| Background:Diabetic nephropathy(DN)is the most common complication in diabetic patients and one of the leading causes of death.The incidence in China is increasing year by year.The pathogenesis of DN is complex and there is currently no effective treatment program to prevent its progress.Each patient is behind a huge cost every year,so early intervention in the development of diabetic nephropathy is beneficial to improving the quality of life of patients and reducing the social burden.Proteinuria is the most prominent clinical manifestation of DN.Studies have shown that the damage of podocytes is the main cause of proteinuria.Some researchers even suggested that DN may also be a kind of "podocyte disease."In our previous study,in a five-fifths nephrectomies model,a classic model of Focal segmental glomerulosclerosis(FSGS),we found a decrease in the number of podocytes that detected a parietal layer.Epithelial cells(parietal epithelial cells,PECs)expressed podocyte markers,and double-positive expression of podocyte markers and PECs markers was found at multiple capillary loops.It suggests that there is a phenomenon of transdifferentiation of PECs into podocytes.This transdifferentiation may be involved in the repair of podocytes after injury.Podocyte is a terminallydifferentiated cell that is difficult to supplement by proliferation and cannot be regenerated once damaged.More and more studies have found that podocytes can be regenerated from other sources and play a crucial role in the repair of podocyte injury.Unlike glomerular epithelial cells,PECs have the potential for proliferation.Some scholars have suggested that PECs may act as progenitor cells of podocytes,thus replacing the repair of damaged podocytes and participating in the repair process of podocyte injury.Dr.Romagmani and Dr.Moeller observed the process of transdifferentiation of PECs into podocytes under electron microscope,suggesting that PECs may be transdifferentiated into podocytes,thereby repairing damaged podocytes.So what are the factors that affect the process of podocyte injury repair?Recent studies have found that miR-193a expression is down-regulated in human micro RNA(mi R)and can mediate high expression of WT1 and other podocyte markers in PECs and promote their transdifferentiation into podocytes [8].miR-193a is specifically expressed in mature PECs and its unregulated levels can induce a series of molecular cascades that can eventually lead to glomerular injury and FSGS in mice and humans.In the previous study on the process of podocyte transdifferentiation of PECs,we also found that there were more transdifferentiation of PECs into podocytes in the intervention group of RAS inhibitor(ARB).It suggests that the ARB group may protect the podocytes by repairing damaged podocytes by promoting PECs-podocyte transdifferentiation.So does ARB promote podocyte injury repair by regulating miR-193a? For this reason,we proposed a hypothesis that ARB inhibits the expression of miR-193a in diabetic nephropathy,thereby promoting the repair of podocyte injury.Objective:Observe the role of miR-193a in the early process of podocyte repair in the early diabetic nephropathy,and provide a new idea for delaying the progression of diabetic nephropathy with miR-193a as a new point of view.Methods:Randomly numbered 40 db/db mice at 5 weeks old into 2 groups: db/db group and db/db-losartan treatment group(db/db-ARB group),20 db/m mice As a normal control group.After one week of quarantine and adaptive rearing,body weight and tail vein blood glucose of each group of mice were measured every four weeks from the age of 6 weeks,and 24-hour urine was collected using a metabolic cage.At 10 weeks of age,the following drug interventions were performed: db/db-ARB group was given ARB Losantan 10 mg/(kg·d);db/m group and db/db group were given equal amounts of drinking water,each group of mice was intragastric administration,kidney biopsies were taken in batches before treatment(10 weeks of age),4 weeks after treatment(14 weeks of age),8 weeks after treatment(18 weeks of age),and 12 weeks after treatment(22 weeks of age).Prepare pathological sections and observe the pathological changes of renal tissue.Expression of WT1 was detected by immunohistochemistry;Expression of synaptopodin in glomeruli was detected by immunofluorescence;Western Blotting was used to detect the expression of WT-1and synaptopodin in renal podocyte markers,claudin1 and Pax-2 in the parietal epithelial cells markers.Total RNA was extracted and the expression of WT-1,synaptopodin,claudin1,and pax-2 m RNA and the expression of miR-193a in kidney tissue were detected by real-time fluorescence quantitative PCR.Results:1.With the increase of age,the body weight,blood glucose,urinary protein,and serum creatinine in db/db mice were significantly higher than those in db/m mice(P<0.05).The level of urinary protein and serum creatinine after intervention with losartan was lower than that of the untreated group(P<0.05).2.In the course of this experiment,compared with db/m mice,db/db mice always maintained hyperglycemia,and there was no significant difference in blood glucose levels between losartan-treated and untreated db/db mice.(P>0.05).3.With the increase of age,the protein and m RNA of podocyte markers WT-1and synaptopodin in renal tissue of db/db and db/db-ARB mice were lower than those of db/m mice of the same age(P<0.05).The protein and m RNA expression of claudin1 and Pax-2 in the parietal epithelial cell markers was higher than that in db/m mice of the same age(P<0.05).After treatment with losartan,the protein and m RNA of the podocyte markers WT-1 and synaptopodin were higher than those of the non-drug group(P<0.05),while the expression of protein and m RNA of claudin1 and Pax-2 in the parietal epithelial cells was decreased compared with that in the non-drug group(P<0.05).It was demonstrated that ARB has a protective effect on podocyte and kidney damage in mice with diabetic nephropathy.It was demonstrated that ARB has a protective effect on podocyte and kidney damage in mice with diabetic nephropathy.4.With the progress of diabetic nephropathy,db/db mice showed thickened glomerular basement membranes,increased mesangial cells and mesangial matrix,and tubular and interstitial fibrosis.After the treatment with losartan,the above pathological changes were alleviated.5.The results of immunohistochemistry showed that with the increase of age,the expression of WT1 in kidneys of db/db mice and db/db-ARB mice was lower than that of db/m mice,and the db/db-ARB mouse kidney tissues were decreased.The expression of WT1 in mice increased compared with db/db mice.6.Immunofluorescence showed that with the increase of age,the expression of synaptopodin in glomeruli of db/db mice and db/db-ARB mice was lower than that of db/m mice.After treatment with ARB,the expression of synaptopodin in db/db-ARB mice was higher than that in db/m mice7.Real-time fluorescence quantitative PCR showed that after treatment with losartan,the expression of miR-193a in db/db-ARB mice was significantly lower than that in db/db mice(P<0.05),suggesting that ARB may inhibit the expression of miR-193a.Conclusion:ARB may inhibit the expression of miR-193a and promote the repair of podocytes in mice with early diabetic nephropathy,thereby protecting the damaged podocytes and delaying the development of diabetic nephropathy. |
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