Modelling APOL1-mediated kidney inflammation and fibrosis using a partially reprogrammed urine derived SIX2-positive renal progenitor cell line.

Abstract

BackgroundCKD affects approximately 850 million people worldwide and is a leading cause of mortality. Podocytes, cells in the kidney are terminally differentiated and incapable of division in vivo making the establishment of primary cultures particularly challenging. The ability of cells to proliferate and avoid senescence is closely linked to telomere length. When telomere length becomes critically reduced, it results in cellular senescence. MethodsWe present the successful rejuvenation of a human SIX2-positive renal progenitor cell line derived from the urine of a 30-year-old West African male (UM30-OSN). To achieve partial reprogramming, plasmids expressing the Yamanaka factors OCT4, SOX2, NANOG, c-Myc, and KLF4 were employed. ResultsUM30-OSN expresses the pluripotency-associated marker SSEA4, renal stem cell markers such as SIX2, CD133 and CD24, determined by immunofluorescence, FACS and qPCR. Expression analysis revealed downregulation of senescence markers p21and p53 and upregulation of proliferation-associated genes PCNA, KI67 and TERT, confirming rejuvenation. Upon podocyte differentiation, UM30-OSN cells expressed podocyte-specific markers NPHS1, NPHS2, SYNPO and CD2AP. Comparative transcriptome analyses revealed a correlation co-efficiency (R2 = 0.88) with the immortal podocyte line AB 8/13. To demonstrate the usefulness of UM30-OSN to model APOL1-mediated kidney disease, we investigated the effects of Interferon-{gamma} (IFN-{gamma}) on UM30-OSN derived podocytes and evaluated the potential of the JAK1/JAK2 inhibitor Baricitinib to mitigate IFN-{gamma}-induced cellular responses. IFN-{gamma} stimulation resulting in increased phosphorylation of STAT1, activation of APOL1, upregulation of pro-inflammatory and fibrotic markers such as, IL-6, TGF-{beta}, Vimentin, Fibronectin, and morphological changes indicative of cell stress. Pre-treatment with Baricitinib effectively inhibited STAT1 phosphorylation, reduced expression of pro-inflammatory and fibrosis-associated genes, and preserved podocyte morphology. ConclusionGiven their robust proliferation capacity, UM30-OSN cells represent a valuable additional model for investigating kidney-associated diseases such the contribution of APOL1 high-risk variants to kidney injury and fibrosis.