Recapitulation of Cellular Senescence, Inflammation, and Fibrosis in Human Kidney-Derived Tubuloids by Repeated Cisplatin Treatment

Abstract

AbstractIn the pursuit of more accurate pathophysiological models for assessing renal drug response, the development of kidney organoids derived from human pluripotent stem cells represents a significant step forward. However, recapitulating aging/senescence-associated pathophysiology within these models remains challenging. Here, we present an innovative approach to generate more homogeneous epithelial-like structures known as “tubuloid” using primary human renal proximal tubular epithelial cells (hRPTECs) cultured from human resected kidneys, as a refined alternative. We evaluated the efficacy of tubuloids using cisplatin treatment at three different concentrations: 0.2, 2.0, and 20.0 µg/mL. Tubuloids showed highly differentiated structures with proximal tubular epithelial cells that expressed lotus tetragonolobus lectin and LRP2/Megalin. Upon exposure to cisplatin, γH2AX expression increased in a dose-dependent manner, indicating DNA damage. Cisplatin treatment also resulted in the expression of Kidney Injury Molecule-1 (KIM-1) and Cleaved Caspase-3, which are indicators of kidney injury and apoptotic signaling, respectively. Repeated cisplatin administration resulted in upregulation of the cellular senescence marker p16, alongside increased secretion of inflammatory cytokines IL-1β and IL-6, indicating the induction of a senescence-associated secretory phenotype (SASP). Furthermore, supernatant collected from cisplatin-treated tubuloids induced myofibroblast activation, indicating the onset of renal fibrosis. We successfully established a tubuloid-based model of cisplatin-induced kidney injury using hRPTECs. Tubuloids provide a novel platform for studying the response of renal epithelial cells to toxins and therapeutics. Tubuloids can replicate cellular senescence, SASP, and fibrosis, making them a promising pathophysiological model for chronic kidney disease (CKD), providing insights into the disease’s fibrotic mechanisms.Translational StatementRecapitulating aging/senescence-associated pathophysiological reaction in kidney organoids remains challenging. Our study reveals that tubuloids could be novel candidate for chronic kidney disease (CKD) model.