Abstract: OBJECTIVE:To investigate the effect of mitofusin 2(Mfn2)and its downstream signaling pathway on glomerular mesangial cells(GMCs)proliferation in IgA nephropathy(IgAN),as well as the mechanism of action of Jixuecao(Herbo Centellae Asiaticae,HCA)in the treatment of IgAN.METHODS:Adenovirus-mediated Mfn2 gene transfection and Mfn2 expression were analyzed by real-time polymerase chain reaction(PCR)and Western blotting.IgA1 induced the proliferation of GMCs,which were then treated with HCA.Cell proliferation was detected with cell counting kit-8(CCK-8),and Mfn2 expression was analyzed by real-time PCR and western blotting.An IgAN animal model was also established and treated with HCA.GMCs proliferation was detected by hematoxylin-eosin staining,mitochondrial structure was analyzed by electron microscopy,mitochondrial function was determined by the Clark oxygen electrode method,and the expression of Mfn2,Phospho-extracellular regulated protein kinases1/2(P-ERK1/2),Cyclin-dependent kinase 2(CDK2),Phospho-p27(p-p27),and cyclin A was analyzed by Western blotting.RESULTS:In vitro,HCA inhibited GMCs in a concentration-dependent manner in association with the upregulation of Mfn2 expression.The overexpression of Mfn2 inhibited IgA1-induced GMCs proliferation and elevated the effect of HCA.In vivo,treatment with HCA could alleviate albuminuria and creatinine and GMCs proliferation.These effects were related to the upregulation of Mfn2,p-p27 and inhibition of p-ERK1/2,CDK2,and cyclinA.Mitochondrial swelling,vacuolar degeneration,and reduction of respiratory control rate were identified in IgAN,but HCA could improve the mitochondrial structure and function.CONCLUSION:HCA inhibited GMCs proliferation via the upregulation Mfn2 and the inhibition of Ras-Raf-ERK/MAPK.We revealed that changes of mitochondrial structure and function are associated with IgAN,but that HCA can improve these mitochondrial features.

Key words: Centella, Glomerulonephritis,IGA, Mesangial cells, Mitofusin 2