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  • Oct-4B isoform is differentially expressed in breast cancer cells: hypermethylation of regulatory elements of Oct-4A suggests an alternative promoter and transcriptional ... 20433421

    The human Oct-4 gene has three isoforms, Oct-4A, Oct-4B and Oct-4B1, which are thought to be derived from alternative splicing. It remains controversial whether the Oct-4 gene is expressed in cancer cells. Expression of Oct-4A is regulated by two elements, the PE (proximal enhancer) and DE (distal enhancer), but the expression and regulation of Oct-4B are not well known. Here, we firstly report that Oct-4B is expressed at low levels in MCF-7 cells, while the Oct-4A gene is inactivated. By analysing the function of different promoter constructs and the DNA methylation status of three regulatory regions, we demonstrate that the Oct-4A gene in MCF-7 cells is repressed by epigenetic control rather than transcriptional control. In addition, we speculate that the transcription of Oct-4B in MCF-7 cells is differentially regulated by additional regulatory elements. This work will enhance the understanding of Oct-4 gene in differential regulation.
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  • Generation of iPS Cells from Human Hair Follice Dermal Papilla Cells. 24772326

    Dermal papilla (DP) cells are unique regional stem cells of the skin that induce formation of a hair follicle and its regeneration cycle. DP are multipotent stem cells; therefore we supposed that the efficiency of DPC reprogramming could exceed that of dermal fibroblasts reprogramming. We generated induced pluripotent stem cells from human DP cells using lentiviral transfection with Oct4, Sox2, Klf4, and c-Myc, and cultivation of cells both in a medium supplemented with valproic acid and at a physiological level of oxygen (5%). The efficiency of DP cells reprogramming was ~0.03%, while the efficiency of dermal fibroblast reprogramming under the same conditions was ~0.01%. Therefore, we demonstrated the suitability of DP cells as an alternative source of iPS cells.
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  • Transcriptional profiling of ectoderm specification to keratinocyte fate in human embryonic stem cells. 25849374

    In recent years, several studies have shed light into the processes that regulate epidermal specification and homeostasis. We previously showed that a broad-spectrum γ-secretase inhibitor DAPT promoted early keratinocyte specification in human embryonic stem cells triggered to undergo ectoderm specification. Here, we show that DAPT accelerates human embryonic stem cell differentiation and induces expression of the ectoderm protein AP2. Furthermore, we utilize RNA sequencing to identify several candidate regulators of ectoderm specification including those involved in epithelial and epidermal development in human embryonic stem cells. Genes associated with transcriptional regulation and growth factor activity are significantly enriched upon DAPT treatment during specification of human embryonic stem cells to the ectoderm lineage. The human ectoderm cell signature identified in this study contains several genes expressed in ectodermal and epithelial tissues. Importantly, these genes are also associated with skin disorders and ectodermal defects, providing a platform for understanding the biology of human epidermal keratinocyte development under diseased and homeostatic conditions.
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  • The intracellular distribution of the ES cell totipotent markers OCT4 and Sox2 in adult stem cells differs dramatically according to commercial antibody used. 19199344

    To characterize ES cells, researchers have at their disposal a list of pluripotent markers, such as OCT4. In their quest to determine if adult stem cell populations, such as MSCs and ASCs, are pluripotent, several groups have begun to report the expression of these markers in these cells. Consistent with this, human ASCs (hASCs) are shown in this study to express a plethora of ES pluripotent markers at the gene and protein level, including OCT4, Sox2, and Nanog. When intracellular distribution is examined in hASCs, both OCT4 and Sox2 are expressed within the nuclei of hASCs, consistent with their expression patterns in ES cells. However, a significant amount of expression can be noted within the hASC cytoplasm and a complete absence of nuclear expression is observed for Nanog. Recent descriptions of OCT4 transcript variants may explain the cytoplasmic expression of OCT4 in hASCs and consistent with this, hASCs do express both the OCT4A and 4B transcript variants at the gene level. However, discrepancies arise when these three pluripotent markers are studied at the protein level. Specifically, distinct differences in intracellular expression patterns were noted for OCT4, Sox2, and Nanog from commercial antibody to commercial antibody. These antibody discrepancies persisted when hMSCs and rat ASCs and MSCs were examined. Therefore, confirming the expression of OCT4, Sox2, and Nanog in adult stem cells with today's commercial antibodies must be carefully considered before the designation of pluripotent can be granted.
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  • Targeting cancer stem cells through L1CAM suppresses glioma growth. 18676824

    Malignant gliomas are lethal cancers that display striking cellular heterogeneity. A highly tumorigenic glioma tumor subpopulation, termed cancer stem cells or tumor-initiating cells, promotes therapeutic resistance and tumor angiogenesis. Therefore, targeting cancer stem cells may improve patient survival. We interrogated the role of a neuronal cell adhesion molecule, L1CAM, in glioma stem cells as L1CAM regulates brain development and is expressed in gliomas. L1CAM(+) and CD133(+) cells cosegregated in gliomas, and levels of L1CAM were higher in CD133(+) glioma cells than normal neural progenitors. Targeting L1CAM using lentiviral-mediated short hairpin RNA (shRNA) interference in CD133(+) glioma cells potently disrupted neurosphere formation, induced apoptosis, and inhibited growth specifically in glioma stem cells. We identified a novel mechanism for L1CAM regulation of cell survival as L1CAM knockdown decreased expression of the basic helix-loop-helix transcription factor Olig2 and up-regulated the p21(WAF1/CIP1) tumor suppressor in CD133(+) glioma cells. To determine if targeting L1CAM was sufficient to reduce glioma stem cell tumor growth in vivo, we targeted L1CAM in glioma cells before injection into immunocompromised mice or directly in established tumors. In each glioma xenograft model, shRNA targeting of L1CAM expression in vivo suppressed tumor growth and increased the survival of tumor-bearing animals. Together, these data show that L1CAM is required for maintaining the growth and survival of CD133(+) glioma cells both in vitro and in vivo, and L1CAM may represent a cancer stem cell-specific therapeutic target for improving the treatment of malignant gliomas and other brain tumors.
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