The Role Of PLA2G12B In Diabetic Nephropathy

Diabetic nephropathy(DN)is one of the microvascular complications of diabetes,which is characterized by a gradual increase in urine protein and a gradual decrease in renal function,and eventually develops into end-stage renal disease.Podocyte damage is a key cellular event in the progression of DN,and it is also an important factor driving kidney damage.Therefore,clarifying the molecular mechanism of podocyte damage in DN is of great significance for the treatment of the disease and the discovery of clinical markers.Previous studies have found that lipid metabolism disorders are an important phenomenon that accompanies the progression of podocyte damage in DN,and lipid disorders will further promote changes in the immune microenvironment in the glomerular region,leading to disease progression.At present,the mechanism of podocyte injury caused by abnormal lipid metabolism is still unclear.It is of great significance to analyze the role of key molecules in DN for the prevention and treatment of DN.PLA2G12B(phospholipase A2 group XIIB)belongs to the secreted phospholipase A2 family.Unlike other members of the same family,the active site of PLA2G12B has changed,which greatly reduces the activity of phospholipase.Previous studies have shown that PLA2G12B is mainly present in the liver,muscle and kidney,and plays an important role in the synthesis and transport of triglycerides in the liver.When kidney injury occurs,it is found that the expression level of PLA2G12B can be significantly increased.This study found for the first time that PLA2G12B can affect the lipid synthesis process in podocytes through the Notch3SREBP1/PPAR-γ pathway.It was found that the expression of PLA2G12B is significantly increased in DN and other chronic kidney diseases.Further silencing PLA2G12B in podocytes can improve podocyte damage and lipid deposition in podocytes.In terms of mechanism,PLA2G12B is achieved through the Notch3-SREBP1/PPAR-γ signaling pathway.Regulation of lipid synthesis in podocytes.Objective1.To clarify the expression pattern of PLA2G12B in DN.2.To study the role of PLA2G12B in DN.3.To explore the mechanism of PLA2G12B in DN.4.To explore whether PLA2G12B can be a simple and efficient biomarker for the identification of DN.Methods and Results:1 Expression pattern of PLA2G12B in DN1.1 Detection of PLA2G12B expression in serum and kidney tissue samples of clinical patientsElisa kit showed that serum PLA2G12B in DN patients was significantly higher than that in normal people,and it was positively correlated with urine protein and serum creatinine,and negatively correlated with glomerular filtration rate,PLA2G12B was significantly increased in the glomerulus in the pathological sections of the patients detected by IHC,after the serum of patients with DN stimulated podocytes,PLA2G12B was significantly increased.1.2 Expression of PLA2G12B in animal models of DNThe results of Western Blot,RT-qPCR and IHC showed that the expression of PLA2G12B was significantly increased in db/db mice,and IF results showed that PLA2G12B was significantly increased in podocytes.The same results were observed in STZ/HFD mice.1.3 In vitro simulated DN to detect the expression of PLA2G12B in podocytesIn vitro,HG,AGEs and PA treatment can induce the expression of PLA2G12B in podocytes increased.2 The role of PLA2G12B in podocyte injury in DN2.1 Construction of PLA2G12B knockout miceThe mouse tail DNA was extracted to identify the mouse genotype,and the results showed that PLA2G12B was successfully silenced.2.2 Deletion of PLA2G12B in mice ameliorates kidney injury from DNCompared with control mice,the ratio of UACR and the expression of IL-6 in STZ/HFD mice were increased,and both were significantly decreased after PLA2G12B was knocked out in vivo.The results of PAS showed that knockout of PLA2G12B could significantly improve the kidney injury induced by STZ/HFD.2.3 Silencing of PLA2G12B in podocytes ameliorates high glucose-induced podocyte injurySilencing PLA2G12B in podocytes can significantly improve the increase of inflammatory factors,podocyte apoptosis,Nephrin/Podocin damage and F-actin damage caused by high glucose stimulation.2.4 PLA2G12B affects lipid deposition in podocytes After silencing PLA2G12B in podocytes,lipid deposition was reduced,whereas it was significantly increased after overexpression.Silencing of PLA2G12B in podocytes ameliorated the elevation of SREBP1 and PPAR-γ induced by HG.2.5 PLA2G12B regulates Notch3 expression in podocytesThe results of RT-qPCR and Western Blot showed that silencing of PLA2G12B in podocytes could significantly reduce the elevation of Notch3 induced by HG stimulation.2.6 PLA2G12B regulates podocyte injury through the Notch3/PPAR-γ pathwayRT-qPCR results showed that silencing of Notch3 in podocytes could significantly reduce the elevation of SREBP1 and PPAR-γ caused by overexpression of PLA2G12B.Nile Red showed that silencing Notch3 in podocytes significantly ameliorated lipid droplet deposition caused by overexpression of PLA2G12B.Silencing Notch3 in podocytes can significantly reduce the rearrangement of cytoskeleton and the elevation of inflammatory factors caused by overexpression of PLA2G12B.Conclusion and innovation:1.The expression of PL A2G12B is increased in serum and kidney of patients with DN,and it is positively correlated with serum creatinine and urine protein,and negatively correlated with glomerular filtration rate.2.This project first discovered the function and role of PLA2G12B in the kidney,found that the expression of PLA2G12B was significantly increased in DN,and it could regulate lipid deposition in podocytes through the Notch3-SREBP1/PPAR-γ pathway.3.First discovery of PLA2G12B as a potential biomarker for kidney disease.