Kim B S, Cho Y, Lee H, Joo D J, Huh K H, Kim M S, Kim Y S
Division of Nephrology, Department of Internal Medicine, Severance Hospital, Yonsei University Health System, Seoul, Republic of Korea; The Research Institute for Transplantation, Yonsei University College of Medicine, Seoul, Republic of Korea.
The Research Institute for Transplantation, Yonsei University College of Medicine, Seoul, Republic of Korea.
Transplant Proc. 2016 May;48(4):1297-301. doi: 10.1016/j.transproceed.2016.01.022.
The mechanism of podocyte injury observed with the use of rapamycin (RPM) remains unclear. The conversion from calcineurin inhibitors (CNIs) to RPM in kidney transplant recipients has been associated with a higher incidence of proteinuria and renal injury. In this study, we performed proteomic analyses to investigate the alteration of protein expression in mouse podocytes treated with RPM in comparison with CNI/RPM combination.
Immortalized mouse podocytes were treated with 20 nmol/L RPM or 20 nmol/L RPM + 1 μg/mL cyclosporine. Podocyte proteins were separated by 2-dimensional polyacrylamide gel electrophoresis (2DE) and identified by matrix-assisted laser desorption time-of-flight (MALDI-TOF) mass spectrometry and peptide fingerprinting. Selected proteins were analyzed by means of Western blot assay.
We identified 36 differently expressed proteins after isolated RPM or CNI/RPM combination treatment in cultured mouse podocytes. There are 3 distinct patterns of protein expression: (1) potentiated down- or upregulation of proteins by CNI/RPM treatment compared with isolated RPM treatment (n = 4); (2) partial offset of down-regulation by CNI/RPM in comparison with RPM treatment (n = 25); (3) no difference in down-regulation between RPM and CNI/RPM treatment (n = 5). We found a significant interplay between RPM and CNI on the expression of the selected proteins in mouse podocytes. This might explain the higher incidence of proteinuria by CNI/RPM combination in clinical settings.
Further study is required to elucidate the target protein associated with RPM-induced podocyte injury.
使用雷帕霉素（RPM）时观察到的足细胞损伤机制仍不清楚。肾移植受者从钙调神经磷酸酶抑制剂（CNI）转换为RPM与蛋白尿和肾损伤的较高发生率相关。在本研究中，我们进行了蛋白质组学分析，以研究与CNI/RPM联合治疗相比，RPM处理的小鼠足细胞中蛋白质表达的变化。
用20 nmol/L RPM或20 nmol/L RPM + 1 μg/mL环孢素处理永生化小鼠足细胞。通过二维聚丙烯酰胺凝胶电泳（2DE）分离足细胞蛋白，并通过基质辅助激光解吸飞行时间（MALDI-TOF）质谱和肽指纹图谱进行鉴定。通过蛋白质印迹分析对选定的蛋白质进行分析。
在培养的小鼠足细胞中，单独使用RPM或CNI/RPM联合处理后，我们鉴定出36种差异表达的蛋白质。有3种不同的蛋白质表达模式：（1）与单独使用RPM处理相比，CNI/RPM处理使蛋白质下调或上调增强（n = 4）；（2）与RPM处理相比，CNI/RPM使下调部分抵消（n = 25）；（3）RPM和CNI/RPM处理之间下调无差异（n = 5）。我们发现RPM和CNI在小鼠足细胞中选定蛋白质的表达上存在显著相互作用。这可能解释了临床环境中CNI/RPM联合使用时蛋白尿发生率较高的原因。
需要进一步研究以阐明与RPM诱导的足细胞损伤相关的靶蛋白。
