Experimental Study On Gal-3 Induced IL-6 Expression Of Endothelial Cells Through ERK1/2 Pathway Promotes The Formation Of DVT

Objective1、To investigate the effect of the recombinant human galectin3 (rGal3) and PD98059 of the ERK1/2 signaling pathway inhibitor on ERK1/2 and down stream inflammatory genes expression of HUVECs and then analysis the relationship between these genes with Gal3.2、To study the effect of rGal-3 and PD98059 on the formation of thrombus length, wet weight and the expression of IL-6 mRNA in the vein wall of mouse DVT model, then analysis the relationship between Gal3 and the ERK1/2 signaling pathway with IL-6 in the DVT model.Methods:1、Cultured HUVECs, divided control group, rGal-3 group, PD98059 group, rGal-3 + PD98059 group; rGal-3 group use rGal-3 to 1μg/ml treated HUVECs 24 hours; PD98059 group with PD98059 to 50μM treated HUVECs 2 hours; rGal -3 + PD98059 treated with 50μM PD98059 to HUVECs 2 hours then use rGal-3 with 1μg/ ml treatment 24 hours; Western-blot to detect the ERK1/2, p-ERK1/2, IL-6 expression; statistical analysis of each group ERK1/2, p-ERK1/2, IL-6 expression changes, then research the relationship between galectin-3 and ERK1/2 with the vein endothelial cells of IL-6 expression.2、90 C57 mices were randomly divided into blank group (n=20), DVT control group (n=25), rGal-3 DVT group(n=15), PD98059 DVT group(n=15), rGal-3 + PD98059 DVT group(n=15). Twenty-four hours before DVT group was built by stenosis inferior vena cava DVT mice model, rGal-3 DVT group mices were given intravenous injection rGal-3 (5μ,g/each) and given intraperitoneal injection of a dose corresponding 10% dimethyl sulfoxide (10% DMSO) with PD98059 DVT group. PD98059 DVT group mices were given intraperitoneal injection of ERK1/2 inhibitor PD98059 (10mg/kg body weight) and given intravenous injection of a corresponding dose of saline with rGal-3 DVT group. rGal-3 + PD98059 DVT mices were given intravenous injection rGal-3 (5μg/each) and intraperitoneal injection of ERK1/2 inhibitor PD98059 (10mg/kg body weight), blank group and DVT control group mices were given intravenous injection of a corresponding dose of saline and intraperitoneal injection of a dose corresponding 10% dimethyl sulfoxide (10% DMSO). And harvested inferior vena cava venous tissue 24 hours after stenosis, observed the DVT formation by HE dyeing, we detected gal-3 mRNA expression for the venous tissue of normal and DVT control group by real-time PCR, and test IL-6 mRNA expression of each group vein wall tissue. Analysis the relationship between IL-6 with Gal3 and ERK1/2, then explore its relationship with the DVT.Results:1、The cell experiment, rGal-3 pretreated HUVECs with 1μg/ml for 24 hours, ERK1/2、p-ERK1/2、IL-6 expression up-regulate compare with control gourp (P<0.01). PD98059 pretreated HUVECs with 50μM for 2 hours, ERK1/2、 p-ERK1/2、IL-6 expression down-regulate compare with control gourp(P<0.01). The ERK1/2、p-ERK1/2、IL-6 expression in rGal-3+PD98059 group was down-regulate compare with rGal-3 group(P<0.01).2、In the animal experiment,the rGal-3 + PD98059 DVT group motality rate 6.67%, the other group was 0%. The thrombosis rate of blank group、DVT control group、rGal-3 DVT group、PD98059 DVT group、rGal-3 + PD98059 DVT group respectively 0%,72.0%,80.0%,66.7%,71.4%. The thrombus length and weight in rGal-3 DVT group was significantly higher compare with the DVT control group (P< 0.05); The thrombus length and weight in rGal-3+PD98059 DVT group was significantly lower compare with the rGal-3 DVT group (P< 0.05)。Real-time PCR detected mice venous tissue gal3 mRNA expression in blank group and DVT control group, gal3 mRNA expression were higher in DVT control group compare with blank group (P<0.05).Real-time PCR test IL-6 mRNA expression of blank group and PD98059 DVT group were lower than the DVT control group(P<0.05); The IL-6 mRNA expression of rGal-3 DVT group were higher than the DVT control group(P<0.05); The IL-6 mRNA expression of rGal-3+ PD98059 DVT group were lower than the rGal-3 DVT group(P<0.05); The IL-6 mRNA expression of rGal-3+ PD98059 DVT group were higher than the PD98059 DVT group(P<0.05);Conclusions:1、In vitro experiments, Gal-3 may regulate IL-6 expression through ERK1/2 signaling pathway in endothelial cells;2、In the DVT model of C57 mice experiments, Gal-3 may regulate IL-6 expression through ERK1/2 signaling pathway in endothelial cells,and promote the formation of DVT.