Study Of The Effect Of SQW On Bladder Dyfunction In DCP Mice

ObjectiveEstablish DCP model to observe the influence of SQW on urination,glucose metabolism states,the tissue structures and function of bladder.And using modern molecular biology technology to study the effect of SQW on the expression of SP,NK1R,P2X2,P2X3 and CHRM3,?3-AR,TRPV1 mRNA,to explore the mechanism of SQW improves bladder dyfunction of DCP mice.Method1.Study aimed on obtain DCP model mice,and to observe the effect of SQW on the urination and glucose metabolism states.Sixty male C57BL/6 mice of SPF level were randomly assigned into normal group(n=8)and modle group.During the whole,normal group mice were fed with general feed and model group mice were fed with high fat feed.After feed for a month,the model group mice were intraperitoneal injection of 2%STZ with dosage of 200mg/kg.Fasting blood glucose was tested after 3 days,mice with fasting blood glucose level ?189mg/dl were considered as model of type 2 diabetes.Modeled mice were allocated into model group,mecobalamin group,high dose of SQW group(SQWH)and low dose of SQW group(SQWL),each group contained 8 mice.Eight weeks after streptozotocin treatment,the model mice were treated with distilled water,mecobalamin or SQW four weeks.During the administration period,changes of weight and fasting blood glucose,amount of feed consumed,water consumed and urine excreted were tested every week.Oral glucose tolerance,plasma glucosylated serum protein and plasma insulin were tested after administration.2.To observe the influence of SQW on bladder function and tissue structures on DCP mice.After administration,all mice were assessed by urodynamic.Then animals were sacrificed,bladder smooth muscle strips were isolated steritily,using powerlab instruments to record the contractile response to ?,?-me-ATP,capsaicin,carbachol and KCl.For histopathology,bladder was fixed in 4%paraformaldehyde solution,disposed by routine procedure,and embedded in paraffin.The processed bladder tissue was sectioned at 5?m.Sections were stained with haematoxylin and eosin(HE)and Masson' s trichrome,and examined under light microscope.3.Study of the effect of SQW on related neuroreceptor and neurotransmitter on DCP mice.Real-Time PCR was performed to quantify the expressions of CHRM3,?3-AR,TRPV1 mRNA,and immunofluorescence was performed to quantify and localize the expressions of SP,NK1R,P2X2 and P2X3,to illuminate the mechanism of SQW improves bladder function of DCP mice.Results1.Results of the urination and glucose metabolism states.Mice with DCP had significantly greater water consumption,feed consumption,urine production,plasma GSP level and plasma insulin level,and had significantly lighter weight than control mice.SQW treated have no effect on water consumption,feed consumption,fasting blood glucose,plasma GSP level and plasma insulin level,SQWH can reduce the excretion of urine.2.Results of urodynamic study,in vitro experiment,and histopathology.DCP caused significant changes urodynamically.Mice with DCP had significantly greater Vv,RV,MBC and BC than control mice.DCP caused a decrease in BLPP,MP and VE compared to controls.The BLPP and MP increased significantly after SQW treatment and SQW can significantly decreased Vv,RV,MBC and BC in DCP mice,but VE was no different after treatment with SQW.Comparing with control group,the ATP emission and the maximum effects of ?,?-me-ATP,capsaicin and KCl were significantly decreased in model group.In contrast,the ATP emission and the bladder responses to ?,?-me-ATP,capsaicin and KCl were significantly greater in SQWH compared with model group.Similarly,SQWL was significantly increased the ATP emission and the responses to ?,?-me-ATP and capsaicin in type 2 diabetic mice bladder,but no difference was observed in contractile response to KClThe morphometric analysis in bladder tissue from C57BL/6 mice revealed a decreased in wall thickness and collagen density in DCP mice,SQW can improve this changes.And smooth muscle density were not significantly modified in model group mice compared with bladder tissue from control mice.Disordered muscle bundle,myocyte hypertrophy,loosen collagen and elastin appeared in model group mice,after SQW treatment the condition of bladder was improved.3.Results of the expression of related neuroreceptor and neurotransmitter.RT-PCR and immunofluorescence experiment demonstrated that the expression of CHRM3,TRPV1,SP,NKIR,P2X2 and P2X3 was significantly decreased,SQW can increase the expression of CHRM3,TRPV1,SP,NK1R,P2X2 and P2X3.ConclusionSQW can improve the bladder function of DCP mice by myogenic and neurogenic mechanism.And it was suggested to be related with the regulation on the levels of CHRM3,TRPV1,NK1R and P2X3 receptor of bladder tissue,stimulating bladder to release ATP and SP,improve the symptoms of diabetic bladder dysfunction.