Immortalized Human Glomerular Podocytes-SV40 (Tet-on)

Voclosporin (VCS) Improved In Vitro Podocyte Viability and Decreased Cellular Stress after PAN Treatment

Voclosporin (VCS) is a second generation calcineurin inhibitor (CNI), which is currently approved for use only in adults with lupus nephritis (LN) on a background of immunosuppression. However, VCS is not yet approved for use in children or adults with any other renal diseases. Based on this, we assessed the ability of VCS to protect conditionally immortalized human podocytes (hiPODs) from puromycin aminonucleoside (PAN)-induced toxicity and stress. Differentiated hiPODs were challenged with 50 µg/ml PAN (or Saline control) and treated with VCS (or DMSO vehicle control) over a dose curve (0.01, 0.03, 0.1, 0.3, 0.9, and 2.7 µg/ml). After 24 h, results showed that PAN treatment significantly reduced cell viability to 49% (Fig. 1a) vs. No PAN + DMSO treated cells (set at 100%). VCS rescued cell viability in PAN-treated cells at all doses and was significantly increased at 0.03 µg/ml (83%) and 0.3 µg/ml (85%) vs. PAN + DMSO-treated cells. For cellular stress analyses, LDH release was significantly increased 1.98-fold in PAN + DMSO treated cells (Fig. 1b) vs. No PAN + DMSO cells. VCS decreased cellular stress in PAN-treated hiPODs at 0.03–0.9 µg/ml, but this was not significant. The dose-response curve showed that at 2.7 µg/ml VCS treatment in PAN-treated cells significantly increased cellular stress as compared to PAN + DMSO-treated cells (1.46-fold) (Fig. 1b). These results demonstrate that VCS protected hiPODs by reducing cell death and cellular stress induced by PAN treatment.