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Hepatoprotective Effects of Semisulcospira libertine Hydrolysate on Alcohol-induced Fatty Liver in Mice

알코올성 지방간 유발 마우스에서 다슬기 유래 가수분해물의 간 보호 효과

  • 송은진 (경상대학교 의과대학 외과학교실 건강과학연구원) ;
  • 조경환 ((재)하농녹차연구소) ;
  • 추호진 ((주)정옥) ;
  • 양은영 ((주)정옥) ;
  • 정윤경 ((주)정옥) ;
  • 서민균 ((주)정옥) ;
  • 김종철 ((재)하농녹차연구소) ;
  • 강은주 ((재)하농녹차연구소) ;
  • 류기형 (공주대학교 식품공학과) ;
  • 박범용 (경상대학교 의과대학 외과학교실 건강과학연구원) ;
  • 하영술 (경상대학교 의과대학 외과학교실 건강과학연구원)
  • Received : 2017.07.24
  • Accepted : 2017.08.22
  • Published : 2017.11.30

Abstract

Alcoholic steatosis is a fundamental metabolic disorder and may precede the onset of more severe forms of alcoholic liver disease. In this study, we isolated enzymatichydrolysate from Semisulcospira libertine by alcalase hydrolysis and investigated the protective effect of Semisulcospira libertine hydrolysate on liver injury induced by alcohol in the mouse model of chronic and binge ethanol feeding (NIAAA). In an in vitro study, the hydrolysate protects HepG2 cells from ethanol toxicity. Liver damage was assessed by histopathological examination, as well as by quantitating activities of serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP). After the administration of S. libertina hydrolysate, fat accumulation and infiltration of inflammatory cells in liver tissues were significantly decreased in the NIAAA mouse model. The elevated levels of serum AST, ALT, and ALP activities, along with the lipid contents of a damaged liver, were recovered in experimental mice administrated with S. libertina hydrolysate, suggesting its role in blood enzyme activation and lipid content restoration within damaged liver tissues. Moreover, treatment with S. libertine hydrolysate reduced the expression rate of cyclooxygenase (COX-2), interleukin $(IL)-1{\beta}$, and IL-6, which accelerate inflammation and induces tissue damage. All data showed that S. libertine hydrolysate has a preventive role against alcohol-induced liver damages by improving the activities of blood enzymes and modulating the expression of inflammation factor, suggesting S. libertine hydrolysate could be a commercially potential material for the restoration of hepatotoxicity.

본 연구는 다슬기의 간 보호 효능을 확인하고자 하였다. 다슬기를 단백질 가수분해효소인 alcalase를 이용하여 가수분해물을 얻은 후 실험을 수행하였다. HepG2 세포에 다슬기 가수분해물을 처리하였을 때 세포독성이 나타나지 않음을 확인하였다. 그리고 다슬기 가수분해물과 에탄올(1 M)을 가하여 24시간 후 세포생존율을 확인했을 때, 세포가 알코올 독성으로부터 보호되는 것을 확인할 수 있었다. 다음으로 다슬기 가수분해물의 간 손상 보호 효과를 Liber-DeCarli 알코올 액체 식이 급여와 동시에 경구 투여를 통한 알코올 단회 폭음(single binge)으로 알코올성 지방간과 염증을 유발시킨 마우스모델(NIAAA)을 사용하여 확인하고자 하였다. 이들 마우스 모델에서 알코올 식이 급여 후 초기에 체중이 감소하는 경향을 보이나 점차 비알코올 식이 급여군과 유사한 체중으로 회복하였다. 조직학적 관찰에서 알코올로 유도된 간 세포내 지방 축적과 지방성 염증이 에탄올과 함께 다슬기 가수분해물을 투여한 군에서 개선되는 것을 관찰 할 수 있었다. 또한, 알코올 투여에 의한 AST, ALT 및 ALP의 효소활성 증가가 다슬기 가수분해물 처리에 의해 억제되었으며, 만성 알코올 섭취에 의한 염증 매개 효소와 염증성 사인토카인의 발현 증가는 다슬기 가수분해물을 투여한 군에서 양성대조군에 비하여 유의하게 감소하였다. 이상의 연구결과를 종합해 보면 다슬기 효소 가수분해물 소재는 알코올성 지방간의 예방 및 개선을 위한 기능성식품용 상업적 소재로서 높은 실용화 가능성을 시사한다.

Keywords

Acknowledgement

Supported by : 중소벤처기업부

References

  1. Angulo P. 2002. Nonalcoholic fatty liver disease. N. Engl. J. Med. 346: 1221-1231. https://doi.org/10.1056/NEJMra011775
  2. Assy N, Kaita K, Mymin D, Levy C, Rosser B, Minuk G. 2000. Fatty infiltration of liver in hyperlipidemic patients. Dig. Dis. Sci. 45: 1929-1934. https://doi.org/10.1023/A:1005661516165
  3. Bellentani S, Saccoccio G, Costa G, Tiribelli C, Manenti F, Sodde M, Saveria Croce L, Sasso F, Pozzato G, Cristianini G, Brandi G. 1997. Drinking habits as cofactors of risk for alcohol induced liver damage. The Dionysos Study Group. Gut. 41: 845-850.
  4. Bertola A, Mathews S, Ki SH, Wang H, Gao B. 2013. Mouse model of chronic and binge ethanol feeding (the NIAAA model). Nat. Protoc. 8: 627-637. https://doi.org/10.1038/nprot.2013.032
  5. Cho JM, Hong KS, Lee DY, Kim KY, Ji SD, Kim EH. 2016. Protective effect of silkworm (Bombyx mori) powder against diethylnitrosamine-induced hepatotoxicity in mice. Food Eng. Prog. 20: 342-348. https://doi.org/10.13050/foodengprog.2016.20.4.342
  6. Cha HO, Baik WA. 1999. Effect of Semisulcospira libertina, Corbicula sp. and Cipangopaludina chinensis malleata extracts on liver function and other biological activities. 45th National Sci. Exhibition of Ministry of Science and Technology, Korea
  7. Cho KH, Choo HJ, Seo MG, Kim JC, Shin YJ, Ryu GH, Cho HY, Jeong CY, Hah YS. 2017. Effect of Semisulcospira libertina extracts from different extraction processes on liver cell toxicity and ethanol metabolism. Food Eng. Prog. 21: 158-166. https://doi.org/10.13050/foodengprog.2017.21.2.158
  8. Choi JH, Kim MS, Yu HJ, Kim KH, Lee HS, Cho HY, Lee SH. 2014. Hepatoprotective effects of lactic acid-fermented garlic extracts on fatty liver-induced mouse by alcohol. J. Korean Soc. Food Sci. Nutr. 43: 1642-1647. https://doi.org/10.3746/jkfn.2014.43.11.1642
  9. Corsetti G, Stacchiotti A, Tedesco L, D'Antona G, Pasini E, Dioguardi FS, Nisoli E, Rezzani R. 2011. Essential amino acid supplementation decreases liver damage induced by chronic ethanol consumption in rats. Int. J. Immunopathol. Pharmacol. 24: 611-619. https://doi.org/10.1177/039463201102400307
  10. D'Antona G1, Ragni M, Cardile A, Tedesco L, Dossena M, Bruttini F, Caliaro F, Corsetti G, Bottinelli R, Carruba MO, Valerio A, Nisoli E. 2010. Branched-chain amino acid supplementation promotes survival and supports cardiac and skeletal muscle mitochondrial biogenesis in middle-aged mice. Cell Metab. 12: 362-372. https://doi.org/10.1016/j.cmet.2010.08.016
  11. Di Luzio N, Hartman A. 1967. Role of lipid peroxidation in the pathogenesis of the ethanol-induced fatty liver. Fed. Proc. 26: 1436-1442.
  12. Haybaeck J, Zeller N, Wolf MJ, Weber A, Wagner U, Kurrer MO, Bremer J, Iezzi G, Graf R, Clavien PA, Thimme R, Blum H, Nedospasov SA, Zatloukal K, Ramzan M, Ciesek S, Pietschmann T, Marche PN, Karin M, Kopf M, Browning JL, Aguzzi A, Heikenwalder M. 2009. A lymphotoxin-driven pathway to hepatocellular carcinoma. Cancer Cell 16: 295-308. https://doi.org/10.1016/j.ccr.2009.08.021
  13. Kim YK, Moon HS, Lee, MH, Park MJ, Lim CW, Paek HY, Park JI, Yoon HD, Kim DH. 2009. Biological activities of seven melania snails in Korea. Kor. J. Fish Aquat. Sci. 42: 434-441.
  14. Kleiner DE, Brunt EM, Van Natta M, Behling C, Contos MJ, Cummings OW, Ferrell LD, Liu YC, Torbenson MS, Unalp-Arida A, Yeh M, McCullough AJ, Sanyal AJ. 2005. Design and validation of a histological scoring system for nonalcoholic fatty liver disease. Hepatology 41: 1313-1321. https://doi.org/10.1002/hep.20701
  15. Lee MS, Park JB, Yoon SH. 2005. Hepatoprotective effect of the water extract from Semisulcospira gottschei against liver injuries induced by carbon tetrachloride in rats. J. Korean Soc. Hyg. Sci. 11: 17-26.
  16. Lefkowitch JH. 2005. Morphology of alcoholic liver disease. Clin. Liver Dis. 9: 37-53. https://doi.org/10.1016/j.cld.2004.11.001
  17. Liaskou E, Wilson DV, Oo YH. 2012. Innate immune cells in liver inflammation. Mediators Inflamm. 2012: 949157.
  18. Lim CW, Kim YK, Kim DH, Park JI, Lee MH, Park HY, Jang MS. 2009. Comparison of quality characteristics of melania snails in Korea. Kor. J. Fish Aquat. Sci. 42: 555-560.
  19. Linder MC. 1991. Nutrition and metabolism of fats. Nutritional biochemistry and metabolism: with clinical applications 2nd ed. Elsevier, Amsterdam. Oxford, NY, USA. pp 79-83.
  20. Lu KH1, Tseng HC, Liu CT, Huang CJ, Chyuan JH, Sheen LY. 2014. Wild bitter gourd protects against alcoholic fatty liver in mice by attenuating oxidative stress and inflammatory responses. Food Funct. 5: 1027-1037. https://doi.org/10.1039/C3FO60449G
  21. Luedde T, Beraza N, Kotsikoris V, van Loo G, Nenci A, De Vos R, Roskams T, Trautwein C, Pasparakis M. 2007. Deletion of NEMO/IKKgamma in liver parenchymal cells causes steatohepatitis and hepatocellular carcinoma. Cancer Cell 11: 119-132. https://doi.org/10.1016/j.ccr.2006.12.016
  22. Mezey E. 1980. Alcoholic liver disease: roles of alcohol and malnutrition. Am. J. Clin. Nutr. 33: 2709-2718. https://doi.org/10.1093/ajcn/33.12.2709
  23. Moscatiello S, Manini R, Marchesini G. 2007. Diabetes and liver disease: an ominous association. Nutr. Metab. Cardiovasc. Dis. 17: 63-70. https://doi.org/10.1016/j.numecd.2006.08.004
  24. Rehm J, Samokhvalov AV, Shield KD. 2013. Global burden of alcoholic liver diseases. J. Hepatol. 59: 160-168. https://doi.org/10.1016/j.jhep.2013.03.007
  25. Nagata K, Suzuki H, Sakaguchi S. 2007. Common pathogenic mechanism in development progression of liver injury caused by non-alcoholic or alcoholic steatohepatitis. J. Toxicol. Sci. 32: 453-468. https://doi.org/10.2131/jts.32.453
  26. Pikaar NA, Wedel M, van der Beek EJ, van Dokkum W, Kempen HJ, Kluft C, Ockhuizen T, Hermus RJ. 1987. Effects of moderate alcohol consumption on platelet aggregation, fibrinolysis, and blood lipids. Metabolism 36: 538-543. https://doi.org/10.1016/0026-0495(87)90163-6
  27. Schneider C, Teufel A, Yevsa T, Staib F, Hohmeyer A, Walenda G, Zimmermann HW, Vucur M, Huss S, Gassler N, Wasmuth HE, Lira SA, Zender L, Luedde T, Trautwein C, Tacke F. 2012. Adaptive immunity suppresses formation and progression of diethylnitrosamine-induced liver cancer. Gut. 61: 1733-1743. https://doi.org/10.1136/gutjnl-2011-301116
  28. Son M, Moon JY, Park S, Cho M. 2016. Hepatoprotective effect of hippocampus abdominalis hydrolysate. J. Appl. Biol. Chem. 59: 265-271. https://doi.org/10.3839/jabc.2016.046
  29. Sozio M, Crabb DW. 2008. Alcohol and lipid metabolism. Am. J. Physiol. Endocrinol. Metab. 295: E10-E16. https://doi.org/10.1152/ajpendo.00011.2008
  30. You M, Fischer M, Deeg MA, Crabb DW. 2002. Ethanol induces fatty acid synthesis pathways by activation of sterol regulatory element-binding protein (SREBP). J. Biol. Chem. 277: 29342-29347. https://doi.org/10.1074/jbc.M202411200
  31. You M, Matsumoto M, Pacold CM, Cho WK, Crabb DW. 2004. The role of AMP-activated protein kinase in the action of ethanol in the liver. Gastroenterology. 127: 1798-1808. https://doi.org/10.1053/j.gastro.2004.09.049
  32. Wu D, Cederbaum AI. 1999. Ethanol cytotoxicity to a transfected HepG2 cell line expressing human cytochrome P450E1. Alcohol. Clin. Exp. Res. 23: 67-76. https://doi.org/10.1111/j.1530-0277.1999.tb04025.x