Electrophysiology, immunophenotype, and gene expression characterization of senescent and cryopreserved human amniotic fluid stem cells

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• 作者：Florin Iordache ; Andrei Constantinescu…
• 关键词：Amniotic fluid stem cell ; Cryopreserved cell ; Differential gene expression ; Immunophenotype ; Patch ; clamp ; RT ; PCR
• 刊名：The Journal of Physiological Sciences
• 出版年：2016
• 出版时间：November 2016
• 年：2016
• 卷：66
• 期：6
• 页码：463-476
• 全文大小：2,826 KB
• 刊物主题：Human Physiology; Neurosciences; Animal Biochemistry; Animal Physiology; Cell Physiology; Neurobiology;
• 出版者：Springer Japan
• ISSN：1880-6562
• 卷排序：66

文摘

We characterized human amniotic fluid stem cells (AFSC) in senescent cultures (6 weeks) versus cryopreserved cells using whole-cell patch-clamp, immunophenotyping, and differential gene expression profiling for senescence genes. We evidenced five ion current components (outward rectifier, A-type, inward rectifier, and big conductance Ca2+-dependent K+ currents, fast voltage-dependent Na+ currents). Senescent AFSC showed reduced expression of CD90, CD44, CD133, over 500-fold increase of interferon gamma and telomerase reverse transcriptase genes, increased cycle-dependent kinase 4 inhibitors, p53-binding protein 1, and decreased calreticulin and CD44. HLA-ABC immune expression was similar, and HLA-DR expression very low in both cell types. A subset of cryopreserved AFSC featured large inward rectifier K+ currents, voltage-dependent Na+ currents, and neural progenitor markers evidenced by immunophenotyping and RT-PCR. In all AFSC, in both culture conditions, at patch rupture the outward currents were very low, and they increased progressively over several minutes upon cytoplasm dialysis with pipette solution.