impaired functioning of the respiratory chain and processes linked to it. Some assay to assess this have been described for KE1, with the limitation that they are not specific for complex I. In the context of overall mitochondrial dysfunction, the same assays provide useful information, when performed under slightly different assay conditions (e.g. without addition of complex III and IV inhibitors). The second approach assesses a ‘non-desirable gain-of-function’, i.e. processes that are usually only present to a very small degree in healthy cells, and that are triggered in a cell, in which mitochondria fail. I. Mitochondrial dysfunction assays assessing a loss-of function. 1. Cellular oxygen consumption. See KE1 for details of oxygen consumption assays. The oxygen consumption parameter can be combined with other endpoints to derive more specific information on the efficacy of mitochondrial function. One approach measures the ADP-to-O ratio (the number of ADP molecules phosphorylated per oxygen atom reduced (Hinkle, 1995 and Hafner et al., 1990). The related P/O ratio is calculated from the amount of ADP added, divided by the amount of O2 consumed while phosphorylating the added ADP (Ciapaite et al., 2005; Diepart et al., 2010; Hynes et al., 2006; James et al., 1995; von Heimburg et al., 2005). 2. Mitochondrial membrane potential (Δψm ). - Revision of AOP3 (Project: <a href="https://www.efsa.europa.eu/en/call/npefsaprev202402-development-aop-network-parkinsonian-motor-symptoms" rel="noreferrer noopener" target="_blank">NP/EFSA/PREV/2024/02</a>): The mitochondrial membrane potential (Δψm) is the electric potential difference across the inner mitochondrial membrane. It requires a functioning respiratory chain in the absence of mechanisms that dissipate the proton gradient without coupling it to ATP production. Quantitative assessment of ΔΨm in living cells is most commonly achieved through the use of cationic, lipophilic fluorescent probes that accumulate within the mitochondrial matrix in proportion to the electrochemical gradient (Leonard et al., 2014). Among these, tetramethylrhodamine derivatives such as TMRE (tetramethylrhodamine ethyl ester) and TMRM (tetramethylrhodamine methyl ester) are widely employed due to their reversible, potential-dependent distribution across the inner mitochondrial membrane (Scaduto and Grotyohann, 1999; Creed and McKenzie, 2019). When applied at non-quenching, nanomolar concentrations, these dyes allow linear and quantitative detection of ΔΨm, as fluorescence intensity directly correlates with mitochondrial polarization. Detection can be performed by flow cytometry for population-level quantification, by high-content microscopy for spatially resolved analysis, or by fluorescence plate readers for higher throughput (Wong and Cortopassi, 2002; Valdebenito and Dunchen, 2022). Quantitative interpretation requires the use of appropriate controls, typically involving treatment with protonophores such as FCCP or CCCP, which fully dissipate ΔΨm and thereby establish baseline fluorescence, and inhibitors such as oligomycin or antimycin A to reveal different components of mitochondrial respiration. In parallel, dyes such as JC-1 are also used, though their ratiometric readout is less sensitive at low potentials and more prone to artifacts compared with TMRE or TMRM (Leonard et al., 2022). For accurate normalization, measurements are often corrected for cell number, mitochondrial content, or total protein, and fluorescence changes are expressed relative to maximal depolarization. In addition to chemical probes, genetically encoded sensors, such as mitochondria-targeted fluorescent proteins fused to potential-sensitive domains, provide complementary tools for ΔΨm monitoring in live-cell and in vivo contexts (Leonard et al., 2022). - Not endorsed  3. Enzymatic activity of the electron transport system (ETS). Determination of ETS activity can be dene following Owens and King's assay (1975). The technique is based on a cell-free homogenate that is incubated with NADH to saturate the mitochondrial ETS and an artificial electron acceptor [l - (4 -iodophenyl) -3 - (4 -nitrophenyl) -5-phenylte trazolium chloride (INT)] to register the electron transmission rate. The oxygen consumption rate is calculated from the molar production rate of INT-formazan which is determined spectrophotometrically (Cammen et al., 1990). 4. ATP content. For the evaluation of ATP levels, various commercially-available ATP assay kits are offered  based on luciferin and luciferase activity. For isolated mitochondria various methods are available to continuously measure ATP with electrodes (Laudet 2005), with luminometric methods, or for obtaining more information on different nucleotide phosphate pools (e.g. Ciapaite et al., (2005). <div> - Revision of AOP3 (Project: <a href="https://www.efsa.europa.eu/en/call/npefsaprev202402-development-aop-network-parkinsonian-motor-symptoms">NP/EFSA/PREV/2024/02</a>): Determination of mitochondrial ATP production based on extracellular flux analysis   The method is based on the detection of OCR (Oxygen Consumption Rate) that represents mitochondrial respiration as well as on the detection of ECAR (extracellular acidification rate) / proton efflux rate (PER): reflects extracellular acidification, a proxy for glycolysis (lactate release) plus contributions from CO₂/HCO₃⁻. PER is preferred over raw ECAR since it corrects for CO₂-derived acidification (Desousa et al., 2023; Espinosa et al., 2022). Application of inhibitors of individual complexes of the respiratory chain allows the detection of ATP-linked OCR: portion of oxygen consumption directly driving ATP synthesis (lost after ATP synthase inhibition) (Yoo et al., 2024). The proton leak & non-mitochondrial OCR represents remaining oxygen consumption after ATP synthase and electron transport chain inhibitor addition. The difference yields the ATP-coupled respiration component.   Calculation of mitochondrial ATP production  Mito ATP production rate (pmol ATP/min) = OCRATP (pmol O2/min) × 2 × P/O    OCR_ATP: ATP-coupled portion of OCR.   Factor 2: each O₂ molecule contains two oxygen atoms.   P/O ratio: number of ATP molecules synthesized per oxygen atom reduced. A mean P/O ≈ 2.75 is typically assumed (validated across many cell types but substrate- and condition-dependent) (Plitzko and Loesgen, 2018; Mookerjee et al., 2017; Motawe et al., 2024).    Limitations   P/O ratio varies by substrate (glucose vs. fatty acids), cell type, and conditions. Fixed values are approximations.   Non-mitochondrial oxygen consumption (oxidases, peroxidases, etc.) can confound OCR, hence use of ETC inhibitors.   PER vs. ECAR: CO₂-driven acidification must be corrected to avoid overestimating glycolytic ATP.   Normalization: results are usually expressed per cell, protein content, DNA, or mitochondrial mass — interpretation depends on normalization method.  - Not endorsed </div> II. Mitochondrial dysfunction assays assessing a gain-of function. 1. Mitochondrial permeability transition pore opening (PTP). The opening of the PTP is associated with a permeabilization of mitochondrial membranes, so that different compounds and cellular constituents can change intracellular localization. This can be measured by assessment of the translocation of cytochrome c, adenylate kinase or AIF from mitochondria to the cytosol or nucleus. The translocation can be assessed biochemically in cell fractions, by imaging approaches in fixed cells or tissues or by life-cell imaging of GFP fusion proteins (Single 1998; Modjtahedi 2006). An alternative approach is to measure the accessibility of cobalt to the mitochondrial matrix in a calcein fluorescence quenching assay in live permeabilized cells (Petronilli et al., 1999). 2. mtDNA damage as a biomarker of mitochondrial dysfunction. Various quantitative polymerase chain reaction (QPCR)-based assays have been developed to detect changes of DNA structure and sequence in the mitochondrial genome. mtDNA damage can be detected in blood after low-level rotenone exposure, and the damage persists even after CI activity has returned to normal. With a more sustained rotenone exposure, mtDNA damage is also detected in skeletal muscle. These data support the idea that mtDNA damage in peripheral tissues in the rotenone model may provide a biomarker of past or ongoing mitochondrial toxin exposure (Sanders et al., 2014a and 2014b). 3. Generation of ROS and resultant oxidative stress. a. General approach. Electrons from the mitochondrial ETS may be transferred ‘erroneously’ to molecular oxygen to form superoxide anions. This type of side reaction can be strongly enhanced upon mitochondrial damage. As superoxide may form hydrogen peroxide, hydroxyl radicals or other reactive oxygen species, a large number of direct ROS assays and assays assessing the effects of ROS (indirect ROS assays) are available (Adam-Vizi, 2005; Fan and Li 2014). Direct assays are based on the chemical modification of fluorescent or luminescent reporters by ROS species. Indirect assays assess cellular metabolites, the concentration of which is changed in the presence of ROS (e.g. glutathione, malonaldehyde, isoprostanes,etc.) At the animal level the effects of oxidative stress are measured from biomarkers in the blood or urine. b. Measurement of the cellular glutathione (GSH) status. GSH is regenerated from its oxidized form (GSSH) by the action of an NADPH dependent reductase (GSSH + NADPH + H+ à 2 GSH + NADP+). The ratio of GSH/GSSG is therefore a good indicator for the cellular NADH+/NADPH ratio (i.e. the redox potential). GSH and GSSH levels can be determined by HPLC, capillary electrophoresis, or biochemically with DTNB (Ellman’s reagent). As excess GSSG is rapidly exported from most cells to maintain a constant GSH/GSSG ratio, a reduction of total glutathione (GSH/GSSG) is often a good surrogate measure for oxidative stress. c. Quantification of lipid peroxidation. Measurement of lipid peroxidation has historically relied on the detection of thiobarbituric acid (TBA)-reactive compounds such as malondialdehyde generated from the decomposition of cellular membrane lipid under oxidative stress (Pryor et al., 1976). This method is quite sensitive, but not highly specific. A number of commercial assay kits are available for this assay using absorbance or fluorescence detection technologies. The formation of F2-like prostanoid derivatives of arachidonic acid, termed F2-isoprostanes (IsoP) has been shown to be more specific for lipid peroxidation. A number of commercial ELISA kits have been developed for IsoPs, but interfering agents in samples requires partial purification before analysis. Alternatively, GC/MS may be used, as robust (specific) and sensitive method. d. Detection of superoxide production. Generation of superoxide by inhibition of complex I and the methods for its detection are described by Grivennikova and Vinogradov (2014). A range of different methods is also described by BioTek (<a class="external free" href="http://www.biotek.com/resources/articles/reactive-oxygen-species.html" rel="nofollow" target="_blank">http://www.biotek.com/resources/articles/reactive-oxygen-species.html</a>). The reduction of ferricytochrome c to ferrocytochrome c may be used to assess the rate of superoxide formation (McCord, 1968). Like in other superoxide assays, specificity can only be obtained by measurements in the absence and presence of superoxide dismutase. Chemiluminescent reactions have been used for their increased sensitivity. The most widely used chemiluminescent substrate is lucigenin. Coelenterazine has also been used as a chemiluminescent substrate. Hydrocyanine dyes are fluorogenic sensors for superoxide and hydroxyl radical, and they become membrane impermeable after oxidation (trapping at site of formation). The best characterized of these probes are Hydro-Cy3 and Hydro-Cy5. generation of superoxide in mitochondria can be visualized using fluorescence microscopy with MitoSOX™ Red reagent (Life Technologies). MitoSOX™ Red reagent is a cationic derivative of dihydroethidium that permeates live cells and accumulates in mitochondria. e. Detection of hydrogen peroxide (H2O2) production. There are a number of fluorogenic substrates, which serve as hydrogen donors that have been used in conjunction with horseradish peroxidase (HRP) enzyme to produce intensely fluorescent products in the presence of hydrogen peroxide (Zhou et al., 1997: Ruch et al., 1983). The more commonly used substrates include diacetyldichloro-fluorescein, homovanillic acid, and Amplex® Red. In these examples, increasing amounts of H2O2 form increasing amounts of fluorescent product (Tarpley et al., 2004). Summing up, mitochondrial dysfunction can be measured by: • ROS production: superoxide (O2-), and hydroxyl radicals (OH−) • Nitrosative radical formation such as ONOO− or directly by: • Loss of mitochondrial membrane potential (MMP) • Opening of mitochondrial permeability transition pores (mPTP) • ATP synthesis • Increase in mitochondrial Ca2+ • Cytochrome c release • AIF (apoptosis inducing factor) release from mitochondria • Mitochondrial Complexes enzyme activity • Measurements of mitochondrial oxygen consumption • Ultrastructure of mitochondria using electron microscope and mitochondrial fragmentation measured by labelling with DsRed-Mito expression (Knott et al, 2008) Mitochondrial dysfunction-induced oxidative stress can be measured by: • Reactive carbonyls formations (proteins oxidation) • Increased 8-oxo-dG immunoreactivity (DNA oxidation) • Lipid peroxidation (formation of malondialdehyde (MDA) and 4- hydroxynonenal (HNE) • 3-nitrotyrosine (3-NT) formation, marker of protein nitration • Translocation of Bid and Bax to mitochondria • Measurement of intracellular free calcium concentration ([Ca2+]i): Cells are loaded with 4 μM fura-2/AM). • Ratio between reduced and oxidized form of glutathione (GSH depletion) (Promega assay, TB369; Radkowsky et al., 1986) • Neuronal nitric oxide synthase (nNOS) activation that is Ca2+-dependent. All above measurements can be performed as the assays for each readout are well established in the existing literature (e.g. Bal-Price and Brown, 2000; Bal-Price et al., 2002; Fujikawa, 2015; Walker et al., 1995). See also KE <a href="/wiki/index.php/Event:209" title="Event:209"> Oxidative Stress, Increase</a> <table border="1" cellpadding="1" cellspacing="1"> <tbody> <tr> <td> Assay Type & Measured Content </td> <td>Description</td> <td>Dose Range Studied</td> <td> Assay Characteristics (Length/Ease of use/Accuracy) </td> </tr> <tr> <td> Rhodamine 123 Assay Measuring Mitochondrial membrane potential (MMP) and its collapse  (Shaki et al., 2012) </td> <td> Mitochondrial uptake of cationic fluorescent dye, rhodamine 123, is used for estimation of mitochondrial membrane potential. The fluorescence was monitored using Schimadzou RF-5000U fluorescence spectrophotometer at the excitation and emission wavelength of 490 nm and 535 nm, respectively. </td> <td>50, 100 and 500 μM of uranyl acetate</td> <td> Short / easy Medium accurancy </td> </tr> <tr> <td> TMRE fluorescence Assay Measuring Mitochondrial permeability transition pore (mPTP) opening (Huser et al., 1998) </td> <td>Laser scanning confocal microscopy in combination with the potentiometric fluorescence dye tetramethylrhodamine ethyl ester to monitor relative changes in membrane potential in single isolated cardiac mitochondria. The cationic dye distributes across the membrane in a voltage-dependent manner. Therefore, the large potential gradient across the inner mitochondrial membrane results in the accumulation of the fluorescent dye within the matrix compartment. Rapid depolarizations are caused by the opening of the transition pore.</td> <td>1 µM cyclosporin A</td> <td> Short / easy Low accurancy </td> </tr> <tr> <td> GSH / GSSG Determination Assay Measuring  cellular glutathione (GSH) status; ratio of GSH/GSSG (Owen & Butterfield, 2010; Shaki et al., 2013) </td> <td>GSH and GSSG levels are determinted biochemically with DTNB (Ellman’s reagent). The developed yellow color was read at 412 nm on a spectrophotometer.</td> <td>100 µM uranyl acetate</td> <td> Short / easy Low accurancy </td> </tr> <tr> <td> TBARS Assay Quantification of lipid peroxidation (Yuan et al., 2016) </td> <td>MDA content, a product of lipid peroxidation, was measured using a thiobarbituric acid reactive substances (TBARS) assay. Briefly, the kidney cells were collected in 1 ml PBS buffer solution (pH 7.4) and sonicated. MDA reacts with thiobarbituric acid forming a colored product which can be measured at an absorbance of 532 nm.</td> <td>200, 400, 800 µM uranyl acetate</td> <td> Medium / medium High accurancy </td> </tr> <tr> <td> Aequorin-based bioluminescence assay Increase in mitochondrial Ca2+ influx (Pozzan & Rudolf, 2009) </td> <td>Together with GFP, the aequorin moiety acts as Ca2+ sensor in vivo, which delivers emission energy to the GFP acceptor molecule in a BRET (Bioluminescence Resonance Energy Transfer) process; the Ca2+ can then be visualized with fluorescence microscopy.</td> <td> </td> <td> Short / easy Low accurancy </td> </tr> <tr> <td> Western blot & immunostaining analyses Measuring cytochrome c release (Chen et al., 2000)</td> <td>Examining the redistribution of Cyto c in cytosolic and mitochondrial cellular fractions. Cells are homogenized and centrifuged, then prepared for immunoblots. Cellular fractions were washed in PBS and lysed in 1% NP-40 buffer. Cellular proteins were separated by SDS–PAGE, transferred onto nitrocellulose membranes, probed using immunoblot analyses with antibodies specific to cyto c (6581A for Western and 65971A for immunostaining; Pharmingen)</td> <td> </td> <td> Short / easy Medium accurancy </td> </tr> <tr> <td> Quantikine Rat/Mouse Cytochrome c Immunoassay Measuring cytochrome c release (Shaki et al., 2012) </td> <td>Cytochrome C release was measured a monoclonal antibody specific for rat/mouse cytochrome c was precoated onto the microplate. Seventy-five microliter of conjugate (containing mono- clonal antibody specific for cytochrome c conjugated to horseradish peroxidase). After 2 h of incubation, the substrate solution (100 μl) was added to each well and incubated for 30 min. After 100 μl of the stop solution was added to each well; the optical density of each well was determined by the aforementioned microplate spectrophotometer set to 450 nm.</td> <td> </td> <td> Short / easy Low accurancy </td> </tr> <tr> <td> Membrane potential and cell viability – Flow Cytometry Measuring cytochrome c release (Kruidering et al., 1997) </td> <td>“Dc and viability were determined by analyzing the R123 and propidium iodide fluorescence intensity with a FACScan flow cytometer (Becton Dickinson, San Jose, CA) equipped with an argon laser, with the Lysis software program (Becton Dickinson). R123 is a cationic dye that accumulates in the negatively charged inner side of the mitochondria. When the potential drops, less R123 accumulates in the mitochondria, which results in a lower fluorescence signal. The potential was measured as follows: at the indicated times, a 500-ml sample of the cell suspension was taken and transferred to an Eppendorf minivial. To this sample, 100 ml of 6 mM R123 in buffer D was added. After incubation for 10 min at 37°C, the cell suspension was centrifuged for 5 min at 80 3 g. The cell pellet was resuspended in 200 ml of buffer D, containing 0.2 mM R123 and 10 mM propidium iodide, to prevent loss of R123 and to stain nonviable cells, respectively. The samples were transferred to FACScan tubes and analyzed immediately. Analysis was performed at a flow rate of 60 ml/min. R123 fluorescence was detected by the FL1 detector with an emission detection limit below 560 nm. Propidium iodide fluorescence was detected by the FL3 detector, with emission detection above 620 nm. Per sample 3,000 to 5,000 cells were counted (Van de Water et al., 1993)”</td> <td> </td> <td> Short / easy Medium accurancy </td> </tr> </tbody> </table>

Mitochondrial dysfunction is a universal event occurring in cells of any species (Farooqui and Farooqui, 2012). Many invertebrate species (drosophila, C, elegans) are considered as potential models to study mitochondrial function. New data on marine invertebrates, such as molluscs and crustaceans and non-Drosophila species, are emerging (Martinez-Cruz et al., 2012). Mitochondrial dysfunction can be measured in animal models used for toxicity testing (Winklhofer and Haass, 2010; Waerzeggers et al., 2010) as well as in humans (Winklhofer and Haass, 2010). - Revision of AOP3 (Project: <a href="https://www.efsa.europa.eu/en/call/npefsaprev202402-development-aop-network-parkinsonian-motor-symptoms" rel="noreferrer noopener" target="_blank">NP/EFSA/PREV/2024/02</a>): Endogenous ROS formation by complex I: In mammals, complex I is a dominant site of mitochondrial ROS, especially via RET. In plants (Senkler et al. 2017; Maldonado), mitochondria contain alternative NAD(P)H dehydrogenases and an alternative oxidase (AOX) that bypass Complex I and III These pathways reduce ROS formation by preventing over-reduction of the ETC. Complex I still produces ROS, but generally less damaging due to AOX. Yeast: S. cerevisiae lacks a canonical Complex I entirely, relying instead on alternative NADH dehydrogenases. Consequently, mitochondrial ROS production from a Complex I-like source is absent. Other fungi with true Complex I (e.g., Neurospora crassa) do generate ROS similar to animals. - Not endorsed

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2026-02-11T07:06:25

Increase, Inflammation

Cellular

Inflammation is complex to define.  Villeneuve et al. (2018) analyzed the varied biological responses, provided guidance to simplify the  process representing inflammation in adverse outcome pathways, and recommended 3 key steps: 1. Tissue resident cell activation 2. Increased Pro-inflammatory mediators 3. Leukocyte recruitment/activation.  Tissue resident cell activation generally occurs when healthy tissue is exposed to a stressor, or when damage occurs, initiating a signal response of pro-inflammatory mediators (ex. cytokines).  Pro-inflammatory mediators result in the production of lipids and proteins, signaling, and initiate leukocyte recruitment/activation.  Leukocyte recruitment/activation initiate inflammation and other morphological changes.  In cancer, inflammation is a cascade of events created by the host in response to the spread of the cancer (Coussens and Werb, 2002). In response to an injury or the presence of cancer, the host heals itself through inflammation. Indeed, the activation and the migration of  leukocytes (neutrophils, monocytes and eosinophils) to the wound induces the healing process. These inflammatory cells provide an extracellular matrix that forms upon which fibroblast and endothelial cells proliferate and migrate in order to recreate a normal environment. Damage to the epithelial layer initiate inflammatory reactions (Palmer et al. 2011).  In cancer, this inflammatory state induces cell proliferation, increases the production of reactive oxygen species leading to oxidative DNA damage, and reduces DNA repair (Coussens and Werb, 2002).  For review of inflammation caused by microplastics in mammals, see Wright and Kelly (2017).     Inflammation can be defined as the response of the organism to a tissue injury (Coussens). Indeed, in order to heal this injury, a multitude of chemical signals initiate and maintain a host response. Leukocytes (neutrophils, monocytes and eosinophils) are recruited to the site of the damage through the attraction by chemokines (TNF-α (tumour necrosis factor-α), interleukines…). A provisional extracellular matrix (ECM) is created, and fibroblast and endothelial cells proliferate and migrate to it. Wound healing is an example of physiological inflammation and is self-limiting (Coussens). In case of a dysregulation, inflammation can lead to pathologies. Inflammation can be caused by physical injury, ischemic injury, infection, exposure to toxins, or other types of trauma (Singh). Inflammation was described as one of the hallmarks of cancer by Hannahan et al. as a response to tumor invasion through mainly two mechanisms: promoting genetic instatbility and supply pro-tumorogenic factors. First, inflammation in cancer promotes genetic instability (Mantovani, colotta). Macrophages, in contact with the inflammatory site can be responsible of a reactive stress oxygen reaction (ROS) (Maeda, Pollard, Grivennikov). Indeed, they generate high levels of reactive oxygen and nitrogen species which produce mutagenic agents (peroxynitrite), which in turn causes DNA mutations. Second, in inflammation, the tumor micro environment plays a critical role (Coussens). Indeed, in can supply growth factors, survival factors, proangiogenic factors, extracellular matrix-modifying enzymes that facilitate angiogenesis, invasion, and metastasis, and inductive signals that lead to activation of EMT and other hallmark-facilitating programs (Hannahan). For example, macrophages can become tumor associated macrophage which promote cell proliferation, angiogenesis, and invasion (Singh, Lin, Qian). Moreover, chronic inflammation can also lead to tumorigenesis (Karin, Singh). Indeed, since 1863, Virchow has hypothesized that chronic inflammation causes cell proliferation (Balkwill). According to Aggarwall, several pro-inflammatory markers such as TNF and members of its superfamily, IL-1alpha, IL-1beta, IL-6, IL-8, IL-18, chemokines, MMP-9, VEGF, COX-2, and 5-LOX mediate suppression of apoptosis, proliferation, angiogenesis, invasion, and metastasis (Aggarwal).

Inflammation is generally detected in histopathological examination of organs (ex. liver, intestines) or in changes in gene expression (ex. interleukins).  Activation of the innate immune response and the release of various inflammatory cytokines can also be assessed (Flake and Morgan, 2017).   Several assays can be used to measure inflammation: <ul> <li style="text-align:justify">Histopathology on samples. Several scoring tools exist (Goeboes)</li> <li style="text-align:justify">Measuring chemokines in the blood (ELISA, multiplex bead assays : interleukines (IL1, IL6), TNF, interferon… ) (Brenner) and histopathology samples</li> <li style="text-align:justify">Measuring Prostaglandin levels, COX-2 (ELISA Liquid chromatography/tandem mass spectrometry, IHC)</li> <li style="text-align:justify">Transcription factors : STAT3 Activation, NF-κB Activation (ELISA RtPCR to measure mRNA</li> <li style="text-align:justify">Biomarkers (white cell count, CRP) ratios, and predictive score using </li> <li style="text-align:justify">Measuring ROS(DCFDA, horseradish peroxidase (HRP)-oxidizing substrates, SOD-inhibitable reduction of cytochrome c) (Murphy).</li> </ul>   Methods are extensively reviewed in Marchand et al and Murphy et al.

Taxonomic: appears to be present broadly, with representative studies focused on mammals (humans, lab mice, lab rats).   Extensive data exists on the presence of inflammation in human (Coussens, Aggarwal, Hannhan, Mantovani..) In human, many examples of chronic inflammation leading to cancer or cancer progression exist. For instance, Helicobacter pylori infection leads to gut cancer (Wang).

CL:0000255

CL eukaryotic cell

High Unspecific

High

All life stages

High High High

Flake, G.P., and Morgan, D.L. 2017. Pathology of diacetyl and 2,3-pentanedione airway lesions in a rat model of obliterative bronchiolitis. Toxicology, 388, 40–47. <a href="https://doi.org/10.1016/j.tox.2016.10.013">https://doi.org/10.1016/j.tox.2016.10.013</a> Palmer, S.M., Flake, G.P., Kelly, F.L., Zhang, H.L., Nugent, J.L., Kirby, P.J., Zhang, H.L., Nugent, J.L., Kirby, P.J., Foley, J.F., Gwinn, W.M., and Morgan, D.L. 2011. Severe airway epithelial injury, aberrant repair and Bronchiolitis obliterans develops after diacetyl instillation in rats. PLoS ONE, 6(3). <a href="https://doi.org/10.1371/journal.pone.0017644">https://doi.org/10.1371/journal.pone.0017644</a>   Wang F, Meng W, Wang B, Qiao L. Helicobacter pylori-induced gastric inflammation and gastric cancer. Cancer Lett. 2014 Apr 10;345(2):196-202. doi: 10.1016/j.canlet.2013.08.016. Epub 2013 Aug 24. PMID: 23981572. Naylor MS, Stamp GW, Foulkes WD, Eccles D, Balkwill FR. Tumor necrosis factor and its receptors in human ovarian cancer. Potential role in disease progression. J Clin Invest. 1993;91:2194–206. Coussens L.M. and Werb Z. Inflammation and cancer. Nature. 2002 Dec 19-26;420(6917):860-7. doi: 10.1038/nature01322. PMID: 12490959; PMCID: PMC2803035. Wright, S.L. and Kelly, F.J.  2017.  Plastic and human health: a micro issue?  Enviromental Science and Technology 51: 6634-6647. Villeneuve, D.L., Landesmann, B., Allavena, P., Ashley, N., Bal-Price, A., Corsini, E., Halappanavar, S., Hussell, T., Laskin, D., Lawrence, T., Nikolic-Paterson, D., Pallary, M., Paini, A., Pietrs, R., Roth, R., and Tschudi-Monnet, F.  2018.  Toxicological Sciences 163(2): 346-352.     Qian BZ, Pollard JW. Macrophage diversity enhances tumor progression and metastasis. Cell. 2010 Apr 2;141(1):39-51. doi: 10.1016/j.cell.2010.03.014. PMID: 20371344; PMCID: PMC4994190. Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell. 2011 Mar 4;144(5):646-74. doi: 10.1016/j.cell.2011.02.013. PMID: 21376230. Karin M. Nuclear factor-kappaB in cancer development and progression. Nature. 2006;441:431–6. Aggarwal BB, Shishodia S, Sandur SK, Pandey MK, Sethi G. Inflammation and cancer: How hot is the link? Biochem Pharmacol. 2006;72:1605–21 Singh N, Baby D, Rajguru JP, Patil PB, Thakkannavar SS, Pujari VB. Inflammation and cancer. Ann Afr Med. 2019 Jul-Sep;18(3):121-126. doi: 10.4103/aam.aam_56_18. PMID: 31417011; PMCID: PMC6704802. Coussens LM, Werb Z. Inflammation and cancer. Nature. 2002;420:860–7 Balkwill F, Mantovani A. Inflammation and cancer: back to Virchow? Lancet. 2001;357:539–545 Mantovani A, Allavena P, Sica A, Balkwill F. Cancer-related inflammation. Nature. 2008;454:436–44 Cancer-related inflammation, the seventh hallmark of cancer: Links to genetic instability.  Maeda H, Akaike T. Nitric oxide and oxygen radicals in infection, inflammation, and cancer. Biochemistry (Mosc) 1998;63:854–65. Pollard JW. Tumour-educated macrophages promote tumour progression and metastasis. Nat Rev Cancer. 2004;4:71–8 Lin, Y., Xu, J. & Lan, H. Tumor-associated macrophages in tumor metastasis: biological roles and clinical therapeutic applications. J Hematol Oncol 12, 76 (2019). <a href="https://doi.org/10.1186/s13045-019-0760-3" style="color:#467886; text-decoration:underline">https://doi.org/10.1186/s13045-019-0760-3</a> Grivennikov SI, Greten FR, Karin M. Immunity, inflammation, and cancer. Cell. 2010 Mar 19;140(6):883-99. doi: 10.1016/j.cell.2010.01.025. PMID: 20303878; PMCID: PMC2866629.   Murphy, M.P., Bayir, H., Belousov, V. et al. Guidelines for measuring reactive oxygen species and oxidative damage in cells and in vivo. Nat Metab 4, 651–662 (2022). <a href="https://doi.org/10.1038/s42255-022-00591-z" style="color:#467886; text-decoration:underline">https://doi.org/10.1038/s42255-022-00591-z</a> Geboes K, Riddell R, Öst A, et al A reproducible grading scale for histological assessment of inflammation in ulcerative colitis Gut 2000;47:404-409. Brenner DR, Scherer D, Muir K, Schildkraut J, Boffetta P, Spitz MR, Le Marchand L, Chan AT, Goode EL, Ulrich CM, Hung RJ. A review of the application of inflammatory biomarkers in epidemiologic cancer research. Cancer Epidemiol Biomarkers Prev. 2014 Sep;23(9):1729-51. doi: 10.1158/1055-9965.EPI-14-0064. Epub 2014 Jun 24. PMID: 24962838; PMCID: PMC4155060. AOPWiki 2016-11-29T18:41:23

2024-02-28T06:33:44

Increase, Blood pressure

Blood pressure

Organ

AOPWiki 2026-07-24T11:18:54

2026-07-24T11:18:54

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AOPWiki 2026-07-09T05:09:32

2026-07-09T05:09:32

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AOPWiki 2026-07-09T07:58:21

2026-07-09T07:58:21

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AOPWiki 2026-07-09T05:11:42

2026-07-09T05:11:42

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AOPWiki 2026-03-29T04:17:19

2026-03-29T04:17:19

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AOPWiki 2026-07-09T05:12:44

2026-07-09T05:12:44

<upstream-id>e6703558-f1f9-499c-9152-2ba6be915f16</upstream-id> <downstream-id>e0382ff7-0130-4477-b512-1ccc3dba1b64</downstream-id>

AOPWiki 2026-07-09T08:11:15

2026-07-09T08:11:15

<upstream-id>d673dad6-0afc-4538-bc91-b211114c8bf7</upstream-id> <downstream-id>53dfa5d8-c50e-4214-85d2-7a52ea34eb54</downstream-id> Inflammation is one consequence of oxidative stress.   Inflammation can be characterized as a multi-step process (Villeneuve et al. 2018): 1. Activation of tissue cells due to stress; 2. Increases in proinflammatory mediator (ex. cytokines); 3. Leukocyte recruitment; 4. Inflammatory response.

This KER was identified as part of an Environmental Protection Agency effort to represent putative AOPs from peer-reviewed literature which were heretofore unrepresented in the AOP-Wiki.  Support for this KER is referenced in publications cited in the originating work of Jeong and Choi (2020).

The biological plausibility linking inflammation to oxidative stress is strong.  Oxidative stress triggers cellular signals, mediated by proinflammatory mediators such as cytokines, which initiates inflammation pathways.  At the cellular level, there are increases in leukocyte recruitment; at the tissue and organ levels, visible inflammation occurs.

Biological, physical, and chemical stressors from environmental sources can increase oxidative stress. Inflammation is one of the most common responses to oxidative stress (for review see Wright and Kelly (2017); Villeneuve et al. (2018); for empirical studies see Gamo et al. (2008); Lu et al. (2016); Jin et al. (2018); Lei et al. (2018)).   Stress triggers increased gene response of proinflammatory signaling mediators (ex. cytokines, interleukins, interferons).  Increased leukocyte response results in inflammation. <table class="MsoTableGrid" style="border-collapse:collapse !msorm; border:none !msorm"> <tbody> <tr> <td style="background-color:#d9d9d9; border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:102px"> Species </td> <td style="background-color:#d9d9d9; border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:71px"> Duration </td> <td style="background-color:#d9d9d9; border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:114px"> Dose </td> <td style="background-color:#d9d9d9; border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:72px"> Increased Oxidative Stress? </td> <td style="background-color:#d9d9d9; border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:72px"> Increased Inflammation? </td> <td style="background-color:#d9d9d9; border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:112px"> Summary </td> <td style="background-color:#d9d9d9; border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:81px"> Citation </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:102px"> Lab mice (Mus musculus) </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:71px"> 56 days </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:114px"> NA </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> yes </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> yes </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:112px"> Seven-week old male mice with surgical brain nerve injury showed changes in inflammatory gene expression (increased interleukin-1beta and interleukin 6), with G-protein coupled receptors mitigating the oxidative stress responses. </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:81px"> Gamo et al. (2018) </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:102px"> Zebrafish (Danio rerio) </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:71px"> 7 days </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:114px"> Aquatic exposure of 20, 200, 2000 μg/L of 70 nm and 5 um polystyrene microplastics. </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> yes </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> yes </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:112px"> Adult 5-month old fish had increased oxidative stress enzyme levels of superoxide dismutase and catalase and liver inflammation. </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:81px"> Lu et al. (2016) </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:102px"> Zebrafish (Danio rerio) </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:71px"> 14 days </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:114px"> Aquatic exposure of 100, 1000 ug/L of 0.5 and 50 um diameter polystyrene microplastic. </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> yes </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> yes </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:112px"> Adult 6-month old male fish increased oxidative stress as measured by statistically significant changes to gut microbiota and changes to inflammatory gene expression, with statistically significant increases of interleukin-1alpha, interleukin-1beta, interferon, interleukin-6. </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:81px"> Jin et al. (2018) </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:102px"> Zebrafish (Danio rerio) </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:71px"> 10 days </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:114px"> Environmental exposure of 1.0 mg/mL of microplastic particles (polyamides (PA), polyethylene (PE), polypropylene (PP), polyvinyl chloride (PVC), and 0.1, 1.0, 5.0 um size polystyrene (PS)). </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> yes </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> yes </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:112px"> Adult fish showed increased oxidative stress in intestinal damage, and increased intestinal inflammation for all but polystyrene (PS) exposure. </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:81px"> Lei et al. (2018) </td> </tr> </tbody> </table>

High Unspecific

High

All life stages

High High High

Life Stage: The life stage applicable to this key event relationship is all life stages.  Sex: This key event relationship applies to both males and females. Taxonomic: This key event relationship appears to be present broadly, with representative studies including mammals (humans, lab mice, lab rats) and teleost fish.

Gamo, K., Kiryu-Seo, S., Konishi, H., Aoki, S., Matushima, K., Wada, K., and Kiyama, H.  2008.  G-protein-coupled receptor screen reveals a role for chemokine recepteor CCR5 in suppressing microglial neurotoxicity.  Journal of Neuroscience 28: 11980-11988. Jin, Y., Xia, J., Pan, Z., Yang, J., Wang, W., and Fu, Z.  2018.  Polystyrene microplastics induce microbiota dysbiosis and inflammation in the gut of adult zebrafish.  Environmental Pollution 235: 322-329. Lei, L., Wu, S., Lu, S., Liu, M., Song, Y., Fu, Z., Shi, H., Raley-Susman, K.M., and He, D.  2018.  Microplastic particles cause intestinal damage and other adverse effects in zebrafish Danio rerio and nematode Caenorhabditis elegans.  Science of the Total Environment 619-620: 1-8. Lu, Y., Zhang, Y., Dengy, Y., Jiang, W., Zhao, Y., Geng, J., Ding, L., Ren, H.  2016.  Uptake and accumulation of polystyrene microplastics in zebrafish (Danio rerio) and toxic effects in liver.  Environmental Science and Technology 50: 4054-4060. Villeneuve, D.L., Landesmann, B., Allavena, P., Ashley, N., Bal-Price, A., Corsini, E., Halappanavar, S., Hussell, T., Laskin, D., Lawrence, T., Nikolic-Paterson, D., Pallary, M., Paini, A., Pietrs, R., Roth, R., and Tschudi-Monnet, F.  2018.  Toxicological Sciences 346:352. Wright, S.L. and Kelly, F.J.  2017.  Plastic and human health: a micro issue?  Enviromental Science and Technology 51: 6634-6647. AOPWiki 2023-07-31T09:35:04

2023-10-19T09:39:43

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AOPWiki 2026-07-09T08:15:02

2026-07-09T08:15:02

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AOPWiki 2026-07-09T08:19:33

2026-07-09T08:19:33

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AOPWiki 2026-07-24T11:20:36

2026-07-24T11:20:36

Binding and activation of AhR lead to cardiovascular aging

Shiheng Gui

Jian Liu, Ruifang Fan, Shiheng Gui

BY-SA

2.8

This AOP aims to elucidate the key biological pathways through which polycyclic aromatic hydrocarbons (PAHs) activate the aryl hydrocarbon receptor (AhR) and promote cardiovascular aging. AhR is a ligand-dependent transcription factor that plays a central role in regulating the expression of xenobiotic metabolism and detoxification-related enzymes (such as the CYP family); simultaneously, AhR is expressed in various tissue types including vascular endothelium, and its activation is closely associated with endothelial dysfunction, inflammatory responses, and vascular remodeling processes. As exogenous pollutants, PAHs may cause multi-system damage after entering the organism. In this AOP, the binding and activation of PAHs to AhR is defined as the Molecular Initiating Event (MIE). Subsequently, AhR signaling can directly or indirectly inhibit the expression or activity of SIRT1/SIRT6, serving as Key Event 1 (KE1), and induce DNA damage responses, further leading to upregulation of p16 and p21, accompanied by aging-related changes including enhanced systemic inflammation and oxidative stress, telomere attrition, and mitochondrial dysfunction. The persistent cellular damage described above ultimately drives structural and functional remodeling of the myocardium and vasculature, manifested as increased vascular calcium deposition, elevated blood pressure, and consequently the formation of cardiovascular aging as the Adverse Outcome (AO). We have identified multiple key events in this pathway and delineated the logical relationships between key events; based on this, we have constructed this AOP to describe the molecular mechanisms by which AhR binding and activation lead to cardiovascular aging. Polycyclic aromatic hydrocarbons (PAHs) refer to hydrocarbon compounds containing two or more benzene rings in their molecular structure, primarily originating from incomplete combustion of organic matter(Zhang, 2017) . PAHs are widely distributed in environmental water, air, soil, and food. Dietary intake, respiration, and dermal contact are the main routes of PAH exposure, characterized by persistence, passivity, and inevitability, increasing cancer risk and causing high incidence of cardiovascular diseases. Numerous population epidemiological studies have shown that PAHs exposure are closely associated with the development of cardiovascular diseases such as hypertension, atherosclerosis, and ischemic heart disease(Curfs et al., 2004; Xu et al., 2021) . Given their widespread environmental distribution and potential toxicity, the United States Environmental Protection Agency (EPA) has listed 16 of them as priority controlled toxic organic pollutants. Therefore, it is necessary to assess the health risks of PAHs based on the AOP (Adverse Outcome Pathway) framework.

adjacent

High

Moderate adjacent

High

High adjacent

High

High adjacent

High

High adjacent

High

High adjacent

High

High adjacent

High

High adjacent

High

High adjacent

Moderate

Moderate adjacent

High

High Male

High

During brain development, adulthood and aging

Low High

As the DNA damage effects of PAHs are widely recognized, previous research has primarily focused on the carcinogenic, teratogenic, and mutagenic effects induced by PAH-mediated DNA damage. However, DNA damage is also an important driving factor in accelerating cellular senescence, and cellular senescence is an important pathway for suppressing cellular malignant transformation; therefore, this Adverse Outcome Pathway (AOP) focuses on the association between PAH exposure and cardiovascular aging. However, current in vivo and in vitro evidence investigating PAH exposure-induced cardiovascular toxic effects remains relatively limited. Moreover, existing in vivo or in vitro studies typically focus on single or a few PAHs, and the exposure doses set are high, making it difficult to reflect the adverse effects of mixed PAHs at environmental doses on the cardiovascular system in the general population. Therefore, low-dose mixed animal exposure, cell experiments, and computer simulation studies are needed to investigate the adverse effects of mixed exposure to 16 priority-controlled PAHs on the cardiovascular system from the perspective of aging. This AOP takes AhR activation as the initiating event; PAHs can directly activate AhR(Das et al., 2016), then directly or indirectly inhibit the expression of SIRT1/SIRT6 in the cardiovascular system, and induce DNA damage responses, further leading to upregulation of p16 and p21, accompanied by aging-related changes including enhanced systemic inflammation and oxidative stress, telomere attrition, and mitochondrial dysfunction (Bin et al., 2010; Malik and Czajka, 2013) . Experimental results can be obtained from various models, including experimental animals, mice, and cell lines. Among these, in vivo animal experiments, in vitro experiments, and computer simulation evidence can confirm the associations between the Molecular Initiating Event (MIE) and Key Event Relationships (KERs). In HUVEC cells, studies have found that exposure to low doses (10-100 µM) of naphthalene, fluoranthene, and fluorene can cause Ca²⁺ ion influx, promote eNOS activation, and further lead to increased NO synthesis(Li et al., 2004) . This suggests that the negative regulatory effect between PAH exposure levels and blood pressure may be closely related to PAH-promoted NO synthesis in endothelial cells. Similarly, in HUVEC cells, exposure to environmental doses of mixed 16 priority-controlled PAHs caused inflammatory responses and oxidative stress, and significantly decreased migration and tube formation capabilities(He et al., 2022). In rat models, after B[a]P exposure, significant increases in systolic blood pressure, diastolic blood pressure, and mean arterial pressure were observed in rats, with significantly decreased maximal acetylcholine-stimulated aortic relaxation responses(Gan et al., 2012) . In mouse models, B[a]P exposure caused systemic inflammation in apolipoprotein-deficient (ApoE⁻/⁻) mice and led to atherosclerosis development; exposure to 16 priority-controlled PAHs promoted atherosclerotic plaque formation in ApoE⁻/⁻ mice by upregulating miR-155 and inhibiting SERPIND1 expression (He et al., 2021) . Additionally, in zebrafish models, exposure to Pyr, B[a]P, and B[k]P all caused cardiac developmental abnormalities (Zhang et al., 2021). Meanwhile, combining epidemiological surveys and experimental research results, PAH exposure has significant effects on cardiovascular aging, particularly on blood pressure, atherosclerosis formation, cardiac development, and heart rate. Existing studies have found that urinary 1-OHPhe levels in the hypertension group (0.152 µg/g) were significantly higher than those in the non-hypertension group (0.128 µg/g) (Lee et al., 2020) . Another study found that for every 1 µg/mmol increase in urinary 4-OHPhe or total -OHPAHs content, the 10-year risk of atherosclerotic cardiovascular disease (ASCVD) increased by 12.63% or 11.91%, respectively (p < 0.05) (Yin et al., 2017) . Pregnancy and infancy are critical time windows for cardiac development, relatively sensitive to pollutant exposure. Maternal occupational PAH exposure during pregnancy increases the risk of congenital heart defects (CHD) in offspring, and among occupationally PAH-exposed populations, PAH metabolite levels are associated with decreased heart rate, with a dose-response relationship (Deng et al., 2022; Li et al., 2012).

MIE1: AhR activation PAHs (such as benzo[a]pyrene) enter cells as ligands and bind to AhR in the cytoplasm, triggering dissociation of AhR from the molecular chaperone complex (HSP90, XAP2, p23), nuclear translocation, and heterodimer formation with ARNT, thereby binding to XRE sequences to initiate target gene transcription. AhR activation not only initiates Phase I metabolic reactions but also lays the foundation for oxidative stress and inflammatory responses. KE1: Oxidative stress Oxidative stress refers to the disruption of redox balance in vivo or within cells, resulting in massive production of ROS (including superoxide anion, hydrogen peroxide, and hydroxyl radicals), while simultaneously consuming intracellular antioxidant substances (GSH, SOD, CAT), exceeding the scavenging capacity of the antioxidant defense system. KE2: Impact on sirtuins-related aging signaling pathway gene expression Among the sirtuins family, SIRT1 and SIRT6 are known as "longevity genes" and are NAD⁺-dependent deacetylases that play important roles in maintaining mitochondrial function and integrity, DNA damage repair, telomere maintenance, and cardiovascular homeostasis. KE3: DNA damage ROS and PAH active metabolites (such as BPDE) directly attack DNA, forming DNA adducts and leading to double-strand breaks or base mutations. KE4: Cell cycle arrest Both p16INK4a and p21Cip1 are cell cycle arrest proteins. When DNA damage occurs within cells, cell cycle checkpoints are activated, cell cycle arrest protein expression increases, causing cell cycle stagnation. KE5: Telomere attrition Telomeres serve as protective structures at chromosome ends; their attrition (length shortening or structural dysfunction) is caused by ROS-mediated oxidative damage and end replication problems during DNA replication. KE6: Mitochondrial dysfunction With increasing age, various adverse factors accumulate within mitochondria, such as mtDNA mutations and changes in mitochondrial dynamics, which can cause mitochondrial dysfunction, increased ROS generation, and further increase mitochondrial membrane permeability, thereby triggering inflammation and cell death. When mtDNAcn is abnormal, mitochondria are facing stress, which is one manifestation of mitochondrial dysfunction. KE7: Systemic inflammation An immune process occurring within cardiovascular tissues characterized by immune cell infiltration, inflammatory factor release, tissue damage, and repair responses. KE8: Myocardial and vascular structural remodeling Myocardial and vascular structural remodeling includes ventricular hypertrophy, thinning and loose structure of ventricular walls, focal vascular proliferation, and disordered arrangement of vascular smooth muscle, providing structural basis for subsequent blood pressure elevation and vascular calcification. KE9: Blood pressure elevation Vascular aging leads to increased vascular stiffness, disordered secretion of vasoactive substances, decreased vascular diastolic regulation capacity, and increased susceptibility to hypertension. Blood pressure elevation is manifested as sustained elevation of arterial systolic or diastolic pressure; hypertension accelerates vascular remodeling and calcification.

<table align="center" cellspacing="0" class="MsoTableGrid" style="border-collapse:collapse; border:none"> <tbody> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:151px"> Essentiality of KE </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:72px"> Definitional Question </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:118px"> High (Strong) </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:106px"> Moderate </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:1px solid black; vertical-align:top; width:106px"> Low (Weak) </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px">   </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> If the upstream KE is blocked, will the downstream KE and/or AO be prevented? </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:118px"> Direct evidence from specifically designed experimental studies indicating that at least one important KE is essential </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:106px"> Indirect evidence suggesting that sufficient modification of the expected modulating factor would weaken or enhance the KE </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:106px"> No or contradictory experimental evidence proving the essentiality of any KE </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px"> KE1: Oxidative stress </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> Moderate </td> <td colspan="3" style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:330px"> Extracellular signaling molecules bind and activate AhR, inducing CYP1A1/1B1 expression, metabolizing PAHs, and producing large amounts of ROS. </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px"> KE2: Impact on sirtuins-related aging signaling pathway gene expression </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> High </td> <td colspan="3" style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:330px"> The AhR/ARNT complex binds to XREs (xenobiotic response elements) in the SIRT1/SIRT6 promoter regions, inhibiting transcription. </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px"> KE3: DNA damage </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> High </td> <td colspan="3" style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:330px"> SIRT1/SIRT6 participate in DNA repair; reduced expression of both leads to decreased repair efficiency and accumulation of DNA damage. PAHs can directly and competitively bind to DNA strand binding sites in SIRT6, limiting its ability to bind damaged DNA strands, causing decreased DNA damage response capability of SIRT6, and exacerbating DNA damage in the cardiovascular system. </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px"> KE4: Cell cycle arrest </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> High </td> <td colspan="3" style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:330px"> DNA damage accumulation activates cell cycle checkpoints, cell cycle arrest proteins p16/p21 expression increases, causing stable cell cycle arrest. </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px"> KE5: Telomere attrition </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> High </td> <td colspan="3" style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:330px"> DNA damage can cause telomere dysfunction, telomere binding protein TRF2 expression is inhibited, and p53 expression is activated, thereby promoting cellular senescence or apoptosis. Telomerase activity is inhibited in senescent cells. The p53/p21 and p16/pRB pathways may synergistically limit telomerase expression. ROS continuously attacks telomere DNA. </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px"> KE6: Mitochondrial dysfunction </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> High </td> <td colspan="3" style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:330px"> DNA damage can cause mitochondrial dysfunction by inhibiting PGC-1α through p53, reducing mitochondrial biogenesis; increased ROS generation further increases mitochondrial membrane permeability, potentially causing abnormal mtDNAcn. </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px"> KE7: Systemic inflammation </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> Moderate </td> <td colspan="3" style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:330px"> Chronic inflammation caused by oxidative stress activates pathways such as NF-κB and releases pro-inflammatory cytokines such as TNF-α, causing cardiovascular system aging. </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px"> KE8: Myocardial and vascular structural remodeling </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> High </td> <td colspan="3" style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:330px"> Decreased SIRT1/SIRT6 expression causes cardiovascular aging and myocardial and vascular remodeling. Inflammatory factors promote myocardial and vascular structural remodeling. Environmental doses of PAHs cause ventricular hypertrophy in rats, while high concentrations of PAH exposure cause thinning of ventricular walls and structural dilation. High concentration PAH exposure causes focal arterial proliferation, disordered smooth muscle arrangement, and increased arterial calcium deposition, causing vascular structural changes. </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px"> KE9: Blood pressure elevation </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> High </td> <td colspan="3" style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:330px"> Decreased vascular compliance leads to increased peripheral resistance. Increased arterial stiffness leads to elevated systolic pressure and increased pulse pressure difference. </td> </tr> <tr> <td style="border-bottom:1px solid black; border-left:1px solid black; border-right:1px solid black; border-top:none; vertical-align:top; width:151px"> AO: Cardiovascular aging </td> <td style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:72px"> High </td> <td colspan="3" style="border-bottom:1px solid black; border-left:none; border-right:1px solid black; border-top:none; vertical-align:top; width:330px"> Cardiovascular aging caused by hypertension is usually associated with cardiovascular structural and functional abnormalities; sustained hypertension accelerates increased cardiac afterload, myocardial hypertrophy, and deterioration of vascular lesions. </td> </tr> </tbody> </table>

<div> <table class="table table-bordered table-fullwidth"> <thead> <tr> <th>Modulating Factor (MF)</th> <th>Influence or Outcome</th> <th>KER(s) involved</th> </tr> </thead> <tbody> <tr> <td> </td> <td> </td> <td> </td> </tr> </tbody> </table> </div>

Not Specified

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