枸杞多糖主要组分甘露糖及其潜在靶标代谢物肌醇对小鼠胰岛β-TC6细胞的影响

Effects of the mannose,the major component of Lycium barbarum polysaccharides,and its potential target metabolite inositol on mouseβ-TC6 cells

目的初步探讨枸杞多糖主要组分甘露糖及其潜在靶标代谢物肌醇对小鼠胰岛β-TC6细胞的影响。方法以不同浓度(0、4.6875、9.375、18.75、37.5、75和150μg/mL)甘露糖或肌醇干预β-TC6细胞24 h,CCK-8(cell counting kit-8)法测定细胞增殖活性;20 mmol/L葡萄糖联合不同浓度(0、18.75、75和150μg/mL)甘露糖或肌醇干预β-TC6细胞60 min,酶联免疫法检测细胞胰岛素分泌量;设立吡格列酮阳性对照组(3.92 mg/L),经(0、9.375、18.75、75和150μg/mL)甘露糖或肌醇干预24 h后,应用实时荧光定量PCR法检测β-TC6细胞胰岛素、葡萄糖激酶(glucose kinase,GK)、葡萄糖转运蛋白4(glucose transporter 4,GLUT4)以及糖原合成酶(glycogen synthase,GS)mRNA表达水平。结果与空白对照组相比,4.6875~150μg/mL甘露糖和18.75~150μg/mL肌醇均呈浓度依赖性促进β-TC6细胞增殖(P<0.05),4.6875和9.375μg/mL肌醇虽然有促进细胞增殖的趋势,但差异无统计学意义(P>0.05)。在20 mmol/L葡萄糖刺激下,18.75、75和150μg/mL甘露糖或肌醇干预后,β-TC6细胞胰岛素分泌量与对照组相比差异均无统计学意义(P>0.05)。与对照组相比,150μg/mL甘露糖组GLUT4 mRNA表达上调(P<0.01),75μg/mL肌醇组GK和GLUT4基因表达水平都上调(P<0.01),18.75μg/mL肌醇仅使GLUT4基因表达上调(P<0.01)。结论甘露糖和肌醇可通过改善GLUT4 mRNA表达水平,从而提高外周细胞对葡萄糖的摄取;此外,肌醇还可以通过上调GK mRNA表达水平以提高胰岛素敏感性,改善葡萄糖代谢。

OBJECTIVE To investigate the effects of mannose,the major component of Lycium barbarum polysaccharide,and its potential target metabolite,inositol,on mouse isletβ-TC6 cells.METHODS Different concentrations(0,4.6875,9.375,18.75,37.5,75 and 150μg/mL)of mannose or inositol were used to interveneβ-TC6 cells for 24 hours,and the proliferation activity of cells was determined by CCK-8 method.Enzyme-linked immunosorbent assay was used to detect insulin secretion after the intervention of theβ-TC6 cells from different concentration of the mannose or inositol(0,18.75,75 and 150μg/mL)combining with glucose stimulation(20 mmol/L)for 60 minutes.Pioglitazone(3.92 mg/L)was set up as positive group,and after intervention of the mannose or inositol(0,9.375,18.75,75 and 150μg/mL)for 24 h,the expression levels of insulin,glucose kinase(GK),glucose transporter 4(GLUT4)and glycogen synthase(GS)mRNA were detected by real-time quantitative PCR.RESULTS Compared with the control group,mannose and inositol promoted the proliferation ofβ-TC6 cells in a concentration-dependent manner(18.75-150μg/mL)(P<0.05).Although the inositol solution of 4.6875μg/mL and 9.375μg/mL had a tendency to promote cell proliferation,there was no statistical difference(P>0.05).After stimulation with 20 mmol/L glucose combining with different intervention concentrations(18.75,75 and 150μg/mL)of mannose or inositol,no significant difference was observed in the insulin secretion of each group(P>0.05)comparing with the control group.RT-qPCR result showed that 150μg/mL mannose increased the expression level of GLUT4(P<0.01)and the expression levels of GK and GLUT4 genes in the 75μg/mL inositol group were significantly increased(P<0.01).The expression level of GLUT4 was improved only when the concentration was decreased to 18.75μg/mL in inositol group(P<0.01).CONCLUSION Mannose and inositol can improve the expression of GLUT4 mRNA,which may help to increase glucose uptake by peripheral cells.In addition,inositol can improve insulin sensitivity and glucose metabolism by increasing the expression level of GK mRNA.