Current Issue : January - March Volume : 2013 Issue Number : 1 Articles : 6 Articles
Introduction: The capacity of bone marrow mesenchymal stromal cells (BMSCs) to be induced into chondrocytes has\r\ndrawn much attention for cell-based cartilage repair. BMSCs represent a small proportion of cells of the bone marrow\r\nstromal compartment and, thus, culture expansion is a necessity for therapeutic use. However, there is no consensus on\r\nhow BMSCs should be isolated nor expanded to maximize their chondrogenic potential. During embryonic\r\ndevelopment pluripotent stem cells differentiate into chondrocytes and form cartilage in a hypoxic microenvironment.\r\nMethods: Freshly harvested human BMSCs were isolated and expanded from the aspirates of six donors, under\r\neither hypoxic conditions (3% O2) or normoxic conditions (21% O2). A colony-forming unit fibroblastic (Cfu-f) assay\r\nwas used to determine the number of cell colonies developed from each donor. BMSCs at passage 2 (P2) were\r\ncharacterized by flow cytometry for the phenotypic expression of cell surface markers on mesenchymal stem cells.\r\nBMSCs at P2 were subsequently cultured in vitro as three-dimensional cell pellets in a defined serum-free\r\nchondrogenic medium under normoxic and hypoxic conditions. Chondrogenic differentiation of the BMSCs was\r\ncharacterized by biochemical and histological methods and by quantitative gene-expression analysis.\r\nResults: After 14 days of culture, the number of BMSC colonies developed under hypoxia was generally higher (8%\r\nto 38% depending on donor) than under normoxia. BMSCs were positive for the cell surface markers CD13, CD29,\r\nCD44, CD73, CD90, CD105 and CD151, and negative for CD34. Regardless of the oxygen tension during pellet\r\nculture, hypoxia-expanded BMSC pellets underwent a more robust chondrogenesis than normoxia-expanded BMSC\r\npellets after three weeks of culture, as judged by increased glycosaminoglycan synthesis and Safranin O staining,\r\nalong with increased mRNA expression of aggrecan, collagen II and Sox9. Hypoxic conditions enhanced the mRNA\r\nexpression of hypoxia inducible factor-2 alpha (HIF-2a) but suppressed the mRNA expression of collagen X in\r\nBMSC pellet cultures regardless of the oxygen tension during BMSC isolation and propagation.\r\nConclusions: Taken together, our data demonstrate that isolation and expansion of BMSCs under hypoxic\r\nconditions augments the chondrogenic potential of BMSCs. This suggests that hypoxia-mediated isolation and\r\nexpansion of BMSCs may improve clinical applications of BMSCs for cartilage repair....
Hypoxic-brain injury is a major cause of neonatal morbidity and mortality. However, melatonin (N-acetyl-5-\r\nmethoxytryptamine) has been identified as an indirect anti-oxidant and direct free radical scavenger that could\r\npossibly reduce the injurious effects of hypoxic-ischemic brain injury in neonatal infants. Hypoxia-ischemia leads\r\nto multiple consequences such as an increase in extracellular glutamate. Yet the many mechanisms involved in\r\nmelatonin-induced neuroprotection are still under investigation. We have hypothesized that melatonin could induce\r\nneuroprotection by increasing levels of cystine glutamate exchanger (xCT), an amino acid transporter as shown in\r\nprevious work in our laboratory. Mouse neural stem cells were used for all in vitro studies for western blot analysis.\r\nIn dose-response studies, melatonin increases xCT expression by 2.43 �± 0.81, 3.58 �± 0.6, 3.21 �± 1.13, 3.30 �± 0.96\r\nand 3.48 �± 0.30 (p < 0.01) folds at 1 nM, 10 nM, 100 nM, 1 �µM and 10 �µM concentrations respectively in neural stem\r\ncells. In time-course studies, melatonin increases xCT by 2.60 �± 0.97, 2.65 �± 0.27, 3.29 �± 0.40, and 3.57 �± 0.60 fold\r\nat 4 hours, 8 hours, 12 hours, and 24 hours. Melatonin increases cystine uptake. System Xc inhibition decreased\r\ncell viability. These results suggest that melatonin may induce neuroprotection by increasing xCT expression and\r\nactivity....
Despite state-of-the-art methods for the early diagnosis and treatment, heart failure resulting from myocardial\r\ninfarction (MI) continues to be a major cause of morbidity and mortality worldwide. Most of the existing treatment\r\nmodalities against heart failure are symptom-based, short-term and do not prolong survival. Stem cell-based therapy\r\nis promising strategy to lead to cardiac repair after MI. Over the last decade, stem cells with diverse origin, identity,\r\nand plasticity have been utilized for the regeneration and repair of damaged myocardium after MI, both in animal\r\nmodels and humans. The major challenges and dilemmas in stem cell therapy after MI included- ethical concerns\r\nand alloreactivity (with embryonic stem cells), malignant transformation and vector contamination (with inducible\r\nprogenitor cells), coronary restenosis (with mobilization of bone marrow stem cells), and cardiac arrhythmias and\r\nstructural heterogeneity due to non-coupling of cardiac and non-cardiac skeletal cells (with skeletal myoblasts).\r\nTherefore, as much as the progress made in the field of cardiac regenerative therapy, questions have been asked on\r\nwhat constitutes the most appropriate source for the stem cells. In particular, the identity, characteristics and ability\r\nof the stem cells to retain their fate while being propagated ex vivo have invited a passionate discussion among\r\ncell-biologists, geneticists and clinicians. This review summarizes the diverse origin of the stem cells and discusses\r\nrecent advances made for the identification, selection and propagation of stem cells for the regeneration or repair of\r\ndamaged myocardium after MI....
Aim: To evaluate the outcomes of Conjunctival Limbal Autograft (CLAU) and Conjunctival Limbal Allograft\r\n(CLAL) transplants for the treatment of partial or total Limbal Stem Cell Deficiency (LSCD).\r\nMethods: Retrospective, cohort study. All eyes treated with limbal stem cell transplant (LSCT) that had 1 year\r\nfollow-up or more were included. Visual success was measured by improvement in VA post operatively. Surgical\r\nsuccess was defined as maintaining a healthy clear corneal surface post operatively.\r\nResults: There were 8 autolimbal and 9 allolimbal transplants. Of the latter, 8 were from living related donors\r\n(Lr-CLAL) and one was Keratolimbal Allograft (KLAL). Fifteen eyes had total LSCD and two eyes had partial LSCD.\r\nPrimary diagnosis included combined chemical & thermal Injury burn (n=13), vernal keratoconjunctivits (n=2), herpes\r\nsimplex infection (n=1) and idiopathic (n=1). Mean post operative follow-up was 50.65 �± 34.68 months (range 12-\r\n108 months). CLAU was successful in 7 out of 8 eyes (87.5%). Mean VA improved from 0.1 �± 0.12 to 0.44 �± 0.28\r\n(measured in decimal fraction). CLAL was successful in 2 out of 9 eyes (22.2%). Mean VA improved from 0.03 �± 0.04\r\nto 0.10 �± 0.22. All eyes with CLAU achieved re-epithelialization and maintained an intact epithelium. Eyes with CLAL\r\nachieved re-epithelialization and maintained an intact epithelium in 66.7% (6).\r\nConclusion: LSCT is an effective modality of treatment in patients with LSCD. CLAU transplant and absence of\r\npost operative complications were associated with statistically higher success rate. Younger patients and wet ocular\r\nsurface had more favorable outcome....
Introduction: Mesenchymal stem cells (MSCs) are multipotent cells able to differentiate into several mesenchymal\r\nlineages, classically derived from bone marrow (BM) but potentially from umbilical cord blood (UCB). Although they\r\nare becoming a good tool for regenerative medicine, they usually need to be expanded in fetal bovine serum\r\n(FBS)-supplemented media. Human platelet lysate (HPL) has recently been proposed as substitute for safety\r\nreasons, but it is not yet clear how this supplement influences the properties of expanded MSCs.\r\nMethods: In the present study, we compared the effect of various media combining autologous HPL with or\r\nwithout FBS on phenotypic, proliferative and functional (differentiation, cytokine secretion profile) characteristics of\r\nhuman BM-derived MSCs.\r\nResults: Despite less expression of adipogenic and osteogenic markers, MSCs cultured in HPL-supplemented media\r\nfully differentiated along osteoblastic, adipogenic, chondrogenic and vascular smooth muscle lineages. The analyses\r\nof particular specific proteins expressed during osteogenic differentiation (calcium-sensing receptor (CaSR) and\r\nparathormone receptor (PTHR)) showed their decrease at D0 before any induction for MSC cultured with HPL\r\nmostly at high percentage (10%HPL). The cytokine dosage showed a clear increase of proliferation capacity and\r\ninterleukin (IL)-6 and IL-8 secretion.\r\nConclusions: This study shows that MSCs can be expanded in media supplemented with HPL that can totally\r\nreplace FBS. HPL-supplemented media not only preserves their phenotype as well as their differentiation capacity,\r\nbut also shortens culture time by increasing their growth rate....
Introduction: The development of reproducible methods for deriving human embryonic stem cell (hESC) lines in\r\ncompliance with good manufacturing practice (GMP) is essential for the development of hESC-based therapies.\r\nAlthough significant progress has been made toward the development of chemically defined conditions for the\r\nmaintenance and differentiation of hESCs, efficient derivation of new hESCs requires the use of fibroblast feeder\r\ncells. However, GMP-grade feeder cell lines validated for hESC derivation are not readily available.\r\nMethods: We derived a fibroblast cell line (NclFed1A) from human foreskin in compliance with GMP standards.\r\nConsent was obtained to use the cells for the production of hESCs and to generate induced pluripotent stem cells\r\n(iPSCs). We compared the line with a variety of other cell lines for its ability to support derivation and self-renewal\r\nof hESCs.\r\nResults: NclFed1A supports efficient rates (33%) of hESC colony formation after explantation of the inner cell mass\r\n(ICM) of human blastocysts. This compared favorably with two mouse embryonic fibroblast (MEF) cell lines.\r\nNclFed1A also compared favorably with commercially available foreskin fibroblasts and MEFs in promoting\r\nproliferation and pluripotency of a number of existing and widely used hESCs. The ability of NclFed1A to maintain\r\nself-renewal remained undiminished for up to 28 population doublings from the master cell bank.\r\nConclusions: The human fibroblast line Ncl1Fed1A, produced in compliance with GMP standards and qualified for\r\nderivation and maintenance of hESCs, is a useful resource for the advancement of progress toward hESC-based\r\ntherapies in regenerative medicine....
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