ABSTRACT
Autosomal dominant polycystic kidney disease (ADPKD) is a common genetic disorder of adults, characterized by uncontrolled cysts formation that causes a gradual impairment of kidney function. We generated a human induced pluripotent stem cell (hiPSC) line from the urinary cells of a patient diagnosed with ADPKD using a non-integrating Epi5™ Episomal iPSC reprogramming strategy. Characterization of the cell line was performed regarding their undifferentiated status, differentiation potential, and quality control for karyotypic integrity, identity, and clearance of reprogramming vectors. The newly derived hiPSC line, namely BCRTi007-A, can be used in vitro for disease modeling of ADPKD as well as testing for novel therapeutic approaches.
Subject(s)
Induced Pluripotent Stem Cells , Polycystic Kidney, Autosomal Dominant , Adult , Humans , Induced Pluripotent Stem Cells/metabolism , Polycystic Kidney, Autosomal Dominant/genetics , Polycystic Kidney, Autosomal Dominant/metabolism , Mutation , Cell Differentiation , Cell LineABSTRACT
Individuals with transient reception potential cation channel 6 (TRPC6) mutation have variable phenotypes, ranging from healthy carriers to focal segmental glomerulosclerosis (FSGS). Human induced pluripotent stem cell (hiPSC) line was generated from the urinary cells of a patient with FSGS with a mutant variant of TRPC6. The cells were reprogrammed with Yamanaka factors (OCT3, SOX2, LIN28, L-MYC, and KLF4) using a commercially available Epi5 Reprogramming Kit. The pluripotency of the hiPSC line was confirmed by the expression of common stem cell markers and by their ability to generate all germ layers in vitro. The line is available and registered in the human pluripotent stem cell registry as BCRTi006-A. The generated line represents a valuable tool for disease modeling and drug development for FSGS.
Subject(s)
Glomerulosclerosis, Focal Segmental , Induced Pluripotent Stem Cells , Pluripotent Stem Cells , Humans , Induced Pluripotent Stem Cells/metabolism , Glomerulosclerosis, Focal Segmental/genetics , TRPC6 Cation Channel/genetics , TRPC6 Cation Channel/metabolism , Mutation , Cell Differentiation , Cellular ReprogrammingABSTRACT
Cells release extracellular vesicles (EVs) to communicate in a paracrine manner with other cells, and thereby influence processes, such as angiogenesis. The conditioned medium of human cardiac-derived adherent proliferating (CardAP) cells was recently shown to enhance angiogenesis. To elucidate whether their released EVs are involved, we isolated them by differential centrifugation from the conditioned medium derived either in the presence or absence of a pro-inflammatory cytokine cocktail. Murine recipient cells internalized CardAP-EVs as determined by an intracellular detection of human proteins, such as CD63, by a novel flow cytometry method for studying EV-cell interaction. Moreover, endothelial cells treated for 24 h with either unstimulated or cytokine stimulated CardAP-EVs exhibited a higher tube formation capability on Matrigel. Interestingly, unstimulated CardAP-EVs caused endothelial cells to release significantly more vascular endothelial growth factor and interleukin (IL)-6, while cytokine stimulated CardAP-EVs significantly enhanced the release of IL-6 and IL-8. By nCounter® miRNA expression assay (NanoString Technologies) we identified microRNA 302d-3p to be enhanced in unstimulated CardAP-EVs compared to their cytokine stimulated counterparts, which was verified by quantitative polymerase chain reaction. This study demonstrates that both CardAP-EVs are pro-angiogenic by inducing different factors from endothelial cells. This would allow to select potent targets for a safe and efficient therapeutic application.
