Reprogramming of T Cells from Human Peripheral Blood

Division of Pediatric Hematology Oncology, Department of Biological Chemistry and Molecular Pharmacology, Children's Hospital Boston and Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115, USA.
Cell stem cell (Impact Factor: 22.27). 07/2010; 7(1):15-9. DOI: 10.1016/j.stem.2010.06.004
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    • "Each of these methods has its advantages and disadvantages. Peripheral blood is an advantageous alternative to skin for hIPSC derivation (Loh et al., 2010; Zhang 2013) since it is widely used in clinical diagnostics, and moreover, the method of blood collection is standardized and relatively less traumatic than skin biopsy. "
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    ABSTRACT: The collection sites of human primary tissue samples and the receiving laboratories, where the human induced pluripotent stem cells (hIPSCs) are derived, are often not on the same site. Thus, the stability of samples prior to derivation constrains the distance between the collection site and the receiving laboratory. To investigate sample stability, we collected blood and held it at room temperature for 5, 24, or 48 hr before isolating peripheral blood mononuclear cells (PBMCs) and reprogramming into IPSCs. Additionally, PBMC samples at 5- and 48-hr time points were frozen in liquid nitrogen for 4 months and reprogrammed into IPSCs. hIPSC lines derived from all time points were pluripotent, displayed no marked difference in chromosomal aberration rates, and differentiated into three germ layers. Reprogramming efficiency at 24- and 48-hr time points was 3- and 10-fold lower, respectively, than at 5 hr; the freeze-thaw process of PBMCs resulted in no obvious change in reprogramming efficiency.
    Full-text · Article · Sep 2015 · Stem Cell Reports
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    • "Dramatic cell-state transitions have been achieved in vitro and in vivo through the enforced expression of transcription factors. For example, differentiated somatic cells—fibroblasts (Takahashi and Yamanaka, 2006), keratinocytes (Aasen et al., 2008), peripheral blood (Loh et al., 2010; Staerk et al., 2010), and neural progenitors (Kim et al., 2009)—have been reprogrammed to pluripotent stem cells; fibroblasts have been converted to cells resembling myoblasts (Davis et al., 1987), motor neurons (Vierbuchen et al., 2010), cardiomyocytes (Ieda et al., 2010), hepatocytes (Huang et al., 2011; Sekiya and Suzuki, 2011), and blood progenitors (Szabo et al., 2010); B cells have been converted to macrophage-like cells (Xie et al., 2004); and exocrine pancreas cells have been converted to insulin-producing beta cells (Zhou et al., 2008). Furthermore , pluripotent stem cells can be coaxed to specific lineages through a combination of defined growth conditions and ectopic gene expression (Murry and Keller, 2008). "
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    ABSTRACT: Somatic cell reprogramming, directed differentiation of pluripotent stem cells, and direct conversions between differentiated cell lineages represent powerful approaches to engineer cells for research and regenerative medicine. We have developed CellNet, a network biology platform that more accurately assesses the fidelity of cellular engineering than existing methodologies and generates hypotheses for improving cell derivations. Analyzing expression data from 56 published reports, we found that cells derived via directed differentiation more closely resemble their in vivo counterparts than products of direct conversion, as reflected by the establishment of target cell-type gene regulatory networks (GRNs). Furthermore, we discovered that directly converted cells fail to adequately silence expression programs of the starting population and that the establishment of unintended GRNs is common to virtually every cellular engineering paradigm. CellNet provides a platform for quantifying how closely engineered cell populations resemble their target cell type and a rational strategy to guide enhanced cellular engineering.
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    • "T cells are clinically suitable for iPSC generation because of their ease of sampling and culturing[5]. T cells have been successfully reprogrammed into a pluripotent state with retrovirus[7], [10], lentivirus[4], [6], or Sendai virus[5] using MEF feeder layers for the reprogramming culture conditions. Here, we report using a combination of SeV and activated T cells to successfully reprogram human T cells into a pluripotent state using defined culture conditions with verification of rearranged TCR genes. "
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    ABSTRACT: Recently, induced pluripotent stem cells (iPSCs) were established as promising cell sources for revolutionary regenerative therapies. The initial culture system used for iPSC generation needed fetal calf serum in the culture medium and mouse embryonic fibroblast as a feeder layer, both of which could possibly transfer unknown exogenous antigens and pathogens into the iPSC population. Therefore, the development of culture systems designed to minimize such potential risks has become increasingly vital for future applications of iPSCs for clinical use. On another front, although donor cell types for generating iPSCs are wide-ranging, T cells have attracted attention as unique cell sources for iPSCs generation because T cell-derived iPSCs (TiPSCs) have a unique monoclonal T cell receptor genomic rearrangement that enables their differentiation into antigen-specific T cells, which can be applied to novel immunotherapies. In the present study, we generated transgene-free human TiPSCs using a combination of activated human T cells and Sendai virus under defined culture conditions. These TiPSCs expressed pluripotent markers by quantitative PCR and immunostaining, had a normal karyotype, and were capable of differentiating into cells from all three germ layers. This method of TiPSCs generation is more suitable for the therapeutic application of iPSC technology because it lowers the risks associated with the presence of undefined, animal-derived feeder cells and serum. Therefore this work will lead to establishment of safer iPSCs and extended clinical application.
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