| Objective:Peritoneal Dialysis(PD)is one of the most important dialysis methods for patients with end-stage renal disease(ESRD)because of its low cost,easy operation,and minimal impact on hemodynamics.The advantages of protecting residual kidney function have become the preferred alternative renal treatment for ESRD patients.However,it is regrettable that as the dialysis time prolongs,the PD's dialysis efficacy gradually decreases,resulting in the occurrence of ultrafiltration failure(UFF).UFF is the main reason for the failure of PD technology and the main obstacle to the further development of PD.The nature of UFF is Peritoneal Fibrosis(PF).PF severely affects the quality of life and long-term survival of patients with PD,and there is no effective treatment in clinical practice.Elucidating the pathogenesis of PF and exploring strategies for the preservation of peritoneal structural integrity have become major challenges in the field of renal replacement therapy(RRT).The epithelial mesenchymal transition(EMT)of peritoneal mesothelial cells is considered to be an early reversible and important pathophysiological process of PF.Therefore,if it is possible to intervene in the peritoneal mesothelial cell EMT at the initial stage,it will be possible to provide a new strategy for suppressing or even reversing PF.At present,EMT has become a research hotspot,but the molecular mechanism of EMT in peritoneal mesothelial cells remains unclear.In recent years,some studies have found that heat shock protein(HSP)plays an important role in the process of EMT,but the relationship between HSP and peritoneal mesothelial EMT has not yet been studied.Recent studies have suggested that HSP27 may participate in the activation of JAK/STAT3 signaling pathway by stabilizing STAT3 phosphorylation,stabilizing the expression of the transcription factor Snail,and thus participating in the maintenance of EMT in prostate cancer cells.To this end,our group established an animal model of peritoneal dialysis to target protein HSP27 as an entry point to observe the expression of HSP27 in rat peritoneal EMT tissue,to explore the relationship between HSP27 and peritoneal EMT,and to explore its downstream JAK/STAT3/Survivin pathway in the role of peritoneal EMT process,looking for some involved in peritoneal mesothelial EMT protein and its related molecular mechanisms explored.Method:1.Take 40 normal SD rats,adaptive feeding for one week,and randomly divide into four groups:control group,2-week dialysis group,4-week dialysis group,and 6-week dialysis group,in which the dialysis group is given 4.25%PD fluid daily.(100mL/kg)was intraperitoneally injected,and the control group received equal doses of 0.9%NS intraperitoneally every day;2.SD rats in each group were sacrificed after 2,4 and 6 weeks respectively,and peritoneal balance experiments were performed to evaluate the peritoneal function.3.The peritoneal tissues of SD rats in each group were stained with HE and Masson,and the morphological changes of the peritoneum were observed by light microscopy.4.q-PCR detection of EMT markers(epi-cadherin,phenotype N-cadherin)and HSP27 mRNA expression;5.Immunohistochemical detection of EMT markers(E ? cadherin,phenotype N-cadherin),and the expression of HSP27,P-STAT3,Survivin protein;6.Western Blot was used to detect the expression of EMT markers(E-cadherin and N-cadherin)and HSP27,P-STAT3,STAT3 and Survivin proteins.Results:1.Dialysis group rats peritoneal ultrafiltration was significantly lower than the control group,and with the prolonged dialysis time,ultrafiltration decreased more significantly,the difference was statistically significant(p<0.01).The above indicators change more significantly2.The solute transfer rate of small molecule in dialysis group was significantly higher than that in the control group,and with the prolonged dialysis time,the ultrafiltration volume increased more significantly,the difference was statistically significant(p<0.05 or p<0.01).3.HE and Masson staining showed that the peritoneal mesothelium was smooth and continuous in the control group,and the subepithelial stromal layer was thin and dense.In the dialysis group,the peritoneal mesothelial layer was detached,a large amount of collagen was deposited in the subepithelial stromal layer,accompanied with inflammatory cell infiltration and neovascularization,showing PF state.The longer the dialysis time,the more serious the degree of PF,and the more significant the increase in peritoneal thickness.The difference between the two groups was statistically significant(p<0.01).4.RT-PCR showed that the relative expression of E-cadherin mRNA in the peritoneal tissue of dialysis group was significantly lower than that of the control group,and the longer the dialysis time was,the more significant the decrease was(p<0.05 or p<0.01);The relative expression of N-cadherin mRNA in the interstitial markers was significantly higher than that in the control group.The longer the dialysis time was,the more significant the increase was.The difference between the two groups was statistically significant(p<0.05 or p<0.01).).5.Immunohistochemistry showed that the expression of E-cadherin protein in the peritoneal tissue of the dialysis group was significantly less than that of the control group,and the decrease was more significant with the extension of the dialysis time.The difference between the two groups was statistically significant.(p<0.05 or p<0.01);N-cadherin,HSP27,P-STAT3,and Survivin protein were highly expressed in the peritoneal tissue of the dialyzed rats,and the expression increased more significantly with the extension of the dialysis time.There was a statistically significant difference between the two groups(p<0.05).6.Western Blot and immunohistochemistry showed that the expression of E-cadherin protein in the peritoneal tissue of the dialysis group was significantly less than that of the control group,and with the extension of the dialysis time,the decrease was more significant,and the difference between the two groups was compared.Statistical significance(p<0.05 or p<0.01);N-cadherin,HSP27,P-STAT3,Survivin protein were highly expressed in the peritoneal tissues of rats in the dialyzed group,and the higher the expression,the more significant the expression was.There was a statistically significant difference between the two groups in the difference between the two groups(p<0.05 or p<0.01);there was no significant difference in the expression of STAT3 among the groups(p>0.05).Conclusion:1.High-dose peritoneal dialysis can cause impaired peritoneal structure and function in rats,which is time-dependent;2.High glucose peritoneal dialysis can induce early EMT changes in rat peritoneal mesothelial cells;The occurrence of EMT in rat peritoneal mesothelial cells may be related to the up-regulation of HSP27 protein and mRNA expression.The occurrence of EMT is related to the activation of JAK2/STAT3 signaling pathway;5.Survivin protein is highly expressed in the rat peritoneal EMT model and is time-dependent. |
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