The Effect And Mechanism Of Leflunomide On Rats With Endometriosis

Objective: In this study, we use autologous transplantation to establishrats models with endometriosis. Observ the entire process of leflunomide onendometriosis (Endometriosis, EMT) in rat model; detect transcription proteinkinase (Transcription, p-STAT) and suppressors of cytokine signaling (cellsignaling inhibitory factor the expression of mRNA, SOCS) on rats modelswith endometriosis. To explore the effect and mechanism of leflunomide onthe development of an experimental endometriosis model. Preliminary discussthe mechanism of leflunomide for endometriosis, and provide new strategiesand targets and experimental basis for the clinical treatment of endometriosis.Methods:1SD of rat endometriosis model9weeks, sexual, maturity, unpregnant female, health SD rats, weight(200±80)g, provided by the hebei province laboratory animal center(qualifiednumber:1011030). Forty SD rats was assessed by surgically transplantingautologous fragments of endometrial tissue onto the inner surface of theabdominal wall and arterial cascades of the small intestines. SD rats wereirrigated benzoic acid estradiol by200mg/kg for five days. Through thisMethod, all rats are going in emotive issue. After receiving anesthesia withconcentration for43g/L of the first chlorine aldehyde300mg/kg byintraperitoneal injection, skins are disinfected in povidone-iodine. Thenstomach had been cut2cm long incision on the open about1cm place. Openlayered abdominal wall of each layer. Separate into the right uterine of the rat,and ligate1.5cm long right uterus since the distal0.5cm place to proximaldirection. Put into the physiological saline, and then rapidly cut longitudinalincision in uterine cavity. Cut5mm x5mm long two pieces and seamrespectively in left and right side of abdominal wall. Make its lining surface facing abdominal wall. Postoperative inject continuously in muscle withgentamycin sulfate0.1ml/d for preventing infection, total five days. Afterpostoperative5days, rats were irrigated benzoic acid estradiol50mg/kg forfive days. To promote the transplant lining growth and increase film rate.2Target detection methods2.1Inspect endometriosis kitchen volume change by Measuring thevolume of ectopic endometrial V2, compared with volume V1(According tothe formula (volume V=0.52xLength cm x Width cm x High cm)2.2Through the application of Western blot method, we can inspectendometriosis foci phosphorylation p-STAT protein.2.3Through the RT-PCR method, we can inspect expression of cellsignaling inhibitory factor (SOCS)-1observation mRNA in ectopic tissueResults:1experimental animal in general:A total of32rats model, confirmed by pathological examination of thegraft showed endometrial epithelial cells, glandular and stromal growth shownin Figure1b, modeling the success of ectopic foci specimens are shown inFigure1a. In medicine, the control group died one, because of incisioninfection; treatment group die two, death due to intestinal obstruction infection.The rest did not form typical of ectopic foci were eliminated. The filmforming rate was80%(32/40).2Effects of leflunomide on rat with endometriosisBefore treatment, ectopic endometrial volume of each group were nosignificant difference (F=3.78, P>0.05), after treatment, ectopic endometrialvolume of each drug group in varying degrees reduce. Endometrial Vesiclefluid diminishes or even disappears. Blank control group bulk increasesapparently, abundant blood vessels; lesion volume of leflunomide in high doseand low dose group and danazol group rat with endometriosis reduced, lesionvolume of four groups change before and after treatment. Through acomparative analysis of each group before and after treatment, there is adifference between the difference volume (F=25.689,P<0.001). Comparison groups, leflunomide in high dose group were significantly reduced in size, andsuccessfully suppressed the growth of ectopic foci. Between high dose groupand danazol group, there are no significant differences. The effect of high dosegroup and danazol group is no differences, but leflunomide is without effecton sex hormone axis, and does not affect the pregnancy; thus better thandanazol group.3p-STAT1protein expression (Western blot):The control group and treatment group in molecular weight between10and50kDa are positive black strip, on behalf of the p-STAT protein. Blankcontrol group p-STAT protein expression increased significantly (1.060+0.028) other groups decreased. Through a single factor analysis of variance,the groups were no differences (F=131.041,P<0.001). Through Group two twocompared analysis, p-STAT1protein expression decreased obviously betweeneach drug group and blank control group to; and the difference wasstatistically significant (P<0.05). Leflunomide inhibits the expression ofp-STAT-1protein. Leflunomide in the high dose group is the most significanton the inhibitory effects of protein expression; between leflunomide in lowdose group and danazol group there wre no differences on p-STAT proteininhibition.4SOCSmRNA expressions:Expression of SOCSmRNA significantly elevated on blank control group(0.656+0.017); other groups decreased. Leflunomide in high dose and lowdose group were (0.264+0.016) and (0.552+0.034), danazol group (0.444+0.023). Through a single factor analysis of variance, difference has statisticssignificance between the groups(F=404.523,P<0.001). Through Group twotwo compared analysis, the expression of SOCSmRNA decreased significantlybetween each drug group and control group; the difference was statisticallysignificant(P<0.5); the treatment group can inhibit SOCSmRNA expression,but effect of eflunomide in high dose group is the most obvious. Conclusion:1The method was feasibility that establish a rat model of endometriosis withautologous uterine transplantation on rat unestrus2Leflunomide in high dose (35mg/kg) can significantly inhibit the expressionof SOCSmRNA and p-STAT protein in ectopic endometrium on rats withendometriosis, and can significantly inhibit the growth of ectopic foci; canheal endometriosis. Leflunomide in10mg/kg was without obvious inhibitoryeffect on rats with endometriosis. Leflunomide in35mg/kg is the besttherapeutic.3endometrium ectopic foci of growth was inhibited on leflunomide group.Leflunomide can inhibit the expression of p-STAT1and prevents the excessactivation of JAK/STAT1signal transduction pathway, which may be one ofthe mechanisms for the therapeutic effects in the experimental endometriosismodel.