The effect of miR-155 targeting SMAD2 in podocyte injury and inflammatory response in mouse lupus nephritis
Objective:Exploring the effects of miR-155 regulation of SMAD2 expression in podocyte injury as well as mac-rophage inflammatory response in mouse lupus nephritis.Methods:The experimental mice were randomly divided into blank,model,AAV9-miR-NC,AAV9-miR-155,AAV9-miR-anti,and AAV9-miR-anti-NC groups,with 8 mice in each group,of which the blank group was C57BL/6 mice,and the rest of the groups were MRL/lpr mice,and the groups were treated and sam-pled accordingly.The presence or absence of a binding site for miR-155 and SMAD2 was predicted by target genes,an online tar-get gene prediction website.Dual luciferase reporter gene analysis system to validate the targeting relationship between miR-155 and SMAD2.Determination of 24 h urinary protein content in each group of mice by Caumas Brilliant Blue method.Hematoxylin-Eosin staining was used to observe the kidney tissues of mice;qRT-PCR to detect the expression level of miR-155 in kidney tissues of each group.Western blot was used to detect nephrin,P-cadherin and SMAD2 protein expression in renal tissues.Immunofluorescence assay to detect the expression of inflammatory factors(IL-6,TNF-α)in mice.Results:Compared with the blank group,mice in the model group had elevated urinary protein,severe renal pathological damage,and decreased expression levels of nephrin,P-cadherin,and SMAD2 while increased expression levels of IL-6 and TNF-α in renal tissues(P＜0.05).Com-pared with the model group and AAV9-miR-NC group,mice in the model group of AAV9-miR-155 group showed elevated uri-nary protein,obvious renal pathological damage,and obvious elevation of miR-155 in renal tissues,whereas the expression levels of nephrin,P-cadherin,and SMAD2 were decreased(P＜0.05)whereas the fluorescence intensity of IL-6 and TNF-α was en-hanced significantly;Compared with the model and AAV9-miR-anti-NC groups,mice in the AAV9-miR-anti group showed de-creased urinary protein,improved renal pathological injury,decreased miR-155 expression,and increased expression levels of nephrin,P-cadherin,and SMAD2 compared to the model group(P＜0.05)whereas the fluorescence intensities of IL-6 and TNF-α decreased;In the model group,AAV9-miR-NC group,and AAV9-miR-anti-NC group mice,the differences in the above indexes were not statistically significant(P＞0.05).Dual luciferase reporter gene assay showed that miR-155 inhibited the fluores-cence activity of wild-type SMAD2 cells and targeted the regulation of SMAD2 expression.Conclusion:High expression of miR-155 exacerbated podocyte injury and inflammatory response in lupus nephritis in mice by regulating SMAD2,whereas inhibi-tion of miR-155 expression attenuated podocyte injury and inflammatory response in mice.
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维普
