Springer Nature is making Coronavirus research free. View research | View latest news | Sign up for updates

9,10-Phenanthrenequinone promotes secretion of pulmonary aldo-keto reductases with surfactant


9,10-Phenanthrenequinone (9,10-PQ), a major quinone in diesel exhaust particles, induces apoptosis via the generation of reactive oxygen species (ROS) because of 9,10-PQ redox cycling. We have found that intratracheal infusion of 9,10-PQ facilitates the secretion of surfactant into rat alveolus. In the cultured rat lung, treatment with 9,10-PQ results in an increase in a lower-density surfactant by ROS generation through redox cycling of the quinone. The surfactant contains aldo-keto reductase (AKR) 1C15, which reduces 9,10-PQ and the enzyme level in the surfactant increases on treatment with 9,10-PQ suggesting an involvement of AKR1C15 in the redox cycling of the quinone. In six human cell types (A549, MKN45, Caco2, Hela, Molt4 and U937) only type II epithelial A549 cells secrete three human AKR1C subfamily members (AKR1C1, AKR1C2 and AKR1C3) with the surfactant into the medium; this secretion is highly increased by 9,10-PQ treatment. Using in vitro enzyme inhibition analysis, we have identified AKR1C3 as the most abundantly secreted AKR1C member. The AKR1C enzymes in the medium efficiently reduce 9,10-PQ and initiate its redox cycling accompanied by ROS production. The exposure of A549 cells to 9,10-PQ provokes viability loss, which is significantly protected by the addition of the AKR1C3 inhibitor and antioxidant enzyme and by the removal of the surfactants from the culture medium. Thus, the AKR1C enzymes secreted in pulmonary surfactants probably participate in the toxic mechanism triggered by 9,10-PQ.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6



Aldo-keto reductase


Bronchoalveolar lavage fluid


Copper, zinc-superoxide dismutase


Diesel exhaust particles


Dulbecco’s modified Eagle medium




Dulbecco’s phosphate-buffered saline


Fetal bovine serum


Polyethylene glycol-conjugated catalase




Reactive oxygen species


Surfactant protein






Reduced nicotinamide-adenine dinucleotide


Reverse transcription-polymerase chain reaction


  1. Andersson S, Kheiter A, Merritt TA (1999) Oxidative inactivation of surfactants. Lung 177:179–189

  2. Anseth JW, Goffin AJ, Fuller GG, Ghio AJ, Kao PN, Upadhyay D (2005) Lung surfactant gelation induced by epithelial cells exposed to air pollution or oxidative stress. Am J Respir Cell Mol Biol 33:161–168

  3. Barski OA, Tipparaju SM, Bhatnagar A (2008) The aldo-keto reductase superfamily and its role in drug metabolism and detoxification. Drug Metab Rev 40:553–624

  4. Bérubé KA, Jones TP, Williamson BJ (1997) Electron microscopy of urban airborn particulate matter. Microsc Anal 49:9–11

  5. Boyland E, Sims P (1962) Metabolism of polycyclic compounds. 21. The metabolism of phenanthrene in rabbits and rats: dihydrodihydroxy compounds and related glucosiduronic acids. Biochem J 84:571–582

  6. Chesis PL, Levin DE, Smith MT, Ernster L, Ames BN (1984)Mutagenicity of quinones: pathways of metabolic activation and detoxification.Proc Natl Acad Sci USA 81:1696–1700

  7. Courter LA, Pereira C, Baird WM (2007) Diesel exhaust influences carcinogenic PAH-induced genotoxicity and gene expression in human breast epithelial cells in culture. Mutat Res 625:72–82

  8. Crowther JE, Kutala VK, Kuppusamy P, Ferguson JS, Beharka AA, Zweier JL, McCormack FX, Schlesinger LS (2004) Pulmonary surfactant protein A inhibits macrophage reactive oxygen intermediate production in response to stimuli by reducing NADPH oxidase activity. J Immunol 172:6866–6874

  9. Delfino RJ (2002) Epidemiologic evidence for asthma and exposure to air toxics: linkages between occupational, indoor, and community air pollution research. Environ Health Perspect 110:573–589

  10. Deyashiki Y, Ogasawara A, Nakayama T, Nakanishi M, Miyabe Y, Sato K, Hara A (1994) Molecular cloning of two human liver 3 alpha-hydroxysteroid/dihydrodiol dehydrogenase isoenzymes that are identical with chlordecone reductase and bile-acid binder. Biochem J 299:545–552

  11. El-Kabbani O, Scammells PJ, Day T, Dhagat U, Endo S, Matsunaga T, Soda M, Hara A (2010) Structure-based optimization and biological evaluation of human 20α-hydroxysteroid dehydrogenase (AKR1C1) salicylic acid-based inhibitors. Eur J Med Chem 45:5309–5317

  12. Endo S, Matsunaga T, Horie K, Tajima K, Bunai Y, Carbone V, El-Kabbani O, Hara A (2007) Enzymatic characteristics of an aldo-keto reductase family protein (AKR1C15) and its localization in rat tissues. Arch Biochem Biophys 465:136–147

  13. Endo S, Matsunaga T, Ohta C, Soda M, Kanamori A, Kitade Y, Ohno S, Tajima K, El-Kabbani O, Hara A (2011) Roles of rat and human aldo-keto reductases in metabolism of farnesol and geranylgeraniol. Chem Biol Interact 191:261–268

  14. Eskelson CD, Chvapil M, Strom KA, Vostal JJ (1987) Pulmonary phospholipidosis in rats respiring air containing diesel particulates. Environ Res 44:260–271

  15. Figueroa JD, Malats N, García-Closas M, Real FX, Silverman D, Kogevinas M, Chanock S, Welch R, Dosemeci M, Lan Q, Tardón A, Serra C, Carrato A, García-Closas R, Castaño-Vinyals G, Rothman N (2008) Bladder cancer risk and genetic variation in AKR1C3 and other metabolizing genes. Carcinogenesis 29:1955–1962

  16. Gilliard N, Heldt GP, Loredo J, Gasser H, Redl H, Merritt TA, Spragg RG (1994) Exposure of the hydrophobic components of porcine lung surfactant to oxidant stress alters surface tension properties. J Clin Invest 93:2608–2615

  17. Haddad IY, Nieves-Cruz B, Matalon S (1997) Inhibition of surfactant function by copper-zinc superoxide dismutase (CuZn-SOD). J Appl Physiol 83:1545–1550

  18. Harada T, Koyama I, Shimoi A, Alpers DH, Komoda T (2002) Identification of pulmonary surfactant that bears intestinal-type and tissue-nonspecific-type alkaline phosphatase in endotoxin-induced rat bronchoalveolar fluid. Cell Tissue Res 307:69–77

  19. Higaki Y, Usami N, Shintani S, Ishikura S, El-Kabbani O, Hara A (2003) Selective and potent inhibitors of human 20alpha-hydroxysteroid dehydrogenase (AKR1C1) that metabolizes neurosteroids derived from progesterone. Chem Biol Interact 143–144:503–513

  20. Hirakawa H, Sawada H, Yamahama Y, Takikawa SI, Shintaku H, Hara A, Mase K, Kondo T, Iino T (2009) Expression analysis of the aldo-keto reductases involved in the novel biosynthetic pathway of tetrahydrobiopterin in human and mouse tissues. J Biochem 146:51–60

  21. Hiyoshi K, Takano H, Inoue KI, Ichinose T, Yanagisawa R, Tomura S, Kumagai Y (2005) Effects of phenanthraquinone on allergic airway inflammation in mice. Clin Exp Allergy 35:1243–1248

  22. Huang LJ, Chen SX, Huang Y, Luo WJ, Jiang HH, Hu QH, Zhang PF, Yi H (2006) Proteomics-based identification of secreted protein dihydrodiol dehydrogenase as a novel serum markers of non-small cell lung cancer. Lung Cancer 54:87–94

  23. Ichinose T, Yajima Y, Nagashima M, Takenoshita S, Nagamachi Y, Sagai M (1997) Lung carcinogenesis and formation of 8-hydroxy-deoxyguanosine in mice by diesel exhaust particles. Carcinogenesis 18:185–192

  24. Jarabak J (1991)Polycyclic aromatic hydrocarbon quinone-mediated oxidation reduction cycling catalyzed by a human placental NADPH-linked carbonyl reductase. Arch Biochem Biophys291:334–338

  25. Jain D, Atochina-Vasserman EN, Tomer Y, Kadire H, Beers MF (2008) Surfactant protein D protects against acute hyperoxic lung injury. Am J Respir Crit Care Med 178:805–813

  26. Jin Y, Stayrook SE, Albert RH, Palackal NT, Penning TM, Lewis M (2001) Crystal structure of human type III 3alpha-hydroxysteroid dehydrogenase/bile acid binding protein complexed with NADP(+) and ursodeoxycholate. Biochemistry 40:10161–10168

  27. Juvin P, Fournier T, Grandsaigne M, Desmonts JM, Aubier M (2002) Diesel particles increase phosphatidylcholine release through a NO pathway in alveolar type II cells. Am J Physiol Lung Cell Mol Physiol 282:L1075–L1081

  28. Kumagai Y, Arimoto T, Shinyashiki M, Shimojo N, Nakai Y, Yoshikawa T, Sagai M (1997) Generation of reactive oxygen species during interaction of diesel exhaust particle components with NADPH-cytochrome P450 reductase and involvement of the bioactivation in the DNA damage. Free Radic Biol Med 22:479–487

  29. Luo DX, Huang MC, Ma J, Gao Z, Liao DF, Cao D (2011) Aldo-keto reductase family 1, member B10 is secreted through a lysosome-mediated non-classical pathway. Biochem J 438:71–80

  30. Lusuardi M, Capelli A, Carli S, Tacconi MT, Salmona M, Donner CF (1992) Role of surfactant in chronic obstructive pulmonary disease: therapeutic implications. Respiration 59 (Suppl 1):28–32

  31. Matsunaga T, Kotamraju S, Kalivendi SV, Dhanasekaran A, Joseph J, Kalyanaraman B (2004) Ceramide-induced intracellular oxidant formation, iron signaling, and apoptosis in endothelial cells: protective role of endogenous nitric oxide. J Biol Chem 279:28614–28624

  32. Matsunaga T, Shintani S, Hara A (2006) Multiplicity of mammalian reductases for xenobiotic carbonyl compounds. Drug Metab Pharmacokinet 21:1–18

  33. Matsunaga T, Kamiya T, Sumi D, Kumagai Y, Kalyanaraman B, Hara A (2008) L-Xylulose reductase is involved in 9,10-phenanthrenequinone-induced apoptosis in human T lymphoma cells. Free Radic Biol Med 44:1191–1202

  34. Matsunaga T, Arakaki M, Kamiya T, Endo S, El-Kabbani O, Hara A (2009) Involvement of an aldo-keto reductase (AKR1C3) in redox cycling of 9,10-phenanthrenequinone leading to apoptosis in human endothelial cells. Chem Biol Interact 181:52–60

  35. Matsunaga T, Shinoda Y, Inoue Y, Shimizu Y, Haga M, Endo S, El-Kabbani O, Hara A (2011) Aldo-keto reductase 1 C15 as a quinone reductase in rat endothelial cell: its involvement in redox cycling of 9,10-phenanthrenequinone. Free Radic Res 45:848–857

  36. Matsuura K, Shiraishi H, Hara A, Sato K, Deyashiki Y, Ninomiya M, Sakai S (1998) Identification of a principal mRNA species for human 3alpha-hydroxysteroid dehydrogenase isoform (AKR1C3) that exhibits high prostaglandin D2 11-ketoreductase activity. J Biochem 124:940–946

  37. McCord JM, Fridovich I (1969) Superoxide dismutase. An enzymic function for erythrocuprein (hemocuprein). J Biol Chem 244:6049–6055

  38. Nagaraj NS, Beckers S, Mensah JK, Waigel S, Vigneswaran N, Zacharias W (2006) Cigarette smoke condensate induces cytochromes P450 and aldo-keto reductases in oral cancer cells. Toxicol Lett 165:182–194

  39. Nkadi PO, Merritt TA, Pillers DA (2009) An overview of pulmonary surfactant in the neonate: genetics, metabolism, and the role of surfactant in health and disease. Mol Genet Metab 97:95–101

  40. Oginuma M, Shimada H, Imamura Y (2005) Involvement of carbonyl reductase in superoxide formation through redox cycling of adrenochrome and 9,10-phenanthrenequinone in pig heart.Chem Biol Interact 155:148–154

  41. Polosa R, Salvi S, Di Maria GU (2002) Allergic susceptibility associated with diesel exhaust particle exposure: clear as mud. Arch Environ Health 57:188–193

  42. Quinn AM, Penning TM (2008) Comparisons of (+/-)-benzo[a]pyrene-trans-7,8-dihydrodiol activation by human cytochrome P450 and aldo-keto reductase enzymes: effect of redox state and expression levels. Chem Res Toxicol 21:1086–1094

  43. Rüstow B, Haupt R, Stevens PA, Kunze D (1993) Type II pneumocytes secrete vitamin E together with surfactant lipids. Am J Physiol 265:L133–L139

  44. Sagai M, Furuyama A, Ichinose T (1996) Biological effects of diesel exhaust particles (DEP). III. Pathogenesis of asthma like symptoms in mice. Free Radic Biol Med 21:199–209

  45. Schuetzle D (1983) Sampling of vehicle emissions for chemical analysis and biological testing. Environ Health Perspect 47:65–80

  46. Schwela D (2000) Air pollution and health in urban areas. Rev Environ Health 15:13–42

  47. Taguchi K, Shimada M, Fujii S, Sumi D, Pan X, Yamano S, Nishiyama T, Hiratsuka A, Yamamoto M, Cho AK, Froines JR, Kumagai Y (2008) Redox cycling of 9,10-phenanthraquinone to cause oxidative stress is terminated through its monoglucuronide conjugation in human pulmonary epithelial A549 cells. Free Radic Biol Med 44:1645–1655

  48. Takeda K, Miyahara N, Rha YH, Taube C, Yang ES, Joetham A, Kodama T, Balhorn AM, Dakhama A, Duez C, Evans AJ, Voelker DR, Gelfand EW (2003) Surfactant protein D regulates airway function and allergic inflammation through modulation of macrophage function. Am J Respir Crit Care Med 168:783–789

  49. Terrasa AM, Guajardo MH, Armas SE de, Catalá A (2005) Pulmonary surfactant protein A inhibits the lipid peroxidation stimulated by linoleic acid hydroperoxide of rat lung mitochondria and microsomes. Biochim Biophys Acta 1735:101–110

  50. Tölle A, Kolleck I, Schlame M, Wauer R, Stevens PA, Rüstow B (1997) Effect of hyperoxia on the composition of the alveolar surfactant and the turnover of surfactant phospholipids, cholesterol, plasmalogens and vitamin E. Biochim Biophys Acta 1346:198–204

  51. Ueda T, Ikegami M, Jobe A (1994) Surfactant subtypes. In vitro conversion, in vivo function, and effects of serum proteins. Am J Respir Crit Care Med 149:1254–1259

  52. Usui S, Matsunaga T, Ukai S, Kiho T (1997) Growth suppressing activity for endothelial cells induced from macrophages by carboxymethylated curdlan. Biosci Biotechnol Biochem 61:1924–1925

  53. Veldhuizen RA, Hearn SA, Lewis JF, Possmayer F (1994) Surface-area cycling of different surfactant preparations: SP-A and SP-B are essential for large-aggregate integrity. Biochem J 300:519–524

  54. Wang X, Chorley BN, Pittman GS, Kleeberger SR, Brothers J 2nd, Liu G, Spira A, Bell DA (2010) Genetic variation and antioxidant response gene expression in the bronchial airway epithelium of smokers at risk for lung cancer. PLoS One 5:e11934

  55. White MK, Strayer DS (2000) Surfactant protein A regulates pulmonary surfactant secretion via activation of phosphatidylinositol 3-kinase in type II alveolar cells. Exp Cell Res 255:67–76

  56. Yu CP, Xu GB (1987) Predictive models for deposition of inhaled diesel exhaust particles in humans and laboratory species. Res Rep Health Eff Inst 10:3–22

Download references

Author information

Correspondence to Toshiyuki Matsunaga.

Additional information

This work was supported in part by a grant for encouragement of young scientists from Gifu Pharmaceutical University.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Matsunaga, T., Haga, M., Watanabe, G. et al. 9,10-Phenanthrenequinone promotes secretion of pulmonary aldo-keto reductases with surfactant. Cell Tissue Res 347, 407–417 (2012).

Download citation


  • 9,10-Phenanthrenequinone
  • Aldo-keto reductase
  • Surfactant
  • Epithelial cell
  • Oxidative stress