, Volume 22, Issue 2, pp 345–351 | Cite as

Absolute requirement for iron in the development of chemically induced uroporphyria in mice treated with 3-methylcholanthrene and 5-aminolevulinate

  • Kenzi Nakano
  • Mayumi Ishizuka
  • Kentaro Q. Sakamoto
  • Shoichi Fujita


Accumulating evidence, including experiments using cytochrome P450 1a2 (Cyp1a2) gene knock-out mice (Cyp1a2(−/−)), indicates that the development of chemically induced porphyria requires the expression of CYP1A2. It has also been demonstrated that iron enhances and expedites the development of experimental uroporphyria, but that iron alone without CYP1A2 expression, as in Cyp1a2(−/−) mice, does not cause uroporphyria. The role of iron in the development of porphyria has not been elucidated. We examined the in vivo effect of iron deficiency on hepatic URO accumulation in experimental porphyria. Mice were fed diets containing low (iron-deficient diet (IDD), 8.5 mg iron/kg) or normal (normal diet (ND), 213.7 mg iron/kg) levels of iron. They were treated with 3-methylcholanthrene (MC), an archetypal inducer of CYP1A, and 5-aminolevulinate (ALA), precursors of porphyrin and heme. We found that uroporphyrin (URO) levels and uroporphyrinogen oxidation (UROX) activity were markedly increased in ND mice treated with MC and ALA, while the levels were not raised in IDD mice with the same treatments. CYP1A2 levels and methoxyresorufin O-demethylase (MROD) activities, the CYP1A2-mediated reaction, were markedly induced in the livers of both ND and IDD mice treated with MC and ALA. UROX activity, supposedly a CYP1A2-dependent activity, was not enhanced in iron-deficient mice in spite of the fact of induction of CYP1A2. We showed that a sufficient level of iron is essential for the development of porphyria and UROX activity.


Porphyria Uroporphyrin CYP1A2 Uroporphyrinogen oxidation Iron 


  1. Becker FT (1965) Porphyria cutanea tarda induced by estrogens. Arch Dermatol 92:252–255. doi: 10.1001/archderm.92.3.252 CrossRefGoogle Scholar
  2. Bonkovsky HL, Wood SG, Howell SK, Sinclair PR, Lincoln B, Healey JF, Sinclair JF (1986) High-performance liquid chromatographic separation and quantitation of tetrapyrroles from biological materials. Anal Biochem 155:56–64. doi: 10.1016/0003-2697(86)90224-1 PubMedCrossRefGoogle Scholar
  3. Burke MD, Thompson S, Elcombe CR, Halpert J, Haaparanta T, Mayer RT (1985) Ethoxy-, pentoxy- and benzyloxyphenoxazones and homologues: a series of substrates to distinguish between different induced cytochromes P-450. Biochem Pharmacol 34:3337–3345. doi: 10.1016/0006-2952(85)90355-7 PubMedCrossRefGoogle Scholar
  4. Chaufan G, Rios de Molina MC, de Viale LCSM (2001) How does hexachlorobenzene treatment affect liver uroporphyrinogen decarboxylase? Int J Biochem Cell Biol 33:621–630. doi: 10.1016/S1357-2725(01)00034-6 PubMedCrossRefGoogle Scholar
  5. Clark MA, Bing BA, Gottschall PE, Williams JF (1995) Differential effect of cytokines on the phenobarbital or 3-methylcholanthrene induction of P450 mediated monooxygenase activity in cultured rat hepatocytes. Biochem Pharmacol 49:97–104. doi: 10.1016/0006-2952(94)00438-R PubMedCrossRefGoogle Scholar
  6. Deam S, Elder GH (1991) Uroporphyria produced in mice by iron and 5-aminolevulinic acid. Biochem Pharmacol 41(12):2019–2022PubMedCrossRefGoogle Scholar
  7. Enriquez de Salamanca R, Mingo D, Chinarro S, Munoz JJ, Perpina J (1982) Patterns of porphyrin-excretion in female estrogen-induced porphyria cutanea tarda. Arch Dermatol Res 274:179–184. doi: 10.1007/BF00510371 PubMedCrossRefGoogle Scholar
  8. Epstein JH, Redeker AG (1965) Porphyria cutanea tarda symptomatica (PCT-S). A study of the effect of phlebotomy therapy. Arch Dermatol 92:286–289. doi: 10.1001/archderm.92.3.286 CrossRefGoogle Scholar
  9. Francis JE, Smith AG (1984) Assay of mouse liver uroporphyrinogen decarboxylase by reverse-phase high-performance liquid chromatography. Anal Biochem 138:404–410. doi: 10.1016/0003-2697(84)90829-7 PubMedCrossRefGoogle Scholar
  10. Gorman N, Walton HS, Bement WJ, Honsinger CP, Szakacs JG, Sinclair JF, Sinclair PR (1999) Role of small differences in CYP1A2 in the development of uroporphyria produced by iron and 5-aminolevulinate in C57BL/6 and SWR strains of mice. Biochem Pharmacol 58:375–382. doi: 10.1016/S0006-2952(99)00088-X PubMedCrossRefGoogle Scholar
  11. Gorman N, Ross KL, Walton HS, Bement WJ, Szakacs JG, Gerhard GS, Dalton TP, Nebert DW, Eisenstein RS, Sinclair JF, Sinclair PR (2002) Uroporphyria in mice: thresholds for hepatic CYP1A2 and iron. Hepatology 35:912–921. doi: 10.1053/jhep.2002.32487 PubMedCrossRefGoogle Scholar
  12. Jarrell JF, Gocmen A, Akyol D, Brant R (2002) Hexachlorobenzene exposure and the proportion of male births in Turkey 1935–1990. Reprod Toxicol 16:65–70. doi: 10.1016/S0890-6238(01)00196-4 PubMedCrossRefGoogle Scholar
  13. Jones KG, Sweeney GD (1977) Association between induction of aryl hydrocarbon hydroxylase and depression of uroporphyrinogen decarboxylase activity. Res Commun Chem Pathol Pharmacol 17:631–637PubMedGoogle Scholar
  14. Lambrecht RW, Jacobs JM, Sinclair PR, Sinclair JF (1990) Inhibition of uroporphyrinogen decarboxylase activity. The role of cytochrome P-450-mediated uroporphyrinogen oxidation. Biochem J 269:437–441PubMedGoogle Scholar
  15. Lambrecht RW, Sinclair PR, Gorman N, Sinclair JF (1992) Uroporphyrinogen oxidation catalyzed by reconstituted cytochrome P450IA2. Arch Biochem Biophys 294:504–510. doi: 10.1016/0003-9861(92)90717-B PubMedCrossRefGoogle Scholar
  16. Lowry OH, Rosebrough HJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275PubMedGoogle Scholar
  17. Nonaka S, Ohgami T, Murayama F, Yamashita K, Yoshida H (1986) Five cases of porphyria cutanea tarda with mild cutaneous changes: evaluation of the efficacy of phlebotomy by the pattern analysis of urinary porphyrins. J Dermatol 13:196–202PubMedGoogle Scholar
  18. Omura T, Sato R (1964) The carbon monooxide-binding pigment of liver microsomes. J Biol Chem 239:370–2378Google Scholar
  19. Phillips JD, Jackson LK, Bunting M, Franklin MR, Thomas KR, Levy JE, Andrews NC, Kushner JP (2001) A mouse model of familial porphyria cutanea tarda. Proc Natl Acad Sci USA 98:259–264. doi: 10.1073/pnas.011481398 PubMedCrossRefGoogle Scholar
  20. Schoenfeld N, Mamet R, Leibovici L, Lanir A (1996) Alcohol-induced changes in urinary aminolevulinic acid and porphyrins: unrelated to liver disease. Alcohol 13:59–63. doi: 10.1016/0741-8329(95)02013-6 PubMedCrossRefGoogle Scholar
  21. Sinclair PR, Gorman N, Dalton T, Walton HS, Bement WJ, Sinclair JF, Smith AG, Nebert DW (1998) Uroporphyria produced in mice by iron and 5-aminolevulinic acid does not occur in Cyp1a2(−/−) null mutant mice. Biochem J 330:149–153PubMedGoogle Scholar
  22. Sinclair PR, Gorman N, Sinclair JF, Walton HS, Bement WJ, Lambrecht RW (1995) Ascorbic acid inhibits chemically induced uroporphyria in ascorbate-requiring rats. Hepatology 22:565–572PubMedGoogle Scholar
  23. Smith AG, Francis JE (1983) Synergism of iron and hexachlorobenzene inhibits hepatic uroporphyrinogen decarboxylase in inbred mice. Biochem J 214:909–913PubMedGoogle Scholar
  24. Smith AG, Francis JE (1993) Genetic variation of iron-induced uroporphyria in mice. Biochem J 291:29–35PubMedGoogle Scholar
  25. Smith AG, Cabral JR, Carthew P, Francis JE, Manson MM (1989) Carcinogenicity of iron in conjunction with a chlorinated environmental chemical, hexachlorobenzene, in C57BL/10ScSn mice. Int J Cancer 43:492–496. doi: 10.1002/ijc.2910430325 PubMedCrossRefGoogle Scholar
  26. Smith AG, Clothier B, Carthew P, Childs NL, Sinclair PR, Nebert DW, Dalton TP (2001) Protection of the Cyp1a2(−/−) null mouse against uroporphyria and hepatic injury following exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin. Toxicol Appl Pharmacol 173:89–98. doi: 10.1006/taap.2001.9167 PubMedCrossRefGoogle Scholar
  27. Smith AG, Clothier B, Robinson S, Scullion MJ, Carthew P, Edwards R, Luo J, Lim CK, Toledano M (1998) Interaction between iron metabolism and 2,3,7,8-tetrachlorodibenzo-p-dioxin in mice with variants of the Ahr gene: a hepatic oxidative mechanism. Mol Pharmacol 53:52–61PubMedGoogle Scholar
  28. Smith AG, Francis JE (1987) Chemically-induced formation of an inhibitor of hepatic uroporphyrinogen decarboxylase in inbred mice with iron overload. Biochem J 246:221–226PubMedGoogle Scholar
  29. Smith AG, Francis JE, Green JA, Greig JB, Wolf CR, Manson MM (1990) Sex-linked hepatic uroporphyria and the induction of cytochromes P450IA in rats caused by hexachlorobenzene and polyhalogenated biphenyls. Biochem Pharmacol 40:2059–2068. doi: 10.1016/0006-2952(90)90236-E PubMedCrossRefGoogle Scholar
  30. Sweeney GD, Jones KG, Cole FM, Basford D, Krestynski F (1979) Iron deficiency prevents liver toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin. Science 204:332–335. doi: 10.1126/science.432648 CrossRefGoogle Scholar
  31. Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci USA 76:4350–4354. doi: 10.1073/pnas.76.9.4350 PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC. 2008

Authors and Affiliations

  • Kenzi Nakano
    • 1
  • Mayumi Ishizuka
    • 1
  • Kentaro Q. Sakamoto
    • 1
  • Shoichi Fujita
    • 1
  1. 1.Laboratory of Toxicology, Department of Environmental Veterinary Sciences, Graduate School of Veterinary MedicineHokkaido UniversityKita-ku, SapporoJapan

Personalised recommendations