| Objective: To explore the correlation phenotype of TM7SF1 gene down-regulation with diabetic nephropathy C57BL/6J mice.Methods:Using Crispr/Cas9 technology and using non-homologous recombination repair to introduce mutations,construct a C57BL/6J mouse model of TM7SF1 gene systemic knockout.The STZ intraperitoneal injection method was used to construct the diabetic nephropathy model of TM7SF1-/-and TM7SF1+/+ mice.The differences in phenotypes of the two genotypes of diabetic nephropathy model mice included urine protein,urinary creatinine,kidney coefficient,serum biochemical index,kidney morphology,podocyte-related protein and autophagy-related expression.Results:1.TM7SF1-/-and TM7SF1+/+ mice construct diabetic nephropathy model at the same time.With the increase of mouse age,the urine protein of both groups of mice shows an increasing trend,but compared with TM7SF1+/+,TM7SF1-/-mice have more urinary protein(most significant at the 11 th week of modeling,P<0.05).2.Both groups of diabetic nephropathy model mice have renal tubule mitochondrial damage,but compared with TM7SF1+/+ mice,TM7SF1-/-mice are more severe.TM7SF1-/-mice also showed podocyte fusion,but TM7SF1+/+ mice did not.Conclusion: Deletion of TM7SF1 will accelerate the course of diabetic nephropathy in C57 BL / 6J mice,and will also cause disturbance of renal autophagy.TM7SF1 downregulation can cause autolysosomes degradation in podocytes.This block of autophagy may be due to a decrease in the number of acid lysosomes.Objective: To investigate the mechanism of TM7SF1 gene down-regulation affecting the autophagy of kidney podocytes in mice.Methods: Use lentivirus-mediated methods to construct a mouse podocyte model that interferes with TM7SF1 gene expression.To study the changes of LC3 BII and P62/SQSTM1 in mouse podocytes with down-regulated TM7SF1 gene and the mechanism of autophagy obstruction by Western blot and immunofluorescence.Results:1.Use Western blot to detect the increase of LC3 BII in podocytes down-regulated by TM7SF1.Chloroquine was then used to block the degradation of autophagy-related proteins.Western blot confirmed that the increase in LC3 BII was due to the accumulation of degradation barriers.2.Using Western blot to detect the increase in P62/SQSTM1 in podocytes downregulated by TM7SF1 was verified by immunofluorescence.After using chloroquine to block the degradation of autophagy-related proteins,Western blot and immunofluorescence confirmed that the increase in P62/SQSTM1 was due to the accumulation of degradation barriers.3.Using acridine orange staining method to detect TM7SF1 down-regulation will reduce the number of acid lysosomes in podocytes.Conclusion: Deletion of TM7SF1 will accelerate the course of diabetic nephropathy in C57BL/6J mice,resulting in increased urinary protein and disorder of autophagy in kidney tissue. |
