The multidifferentiation ability of mesenchymal stem cells holds great promise for cell therapy. Numerous studies have focused on the establishment of differentiation protocols, whereas little attention has been paid to the metabolic changes during the differentiation process. Mitochondria, the powerhouse of mammalian cells, vary in their number and function in different cell types with different energy demands, but how these variations are associated with cell differentiation remains elusive. In this study, we investigated the changes of mitochondrial biogenesis and bioenergetic function using human mesenchymal stem cells (hMSCs) because of their well-defined differentiation potentials. Upon osteogenic induction, the copy number of mitochondrial DNA, protein subunits of the respiratory enzymes, oxygen consumption rate, and intracellular ATP content were increased, indicating the upregulation of aerobic mitochondrial metabolism. On the other hand, undifferentiated hMSCs showed higher levels of glycolytic enzymes and lactate production rate, suggesting that hMSCs rely more on glycolysis for energy supply in comparison with hMSC-differentiated osteoblasts. In addition, we observed a dramatic decrease of intracellular reactive oxygen species (ROS) as a consequence of upregulation of two antioxidant enzymes, manganesedependent superoxide dismutase and catalase. Finally, we found that exogenous H 2 O 2 and mitochondrial inhibitors could retard the osteogenic differentiation. These findings suggested an energy production transition from glycolysis to oxidative phosphorylation in hMSCs upon osteogenic induction. Meanwhile, antioxidant enzymes were concurrently upregulated to prevent the accumulation of intracellular ROS. Together, our findings suggest that coordinated regulation of mitochondrial biogenesis and antioxidant enzymes occurs synergistically during osteogenic differentiation of hMSCs. STEM CELLS 2008;26:960 -968 Disclosure of potential conflicts of interest is found at the end of this article.
a b s t r a c tThe helicase domain of dengue virus NS3 protein (DENV NS3H) contains RNA-stimulated nucleoside triphosphatase (NTPase), ATPase/helicase, and RNA 5 0 -triphosphatase (RTPase) activities that are essential for viral RNA replication and capping. Here, we show that DENV NS3H unwinds 3 0 -tailed duplex with an RNA but not a DNA loading strand, and the helicase activity is poorly processive. The substrate of the divalent cation-dependent RTPase activity is not restricted to viral RNA 5 0 -terminus, a protruding 5 0 -terminus made the RNA 5 0 -triphosphate readily accessible to DENV NS3H. DENV NS3H preferentially binds RNA to DNA, and the functional interaction with RNA is sensitive to ionic strength.
Tumor cells have long been observed to share several biological characteristics with normal stem/progenitor cells; however, the oncogenic mechanisms underlying the lung stem/progenitor cell signaling remain elusive. Here, we report that SOX2, a self-renewal factor in lung stem/progenitor cells, is highly expressed in a subclass of lung cancer cells, the proliferation, survival, and chemoresistance of which are dependent on SOX2 signaling. Overexpression of SOX2 promotes oncogenic phenotypes in lung cancer cells; knockdown of SOX2 attenuated cell proliferation. We observed that SOX2 increased the expression of epidermal growth factor receptor (EGFR), and EGFR activation further upregulated SOX2 levels, forming a positive feedback loop. SOX2 expression promoted chemoresistance, and silencing of SOX2 perturbed mitochondrial function, causing marked apoptosis and autophagy. SOX2 induced BCL2L1, the ectopic expression of which rescued the effects of SOX2 silencing on apoptosis, autophagy, and mitochondrial function. SOX2 promoted tumor formation, along with increased cell proliferation in a xenograft mouse model. SOX2 expression is associated with poor prognosis in lung cancer patients; moreover, SOX2, EGFR, and BCL2L1 expression levels were significantly correlated in lung tumors. Our findings support the emerging role of SOX2 in cell proliferation and survival by eliciting oncogenic EGFR and BCL2L1 signaling with potential applications as a prognosis marker and a therapeutic target in lung cancer. STEM CELLS
Myoclonus epilepsy associated with ragged-red fibers (MERRF) is a mitochondrial disorder characterized by myoclonus epilepsy, generalized seizures, ataxia and myopathy. MERRF syndrome is primarily due to an A to G mutation at mtDNA 8344 that disrupts the mitochondrial gene for tRNA(Lys). However, the detailed mechanism by which this tRNA(Lys) mutation causes mitochondrial dysfunction in cardiomyocytes or neurons remains unclear. In this study, we generated human induced pluripotent stem cells (hiPSCs) that carry the A8344G genetic mutation from patients with MERRF syndrome. Compared with mutation-free isogenic hiPSCs, MERRF-specific hiPSCs (MERRF-hiPSCs) exhibited reduced oxygen consumption, elevated reactive oxygen species (ROS) production, reduced growth, and fragmented mitochondrial morphology. We sought to investigate the induction ability and mitochondrial function of cardiomyocyte-like cells differentiated from MERRF-hiPSCs. Our data demonstrate that that cardiomyocyte-like cells (MERRF-CMs) or neural progenitor cells (MERRF-NPCs) differentiated from MERRF-iPSCs also exhibited increased ROS levels and altered antioxidant gene expression. Furthermore, MERRF-CMs or -NPCs contained fragmented mitochondria, as evidenced by MitoTracker Red staining and transmission electron microscopy. Taken together, these findings showed that MERRF-hiPSCs and MERRF-CM or –NPC harboring the A8344G genetic mutation displayed contained mitochondria with an abnormal ultrastructure, produced increased ROS levels, and expressed upregulated antioxidant genes.
MERRF syndrome is predominantly caused by A8344G mutation in the mitochondrial DNA (mtDNA), affecting MT-TK gene, which impairs the mitochondrial electron transport chain function. Here, we report the generation of two isogenic induced pluripotent stem cell (iPSC) lines, TVGH-iPSC-MRF-M and TVGH-iPSC-MRF-M, from the skin fibroblasts of a female MERRF patient harboring mtDNA A8344G mutation by using retrovirus transduction system. Both cell lines share the same genetic background except containing different proportions of mtDNA with the A8344G mutation. Both cell lines exhibited the pluripotency and capacity to differentiate into three germ layers.
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