Molecular and Cellular Biochemistry

, Volume 385, Issue 1–2, pp 297–307 | Cite as

IFN-γ (+874) and not TNF-α (−308) is associated with HBV-HCC risk in India

  • Roli Saxena
  • Yogesh Kumar Chawla
  • Indu Verma
  • Jyotdeep Kaur


Tumor necrosis factor (TNF)-α and interferon (IFN)-γ, the pro-inflammatory Th1 cytokines are the indispensable coordinators of the inflammatory responses involved in hepatitis B virus (HBV) pathogenesis. This study attempted to evaluate any possible association among TNF-α (−308G>A) and IFN-γ (+874T/A) genotypes, the spontaneous blood and mRNA levels and expression of their major signal transducers, namely STAT1 and NF-кB with hepatitis B virus-induced hepatocellular carcinoma (HCC) susceptibility in India. For this, 398 subjects (146 controls, 68 inactive-HBV-carriers, 64 chronic-active HBV patients, 61 HBV-cirrhotics, and 59 HBV-HCC subjects) were enrolled. Polymerase chain reaction–restriction fragment length polymorphism, allele-specific PCR, enzyme-linked immunosorbent assay, reverse transcriptase-PCR, and Western blot analysis were done for assessing polymorphism, blood levels, mRNA expression, and protein expression of signal transducers, respectively, of TNF-α and IFN-γ. The study revealed no significant association of TNF-α (−308) GA genotype, while a significant negative association of IFN-γ (+874) TA and AA genotypes, in HBV-HCC risk. Moreover, blood levels of TNF-α were significantly elevated as disease progresses to HCC, while IFN-γ levels were raised in HCC patients only. Besides, IFN-γ mRNA levels were significantly elevated in cirrhotics, with no change observed in TNF-α transcript levels. Moreover, NF-кB expression also consistently increased during HCC progression. These observations suggest a vital negative association of IFN-γ (+874) with HBV-HCC risk, with no significant association evident in TNF-α (−308). However, the TNF-α and IFN-γ levels markedly increased in HCC development.





The financial assistance by Indian Council of Medical Research (ICMR) is highly acknowledged.


No competing financial interests exist.


  1. 1.
    Wang B, Wang J, Zheng Y, Zhou S, Zheng J, Wang F, Ma X, Zeng Z (2010) A study of TNF-alpha−238 and −308 polymorphisms with different outcomes of persistent hepatitis B virus infection in China. Pathology 42:674–680. doi: 10.3109/00313025.2010.523696 CrossRefGoogle Scholar
  2. 2.
    Saxena R, Chawla YK, Verma I, Kaur J (2012) Interleukin-1 polymorphism and expression in hepatitis B virus-mediated disease outcome in India. J Interferon Cytokine Res. doi: 10.1089/jir.2012.0093 PubMedGoogle Scholar
  3. 3.
    Sun QL, Ran W (2004) Review of cytokine profiles in patients with hepatitis. World J Gastroenterol 10:1709–1715PubMedCentralCrossRefPubMedGoogle Scholar
  4. 4.
    Hohler T, Kruger A, Gerken G, Schneider PM, Meyer zum Buschenefelde KH, Rittner C (1998) A tumor necrosis factor-alpha (TNF-alpha) promoter polymorphism is associated with chronic hepatitis B infection. Clin Exp Immunol 111:579–582PubMedCentralCrossRefPubMedGoogle Scholar
  5. 5.
    Tekeli O, Turacli ME, Egin Y, Akar N, Elhan AH (2008) Tumor necrosis factor alpha-308 gene polymorphism and pseudoexfoliation glaucoma. Mol Vis 14:1815–1818PubMedCentralPubMedGoogle Scholar
  6. 6.
    Kumar A, Gupta V, Changotra H, Sarin BC, Sehajpal PK (2008) Tumor necrosis factor-alpha and transforming growth factor-beta1 polymorphisms in bronchial asthma. Indian J Med Sci 62:323–330CrossRefPubMedGoogle Scholar
  7. 7.
    Cuenca J, Cuchacovich M, Perez C, Ferreira L, Aguirre A, Schiattino I, Soto L, Cruzat A, Salazar-Onfray F, Aguillon JC (2003) The −308 polymorphism in the tumour necrosis factor (TNF) gene promoter region and ex vivo lipopolysaccharide-induced TNF expression and cytotoxic activity in Chilean patients with rheumatoid arthritis. Rheumatology (Oxford) 42:308–313CrossRefGoogle Scholar
  8. 8.
    Costeas PA, Koumas L, Koumouli A, Kyriakou-Giantsiou A, Papaloizou A (2003) Cytokine polymorphism frequencies in the Greek Cypriot population. Eur J Immunogenet 30:341–343CrossRefPubMedGoogle Scholar
  9. 9.
    Grove J, Daly AK, Bassendine MF, Day CP (1997) Association of a tumor necrosis factor promoter polymorphism with susceptibility to alcoholic steatohepatitis. Hepatology 26:143–146. doi: 10.1002/hep.510260119 CrossRefPubMedGoogle Scholar
  10. 10.
    Ben-Ari Z, Mor E, Papo O, Kfir B, Sulkes J, Tambur AR, Tur-Kaspa R, Klein T (2003) Cytokine gene polymorphisms in patients infected with hepatitis B virus. Am J Gastroenterol 98:144–150. doi: 10.1111/j.1572-0241.2003.07179.x CrossRefPubMedGoogle Scholar
  11. 11.
    Nieters A, Yuan JM, Sun CL, Zhang ZQ, Stoehlmacher J, Govindarajan S, Yu MC (2005) Effect of cytokine genotypes on the hepatitis B virus-hepatocellular carcinoma association. Cancer 103:740–748. doi: 10.1002/cncr.20842 CrossRefPubMedGoogle Scholar
  12. 12.
    Kusumoto K, Uto H, Hayashi K, Takahama Y, Nakao H, Suruki R, Stuver SO, Ido A, Tsubouchi H (2006) Interleukin-10 or tumor necrosis factor-alpha polymorphisms and the natural course of hepatitis C virus infection in a hyperendemic area of Japan. Cytokine 34:24–31. doi: 10.1016/j.cyto.2006.03.011 CrossRefPubMedGoogle Scholar
  13. 13.
    Zhang W, Yue B, Wang GQ, Lu SL (2002) Serum and ascites levels of macrophage migration inhibitory factor, TNF-alpha and IL-6 in patients with chronic virus hepatitis B and hepatitis cirrhosis. Hepatobiliary Pancreat Dis Int 1:577–580. doi: [pii]Google Scholar
  14. 14.
    Ataseven H, Bahcecioglu IH, Kuzu N, Yalniz M, Celebi S, Erensoy A, Ustundag B (2006) The levels of ghrelin, leptin, TNF-alpha, and IL-6 in liver cirrhosis and hepatocellular carcinoma due to HBV and HDV infection. Mediators Inflamm 2006:78380. doi: 10.1155/MI/2006/78380 PubMedCentralCrossRefPubMedGoogle Scholar
  15. 15.
    Pravica V, Perrey C, Stevens A, Lee JH, Hutchinson IV (2000) A single nucleotide polymorphism in the first intron of the human IFN-gamma gene: absolute correlation with a polymorphic CA microsatellite marker of high IFN-gamma production. Hum Immunol 61:863–866CrossRefPubMedGoogle Scholar
  16. 16.
    Bendicho MT, Guedes JC, Silva NN, Santana GO, dos Santos RR, Lyra AC, Lyra LG, Meyer R, Lemaire DC (2005) Polymorphism of cytokine genes (TGF-beta1, IFN-gamma, IL-6, IL-10, and TNF-alpha) in patients with chronic pancreatitis. Pancreas 30:333–336CrossRefGoogle Scholar
  17. 17.
    Farhat K, Hassen E, Gabbouj S, Bouaouina N, Chouchane L (2008) Interleukin-10 and interferon-gamma gene polymorphisms in patients with nasopharyngeal carcinoma. Int J Immunogenet 35:197–205. doi: 10.1111/j.1744-313X.2008.00752.x CrossRefPubMedGoogle Scholar
  18. 18.
    Vidyarani M, Selvaraj P, Prabhu Anand S, Jawahar MS, Adhilakshmi AR, Narayanan PR (2006) Interferon gamma (IFNgamma) & interleukin-4 (IL-4) gene variants & cytokine levels in pulmonary tuberculosis. Indian J Med Res 124:403–410PubMedGoogle Scholar
  19. 19.
    Hoffmann SC, Stanley EM, Cox ED, DiMercurio BS, Koziol DE, Harlan DM, Kirk AD, Blair PJ (2002) Ethnicity greatly influences cytokine gene polymorphism distribution. Am J Transplant 2:560–567CrossRefPubMedGoogle Scholar
  20. 20.
    Falasca K, Ucciferri C, Dalessandro M, Zingariello P, Mancino P, Petrarca C, Pizzigallo E, Conti P, Vecchiet J (2006) Cytokine patterns correlate with liver damage in patients with chronic hepatitis B and C. Ann Clin Lab Sci 36:144–150PubMedGoogle Scholar
  21. 21.
    Zhou L, Liu J, Luo F (2006) Serum tumor markers for detection of hepatocellular carcinoma. Methods Enzymol 12:1175–1181Google Scholar
  22. 22.
    Daly AK, Steen VM, Fairbrother KS, Idle JR (1996) CYP2D6 multiallelism. Methods Enzymol 272:199–210CrossRefPubMedGoogle Scholar
  23. 23.
    Chen TY, Hsieh YS, Wu TT, Yang SF, Wu CJ, Tsay GJ, Chiou HL (2007) Impact of serum levels and gene polymorphism of cytokines on chronic hepatitis C infection. Transl Res 150:116–121. doi: 10.1016/j.trsl.2007.01.007 CrossRefPubMedGoogle Scholar
  24. 24.
    Boyum A (1968) Isolation of leucocytes from human blood. A two-phase system for removal of red cells with methylcellulose as erythrocyte-aggregating agent. Scand J Clin Lab Invest Suppl 97:9–29PubMedGoogle Scholar
  25. 25.
    Towbin H, Staehelin T, Gordon J (1992) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. 1979. Biotechnology 24:145–149PubMedGoogle Scholar
  26. 26.
    Guidotti LG, Rochford R, Chung J, Shapiro M, Purcell R, Chisari FV (1999) Viral clearance without destruction of infected cells during acute HBV infection. Science 284:825–829CrossRefPubMedGoogle Scholar
  27. 27.
    Vikram NK, Bhatt SP, Bhushan B, Luthra K, Misra A, Poddar PK, Pandey RM, Guleria R (2011) Associations of −308G/A polymorphism of tumor necrosis factor (TNF)-alpha gene and serum TNF-alpha levels with measures of obesity, intra-abdominal and subcutaneous abdominal fat, subclinical inflammation and insulin resistance in Asian Indians in north India. Dis Markers 31:39–46. doi: 10.3233/DMA-2011-0802 PubMedCentralCrossRefPubMedGoogle Scholar
  28. 28.
    Migita K, Miyazoe S, Maeda Y, Daikoku M, Abiru S, Ueki T, Yano K, Nagaoka S, Matsumoto T, Nakao K, Hamasaki K, Yatsuhashi H, Ishibashi H, Eguchi K (2005) Cytokine gene polymorphisms in Japanese patients with hepatitis B virus infection—association between TGF-beta1 polymorphisms and hepatocellular carcinoma. J Hepatol 42:505–510. doi: 10.1016/j.jhep.2004.11.026 CrossRefPubMedGoogle Scholar
  29. 29.
    Kaur G, Rapthap CC, Kumar N, Kumar S, Neolia S, Mehra NK (2007) Frequency distribution of cytokine gene polymorphisms in the healthy North Indian population. Tissue Antigens 69:113–120. doi: 10.1111/j.1399-0039.2006.00740.x CrossRefPubMedGoogle Scholar
  30. 30.
    Sharma S, Ghosh B, Sharma SK (2008) Association of TNF polymorphisms with sarcoidosis, its prognosis and tumour necrosis factor (TNF)-alpha levels in Asian Indians. Clin Exp Immunol 151:251–259. doi: 10.1111/j.1365-2249.2007.03564.x PubMedCentralCrossRefPubMedGoogle Scholar
  31. 31.
    Somi MH, Najafi L, Noori BN, Alizadeh AH, Aghah MR, Shavakhi A, Ehsani MJ, Aghazadeh R, Masoodi M, Amini S, Baladast M, Zali MR (2006) Tumor necrosis factor-alpha gene promoter polymorphism in Iranian patients with chronic hepatitis B. Indian J Gastroenterol 25:14–15PubMedGoogle Scholar
  32. 32.
    Zein NN, Germer JJ, El-Zayadi AR, Vidigal PG (2004) Ethnic differences in polymorphisms of tumor necrosis factor-alpha, interleukin-10, and transforming growth factor-beta1 genes in patients with chronic hepatitis C virus infection. Am J Trop Med Hyg 70:434–437PubMedGoogle Scholar
  33. 33.
    Trajkov D, Arsov T, Petlichkovski A, Strezova A, Efinska-Mladenovska O, Spiroski M (2005) Cytokine gene polymorphisms in population of ethnic Macedonians. Croat Med J 46:685–692PubMedGoogle Scholar
  34. 34.
    Miyazoe S, Hamasaki K, Nakata K, Kajiya Y, Kitajima K, Nakao K, Daikoku M, Yatsuhashi H, Koga M, Yano M, Eguchi K (2002) Influence of interleukin-10 gene promoter polymorphisms on disease progression in patients chronically infected with hepatitis B virus. Am J Gastroenterol 97:2086–2092. doi: 10.1111/j.1572-0241.2002.05926.x CrossRefPubMedGoogle Scholar
  35. 35.
    Chan HL, Tse AM, Chim AM, Wong VW, Choi PC, Yu J, Zhang M, Sung JJ (2008) Association of cytokine gene polymorphisms and liver fibrosis in chronic hepatitis B. J Gastroenterol Hepatol 23:783–789. doi: 10.1111/j.1440-1746.2007.05110.x CrossRefPubMedGoogle Scholar
  36. 36.
    Jeng JE, Tsai JF, Chuang LY, Ho MS, Lin ZY, Hsieh MY, Chen SC, Chuang WL, Wang LY, Yu ML, Dai CY, Chang JG (2007) Tumor necrosis factor-alpha 308.2 polymorphism is associated with advanced hepatic fibrosis and higher risk for hepatocellular carcinoma. Neoplasia 9:987–992CrossRefPubMedGoogle Scholar
  37. 37.
    Kohaar I, Thakur N, Salhan S, Batra S, Singh V, Sharma A, Sodhani P, Das BC, Sarkar DP, Bharadwaj M (2007) TNF alpha-308G/A polymorphism as a risk factor for HPV associated cervical cancer in Indian population. Cell Oncol 29:249–256PubMedCentralPubMedGoogle Scholar
  38. 38.
    Kohaar I, Tiwari P, Kumar R, Nasare V, Thakur N, Das BC, Bharadwaj M (2009) Association of single nucleotide polymorphisms (SNPs) in TNF-LTA locus with breast cancer risk in Indian population. Breast Cancer Res Treat 114:347–355. doi: 10.1007/s10549-008-0006-5 CrossRefPubMedGoogle Scholar
  39. 39.
    Budak F, Goral G, Heper Y, Yilmaz E, Aymak F, Basturk B, Tore O, Ener B, Oral HB (2007) IL-10 and IL-6 gene polymorphisms as potential host susceptibility factors in Brucellosis. Cytokine 38:32–36. doi: 10.1016/j.cyto.2007.04.008 CrossRefPubMedGoogle Scholar
  40. 40.
    Yilmaz V, Demirbilek V, Gurses C, Yentur SP, Uysal S, Yapici Z, Yilmaz G, Muncey A, Cokar O, Onal E, Gokyigit A, Saruhan-Direskeneli G (2007) Interleukin (IL)-12, IL-2, interferon-gamma gene polymorphisms in subacute sclerosing panencephalitis patients. J Neurovirol 13:410–415. doi: 10.1080/13550280701455383 CrossRefPubMedGoogle Scholar
  41. 41.
    Colakogullari M, Ulukaya E, Yilmaztepe Oral A, Aymak F, Basturk B, Ursavas A, Oral HB (2008) The involvement of IL-10, IL-6, IFN-gamma, TNF-alpha and TGF-beta gene polymorphisms among Turkish lung cancer patients. Cell Biochem Funct 26:283–290. doi: 10.1002/cbf.1419 CrossRefPubMedGoogle Scholar
  42. 42.
    Cheong JY, Cho SW, Chung SG, Lee JA, Yeo M, Wang HJ, Lee JE, Hahm KB, Kim JH (2006) Genetic polymorphism of interferon-gamma, interferon-gamma receptor, and interferon regulatory factor-1 genes in patients with hepatitis B virus infection. Biochem Genet 44:246–255. doi: 10.1007/s10528-006-9029-y CrossRefPubMedGoogle Scholar
  43. 43.
    Kamali-Sarvestani E, Merat A, Talei AR (2005) Polymorphism in the genes of alpha and beta tumor necrosis factors (TNF-alpha and TNF-beta) and gamma interferon (IFN-gamma) among Iranian women with breast cancer. Cancer Lett 223:113–119. doi: 10.1016/j.canlet.2004.09.025 CrossRefPubMedGoogle Scholar
  44. 44.
    Wu WS, McClain KL (1997) DNA polymorphisms and mutations of the tumor necrosis factor-alpha (TNF-alpha) promoter in Langerhans cell histiocytosis (LCH). J Interferon Cytokine Res 17:631–635CrossRefPubMedGoogle Scholar
  45. 45.
    Wilson AG, Symons JA, McDowell TL, McDevitt HO, Duff GW (1997) Effects of a polymorphism in the human tumor necrosis factor alpha promoter on transcriptional activation. Proc Natl Acad Sci USA 94:3195–3199PubMedCentralCrossRefGoogle Scholar
  46. 46.
    Kroeger KM, Carville KS, Abraham LJ (1997) The -308 tumor necrosis factor-alpha promoter polymorphism effects transcription. Mol Immunol 34:391–399CrossRefPubMedGoogle Scholar
  47. 47.
    Braun N, Michel U, Ernst BP, Metzner R, Bitsch A, Weber F, Rieckmann P (1996) Gene polymorphism at position -308 of the tumor-necrosis-factor-alpha (TNF-alpha) in multiple sclerosis and its influence on the regulation of TNF-alpha production. Neurosci Lett 215:75–78PubMedGoogle Scholar
  48. 48.
    Lichtenstein M, Zimran A, Horowitz M (1997) Cytokine mRNA in Gaucher disease. Blood Cells Mol Dis 23:395–401. doi: 10.1006/bcmd.1997.0156 CrossRefPubMedGoogle Scholar
  49. 49.
    Kothari ST, Deshmukh RA (2006) Estimation of mRNA levels of Interleukin (IL)-10, Tumor Necrosis Factor (TNF)-α, IL-4 and Interferon (IFN)-γ in HIV infected children in Mumbai. Indian J Clin Biochem 21:15–26PubMedCentralCrossRefPubMedGoogle Scholar
  50. 50.
    Fan XG, Liu WE, Li CZ, Wang ZC, Luo LX, Tan DM, Hu GL, Zhang Z (1998) Circulating Th1 and Th2 cytokines in patients with hepatitis C virus infection. Mediators Inflamm 7:295–297. doi: 10.1080/09629359890992 PubMedCentralCrossRefPubMedGoogle Scholar
  51. 51.
    le Song H, Binh VQ, Duy DN, Kun JF, Bock TC, Kremsner PG, Luty AJ (2003) Serum cytokine profiles associated with clinical presentation in Vietnamese infected with hepatitis B virus. J Clin Virol 28:93–103CrossRefGoogle Scholar
  52. 52.
    Gramenzi A, Andreone P, Loggi E, Foschi FG, Cursaro C, Margotti M, Biselli M, Bernardi M (2005) Cytokine profile of peripheral blood mononuclear cells from patients with different outcomes of hepatitis C virus infection. J Viral Hepat 12:525–530. doi: 10.1111/j.1365-2893.2005.00634.x CrossRefPubMedGoogle Scholar
  53. 53.
    Ayada M, Ishikawa T, Okumura A, Tanabe J, Ito H, Ohashi T, Matsumoto E, Sato K, Hotta N, Fukuzawa Y, Kakumu S (2006) Alteration of serum cytokine balances among different phases of chronic hepatitis B virus infection. Hepatol Res 34:214–221. doi: 10.1016/j.hepres.2006.01.001 CrossRefPubMedGoogle Scholar
  54. 54.
    Forte GI, Scola L, Misiano G, Milano S, Mansueto P, Vitale G, Bellanca F, Sanacore M, Vaccarino L, Rini GB, Caruso C, Cillari E, Lio D, Mansueto S (2009) Relevance of gamma interferon, tumor necrosis factor alpha, and interleukin-10 gene polymorphisms to susceptibility to Mediterranean spotted fever. Clin Vaccine Immunol 16:811–815. doi: 10.1128/CVI.00121-09 PubMedCentralCrossRefPubMedGoogle Scholar
  55. 55.
    Luo JL, Maeda S, Hsu LC, Yagita H, Karin M (2004) Inhibition of NF-kappaB in cancer cells converts inflammation- induced tumor growth mediated by TNFalpha to TRAIL-mediated tumor regression. Cancer Cell 6:297–305. doi: 10.1016/j.ccr.2004.08.012 CrossRefPubMedGoogle Scholar
  56. 56.
    Lin WW, Karin M (2007) A cytokine-mediated link between innate immunity, inflammation, and cancer. J Clin Invest 117:1175–1183. doi: 10.1172/JCI31537 PubMedCentralCrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Roli Saxena
    • 1
  • Yogesh Kumar Chawla
    • 2
  • Indu Verma
    • 1
  • Jyotdeep Kaur
    • 1
  1. 1.Department of BiochemistryPostgraduate Institute of Medical Education and ResearchChandigarhIndia
  2. 2.Department of HepatologyPostgraduate Institute of Medical Education and ResearchChandigarhIndia

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