Protective effects of MSC-conditioned medium on DFO-induced mimetic-hypoxia injury of renal tubular epithelial cells

Abstract

Background: Acute kidney injury (AKI) is mainly characterized by inflammatory infiltration, the damage and death of renal tubular epithelial cells (RTECs), in which hypoxia plays an important role. Deferoxamine (DFO) inducing cells’ mimetic-hypoxia injury is a well-accepted renal injury model in vitro. Mesenchymal stem cell - conditioned medium (MSC-CM) can reduce local inflammation and repair tissue. In this study, we explored the effect of MSC-CM on RTECs under DFO mimetic-hypoxia and its possible molecular mechanism. Methods: NRK-52E cells with the treatment of DFO25uM for 24 hours could be well-accepted RTECs’ injury model using Cell Counting Kit-8 (CCK-8) to detect cell viability and Western Blot to evaluate the expression of transforming growth factor-beta 1 (TGF-β1), α-smooth muscle actin (α-SMA), hypoxia-inducible factor-1 alpha (HIF-1α) in NRK-52E cells with the treatment of different concentrates of DFO for 24 hours. Then three groups of NRK-52E cells were used in experiments, including normal control (NC), DFO25uM, DFO25uM+MSC-CM. MSC-CM was used to culture cells for 12 hours before DFO treatment, then fresh MSC-CM and DFO 25uM cocultured cells for another 24 hours. Then the cell samples were collected. Results: Using Western Blot and cellular immunofluorescence, we found MSC-CM could reverse the DFO-induced up-regulation of TGF-β1, α-SMA, HIF-1α and steroid receptor coactivator 1 (SRC-1). DFO 25uM coculturing NRK-52E cells for 24 hours could simulate hypoxia well; HIF-1α and SRC-1 might be the harmful factors promoting EMT in NRK-52E cells during DFO mimetic-hypoxia.Conclusions: Our results suggest that MSC-CM have a protective effect on RTECs under DFO mimetic-hypoxia by down-regulating HIF-1α and SRC-1 which may be the harmful factors in renal injury.