Tissue Inhibitor of Matrix Metalloproteinase 2-Loaded Polylactic Acid-Polyethylene Glycol Nanoparticles Directly Regulates Renal Tubular Epithelial Cell Proliferation via Protein Tyrosine Kinase KIT

Abstract

TIMP2 has been previously reported to be associated with acute kidney injury (AKI); however, the underlying mechanism remains unclear. Therefore, the present study investigated the regulation of TIMP2 in human tubular epithelial cells HK-2 cells. Proliferation of HK-2 cells treated by TIMP2 at normal expression level was detected. GST pulldown and mass spectrometry were performed to investigate the interacting protein of TIMP2 and immunofluorescence test was used to determine the location of the protein in HK-2 cells. Cell viability as well as the expression level of CCND1, C-FOS, MAPK1 and P-MAPK1 were detected in the samples treated by overexpressed TIMP2 and the inhibitor of the interacting protein KIT. TIMP2 significantly inhibited cell proliferation compared with the control and BB-94-treated groups (P < 0.05). KIT was identified as the interacting protein of TIMP2, and was located in both the cytoplasm and membrane of HK-2 cells. Inhibited KIT and the overexpressed of TIMP2 both significantly suppressed cell proliferation and decreased the expression levels of CCND1, MAPK1, and P-MAPK1 compared with the control (P < 0.05). No significant difference was observed in cell proliferation and the expression level of aforementioned proteins between overexpressed TIMP2 and KIT-inhibited group. The results revealed that TIMP2 regulates cell proliferation by reducing the expression levels of CCND1, MAPK1, and P-MAPK1 via KIT, indicating that TIMP2 directly regulates cell proliferation without inhibiting matrix metalloproteinases in AKI. Furthermore, PLA-PEG nanoparticles successfully transported TIMP2 to the target with no significant effect on cell proliferation.