| Introduction: Membranous nephropathy (MN) is a kind of primary kidney diseases caused by autoimmune abnormalities, and its pathogenesis is not clear. Anti-M-type phospholipase A2 receptor antibody has been identified as the specific autoantibody in about 70% of patients with MN. But physiological changes after autoantibodies binding to glomerular remain unclear. The purpose of this study is to screen proteins in renal tissues of membranous nephropathy with proteomics techniques, and to explore and explain possible pathways of pathogenesis of membranous nephropathy, expecting to lay the foundation for further study of the pathogenesis of membranous nephopathy.Materials and methods: (1) Proteins of membranous nephropathy such as cytokines are always low-abundance and low molecular weight protein, so this study chose protein chip technology to screen it. In order to find a protein chip suitable for kidney tissue detection,three kinds of protein chips, quantitative chip QAH-INF-3 and semi-quantitative chips L-493 and L-507 were evaluated by normal renal tissue in the first part of this study. These chips were used to detect the 2: 1 dilutions of the same sample, and the signal value of the measured ratio of the sample between 1.3 and 3.0 was defined as a detectable protein. The number and proportion of detectable proteins were used to evaluate the availability of the chip.(2) The first part of this study confirms that chip L-493 is the most suitable one for the detection of protein in kidney tissue. In the second part, three cases of membranous nephropathy and three normal renal tissues were detected by L-493. Proteins with MN / N signal ratio > 1.35 were defined as differential proteins, bioinformatics analysis was used to screen the signaling pathways associated with pathogenesis of membranous nephropathy.And the protein with higher expression was verified by enzyme - linked immunosorbent assay (ELISA) and immunohistochemical staining (IHC).Results and Results: (1) Chips availability evaluation: There were 9 detectable proteins(22.5%) in chip QAH-INF-3, 61 (12.03%) detectable proteins in chip L-507, 214 (43.41%)detectable proteins in L-493. The numbers of proteins detected by chip L-493 are the most,so, L-493 is more suitable for the detection of low molecular weight and low-abundance proteins in kidney tissues.(2) Membranous nephropathy detection: Chip L-493 detected a total of 66 differential proteins, GO analysis and STRING analysis found a variety of signal pathways possible associated with membranous nephropathy, such as immune activation, cell metabolism,cell adhesion, Cell signal transduction and extracellular matrix remodeling. Further analysis found that multiple proteins have diverse functions, they play essential role in the pathways, making functional areas of protein linked to fonn a protein network, the network work together to promote the development of membranous nephropathy. |
