Current Issue : July - September Volume : 2014 Issue Number : 3 Articles : 5 Articles
Introduction: Human induced pluripotent stem cells (hiPSCs) offer great promise for regenerative therapies or\r\nin vitro modelling of neurodegenerative disorders like Parkinsonâ��s disease. Currently, widely used cell sources for the\r\ngeneration of hiPSCs are somatic cells obtained from aged individuals. However, a critical issue concerning the\r\npotential clinical use of these iPSCs is mutations that accumulate over lifetime and are transferred onto iPSCs\r\nduring reprogramming which may influence the functionality of cells differentiated from them. The aim of our\r\nstudy was to establish a differentiation strategy to efficiently generate neurons including dopaminergic cells from\r\nhuman cord blood-derived iPSCs (hCBiPSCs) as a juvenescent cell source and prove their functional maturation\r\nin vitro.\r\nMethods: The differentiation of hCBiPSCs was initiated by inhibition of transforming growth factor-�Ÿ and bone\r\nmorphogenetic protein signaling using the small molecules dorsomorphin and SB 431542 before final maturation\r\nwas carried out. hCBiPSCs and differentiated neurons were characterized by immunocytochemistry and quantitative\r\nreal time-polymerase chain reaction. Since functional investigations of hCBiPSC-derived neurons are indispensable\r\nprior to clinical applications, we performed detailed analysis of essential ion channel properties using whole-cell\r\npatch-clamp recordings and calcium imaging.\r\nResults: A Sox1 and Pax6 positive neuronal progenitor cell population was efficiently induced from hCBiPSCs using\r\na newly established differentiation protocol. Neuronal progenitor cells could be further maturated into dopaminergic\r\nneurons expressing tyrosine hydroxylase, the dopamine transporter and engrailed 1. Differentiated hCBiPSCs exhibited\r\nvoltage-gated ion currents, were able to fire action potentials and displayed synaptic activity indicating synapse\r\nformation. Application of the neurotransmitters GABA, glutamate and acetylcholine induced depolarizing calcium signal\r\nchanges in neuronal cells providing evidence for the excitatory effects of these ligand-gated ion channels during\r\nmaturation in vitro.\r\nConclusions: This study demonstrates for the first time that hCBiPSCs can be used as a juvenescent cell source to\r\ngenerate a large number of functional neurons including dopaminergic cells which may serve for the development of\r\nnovel regenerative treatment strategies....
Introduction: Congenital muscular dystrophies (CMD) are a clinically and genetically heterogeneous group of\r\nneuromuscular disorders characterized by muscle weakness within the first two years of life. Collagen VI-related\r\nmuscle disorders have recently emerged as one of the most common types of CMD. COL6 CMD is caused by\r\ndeficiency and/or dysfunction of extracellular matrix (ECM) protein collagen VI. Currently, there is no specific\r\ntreatment for this disabling and life-threatening disease. The primary cellular targets for collagen VI CMD therapy\r\nare fibroblasts in muscle, tendon and skin, as opposed to muscle cells for other types of muscular dystrophies.\r\nHowever, recent advances in stem cell research have raised the possibility that use of adult stem cells may provide\r\ndramatic new therapies for treatment of COL6 CMD.\r\nMethods: Here, we developed a procedure for isolation of human stem cells from the adipose layer of neonatal\r\nskin. The adipose-derived stem cells (ADSC) were examined for expression of ECM and related genes using gene\r\nexpression array analysis. The therapeutic potential of ADSC was assessed after a single intramuscular transplantation\r\nin collagen VI-deficient mice.\r\nResults: Analysis of primary cultures confirmed that established ADSC represent a morphologically homogenous\r\npopulation with phenotypic and functional features of adult mesenchymal stem cells. A comprehensive gene\r\nexpression analysis showed that ADSC express a vast array of ECM genes. Importantly, it was observed that ADSC\r\nsynthesize and secrete all three collagen VI chains, suggesting suitability of ADSC for COL6 CMD treatment.\r\nFurthermore, we have found that a single intramuscular transplantation of ADSC into Col6a1-/-Rag1-/- mice under\r\nphysiological and cardiotoxin-induced injury/regeneration conditions results in efficient engraftment and migration\r\nof stem cells within the skeletal muscle. Importantly, we showed that ADSC can survive long-term and continuously\r\nsecrete the therapeutic collagen VI protein missing in the mutant mice.\r\nConclusions: Overall, our findings suggest that stem cell therapy can potentially provide a new avenue for the\r\ntreatment of COL6 CMD and other muscular disorders and injuries....
Introduction: The prevalence of nonhealing wounds is predicted to increase due to the growing aging population.\r\nDespite the use of novel skin substitutes and wound dressings, poorly vascularized wound niches impair wound\r\nrepair. Mesenchymal stem cells (MSCs) have been reported to provide paracrine signals to promote wound healing,\r\nbut the effect of human Whartonâ��s jelly-derived MSCs (WJ-MSCs) has not yet been described in human normal skin.\r\nThe aim of this study is to examine the effects of human WJ-MSC paracrine signaling on normal skin fibroblasts\r\nin vitro, and in an in vivo preclinical model.\r\nMethods: Human WJ-MSCs and normal skin fibroblasts were isolated from donated umbilical cords and normal\r\nadult human skin. Fibroblasts were treated with WJ-MSC-conditioned medium (WJ-MSC-CM) or nonconditioned\r\nmedium.\r\nResults: Expression of genes involved in re-epithelialization (transforming growth factor-�Ÿ2), neovascularization\r\n(hypoxia-inducible factor-1a) and fibroproliferation (plasminogen activator inhibitor-1) was upregulated in WJ-MSCCM-\r\ntreated fibroblasts (P = 0.05). WJ-MSC-CM enhanced normal skin fibroblast proliferation (P = 0.001) and migration\r\n(P = 0.05), and promoted wound healing in an excisional full-thickness skin murine model.\r\nConclusions: Under our experimental conditions, WJ-MSCs enhanced skin wound healing in an in vivo mouse model....
Introduction: Rapid establishment of functional blood vessels is a prerequisite for successful tissue engineering.\r\nDuring vascular development, endothelial cells (ECs) and perivascular cells assemble into a complex regulating\r\nproliferation of ECs, vessel diameter and production of extracellular matrix proteins. The aim of this study was to\r\nevaluate the ability of mesenchymal stem cells (MSCs) to establish an endothelial-perivascular complex in\r\ntissue-engineered constructs comprising ECs and MSCs.\r\nMethods: Primary human ECs and MSCs were seeded onto poly(L-lactide-co-1,5-dioxepan-2-one) (poly(LLAco-\r\nDXO)) scaffolds and grown in dynamic culture before subcutaneous implantation in immunocompromised mice\r\nfor 1 and 3 weeks. Cellular activity, angiogenic stimulation and vascular assembly in cell/scaffold constructs seeded\r\nwith ECs or ECs/MSCs in a 5:1 ratio was monitored with real-time RT-PCR, ELISA and immunohistochemical\r\nmicroscopy analysis.\r\nResults: A quiescent phenotype of ECs was generated, by adding MSCs to the culture system. Decreased\r\nproliferation of ECs, in addition to up-regulation of selected markers for vascular maturation was demonstrated.\r\nBaseline expression of VEGFa was higher for MSCs compared with EC (P <0.001), with subsequent up-regulated\r\nVEGFa-expression for EC/MSC constructs before (P <0.05) and after implantation (P <0.01). Furthermore, an\r\ninflammatory response with CD11b + cells was generated from implantation of human cells. At the end of the\r\n3 week experimental period, a higher vascular density was shown for both cellular constructs compared with empty\r\ncontrol scaffolds (P <0.01), with the highest density of capillaries being generated in constructs comprising both ECs\r\nand MSCs....
Introduction: Stem cells are being investigated as catalysts of tissue regeneration to either directly replace or\r\npromote cellularity lost as a result of traumatic injury or degenerative disease. In many reports, despite low\r\nnumbers of stably integrated cells, the transient presence of cells delivered or recruited to sites of tissue\r\nremodeling globally benefits functional recovery. Such findings have motivated the need to determine how\r\nparacrine factors secreted from transplanted cells may be capable of positively impacting endogenous repair\r\nprocesses and somatic cell responses.\r\nMethods: Embryonic stem cells were differentiated as embryoid bodies (EBs) in vitro and media conditioned by EBs\r\nwere collected at different intervals of time. Gene and protein expression analysis of several different growth factors\r\nsecreted by EBs were examined by polymerase chain reaction and enzyme-linked immunosorbent assay analysis,\r\nrespectively, as a function of time. The proliferation and migration of fibroblasts and endothelial cells treated with\r\nEB conditioned media was examined compared with unconditioned and growth media controls.\r\nResults: The expression of several growth factors, including bone morphogenic protein-4, insulin-like growth factors\r\nand vascular endothelial growth factor-A, increased during the course of embryonic stem cell (ESC) differentiation\r\nas EBs. Conditioned media collected from EBs at different stages of differentiation stimulated proliferation and\r\nmigration of both fibroblasts and endothelial cells, based on 5-bromo-2'-deoxyuridine incorporation and transwell\r\nassays, respectively.\r\nConclusions: Overall, these results demonstrate that differentiating ESCs express increasing amounts of various\r\ngrowth factors over time that altogether are capable of stimulating mitogenic and motogenic activity of exogenous\r\ncell populations....
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