POS-381 Podocyte B7-1 Mediates Cell Injury and Glomerulosclerosis through Communicating with β-catenin Pathway

Abstract

Podocyte injury is a hallmark of glomerular diseases, however, the intrinsic mechanisms are not elucidated. Previous studies have shown that B7-1 may be an inducer to podocyte injury. However, the intrinsic role of B7-1 in podocyte injury is controversial. In this study, we have identified that podocyte B7-1 definitely contributes to cell injury and glomerulosclerosis, and the underlying mechanisms are intimately correlated with activation of β-catenin pathway. Our studies provide the importance evidence to the role of B7-1 in podocyte injury and development of glomerulosclerosis. The expression of B7-1 were assessed in various CKD models, human biopsies and isolated glomeruli. Podocyte-specific B7-1 transgenic mice (podo-B7-Tg) were constructed. Podocyte, glomerular injury and β-catenin signaling were assessed in podo-B7-Tg mice at 2, 3 and 6-month age, and advanced oxidative protein products (AOPPs)-administrated mice. In ADR and 5/6 nephrectomized mice, B7-1 interference plasmids (B7-shR) were injected through hydrodynamic-based gene delivery approach. In cultured podocyte cell line (MPC5), ectopic expression of B7-1 was by transient transfection and lentivirus-mediated transduction. The podocyte injury and β-catenin signaling were assessed. The binding of NF-κB-p65 and B7-1 was tested by ChIP assay, and testified by luciferase reporter assessment. The protein binding with B7-1 was assessed by mass spectrum and defined by co-immunoprecipitation. The binding of β-catenin and B7-1 was also determined by ChIP assay. The expression of B7-1 was upregulated in various CKD models (ADR, db/db, 5/6 nephrectomyin and human biopsies), and increased predominantly in podocytes. In vivo, there is a slightly but significantly increase in urinary albumin secretion and podocyte injury in podo-B7-Tg mice at age of 3-month, and further elevated at 6-month-age. Compared with wild-type mice, administration of AOPPs in podo-B7-Tg mice further enhanced the expression of B7-1, which resulting in the acceleration of renal fibrosis, and the further elevated activation of β-catenin pathway. Knockdown of B7-1 could restore podocyte integrity and retard renal fibrosis in ADR and 5/6 nephrectomized mice. The activation of β-catenin pathway was also apparently abolished by B7-1 interference. In cultured cells, exogenous expression of B7-1 induced podocyte injury and β-catenin pathway activation. The transcriptional activity of B7-1 was regulated by NF-κB-p65 through directly triggering the gene promoter. Moreover, B7-1 was identified to bind with HSP90AB1 and LRP5, forming a complex to trigger the activation of β-catenin pathway. Interestingly, there were also several TCF/LEF binding consensus sites in the promoter region of B7-1. Ectopic expression of β-catenin also induced the mRNA and protein expression of B7-1. Podocyte B7-1 which depends on the regulation of NF-κB pathway, plays an important role in maintaining cellular homeostasis. Additionally, B7-1 binds to HSP90AB1 and LRP5 to form a complex, further inducing the activation of β-catenin. While β-catenin can also promote B7-1 transcription through TCF/LEF transcription factors. Our studies provide a new theory in the pathogenesis of glomerulosclerosis, and give an important clue to the therapeutic strategy to podocyte injury through targeting the two-way networks of B7-1 and β-catenin.