The Mechanism Study Of Fushen Recipe Inhibiting Peritoneal Dialysis-related Peritoneal Angiogenesis Through PI3K/Akt/mTOR Pathway

ObjectiveIn this study,by observing the effect of Fushen recipe on peritoneal dialysis-related peritoneal angiogenesis in uremic rats and its possible mechanism,it is clear that Fushen recipe has the effect of inhibiting peritoneal angiogenesis and provides new treatment ideas for clinical prevention and treatment of ultrafiltration failure.MethodsFifty male SD rats were divided into sham operation group,uremic dialysis group,Fushen recipe low-dose group,Fushen recipe high-dose group and positive drug control group(n=10).A rat model of uremia was constructed by 5/6 nephrectomy,and the model was identified in the 4th week after the operation: the morphological changes of the kidney were observed by HE staining method,and the serum creatinine and urea nitrogen levels were measured to evaluate the renal function.On this basis,the intraperitoneal injection of 4.25%glucose peritoneal dialysis solution was used to establish a rat uremic peritoneal dialysis model.At the same time,different doses of Fushen recipe or western medicine celecoxib were given for drug intervention for a total of 4 weeks.After 4 weeks,the uremic peritoneal dialysis model and the efficacy of the Fushen recipe were evaluated: the peritoneal morphology was detected by HE and Masson staining;the peritoneal function was tested by the 2h peritoneal balance experiment to calculate the ultrafiltration volume,glucose absorption rate and creatinine Transport level;peritoneal angiogenesis.Immunohistochemistry was used to detect the expression levels of CD105,HIF-1α,and VEGF in peritoneal tissue.CD105 was used to calculate the number of new blood vessels,and HIF-1α and VEGF were used to assess the promotion of angiogenesis in peritoneal tissue.The protein expression level of factors;RT-PCR technology detects the promotion and inhibition of angiogenesis-related factors VEGF,HIF-1α,COX-2,IL-1β,IL-6,MMP-9,TIMP-1,IL-12 in peritoneal tissue The level of gene expression.After clarifying the efficacy of Fushen recipe,the possible mechanism of its inhibitory effect on angiogenesis was explored: the gene expression levels of PI3K/Akt/mTOR pathway related factors PI3 K,Akt,mTOR were detected by RT-PCR technology,and the immunohistochemical method was used to detect The protein expression levels of PI3 K,p-Akt,and p-mTOR in peritoneal tissues.Results1 Uremia model identification:(1)HE staining of residual kidney tissue: the number of glomeruli in the uremic group decreased,the mesangial cells and matrix components in the spheroid increased,the renal tubules were dilated with casts,and the renal interstitial inflammatory cell infiltration and fibrosis Coexistence;(2)Renal function test: Compared with the sham operation group,the serum creatinine and urea nitrogen levels in the uremic group increased significantly(P<0.01).Peritoneal dialysis model identification:(1)Parietal peritoneum HE and Masson staining: The thickness of the peritoneum in the model group was significantly increased compared with the sham operation group(P<0.01),and inflammatory cell infiltration,fibrosis and angiogenesis were seen in the submesothelial area;(2)Peritoneal function Detection: Compared with the sham operation group,the ultrafiltration volume and DGlu/D0 Glu of the model group were significantly decreased,and the DCr/PCr was significantly increased(P<0.05,P<0.01).2 Observation of the efficacy of Fushen Recipe:(1)Parietal peritoneum HE and Masson staining: Compared with the model group,the peritoneal thickness of each drug intervention group was significantly reduced(P<0.01),and related pathological changes were also significantly improved;(2)Peritoneal function detection: Compared with the model group,the ultrafiltration volume and DGlu/D0 Glu of each drug intervention group were significantly increased(P<0.05);(3)Immunohistochemical staining: Compared with the model group,HIF-1α,HIF-1α,DGlu/D0 Glu in the peritoneal tissue of each drug intervention group The positive expression of VEGF and the number of new blood vessels were significantly reduced(P<0.01);(4)RT-PCR results: Compared with the model group,the m RNA expressions of pro-angiogenesis factors VEGF,HIF-1α,COX-2,IL-1β,IL-6,and MMP-9 in the peritoneal tissues of each drug intervention group were significantly down-regulated.The expressions of neonatal inhibitors TIMP-1 and IL-12 were significantly up-regulated(P<0.05,P<0.01).3 Discussion on the mechanism of Fushen Decoction inhibiting peritoneal neoplasia:(1)RT-PCR results: Compared with the sham operation group,the m RNA expression of PI3 K,Akt and mTOR in the model group was significantly up-regulated(P<0.01).Compared with the model group,the drug intervention groups The expression of PI3 K and Akt was significantly down-regulated(P<0.05,P<0.01),and the expression of mTOR in the Fushenfang high-dose group was significantly down-regulated(P<0.05).(2)Immunohistochemical staining: Compared with the sham operation group,the positive expressions of PI3 K,p-mTOR,and p-Akt in the model group were significantly increased(P<0.01).Compared with the model group,the positive expressions of PI3 K,p-mTOR,and p-Akt in each drug intervention group were significantly reduced(P<0.01).Conclusions1 5/6 nephrectomy combined with intraperitoneal injection of high-sugar peritoneal dialysis fluid can successfully establish a rat uremic peritoneal dialysis model.2(1)Fushen recipe can effectively improve the peritoneal function,increase the amount of ultrafiltration,inhibit peritoneal angiogenesis,and have a certain protective effect on the peritoneal structure.(2)Fushen recipe can inhibit the formation of pathological new blood vessels by regulating,promoting and inhibiting the expression of angiogenesis-related cytokines.3 The effect of Fushen recipe on inhibiting peritoneal angiogenesis may be achieved by inhibiting the PI3K/Akt/mTOR signaling pathway.