The Zuoguiwan Right The Pmop Renal Tissue Of Rats Tgf-¦Â1/smad4 Signal Transduction Pathway Of Experimental Research

Purpose:By exploring the regulative of the Zuogui pill on the signal transduction pathway of TGF-β1/Smad4 in the renal tissue of post-menopause osteoporosis rat.We discussed the mechanism of Zuogui pill in treatment of postmenopausal osteoporosis (PMOP).Material and method:Selecting totle seventy-one female SD rats aged 4 months,we simulated the rat model of PMOP by using ovariectomy. The rats were divided into 7 groups at random:normal group,sham-OVX group , ovariectomized group(OVX),Zuogui pill at high dosage, Zuogui pill at middle dosage,Zuogui pill at low dosage and Nilestr- iol-treated group.Taking nylestriol tab as the positive control drug,we observed the therapeutic effects of Zuogui pill on the rats with PMOP by 60-day gav- age.Collected 24h urine sample of rats before drawing materials. The next day all rats were killed to collect blood sample after drug withdrawal. We observ- ed the activity of serum alkaline phosphatase and tartrate-resistant acid p- hosphatase and the ratio changes of hydroxyproline and creatinine in urine by a colorimetric method.We carefully isolated rat right femur as a whole and determined the bone mineral density(BMD) by using dual energy X-ray absorp- tiometry.Taking the 1/3 quarters of the left distal femur near joints,we maked decalcified bone sectios for the measurement of bone tissue by using micro-image analysis system .Then we weighed the uterus wet weight to calculate the uterus index (the ratio of uterine wet weight and body weight), and observesed the changes in endometrium and glands under light microscope by routine HE staining. In order to study the mechanism of Zuogui pill in treatment of postmenopausal osteoporosis (PMOP), we taked the bilateral renal with asepsis and observed the pathological changes of that by means of optical microscope. Specimens from the right renal medulla is about 100g. We determined the gene expression level of renal tissue of TGF-β1 and Smad4 to observed the effects of Zuogui pill on the signal transduction pathway of TGF-β1/Smad4 in renal tissue of PMOP rats with reverse transcription polymerase chain reaction (RT-PCR).Results:1.Detection of uterine tissues:Compared with normal group, the uterine index in OVX decreased significantly (P<0.05).The Zuogui pill (high and middle and low-dosage) had slightly increased.But there was no obvious increasing trend compared with the that of OVX (P>0.05).The increase of uterus index in Niles- triol-treated group was significant(P<0.05).The OVX had metratrophia and sm- aller uterine cavity and thinner endometrial thinning than that of OVX under optical microscope.The endometrial thickness and gland hyperplasia is not ob- vious in Zuogui pill at high and middle and low dosage.But the number of gland increased in Nilestriol-treated group with little metratrophia.2.Testing of bone metabolic markers:①bone formation marker:Compared with normal group,the serum levels of BGP and ALP in OVX were obviously increased(P <0.05). Zuogui pill(high and middle and low-dosage) could decrease the con- tent of them ,with marked difference as compared with the OVX. But the trend of decreasing in Zuogui pill(high and middle-dosage)appeared obviously(P< 0.05).②bone resorption markers:the serum levels of StrACP in OVX were obv- iously increased(P<0.05),as well as the HOP/Cr of urine(P<0.05). The content of them in Zuogui pill at high and middle and low dosage group seemd lower than that in OVX,which was the most significant(P<0.05) in Zuogui pill at high d- osage.3.The pathological change of bone and measurement of bone tissue morphometry: Compared with normal group,the trabecular bone of OVX was thinner and rare- faction and less solid. But the number of trabecular bone in Zuogui pill at high and middle and low dosage had increased and the structure became closer and completer。We observed the measurement of bone tissue morphometric dete- ction under optical microscope.The percentage of trabecular area (Tb.Ar) in OVX significantly reduced(P<0.05),with that increasing obviously (P<0.05)in Z- uogui pill at high and middle dosage.There was no significant difference between Zuogui pill at high dosage and Nilestriol-treated group(P>0.05).4.Testing of Femur BMD:The BMD levels of the blank model group was sign- ificantly lowered (P<0.05)as compared to that of OVX.The BMD of proximal femur and distal femur in Zuogui pill(high and middle-dosage ) was significantly higher than OVX (P<0.05) .But there was no significant difference between the Zuogui pill(high and middle-dosage) and Nilestriol-treated group(P>0.05).5.The pathological examination of renal tissue:Compared with normal group,the OVX became no significant pathological changes such as sedimentary of collagen fiber.The structure of renal tissue was clear and there was no significant p- rogression of tubulointerstitial fibrosis in Zuogui pill( high and middle and low-dosage) and Nilestriol-treated group.6.The gene expression of TGF-β1 and Smad4 in renal tissue:Compared with the normal group, the gene expression of TGF-β1 and Smad4 of renal tissue in OVX had significantly increased(P<0.05). The Zuogui pill(high and middle and lo- wdosage) can all evidently down-regulate the level of the gene expression of TGF-β1 and Smad4 in renal tissue of PMOP rats(P<0.05),of which the effect was the best in Zuogui pill at low dosage.Conclusion:1.The animol model of PMOP was induced successfully after ovariectomy.2.Zuogui pill can effectively prevent and treat PMOP.3.Zuogui pill can evidently down-regulate the level of the gene expression of TGF-β1 and Smad4 in renal tissue of PMOP rats.4.Zuogui pill can obviously prevent and treat the rats with PMOP .The mechanism might be the intervenient effect of Zuogui pill on the pathway of TGF-β1/Smad4 in renal tissue.