Comparative Proteomic Analysis of Rapamycin Versus Cyclosporine Combination Treatment in Mouse Podocytes.
BACKGROUND
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.


METHODS
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.


RESULTS
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.


CONCLUSIONS
Further study is required to elucidate the target protein associated with RPM-induced podocyte injury.
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.
B S
BS
Kim
The Research Institute for Transplantation, Yonsei University College of Medicine, Seoul, Republic of Korea.
Y
Y
Cho
The Research Institute for Transplantation, Yonsei University College of Medicine, Seoul, Republic of Korea.
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H
Lee
The Research Institute for Transplantation, Yonsei University College of Medicine, Seoul, Republic of Korea; Department of Transplantation Surgery, Severance Hospital, Yonsei University Health System, Seoul, Republic of Korea.
D J
DJ
Joo
The Research Institute for Transplantation, Yonsei University College of Medicine, Seoul, Republic of Korea; Department of Transplantation Surgery, Severance Hospital, Yonsei University Health System, Seoul, Republic of Korea.
K H
KH
Huh
The Research Institute for Transplantation, Yonsei University College of Medicine, Seoul, Republic of Korea; Department of Transplantation Surgery, Severance Hospital, Yonsei University Health System, Seoul, Republic of Korea.
M S
MS
Kim
The Research Institute for Transplantation, Yonsei University College of Medicine, Seoul, Republic of Korea; Department of Transplantation Surgery, Severance Hospital, Yonsei University Health System, Seoul, Republic of Korea. Electronic address: yukim@yuhs.ac.
Y S
YS
Kim
D058186:Acute Kidney Injury / Q000139:chemically induced; D000818:Animals; D065095:Calcineurin Inhibitors / Q000493:pharmacokinetics* / Q000633:toxicity; D002478:Cells, Cultured; D016572:Cyclosporine / Q000494:pharmacology* / Q000633:toxicity; D004338:Drug Combinations; D007166:Immunosuppressive Agents / Q000494:pharmacology* / Q000633:toxicity; D007668:Kidney / Q000378:metabolism; D051379:Mice; D050199:Podocytes / Q000187:drug effects* / Q000378:metabolism; D011506:Proteins / Q000187:drug effects / Q000378:metabolism*; D011507:Proteinuria / Q000378:metabolism; D040901:Proteomics / Q000379:methods; D020123:Sirolimus / Q000494:pharmacology* / Q000633:toxicity; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors
D065095:Calcineurin Inhibitors; D004338:Drug Combinations; D007166:Immunosuppressive Agents; D011506:Proteins; D016572:Cyclosporine; C546843:mTOR protein, mouse; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus