Project description:Analysis of KhES-1 and H9 human ES cells in growth factors-dependent (E8) and -independent (AKIT) medium in feeder-free culture condition and KSR/bFGF medium on a feeder-layer. Results provide insight into genetic stability in different culture media/conditions.
Project description:Reliable, scalable and time-efficient culture methods are required to fully realize the clinical and industrial applications of human pluripotent stem (hPS) cells. Here we present a completely defined, xeno-free medium that supports long-term propagation of hPS cells on uncoated tissue-culture plastic. The medium consists of the Essential 8 (E8) formulation supplemented with Inter-α-inhibitor (IαI), a human serum-derived protein, recently demonstrated to activate key pluripotency pathways in mouse PS cells. IαI efficiently induces attachment and long-term growth of both embryonic and induced hPS cell lines when added as a soluble protein to the medium at seeding. IαI-supplementation efficiently supports adaptation of feeder-dependent hPS cells to xeno-free conditions, clonal growth as well as single-cell survival in the absence of Rho-associated kinase inhibitor (ROCKi). This time-efficient and simplified culture method paves the way for large-scale, high-throughput hPS cell culture, and will be valuable for both basic research and commercial applications.
Project description:Characterization of aortic valve interstitial cells (VICs) maintained in an optimized culture is es-sential for understanding the molecular mechanism underlying aortic valve stenosis. Here, we propose 2% hypoxia as an optimum VIC culture condition. Harvested leaflets from patients with aortic valve regurgitation were digested using collagenase and VICs were cultured under the 2% hypoxic condition. Significant elevation in VIC growth was observed in the 2% hypoxia group (hypo-VICs), compared to the normoxia group (normo-VICs). RNA-sequencing revealed that downregulation of oxidative stress-marker genes (such as superoxide dismutase) and upregula-tion of cell cycle accelerators (such as cyclins) were detected in hypo-VICs.