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Anti-diabetic mechanism of melania snail (Semisulcospira libertina) protamex hydrolysates

다슬기 protamex 가수분해물(MPH)의 항당뇨 기작 연구

  • Pyo, Sang-Eun (Major in Food Biotechnology, Division of Bioindustry, Silla University) ;
  • Choi, Jae-Suk (Major in Food Biotechnology, Division of Bioindustry, Silla University) ;
  • Kim, Mi-Ryung (Major in Food Biotechnology, Division of Bioindustry, Silla University)
  • 표상은 (신라대학교 바이오산업학부 식품공학전공) ;
  • 최재석 (신라대학교 바이오산업학부 식품공학전공) ;
  • 김미령 (신라대학교 바이오산업학부 식품공학전공)
  • Received : 2017.10.30
  • Accepted : 2017.11.22
  • Published : 2017.11.30

Abstract

Melania snail (Semisulcospira libertina) was traditionally used as the healthy food in Korea. It was generally known to improve liver function and heal a diabetes. The aim of this study was to elucidate the anti-diabetic mechanism of melanian snail hydrolysates treated with protamex (MPH) by investigating the inhibitory action on protein tyrosine phosphatase 1B (PTP1B), the improving effect on the insulin resistance in C2C12 myoblast and the protective effect for pancreatic beta-cell (INS-1) under the glucose toxicity. The melania snail hydrolysates treated with protamex (MPH), which showed the highest degree of hydrolysis (43%), and inhibited effectively PTP1B activity ($IC_{50}=15.42{\pm}1.1{\mu}g/mL$), of which inhibitory effect was higher than usolic acid, positive control ($IC_{50}=16.65{\mu}g/mL$). MPH increased the glucose uptake in C2C12 myoblast treated with palmitic acid. In addition, MPH increased insulin mRNA expression level by over 160% with enhanced cell viability in INS-1 cell under the high glucose concentration (30 mM). These results suggest that MHP may improve the diabetic symptom by the inhibiting the PTP1B activity, increasing the glucose uptake in muscle cell and protecting the pancreatic beta-cell from glucose toxicity.

다슬기는 예로부터 간염, 간경화, 지방간 등의 치료 및 개선에 이용되어 왔으며, 특히 소변불통, 소갈증(당뇨) 등의 약용으로 이용되어 왔다. 본 연구에서는 이러한 다슬기를 대상으로 항당뇨에 대한 효능을 과학적으로 검증하고 그 기작을 규명하고자 하였다. 먼저 다슬기의 생물학적 기능성을 높이기 위해 효소 가수분해를 실시하였으며, protamex에 의한 가수분해도는 10시간 후 약 43% 수준을 나타내었다. PTP1B는 인슐린 신호전달기전에서 IRS-1의 인산화를 방해하여 인슐린 민감성을 저해시키는 효소이다. protamex를 이용한 다슬기 가수분해물(MPH)의 PTP1B에 대한 저해활성은 $15.42{\pm}1.1{\mu}g/mL$$IC_{50}$ 값을 나타내어 양성대조군 ursolic acid의 $16.7{\mu}g/mL$ 보다 높은 저해활성을 보이면서 강한 항당뇨 활성 소재로서의 가능성을 보였다. 이에 따라 유리지방산을 이용하여 C2C12 myoblast에서 인슐린 저항성을 유도하고, MPH에 의한 포도당 흡수 정도를 확인하였다. 그 결과, 1 mM PA 처리에 의해 약 32% 수준으로 떨어진 포도당 흡수율은 MPH 처리에 의해 약 199% 수준으로 증가하였다. 또한 장기간 고농도의 포도당(30 mM)에 의해 유도된 당독성 조건에서 MPH는 췌장의 베타세포 INS-1 세포의 생존율을 증가시키고, 대조군에 비해 약 160% 인슐린 mRNA 발현량을 증가시켰다. 이러한 결과에서 MPH는 PTP1B 활성을 저해함으로써 인슐린 신호전달 기작을 활성화하고, 인슐린저항성 환경에서 포도당 흡수를 증진시켜 인슐린저항성을 개선하며, 나아가 고농도 포도당에 의해 유도되는 당독성환경에서 췌장 베타세포를 보호하고 인슐린 mRNA발현량을 정상화할 수 있다는 것을 확인할 수 있었다.

Keywords

References

  1. Jeong HJ, Lee SG, Lee EJ, Park WD, Kim JB, Kim HJ (2010) Antioxidant activity and anti-hyperglycemic activity of medicinal herbal extracts according to extraction methods. Korean J Food Sci Technol, 42, 571-577
  2. Xu ML, Hu JH, Wang L, Kim HS, Jin CW, Cho DH (2010) Antioxidant and anti-diabetes activity of extracts from Machilus thunbergii S. et Z. Korean J Med Crop Sci, 18, 34-39
  3. World Health Organization. Global report on diabetes. http://www.who.int/diabetes/global-report/en/ (accessed 2006).
  4. Hwang JT, Kim SH (2012) Evaluation of anti-diabetic effect of biochanin A in C2C12 myotube. Korean Soc Biotechnol Bioeng J, 27, 57-60
  5. Yoo HJ (2012) Pharmacotherapy for postprandial hyperglycemia in Type 2 diabetes. J Korean Diabetes, 13, 39-43 https://doi.org/10.4093/jkd.2012.13.1.39
  6. Park KS, Ko SK, Chung SH (2003) Comparisons of antidiabetic effect between ginseng radix alba, ginseng radix rubra and panax quinquefoli radix in MLD STZ-induced diabetic Rats. J Ginseng Res, 27, 56-61 https://doi.org/10.5142/JGR.2003.27.2.056
  7. Lee JM (2014) Antihyperglycemic agent combination therapy for patients with type 2 diabets mellitus. J Korean Med Assoc, 57, 435-443 https://doi.org/10.5124/jkma.2014.57.5.435
  8. Kwon EJ, Hong SG, Kim MM, Kim JW, Kim DW, Chung KT (2014) Effects of ginseng berry water extract on the polysaccharide hydrolysis of extracellular enzymes and intracellular PTP1B and AKT1. J Life Sci, 24, 1006-1011 https://doi.org/10.5352/JLS.2014.24.9.1006
  9. Im SA, Kim KH, Shin EJ, Do SG, Jo TH, Park YI, Lee CK (2013) Effects of antidiabetic agent, aloe QDM complex, on intracellular glucose uptake. Korean J Pharmacogn, 44, 75-82
  10. Cho EK, Choi YJ (2013) Antioxidant, antidiabetic and anti-inflammatory effects of extracts and fractions from Parthenocissus tricuspidata stems. J Life Sci, 23, 399-405 https://doi.org/10.5352/JLS.2013.23.3.399
  11. Kang TS, Kang MS, Sung JM, Kang AS, Shon HR, Lee SY (2001) Effects of Pleurotus eryngii on the blood glucose and cholesterol in diabetic rats. Korean J Mycol, 29, 86-90
  12. Kim YH, Lee TK, Cha YS (1985) Studies on the Nutritive Component of black snail (Semisulcospira libertina). Bull Agric College Chonbuk Univ, 16, 101-105
  13. Kim YK, Moon HS, Lee MH, Park MJ, Lim CW, Park HY, Park JI, Yoon HD, Kim DH (2009) Biological Activities of seven melania snails in Korea. Korean J Fish Aquat Sci, 42, 434-441
  14. Lee MS, Park JB, Yoon SH (2005) Hepatoprotective effects of the water extract from Semisulcospira gottschei against liver injuries induced by carbon tetrachloride in rats. J Korean Soc Hyg Sci, 11, 17-26
  15. Park YM, Lim JH (2015) Protective effect of Semisulcospira libertina extract on induced hepatitis in rats. J Life Sci, 25, 539-547 https://doi.org/10.5352/JLS.2015.25.5.539
  16. Lim CW, Kim YK, Kim DH, Park JI, Lee MH, Park HY, Jang MS (2009) Comparison of quality characteristics of melania snails in Korea. Korean J Fish Aquat Sci, 42, 555-560
  17. Lee MH, Kim YK, Moon HS, Kim YA, Yoon NY, Lim CW, Park HY, Kim DH (2010) Antioxidant activities of five melania snails of the genus Semisulcospira in Korea. Korean J Fish Aquat Sci, 43, 188-194
  18. Choi JS, Kim JW, Park JB, Pyo SE, Hong YK, Ku SK, Kim MR (2017) Blood glycemia-modulating effects of melanian snail protein hydrolysates in mice with type II diabetes. Int J Mol Med, 39, 1437-1451 https://doi.org/10.3892/ijmm.2017.2967
  19. AOAC (1995) Official Methods of Analysis 16th ed, Association of Official Analytical Chemists, Washington DC, USA, p 69-74
  20. Scopes RK (1987) Protein Purification: Principles and Practice. 2nd ed, Springer-Verlag, New York, NY, USA, p 279-280
  21. Shim TH, Han KS, Lee TJ, Cheong EH, Lee HK (1994) Comparison of lipid and amino acid in Semisulcospira gorrschei tissue. J Food Hyg Saf, 9, 81-87
  22. Shin MJ, Park MJ, Youn MS, Lee YS, Nam MS, Park IS, Jeong YH (2006) Effects of silk protein hydrolysates on blood glucose and serum lipid in db/db diabetic mice. J Korean Soc Food Sci Nutr, 35, 1343-1348 https://doi.org/10.3746/jkfn.2006.35.10.1343
  23. Jang SY, Gu YA, Park NY, Kim IS, Jeong YJ (2007) Physicochemical property changes of whole soymilk dependent on hydrolysis conditions. Korean J Food Preserv, 14, 394-399
  24. Yoon SJ (2013) Diabete and therapeutic agents for glycemic control in diabetes mellitus, Report of Korea Drug Development Fund http://kddf.org/bbs/bbs.asp? mode=view&IDX=785&p=9&cateId=39
  25. Koren S, Fantus IG (2007) Inhibition of the protein tyrosine phosphatase PTP1B: potential therapy for obesity, insulin resistance and type-2 diabetes mellitus. Best Pract Res Clin Endocrinol Metab, 21, 621-640 https://doi.org/10.1016/j.beem.2007.08.004
  26. Kulkarni RN, Bruning JC, Winnay JN, Postic C, Magnuson MA, Kahn CR (1999) Tissue-specific knockout of the insulin receptor in pancreatic ${\beta}$ cells creates an insulin secretory defect similar to that in type 2 diabetes. Cell, 96, 329-339 https://doi.org/10.1016/S0092-8674(00)80546-2
  27. Lee J, Pilch P (1994) The insulin receptor: structure, function, and signaling. Am J Physiol, 266, C319-C334 https://doi.org/10.1152/ajpcell.1994.266.2.C319
  28. Saltiel AR, Kahn CR (2001) Insulin signalling and the regulation of glucose and lipid metabolism. Nature, 414, 799-806 https://doi.org/10.1038/414799a
  29. Kim SM, Lee YM, Kim MJ, Nam SY, Kim SH, Jang HH (2013) Effects of Agrimonia pilosa Ledeb. water extract on ${\alpha}$-glucosidase inhibition and glucose uptake in C2C12 skeletal muscle cells. Korean J Food Nutr, 26, 806-813 https://doi.org/10.9799/ksfan.2013.26.4.806
  30. Yip SC, Saha S, Chernoff J (2010) PTP1B: a double agent in metabolism and oncogenesis. Trends Biochem Sci, 35, 442-449 https://doi.org/10.1016/j.tibs.2010.03.004
  31. Panzhinskiy E, Ren J, Nair S (2013) Pharmacological inhibition of protein tyrosine phosphatase 1B: a promising strategy for the treatment of obesity and type 2 diabetes mellitus. Curr Med Chem, 20, 2609-2625 https://doi.org/10.2174/0929867311320210001
  32. van Huijsduijnen RH, Sauer WHB, Bombrun A, Swinnen D (2004) Prospects for inhibitors of protein tyrosine phosphatase 1B as antidiabetic drugs. J Med Chem, 247, 4142-4146
  33. Elchebly M, Payette P, Michaliszyn E, Cromlish W, Collins S, Loy AL, Normandin D, Cheng A, Himms-Hagen J, Chan CC, Ramachandran C, Gresser MJ, Tremblay ML, Kennedy BP (1999) Increased insulin sensitivity and obesity resistance in mice lacking the protein tyrosine phosphatase-1B gene. Sci, 283, 1544-1548 https://doi.org/10.1126/science.283.5407.1544
  34. Malamas MS, Sredy J, Gunawan I, Mihan B, Sawicki DR, Seestaller L, Sullivan D, Flam DR (2000) New azolidinediones as inhibitors of protein tyrosine phosphatase 1B with antihyperglycemic properties. J Med Chem, 43, 995-1010 https://doi.org/10.1021/jm990476x
  35. Choi CS (2009) Pathogenesis of insulin resistance. Korean J Med, 77, 171-177
  36. Winer DA, Luck H, Tsai S, Winer S (2016) The intestinal immune system in obesity and insulin resistance. Cell Metab, 23, 413-426 https://doi.org/10.1016/j.cmet.2016.01.003
  37. Boden G (2011) Obesity, insulin resistance and free fatty acids. Curr Opin Endocrinol Diabetes Obes, 18, 139-143 https://doi.org/10.1097/MED.0b013e3283444b09
  38. Li HB, Yang YRY, Mo ZJ, Ding Y, Jiang WJ (2015) Silibinin improves palmitate-induced insulin resistance in C2C12 myotubes by attenuating IRS-1/PI3K/Akt pathway inhibition. Braz J Med Biol Res, 48, 440-446 https://doi.org/10.1590/1414-431X20144238
  39. Pauli JR, Ropelle ER, Cintra DE, De Souza CT, da Silva AS, Moraes JC, Prada PO, de Almeida Leme JA, Luciano E, Velloso LA, Carvalheira JB, Saad MJ (2010) Acute exercise reverses aged-induced impairments in insulin signaling in rodent skeletal muscle. Mech Ageing Dev, 131, 323-329 https://doi.org/10.1016/j.mad.2010.03.004
  40. Gonzalez-Rodriguez A, Gutierrez JAM, Sanz-Gonzalez S, Ros M, Burks DJ, Valverde AM (2010) Inhibition of PTP1B restores IRS1-mediated hepatic insulin signaling in IRS2-deficient mice. Diabetes, 59, 588-599 https://doi.org/10.2337/db09-0796
  41. Zabolotny JM, Kim YB, Welsh LA, Kershaw EE, Neel BG, Kahn BB (2008) Protein-tyrosine phosphatase 1B expression is induced by inflammation in vivo. J Biol Chem, 283, 14230-14241 https://doi.org/10.1074/jbc.M800061200
  42. Zhao M, Zhang ZF, Ding Y, Wang JB, Li Y (2012) Astragalus polysaccharide improves palmitate-induced insulin resistance by inhibiting PTP1B and NF-kappaB in C2C12 myotubes. Molecules, 17, 7083-7092 https://doi.org/10.3390/molecules17067083
  43. Nieto-Vazquez I, Fernandez-Veledo S, de Alvaro C, Rondinone CM, Valverde AM, Lorenzo M (2007) Protein-tyrosine phosphatase 1B-deficient myocytes show increased insulin sensitivity and protection against tumor necrosis factor-${\alpha}$-induced insulin resistance. Diabetes, 56, 404-413 https://doi.org/10.2337/db06-0989
  44. Delibegovic M, Bence KK, Mody N, Hong EG, Ko HJ, Kim JK, Kahn BB, Neel BG (2007) Improved glucose homeostasis in mice with muscle-specific deletion of protein-tyrosine phosphatase 1B. Mol Cell Biol, 27, 7727-7734 https://doi.org/10.1128/MCB.00959-07
  45. Unger RH, Grundy S (1985) Hyperglycaemia as an inducer as well as a consequence of impaired islet cell function and insulin resistance: implications for the management of diabetes. Diabetologia, 28, 119-121