Effects Of Glucose And Insulin In Different Concentrations On The Proliferation And Activation Of HSC In Rat

Objective:To investigate the mechanism and provide better clinical treatment Program for hepatic fibrosis caused by diabetes, the effects of glucose and insulin in different concentrations on the proliferation and the expression of p65 protein in rat HSC were observed. Methods:Hepatic stellate cell strain were cultured with different concentration insulin, high glucose alone or high glucose added insulin in different concentration, while cultured with the glucose in normal concentration(5.6mmol/L) and mannitol (25mmol/L) as the normal control and the hyperosmosis control,respectivey. Groups as follows:A: normal glucose (5.6mmol/L), B:high glucose (25 mmol/L), C:normal glucose (5.6mmol/L)+insulin group (10-6 mmol/L), D:High glucose (25 mmol/L)+insulin group (10-6 mmol/L), E:normal glucose (5.6mmol/L)+insulin group (10-8 mmol/L), F:High glucose (25 mmol /L)+insulin group (10-8 mmol/L), G:normal glucose (5.6mmol/L)+ insulin group (10-9 mmol/L), H:High glucose (25 mmol/L)+insulin group (10-9 mmol/L), I:mannitol (25mmol/L).①All of the rat HSC were cultured for 24,48,72 hours, the proliferation of HSC and the growth curve were detected by Cell Counting Kit-8 (CCK-8).②The p65 expression in rat HSC cultured for 24hours were observed by Western blot. Results:①There were no significant differences in the absorbance value between normal glucose(A) group and other groups cultured for 24 hours (P>0.05). After cultured for 48 hours, the increased absorbance value in D and F group and the significant differences were found while compared it with that in A group(P<0.05), but no significant differences were discovered while compared the absorbance value in the B, C, E, G, H and I group with it in A group(P>0.05). After cultured for 72 hours, the eleavated absorbance value in F,G,H and I group were found, and the significant differences were observed while compared it with that in A group (P<0.05), no significant differences were found while compared the absorbance value between B, C, D, E group (P>0.05).②After cultured for 24 hour, the expression(INT ratio) of NF-KBp65 in all groups, showed by western blot, were:A:0.549±0.044;B:0.893±0.044; C:1.360±0.038; D:0.810±0.044; E:1.036±0.053; F:0.613±0.040; G:0.840±0.055; H:0.846±0.060; I:0.572±0.032. There were obvious significant differences in the INT ratio between normal group and other groups (P<0.01). The expression of p65 protein was significantly increased in C,E,G group (P<0.01), but no significantly different were found compared them with that in B group (P>0.05). It is that the expression of p65 protein was increase when the concentration of insulin increased, and the positive correlation between the expression of p65 protein and the concentration of insulin were obsvered. (P<0.01), but no significant differences were found in the expression of p65 protein between A group and I group(P>0.05).Conclusions:1. Insulin, high glucose and insulin add high glucose could induce the activation of rat HSC in vitro, increase the p65 protein expression, but the number of HSC proliferation was not obvious.2. NF-κB signaling pathway may involve in HSC activation.