Alloxan에 의한 HIT-T15 세포 손상에 대한 쑥갓주정추출물의 세포보호효과

Abstract

본 연구는 췌장베타세포인 HIT-T15 세포를 이용하여 쑥갓주정추출물(CSE)의 alloxan에 의한 산화스트레스로부터의 세포보호, 인슐린 분비능 및 항산화 효소 활성을 평가하였다. CSE 는 alloxan에 의해 유발된 산화스트레스로부터 세포를 보호하여 세포생존율을 증가시켰다. CSE는 세포막 손상지표인 LDH 방출을 감소시켰고 NAD+/NADH ratio를 유의적으로 증가시켜 세포사멸을 억제하였다. 또한 alloxan 단독처리군에 비해100 ㎍/㎖ CSE 처리농도에서 인슐린 분비량이 유의적으로 증가하였다. CSE 처리는 HIT-T15 세포 내 항산화효소 활성을유의적으로 상승시켰다. 이상의 연구결과로부터 CSE는 alloxan 에 의해 유발된 산화스트레스로부터 췌장베타세포의 항산화효소 활성을 증가시킴으로써 세포괴사 및 DNA fragmentation 을 억제하여 세포사멸을 억제하고 생존률을 증가시키고, 이에 따라 인슐린 분비능 조절에도 영향을 미치는 것으로 생각된다.

The objective of the present study was to evaluate the potential antidiabetic and antioxidant effect of the ethanol extract from Chrysanthemum cornarium L. var. spatiosum(CSE) against alloxan-induced oxidative stress in pancreatic β-cells,HIT-T15. In this study, the antidiabetic effect of CSE was examined using the 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazoliu bromide(MTT) cell proliferation assay, lactate dehydrogenase(LDH) release assay, NAD+/NADH ratio and insulin secretion. To further investigate whether CSE is involved in the antioxidant activity of alloxan-damaged HIT-T15 cells, its antioxidant effect against alloxan-induced oxidative stress was measured in HIT-T15 cells by determining the levels of antioxidant enzymes including superoxide dismutase(SOD), glutathione S-transferase(GST), glutathione reductase(GR) and glutathione peroxidase(GPx). The results of this analysis showed that alloxan significantly decreased cell viability, increased LDH leakage, and lowered NAD+/NADH ratio and insulin secretion in HIT-T15 cells. However, CSE significantly increased the viability of alloxan-treated cells and lowered LDH leakage. The intracellular NAD+/NADH ratio and insulin secretion were also significantly increased by 1.7-fold and 1.3-fold, respectively, after treatment with 100 ㎍/㎖ CSE. The HIT-T15 cells treated with alloxan showed significant decreases in the activities of antioxidant enzymes, while CSE significantly elevated the levels of antioxidant enzymes. These findings suggest that CSE could have a protective effect against cytotoxicity and dysfunction of pancreatic cells in the presence of alloxan-induced oxidative stress.