The Effect Of Berberine On Insulin Secretion And Autophagy In NIT-1Cells Induced By High Glucose And Palmitic Acid

Section Ⅰ The effect of Berberine on insulin secretion in NIT-1cells induced by high glucose and palmitic acidObjectiveTo investigate the effect of berberine on insulin secretion of NIT-1cell induced by high glucose and palmitic acid.Methods3-(4,5)-dimethylthiahiazo(-z-yl)-3,5-di-phenytetrazoliumromide (MTT) colorimetric assay was used to examine cell growth and determine the proper concentrations of berberine in ordinary, high glucose (25mmol/L) and palmitic acid (PA,0.25mmol/L) contained culture medium. Insulin secretion of NIT-1cells was induced by adding high glucose or palmitic acid into the cultured medium. The acute and chronic effects of berberine on insulin secretion of NIT-1cell were examined by radioimmunoassay.ResultsIn ordinary culture medium, berberine at the concentration of100μmol/L significantly inhibited the proliferation of NIT-1cells (P<0.05). However, in palmitic acid contained cultured medium, berberine inhibited the proliferation of NIT-1cells just at the concentration of30μmol/L and in high glucose contained cultured medium, berberine inhibited the proliferation of NIT-1cells at100μmol/L (P<0.05). Berberine, at the concentration of3μmol/L,10μmo/L and30μmol/L, decreased the basic, glucose-stimulated insulin secretion (GSIS) and palmate-stimulated insulin secretion (PSIS)(P<0.05). Furthermore, berberine, at the concentration of lOμmol/L, acutely (at1hour incubation) inhibited but chronically (at24hours incubation) increased insulin secretion from NIT-1cells in high glucose and palmitic acid contained culture medium (P<0.01,P<0.05).ConclusionIt is suggested that the short time incubation with berberine might inhibit but the sustained incubation with berberine could increase insulin secretion from NIT-1cells in high glucose and palmitic acid contained culture medium. Section II The Effect of Berberine on Autophagy in NIT-1Cells Induced by High Glucose and Palmitic AcidObjectiveTo investigate the effect of berberine on autophagy of NIT-1cells induced by high glucose and palmitic acid.MethodsThe autophagy of NIT-1cells was induced by adding high glucose (Glu) or palmitic acid (PA) into the cultured medium. Then the cells were divided into control group, model group, berberine group,5-Aminoimidazole-4-carboxamide1-β-D-ribofuranoside (AICAR) group, Compound C group, and rapamycin (RAPA) group. The cultured cells in different groups were administered with corresponding intervention. The expressions of autophagy-associated proteins such as AMP-activated protein kinase (AMPK), tuberous sclerosis complex1(TSC1) and microtubule-associated protein1light chain3(LC3-II) in NIT-1cells were examined by Western blot analysis. The number of autophagosomes was observed by transmission electron microscopy.ResultsCompared with control group, the expression AMPK protein decreased significantly in NIT-1cells incubated with high glucose or PA while the expression of TSC1and LC3-II proteins increased (P<0.01,P<0.05). However, in comparison with model group the addition of berberine, AICAR and RAP A increased the expression of AMPK, TSC1and LC3-II proteins in NIT-1cells (P<0.01,P<0.05). Compared with model group, the addition of compound C decreased the expression of AMPK, TSC1and LC3-Ⅱ protein (P<0.01, P<0.05). The appearance of autophagic vesicles were examined under a transmission electron microscope. Compared with control group, Autophagosomes were stimultaneously observed in NIT-1cells incubated with high glucose or PA (P<0.05). Compared with model group, the addition of berberine stimulated the formation of autophagosomes (P<0.05).ConclusionIt is suggested that berberine may increase autophagy in NIT-1cells incubated with high glucose or palmitic acid. The mechanism might be associated with increasing the expression of AMPK, TSC1and LC3-Ⅱ proteins in NIT-1cells.