68 research outputs found

    Culture of human pluripotent stem cells using completely defined conditions on a recombinant E-cadherin substratum

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    <p>Abstract</p> <p>Background</p> <p>To maintain pluripotency of human embryonic stem (huES) cells in feeder-free culture it has been necessary to provide a Matrigel substratum, which is a complex of poorly defined extracellular matrices and growth factors derived from mouse Engelbreth-Holm-Swarm sarcoma cells. Culture of stem cells under ill-defined conditions can inhibit the effectiveness of maintaining cells in a pluripotent state and reduce reproducibility of differentiation protocols. Moreover recent batches of Matrigel have been found to be contaminated with the single stranded RNA virus, Lactate Dehydrogenase Elevating Virus (LDEV), raising concerns regarding the safety of using stem cells that have been cultured on Matrigel in a therapeutic setting. To circumvent such concerns, we attempted to identify a recombinant matrix that could be used as an alternative to Matrigel for the culture of human pluripotent stem cells. huES and human induced pluripotent stem (hiPS) cells were grown on plates coated with a fusion protein consisting of E-cadherin and the IgG Fc domain using mTeSR1 medium.</p> <p>Results</p> <p>Cells grown under these conditions maintained similar morphology and growth rate to those grown on Matrigel and retained all pluripotent stem cell features, including an ability to differentiate into multiple cell lineages in teratoma assays. We, therefore, present a culture system that maintains the pluripotency of huES and hiPS cells under completely defined conditions.</p> <p>Conclusions</p> <p>We propose that this system should facilitate growth of stem cells using good manufacturing practices (GMP), which will be necessary for the clinical use of pluripotent stem cells and their derivatives.</p

    E-Cadherin-Coated Plates Maintain Pluripotent ES Cells without Colony Formation

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    Embryonic stem (ES) cells cultured on gelatin-coated plates or feeder layers form tight aggregated colonies by the E-cadherin-mediated cell-cell adhesions. Here we show that murine ES cells do not make cell-cell contacts or form colonies when cultured on the plate coated with a fusion protein of E-cadherin and IgG Fc domain. The cells in culture retain all ES cell features including pluripotency to differentiate into cells of all three germ layers and germ-line transmission after extended culture. Furthermore, they show a higher proliferative ability, lower dependency on LIF, and higher transfection efficiency than colony-forming conditions. Our results suggest that aggregated colony formation might inhibit diffusion of soluble factors and increase cell-cell communication, which may result in a heterogeneous environment within and between surrounding cells of the colony. This method should enable more efficient and scalable culture of ES cells, an important step towards the clinical application of these cells

    Being Praised for Prosocial Behaviors Longitudinally Reduces Depressive Symptoms in Early Adolescents: A Population-Based Cohort Study

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    BackgroundDepression is highly prevalent and causes a heavy burden in adolescent life. Being praised for prosocial behavior might be a preventive factor because both being praised and prosocial behavior are protective against depression. Here, we investigated the longitudinal relationship between being praised for prosocial behavior and depressive symptoms in adolescents.MethodsIn Tokyo Teen Cohort study (TTC), an ongoing prospective population-based cohort study, we collected 3,171 adolescents' data on self-reported experiences of being praised for prosocial behavior, depressive symptoms, and caregiver-evaluated prosocial behavior. Ten-year-old children were asked to freely describe answers to the question β€œWhat are you praised for?”. Only children who clearly answered that they were praised for their prosocial behavior were designated the β€œprosocial praise group.” The degree of depression at ages 10 and 12 was measured with the Short Mood and Feelings Questionnaire (SMFQ), a self-report questionnaire about depression. Objective prosocial behavior of the 10 year-old children was assessed by the Strength and Difficulty Questionnaire (SDQ). Multiple linear regression analysis was performed using the SMFQ score at age 12 as the objective variable and being praised for prosocial behavior as the main explanatory variable, and the SMFQ score at age 10 and the objective prosocial behavior at age 10 were included as confounders.ResultsDepressive symptoms (SMFQ scores) in the β€œprosocial praise group” were significantly lower than those in the other group both at age 10 (4.3 Β± 4.4 vs. 4.9 Β± 4.6, p &lt; 0.001) and at age 12 (3.4 Β± 4.2 vs. 4.0 Β± 4.6, p &lt; 0.01). In the single regression analysis, the children who reported being praised for prosocial behavior at age 10 had significantly lower depressive symptoms at age 12 (partial regression variable: βˆ’0.57, 95% confidence interval (CI) [βˆ’0.96, βˆ’0.17]). This association remained significant after adjusting for confounders, including baseline depressive symptoms (partial regression variable: βˆ’0.44, 95% CI [βˆ’0.80, βˆ’0.08]). Prosocial behavior alone was not associated with depressive symptoms.ConclusionsBeing praised for prosocial behavior rather than objective prosocial behavior at 10 years of age predicted lower depressive symptoms 2 years later. Praise for adolescents' prosocial behavior can be encouraged to prevent depression

    Influenza A (H3N2) infection followed by anti-signal recognition particle antibody-positive necrotizing myopathy: A case report

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    A 60-year-old Japanese woman presented with subacute progressive muscle pain and weakness in her proximal extremities. She was diagnosed with influenza A (H3N2) infection a week before the onset of muscle pain. At the time of admission, she exhibited weakness in the proximal muscles of the upper and lower limbs, elevated serum liver enzymes and creatinine kinase, and myoglobinuria. She did not manifest renal failure and cardiac abnormalities, indicating myocarditis.Electromyography revealed myogenic changes, and magnetic resonance imaging of the upper limb showed abnormal signal intensities in the muscles, suggestive of myopathy. Muscle biopsy of the biceps revealed numerous necrotic regeneration fibers and mild inflammatory cell infiltration, suggesting immune-mediated necrotizing myopathy (IMNM). Necrotized muscle cells were positive for human influenza A (H3N2). Autoantibody analysis showed the presence of antibodies against the signal recognition particle (SRP), and the patient was diagnosed with anti-SRP-associated IMNM. She was resistant to intravenous methylprednisolone pulse therapy but recovered after administration of oral systemic corticosteroids and immunoglobulins. We speculate that the influenza A (H3N2)infection might have triggered her IMNM. Thus, IMNM should be considered as a differential diagnosis in patients with proximal muscle weakness that persists after viral infections

    Involvement of SIK3 in Glucose and Lipid Homeostasis in Mice

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    Salt-inducible kinase 3 (SIK3), an AMP-activated protein kinase-related kinase, is induced in the murine liver after the consumption of a diet rich in fat, sucrose, and cholesterol. To examine whether SIK3 can modulate glucose and lipid metabolism in the liver, we analyzed phenotypes of SIK3-deficent mice. Sik3βˆ’/βˆ’ mice have a malnourished the phenotype (i.e., lipodystrophy, hypolipidemia, hypoglycemia, and hyper-insulin sensitivity) accompanied by cholestasis and cholelithiasis. The hypoglycemic and hyper-insulin-sensitive phenotypes may be due to reduced energy storage, which is represented by the low expression levels of mRNA for components of the fatty acid synthesis pathways in the liver. The biliary disorders in Sik3βˆ’/βˆ’ mice are associated with the dysregulation of gene expression programs that respond to nutritional stresses and are probably regulated by nuclear receptors. Retinoic acid plays a role in cholesterol and bile acid homeostasis, wheras ALDH1a which produces retinoic acid, is expressed at low levels in Sik3βˆ’/βˆ’ mice. Lipid metabolism disorders in Sik3βˆ’/βˆ’ mice are ameliorated by the treatment with 9-cis-retinoic acid. In conclusion, SIK3 is a novel energy regulator that modulates cholesterol and bile acid metabolism by coupling with retinoid metabolism, and may alter the size of energy storage in mice
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