Effects Of Octreotide On The Proliferation,apoptosis And Expressions Of STAT3、p-STAT3 And SOCS3 In Human Hepatic Stellate Cells

Hepatic fibrosis is a compensative reaction caused by a variety of chronic liver injury, as a necessary stage of liver cirrhosis. A number of studies have shown that liver fibrosis is reversible after removing damage factors[1]. Hepatic stellate cells are the main cells which produce extracellular matrix, and the key cells during the development of liver fibrosis. Janus kinase/signal transducer and activator of transcription-3(JAK/STAT3) signaling pathway plays a role in promoting proliferation and inhibiting apoptosis of cells. SOCS3 as a negative regulator of JAK/STAT3 signaling pathway, participates in human malignant proliferative disease. Our previous study reported, octreotide could inhibit the proliferation and promote apoptosis of hepatic stellate cells, and finally inhibit the progress of liver fibrosis. Whether the effects of octreotide on human liver fibrosis related to JAK/STAT3 signal pathway and negatively regulating factor SOCS3 is still unclear.Part1 Effects of different concentrations of octreotide on proliferation and apoptosis of LX-2 cellsAbstrat Objective: To study the effects of octreotide on the proliferation and apoptosis of STAT3 and SOCS3 in human hepatic stellate cells. Methods: Human hepatic stellate cell line LX-2 were cultured in vitro, LX-2 were identified by immunocytochemistry of α-SMA,Desmin protein, and then using the different concentrations of octreotide(10-3, 10-4, 10-5, 10-6mmol/L) intervene LX-2 for 24 h, 48 h, 72 h.Cell proliferation were detected by MTT,cell apoptosis were detected by TUNEL fluorescence method. Results:(1) α-SMA and Desmin protein werefound in cytoplasm of LX-2 by immunohistochemistry.(2)Octreotide could inhibit the proliferation of LX-2 in a dose- and time-depedent manner, and the inhibitory effect was higher in 72 h 10-3mmol/L group than in other groups.(3)Apoptosis rates for 24 h or 48 h in OCT groups were significantly higher than in control groups,and apoptosis rates in high concentration of octreotide groups were higer than in low concentration groups and the 48 h groups were markedly higher than that of 24 h groups. Conclusion:(1) LX-2 cells cultured in vitro are activated human hepatic stellate cells.(2) Octreotide may inhibit the proliferation of LX-2 and promote its apoptosis in a dose and time dependent manner.Part 2 Effects of octreotide on the expression of STAT3、p-STAT3、SOCS3 in LX-2 cells.Abstrat Objective: To investigate the effects of octreotide on the expressions of STAT3、p-STAT3 and SOCS3 in human hepatic stellate cells, and further explore the anti-fibrosis mechanism of octreotide. Methods: STAT3 m RNA and SOCS3 m RNA were detected by RT-PCR,STAT3 、 SOCS3 and phosphorylated STAT3(P-STAT3) were analyzed by immunocytochemistry and Western-blot. Results:(1)LX-2 were incubated with the different concentration of octreotide for 24h、48h、72h.The immunocytochemistry indicated that the expression of STAT3 protein and SOCS3 protein were significantly lower in OCT groups than in control groups. At the same time, the expression of STAT3 protein increased with the increased concentration of octreotide, but were lower than that in control group(P<0.05); the most expression of SOCS3 protein was in the 10-4 or 10-5mmol/L group, and were all lower than that in control group(P<0.05). With the same concentration, the expression of STAT3 protein had no significant changable when time pass by, while theexpression of SOCS3 gradually decreased(P<0.05).(2) The RT-PCR indicated that the expression of STAT3 m RNA gradually increased with the increased concentration of octreotide at the same time, and the expression of STAT3 m RNA were lower than that in control groups(P<0.05);with the same concentration, the expression of STAT3 m RNA were not significant difference(P>0.05). At the same time, the expression of SOCS3 m RNA increased with the drug concentration increasing, and were lower than that in control group(P<0.05); with the same drug concentration, the expression of SOCS3 m RNA decreased with the increasing incubation time(P<0.05).(3) The Western-blot indicated that the expression of STAT3 protein and SOCS3 protein in OCT groups were apparently decreased, compared with the control groups. There were significant difference in goups(P<0.05). The results indicated that octreotide could down-regulation the expression of STAT3 protein and it’s negative regulator factor SOCS3 protein, and they might be associated with the mechanism of octreotide inhibiting liver fibrosis. Conclusion:(1) Octreotide can inhibit the expression of STAT3 、p-STAT3 and SOCS3 in LX-2.(2) The JAK/STAT3 pathway and its negative regulatory factor SOCS3 may be involved in the mechanism that octreotide inhibit the proliferation and promote the apoptosis of LX-2, and it may be associated with down-regulation of STAT3 expression.