| Physician-scientist proves stem cells heal lungs of newborn animals
November 26, 2009 at 1:11 pm
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| Potential role of dental stem cells in the cellular therapy of cerebral ischemia.
November 26, 2009 at 7:33 am
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| | Potential role of dental stem cells in the cellular therapy of cerebral ischemia. Curr Pharm Des. 2009;15(33):3908-16 Authors: Yalvac ME, Rizvanov AA, Kilic E, Sahin F, Mukhamedyarov MA, Islamov RR, Palotás A Stem cell based therapies for cerebral ischemia (CI) utilize different cell sources including embryonic stem cells (ESCs), neural stem cells (NSCs), umbilical cord blood cells (UCBCs), mesenchymal stem cells (MSCs), and some immortalized cell lines. To date, experimental studies showed that all of these cell sources have been successful to some extent in attenuating the ischemic damage and improving functional recovery after brain injury. Bone marrow derived MSCs seem to be the most widely used and well characterized cell source, which can be also employed for autologous transplantation. Currently, there are two main theories behind the therapeutic effect of stem cell transplantation for treating CIs. The first concept is cell replacement theory in which transplanted stem cells differentiate into progenitor and specialized somatic cells to supersede dying cells. The other hypothesis is based on immuno-modulatory, neuro-protective and neuro-trophic abilities of stem cells which help reducing stroke size and increasing the recovery of behavioral functions. Recent studies focusing on alternative stem cell sources have revealed that dental stem cells (DSCs), including dental pulp stem cells (DPSCs) and dental follicle cells (DFCs) possess properties of MSCs and NSCs. They differentiate into neural linage cells and some other cell types such as osteocytes, adipocytes, chondrocytes, muscle cells and hepatocytes. This review is intended to examine stem cell therapy approaches for CI and emphasize potential use of DSCs as an alternative cell source for the treatment of brain ischemia. PMID: 19938343 [PubMed - in process] |
| mimiRNA: a microRNA expression profiler and classification resource designed to identify functional correlations between microRNAs and their targets.
November 26, 2009 at 7:33 am
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| | mimiRNA: a microRNA expression profiler and classification resource designed to identify functional correlations between microRNAs and their targets. Bioinformatics. 2009 Nov 17; Authors: Ritchie W, Flamant S, Rasko JE MOTIVATION: microRNAs (miRNAs) are short non-coding RNAs that regulate gene expression by inhibiting target mRNA genes. Their tissue-specific and disease-specific expression patterns have immense therapeutic and diagnostic potential. To understand these patterns, a reliable compilation of miRNA and mRNA expression data is required to compare multiple tissue types. Moreover, with the appropriate statistical tools, such a resource could be interrogated to discover functionally related miRNA-mRNA pairs. RESULTS: We have developed mimiRNA, an on-line resource that integrates expression data from 1483 samples and permits visualization of the expression of 635 human miRNAs across 188 different tissues or cell types. mimiRNA incorporates a novel sample classification algorithm, ExParser, that groups identical miRNA or mRNA experiments from separate sources. This enables mimiRNA to provide reliable expression profiles and to discover functional relations between miRNAs and mRNAs such as miRNA targets. Additionally, mimiRNA incorporates a decision tree algorithm to discover distinguishing miRNA features between two tissue or cell types. We validate the efficacy of our resource on independent experimental data and through biologically relevant analyses. AVAILABILITY: http://mimirna.centenary.org.au CONTACT: j.rasko@centenary.org.au SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online. PMID: 19933167 [PubMed - as supplied by publisher] |
| Human embryonic stem-cell derivatives for full reconstruction of the pluristratified epidermis: a preclinical study.
November 26, 2009 at 7:33 am
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| | Human embryonic stem-cell derivatives for full reconstruction of the pluristratified epidermis: a preclinical study. Lancet. 2009 Nov 21;374(9703):1745-1753 Authors: Guenou H, Nissan X, Larcher F, Feteira J, Lemaitre G, Saidani M, Del Rio M, Barrault CC, Bernard FX, Peschanski M, Baldeschi C, Waksman G BACKGROUND: Cell therapy for large burns is dependent upon autologous epidermis reconstructed in vitro. However, the effectiveness of current procedures is limited by the delay needed to culture the patient's own keratinocytes. To assess whether the keratinocyte progeny of human embryonic stem cells (hESCs) could be used to form a temporary skin substitute for use in patients awaiting autologous grafts, we investigated the cells' capability of constructing a pluristratified epidermis. METHODS: hESCs from lines H9 and SA01 were seeded at least in triplicate on fibroblast feeder cells for 40 days in a medium supplemented with bone morphogenetic protein 4 and ascorbic acid. Molecular characterisation of cell differentiation was done throughout the process by quantitative PCR, fluorescence-activated cell sorting, and immunocytochemical techniques. Keratinocyte molecular differentiation and functional capacity to construct a human epidermis were assessed in vitro and in vivo. FINDINGS: From hESCs, we generated a homogeneous population of cells that showed phenotypic characteristics of basal keratinocytes. Expression levels of genes encoding keratin 14, keratin 5, integrin alpha6, integrin beta4, collagen VII, and laminin 5 in these cells were similar to those in basal keratinocytes. After seeding on an artificial matrix, keratinocytes derived from hESCs (K-hESCs) formed a pluristratified epidermis. Keratin-14 immunostaining was seen in the basal compartment, with keratin 10 present in layers overlying the basal layer. Involucrin and filaggrin, late markers of epidermal differentiation, were detected in the uppermost layers only. 12 weeks after grafting onto five immunodeficient mice, epidermis derived from K-hESCs had a structure consistent with that of mature human skin. Human involucrin was appropriately located in spinous and granular layers and few Ki67-positive cells were detected in the basal layer. INTERPRETATION: hESCs can be differentiated into basal keratinocytes that are fully functional-ie, able to construct a pluristratified epidermis. This resource could be developed to provide temporary skin substitutes for patients awaiting autologous grafts. FUNDING: Institut National de la Santé et de la Recherche Médicale, University Evry Val d'Essonne, Association Française contre les Myopathies, Fondation René Touraine, and Genopole. PMID: 19932355 [PubMed - as supplied by publisher] |
| Marginal expression of CXCR4 on c-kit(+)Sca-1 (+)Lineage (-) hematopoietic stem/progenitor cells.
November 26, 2009 at 7:11 am
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| | Marginal expression of CXCR4 on c-kit(+)Sca-1 (+)Lineage (-) hematopoietic stem/progenitor cells. Int J Hematol. 2009 Nov 26; Authors: Sasaki Y, Matsuoka Y, Hase M, Toyohara T, Murakami M, Takahashi M, Nakatsuka R, Uemura Y, Sonoda Y Stromal cell-derived factor 1 (SDF-1) and its receptor CXCR4 are the key regulatory molecules of hematopoietic stem cell (HSC) migration and engraftment to the bone marrow (BM) microenvironment. However, the significance of the ligand-receptor complex on HSC in steady-state BM is not clear. There is currently a lack of information as to how CXCR4 is expressed on HSCs. We herein demonstrate that c-kit(+)Sca-1(+)Lineage(-) (KSL) cells freshly isolated from BM expressed very low to undetectable levels of CXCR4. Two hours of incubation at 37 degrees C quickly up-modulated the receptor expression on KSL cells. Protein synthesis was not required for this early stage up-regulation, thus suggesting the emergence of intracellularly pooled receptors to the cell surface. However, protein synthesis was involved at the later stage of up-regulation. The up-regulated CXCR4 was functional, as evidenced by the fact that the incubated KSL cells more efficiently migrated to the SDF-1 gradient in vitro. Therefore, although KSL cells are able to express functional CXCR4, the receptors are only marginally expressed in the steady-state BM microenvironment. These observations therefore indicate the limited role of the SDF-1-CXCR4 axis on HSC functionality in a static BM environment. PMID: 19937482 [PubMed - as supplied by publisher] |
| Genetic Factors on Mouse Chromosome 18 Affecting Susceptibility to Testicular Germ Cell Tumors and Permissiveness to Embryonic Stem Cell Derivation.
November 26, 2009 at 7:11 am
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| | Genetic Factors on Mouse Chromosome 18 Affecting Susceptibility to Testicular Germ Cell Tumors and Permissiveness to Embryonic Stem Cell Derivation. Cancer Res. 2009 Nov 24; Authors: Anderson PD, Nelson VR, Tesar PJ, Nadeau JH Despite strong heritability, little is known about the genetic control of susceptibility to testicular germ cell tumors (TGCT) in humans or mice. Although the mouse model of spontaneous TGCTs has been extensively studied, conventional linkage analysis has failed to locate the factors that control teratocarcinogenesis in the susceptible 129 family of inbred strains. As an alternative approach, we used both chromosome substitution strains (CSS) to identify individual chromosomes that harbor susceptibility genes and a panel of congenic strains derived from a selected CSS to determine the number and location of susceptibility variants on the substituted chromosome. We showed that 129-Chr 18(MOLF) males are resistant to spontaneous TGCTs and that at least four genetic variants control susceptibility in males with this substituted chromosome. In addition, early embryonic cells from this strain fail to establish embryonic stem cell lines as efficiently as those from the parental 129/Sv strain. For the first time, 129-derived genetic variants that control TGCT susceptibility and fundamental aspects of embryonic stem cell biology have been localized in a genetic context in which the genes can be identified and functionally characterized. [Cancer Res 2009;69(23):9112-7]. PMID: 19934337 [PubMed - as supplied by publisher] |
| The Modulation of MicroRNAs by Type I IFN through the Activation of Signal Transducers and Activators of Transcription 3 in Human Glioma.
November 26, 2009 at 7:11 am
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| | The Modulation of MicroRNAs by Type I IFN through the Activation of Signal Transducers and Activators of Transcription 3 in Human Glioma. Mol Cancer Res. 2009 Nov 24; Authors: Ohno M, Natsume A, Kondo Y, Iwamizu H, Motomura K, Toda H, Ito M, Kato T, Wakabayashi T Type I IFNs are involved in double-stranded RNA responses. Here, we investigated the possibility that IFN-beta may induce or downregulate cellular microRNAs (miRNA) in human neoplasms and thereby use the RNA interference system to show antitumor effects. Because of its known connection to glioma biology, we focused on miR-21 among seven miRNAs influenced by IFN-beta. We analyzed the effect of IFN-beta treatment on miR-21 expression in glioma cells and intracranial glioma xenografts. IFN-beta treatment reduced miR-21 expression in glioma cells markedly, and IFN-beta administration suppressed the growth of glioma-initiating cell-derived intracranial tumors. The levels of primary miR-21 gene transcripts, precursor miR-21, and mature miR-21 decreased 6 hours after the addition of IFN-beta, indicating that the reduction in miR-21 levels was due to transcriptional suppression. We did reporter assays to elucidate the IFN-beta-mediated suppression of miR-21; the addition of signal transducers and activators of transcription 3 (STAT3)-expressing vectors induced the IFN-beta-mediated suppression of miR-21, whereas STAT3-inhibiting agents inhibited the miR-21 suppression. Thus, the results of our study show that the downregulation of miR-21 contributes to the antitumor effects of IFN-beta and that miR-21 expression is negatively regulated by STAT3 activation. These results highlight the importance of understanding the transcriptional regulation of the miRNAs involved in oncogenesis. (Mol Cancer Res 2009;7(12):OF1-9). PMID: 19934272 [PubMed - as supplied by publisher] |
| Bone Marrow Mesenchymal Stem Cells Reduce Intestinal Ischemia/Reperfusion Injuries in Rats.
November 26, 2009 at 7:11 am
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| | Bone Marrow Mesenchymal Stem Cells Reduce Intestinal Ischemia/Reperfusion Injuries in Rats. J Surg Res. 2009 Aug 22; Authors: Jiang H, Qu L, Li Y, Gu L, Shi Y, Zhang J, Zhu W, Li J BACKGROUND: Adult stem cells are promising novel therapies in regenerative medicine. We investigated effects of bone marrow-derived mesenchymal stem cells (MSCs) on intestinal mucosal permeability impaired by ischemia/reperfusion (I/R). METHODS: We used a common I/R model in rats to induce intestinal injury by clamping and unclamping the superior mesenteric artery (SMA) in female Sprague-Dawley rats. MSCs were directly injected into the small intestinal submucosa of the syngenic female rats. Control group were injected with the same volume of 0.9% sodium chloride. Small intestine samples were examined for the engraftment of donor-derived mesenchymal stem cells (MSCs) by Y chromosome in situ hybridization analysis. The small intestinal permeability and histomorphologic alternations were measured to evaluate the therapeutic effect of MSCs transplantation. RESULTS: Small intestinal permeability and villi injuries were significantly reduced in the MSCs administrated group compared with control group. MSCs administration accelerated the recovery of the intestinal barrier dysfunction. CONCLUSION: We concluded that submucosal infusion of MSCs might exert protective effects on the integrity of intestinal barrier. PMID: 19932900 [PubMed - as supplied by publisher] |
| Engineering Considerations for Process Development in Mammalian Cell Cultivation.
November 26, 2009 at 7:11 am
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| | Engineering Considerations for Process Development in Mammalian Cell Cultivation. Curr Pharm Biotechnol. 2009 Nov 23; Authors: Zhang H, Wang W, Quan C, Fan S Mammalian cell cultivation plays a great role in producing protein therapeutics in the last decades. Many engineering parameters are considered for optimization during process development in mammalian cell cultivation, only shear and mixing are especially highlighted in this paper. It is believed that shear stress due to agitation has been over-estimated to damage cells, but shear may result in nonlethal physiological responses. There is no cell damage in the regions where bubbles form, break up and coalescence, but shear stress becomes significant in the wake of rising bubbles and causes great damage to cells in bubble burst regions. Mixing is not sufficient to provide homogeneous dissolved oxygen tension, pH, CO(2) and nutrients in large-scale bioreactors, which can bring severe problems for cell growth, product formation and process control. Scale-down reactors have been developed to address mixing and shear problems for parallel operations. Engineering characterization in conventional and recently developed scale-down bioreactors has been briefly introduced. Process challenges for cultivation of industrial cell lines in high cell densities as well as cultivation of stem cells and other human cells for regenerative medicine, tissue engineering and gene therapy are prospected. Important techniques, such as micromanipulation and nanomanipulation (optical tweezers) for single cell analysis, computational fluid dynamics (CFD) for shear and mixing characterization, and miniaturized bioreactors, are being developed to address those challenges. PMID: 19929819 [PubMed - as supplied by publisher] |
| The Potential of Embryonic Stem Cells Combined with -Omics Technologies as Model Systems for Toxicology.
November 26, 2009 at 7:11 am
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| | The Potential of Embryonic Stem Cells Combined with -Omics Technologies as Model Systems for Toxicology. Curr Med Chem. 2009 Nov 17; Authors: Winkler J, Sotiriadou I, Chen S, Hescheler J, Sachinidis A The derivation of pluripotent embryonic stem (ES) cell lines has opened up new areas of research in basic and applied science, most significantly in developmental biology and regenerative medicine. While application-oriented research has for the most part focussed on obtaining differentiated, organotypic cells from ES cells for future cell grafting therapies, ES cells have more immediate potential for use in toxicological in vitro assays used during drug development. ES cells are derived from blastocyst-stage embryos and offer an in vitro model for early development, thus enabling tests for teratogenicity testing in a human cell culture system and avoiding the pitfalls of inter-species differences. Differentiated, organotypic cells obtained from ES cells can potentially replace the primary cells and cell lines currently used for in vitro toxicology by offering a more consistent and potentially limitless source of differentiated cells. This can facilitate the establishment of comprehensive toxicogenomics and -proteomics databases and complement current databases that rely on data obtained from animal experiments. More recently, induced pluripotent stem (iPS) cells with ES cell-like properties have been obtained through reprogramming of somatic cells, thus enabling the generation of disease-specific cell lines. We review the potential of combining ES cells and ES cell-derived somatic cells with "omics" technologies for in vitro toxicology with a particular emphasis on the development of toxicogenomics and toxicoproteomics signatures. A separate section describes the potential of iPS cells for toxicology. PMID: 19929785 [PubMed - as supplied by publisher] |
| Human adipose-derived mesenchymal stem cells in vitro: evaluation of an optimal expansion medium preserving stemness.
November 26, 2009 at 7:11 am
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| | Human adipose-derived mesenchymal stem cells in vitro: evaluation of an optimal expansion medium preserving stemness. Cytotherapy. 2009 Nov 24; Authors: Baer PC, Griesche N, Luttmann W, Schubert R, Luttmann A, Geiger H Background aims. The potential of cultured adipose-derived stem cells (ASC) in regenerative medicine and new cell therapeutic concepts has been shown recently by many investigations. However, while the method of isolation of ASC from liposuction aspirates depending on plastic adhesion is well established, a standard expansion medium optimally maintaining the undifferentiated state has not been described. Methods. We cultured ASC in five commonly used culture media (two laboratory-made media and three commercially available media) and compared them with a standard medium. We analyzed the effects on cell morphology, proliferation, hepatocyte growth factor (HGF) expression, stem cell marker profile and differentiation potential. Proliferation was measured with a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and a fluorescent assay. Release of HGF was assessed by an immunoassay. Expression of characteristic stem cell-related transcription factors and markers was evaluated by quantitative polymerase chain reaction (qPCR) (Nanog, Sox-2, Rex-1, nestin and Oct-4) and flow cytometry (CD44, CD73, CD90, CD105 and CD166), and differentiation was shown by adipogenic medium. Results. The morphology and expansion of ASC were significantly affected by the media used, whereas none of the media influenced the ASC potential to differentiate into adipocytes. Furthermore, two of the media induced an increase in expression of transcription factors, an increased secretion of HGF and a decrease in CD105 expression. Conclusions. Culture of ASC in one of these two media before using the cells in cell therapeutic approaches may have a benefit on their regenerative potential. PMID: 19929458 [PubMed - as supplied by publisher] |
| Anti-L-NGFR and -CD34 Monoclonal Antibodies identify multipotent mesenchymal stem cells in human adipose tissue.
November 26, 2009 at 7:11 am
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| | Anti-L-NGFR and -CD34 Monoclonal Antibodies identify multipotent mesenchymal stem cells in human adipose tissue. Stem Cells Dev. 2009 Nov 23; Authors: Quirici N, Scavullo C, de Girolamo L, Lopa S, Arrigoni E, Lambertenghi Deliliers G, Brini AT Stem cells hold great promise in tissue engineering for repairing tissues damaged by disease or injury. Mesenchymal stem cells (MSCs) are multipotent cells able to proliferate and differentiate into multiple mesodermal tissues such as bone, cartilage, muscle, tendon and fat. We have previously reported that the low-affinity nerve growth factor receptor (L-NGFR or CD271) defines a subset of cells with high proliferative, clonogenic and multipotential differentiation ability in adult bone marrow (BM). It has been recently shown that adipose tissue is an alternative source of adult multipotent stem cells and human adipose-derived stem cells, selected by plastic-adherence (PA hASCs), have been extensively characterized for their functional potentials in vitro. In this study, immuno-selected L-NGFR+ and CD34+ subpopulations have been analyzed and compared with the PA hASCs. Phenotypic profile of freshly purified subpopulations showed an enrichment in the expression of some stem cell markers: indeed, a great percentage of L-NGFR+ cells co-expressed CD34 and CD117 antigens, whereas the endothelial-committed progenitor markers KDR and P1H12 were mainly expressed on CD34+ cells. Differently from PA hASCs, the immuno-separated fractions showed high increments in cell proliferation, and the fibroblast colony-forming activity (CFU-F) was maintained throughout the time of culture. Furthermore, the immuno-selected populations showed a greater differentiative potential towards adipocytes, osteoblasts and chondrocyte-like cells, compared to PA hASCs. Our data suggest that both CD34+ and L-NGFR+ hASCs can be considered alternative candidates for tissue engineering and regenerative medicine applications. PMID: 19929314 [PubMed - as supplied by publisher] |
| Slowed progression in models of huntington disease by adipose stem cell transplantation.
November 26, 2009 at 6:57 am
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| | Slowed progression in models of huntington disease by adipose stem cell transplantation. Ann Neurol. 2009 Jun 29;66(5):671-681 Authors: Lee ST, Chu K, Jung KH, Im WS, Park JE, Lim HC, Won CH, Shin SH, Lee SK, Kim M, Roh JK OBJECTIVE: Adipose-derived stem cells (ASCs) are readily accessible and secrete multiple growth factors. Here, we show that ASC transplantation rescues the striatal pathology of Huntington disease (HD) models. METHODS: ASCs were isolated from human subcutaneous adipose tissue. In a quinolinic acid (QA)-induced rat model of striatal degeneration, human ASCs (1 million cells) were transplanted into the ipsilateral striatal border immediately after the QA injection. In 60-day-old R6/2 mice transgenic for HD, ASCs (0.5 million cells) were transplanted into each bilateral striata. In in vitro experiments, we treated mutant huntingtin gene-transfected cerebral neurons with ASC-conditioned media. RESULTS: In the QA model, human ASCs reduced apomorphine-induced rotation behavior, lesion volume, and striatal apoptosis. In R6/2 transgenic mice, transplantation of ASCs improved Rota-Rod performance and limb clasping, increased survival, attenuated the loss of striatal neurons, and reduced the huntingtin aggregates. ASC-transplanted R6/2 mice expressed elevated levels of peroxisome proliferator-activated receptor gamma coactivator-1alpha (PGC-1alpha) and reactive oxygen defense enzymes and showed activation of the Akt/cAMP-response element-binding proteins. ASC-conditioned media decreased the level of N-terminal fragments of mutant huntingtin and associated apoptosis, and increased PGC-1alpha expression. INTERPRETATION: Collectively, ASC transplantation slowed striatal degeneration and behavioral deterioration of HD models, possibly via secreted factors. Ann Neurol 2009;66:671-681. PMID: 19938161 [PubMed - as supplied by publisher] |
| Freshly isolated stromal cells from the infrapatellar fat pad are suitable for a one-step surgical procedure to regenerate cartilage tissue.
November 26, 2009 at 6:57 am
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| | Freshly isolated stromal cells from the infrapatellar fat pad are suitable for a one-step surgical procedure to regenerate cartilage tissue. Cytotherapy. 2009;11(8):1052-64 Authors: Jurgens WJ, van Dijk A, Doulabi BZ, Niessen FB, Ritt MJ, van Milligen FJ, Helder MN Background aims Stem cell therapies are being evaluated as promising alternatives for cartilage regeneration. We investigated whether stromal vascular fraction cells (SVF) from the infrapatellar (Hoffa) fat pad are suitable for a one-step surgical procedure to treat focal cartilage defects. Methods SVF was harvested from patients undergoing knee arthroplasty (n = 53). Colony-forming unit (CFU) assays, growth kinetics and surface marker profiles were determined, and the chondrogenic differentiation capacity of freshly isolated SVF was assessed after seeding in three-dimensional poly (l-lactic-co--caprolactone) scaffolds. Results SVF yield per fat pad varied between 0.55 and 16 x 10(6) cells. CFU frequency and population doubling time were 2.6 +/- 0.6% and +/-2 days, respectively. Surface marker profiles matched those of subcutaneous-derived adipose-derived stem cells (ASC). CFU from Hoffa SVF showed differentiation toward osteogenic and adipogenic lineages. Cartilage differentiation was confirmed by up-regulation of the cartilage genes sox9, aggrecan, collagen type II and cartilage oligomeric matrix protein (COMP), collagen II immunostaining, Alcian Blue staining and glycosaminoglycan production. Compared with passaged cells, SVF showed at least similar chondrogenic potential. Conclusions This study demonstrates that SVF cells from the infrapatellar fat pad are suitable for future application in a one-step surgical procedure to regenerate cartilage tissue. SVF shows similar favorable characteristics as cultured ASC, and chondrogenic differentiation even appears to be slightly better. However, because of variable harvesting volumes and yields, SVF from the infrapatellar fat pad might only be applicable for treatment of small focal cartilage defects, whereas for larger osteoarthritic defects subcutaneous adipose tissue depot would be preferable. PMID: 19929469 [PubMed - in process] |
| Isolation of stem cell populations with trophic and immunoregulatory functions from human intestinal tissues: potential for cell therapy in inflammatory bowel disease.
November 26, 2009 at 6:57 am
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| | Isolation of stem cell populations with trophic and immunoregulatory functions from human intestinal tissues: potential for cell therapy in inflammatory bowel disease. Cytotherapy. 2009;11(8):1020-31 Authors: Lanzoni G, Alviano F, Marchionni C, Bonsi L, Costa R, Foroni L, Roda G, Belluzzi A, Caponi A, Ricci F, Luigi Tazzari P, Pagliaro P, Rizzo R, Lanza F, Roberto Baricordi O, Pasquinelli G, Roda E, Paolo Bagnara G Background aims Bone marrow (BM)- and adipose tissue (AT)-derived mesenchymal stromal cells (MSC) are currently under evaluation in phase III clinical trials for inflammatory bowel disease and other intestinal disease manifestations. The therapeutic efficacy of these treatments may derive from a combination of the differentiation, trophic and immunomodulatory abilities of the transplanted cells. We investigated intestinal tissues as sources of MSC: such cells may support tissue-specific functions and hold advantages for engraftment and contribution in the gastrointestinal environment. Methods Intestinal specimens were collected, and the mucosa and submucosa mechanically separated and enzymatically digested. Mesenchymal stromal populations were isolated, expanded and characterized under conditions commonly used for MSC. The differentiation potential, trophic effect and immunomodulatory ability were investigated. Results We successfully isolated and extensively expanded populations showing the typical MSC profile: CD29(+), CD44(+), CD73(+), CD105(+) and CD166(+), and CD14(-), CD34(-) and CD45(-). Intestinal mucosal (IM) MSC were also CD117(+), while submucosal cultures (ISM MSC) showed CD34(+) subsets. The cells differentiated toward osteogenic, adipogenic and angiogenic commitments. Intestinal-derived MSC were able to induce differentiation and organization of intestinal epithelial cells (Caco-2) in three-dimensional collagen cultures. Immunomodulatory activity was evidenced in co-cultures with normal heterologous phytohemagglutinin-stimulated peripheral blood mononuclear cells. Conclusions Multipotent MSC can be isolated from intestinal mucosal and submucosal tissues. IM MSC and ISM MSC are able to perform trophic and immunomodulatory functions. These findings could open a pathway for novel approaches to intestinal disease treatment. PMID: 19929466 [PubMed - in process] |
| Human adipose-derived mesenchymal stem cells in vitro: evaluation of an optimal expansion medium preserving stemness.
November 26, 2009 at 6:57 am
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| | Human adipose-derived mesenchymal stem cells in vitro: evaluation of an optimal expansion medium preserving stemness. Cytotherapy. 2009 Nov 24; Authors: Baer PC, Griesche N, Luttmann W, Schubert R, Luttmann A, Geiger H Background aims. The potential of cultured adipose-derived stem cells (ASC) in regenerative medicine and new cell therapeutic concepts has been shown recently by many investigations. However, while the method of isolation of ASC from liposuction aspirates depending on plastic adhesion is well established, a standard expansion medium optimally maintaining the undifferentiated state has not been described. Methods. We cultured ASC in five commonly used culture media (two laboratory-made media and three commercially available media) and compared them with a standard medium. We analyzed the effects on cell morphology, proliferation, hepatocyte growth factor (HGF) expression, stem cell marker profile and differentiation potential. Proliferation was measured with a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and a fluorescent assay. Release of HGF was assessed by an immunoassay. Expression of characteristic stem cell-related transcription factors and markers was evaluated by quantitative polymerase chain reaction (qPCR) (Nanog, Sox-2, Rex-1, nestin and Oct-4) and flow cytometry (CD44, CD73, CD90, CD105 and CD166), and differentiation was shown by adipogenic medium. Results. The morphology and expansion of ASC were significantly affected by the media used, whereas none of the media influenced the ASC potential to differentiate into adipocytes. Furthermore, two of the media induced an increase in expression of transcription factors, an increased secretion of HGF and a decrease in CD105 expression. Conclusions. Culture of ASC in one of these two media before using the cells in cell therapeutic approaches may have a benefit on their regenerative potential. PMID: 19929458 [PubMed - as supplied by publisher] |
| Anti-L-NGFR and -CD34 Monoclonal Antibodies identify multipotent mesenchymal stem cells in human adipose tissue.
November 26, 2009 at 6:57 am
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| | Anti-L-NGFR and -CD34 Monoclonal Antibodies identify multipotent mesenchymal stem cells in human adipose tissue. Stem Cells Dev. 2009 Nov 23; Authors: Quirici N, Scavullo C, de Girolamo L, Lopa S, Arrigoni E, Lambertenghi Deliliers G, Brini AT Stem cells hold great promise in tissue engineering for repairing tissues damaged by disease or injury. Mesenchymal stem cells (MSCs) are multipotent cells able to proliferate and differentiate into multiple mesodermal tissues such as bone, cartilage, muscle, tendon and fat. We have previously reported that the low-affinity nerve growth factor receptor (L-NGFR or CD271) defines a subset of cells with high proliferative, clonogenic and multipotential differentiation ability in adult bone marrow (BM). It has been recently shown that adipose tissue is an alternative source of adult multipotent stem cells and human adipose-derived stem cells, selected by plastic-adherence (PA hASCs), have been extensively characterized for their functional potentials in vitro. In this study, immuno-selected L-NGFR+ and CD34+ subpopulations have been analyzed and compared with the PA hASCs. Phenotypic profile of freshly purified subpopulations showed an enrichment in the expression of some stem cell markers: indeed, a great percentage of L-NGFR+ cells co-expressed CD34 and CD117 antigens, whereas the endothelial-committed progenitor markers KDR and P1H12 were mainly expressed on CD34+ cells. Differently from PA hASCs, the immuno-separated fractions showed high increments in cell proliferation, and the fibroblast colony-forming activity (CFU-F) was maintained throughout the time of culture. Furthermore, the immuno-selected populations showed a greater differentiative potential towards adipocytes, osteoblasts and chondrocyte-like cells, compared to PA hASCs. Our data suggest that both CD34+ and L-NGFR+ hASCs can be considered alternative candidates for tissue engineering and regenerative medicine applications. PMID: 19929314 [PubMed - as supplied by publisher] |
| Additive and synergistic effects of bFGF and hypoxia on leporine meniscus cell-seeded PLLA scaffolds.
November 26, 2009 at 6:24 am
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| | Additive and synergistic effects of bFGF and hypoxia on leporine meniscus cell-seeded PLLA scaffolds. J Tissue Eng Regen Med. 2009 Nov 24; Authors: Gunja NJ, Athanasiou KA Injuries to avascular regions of menisci do not heal and result in significant discomfort to patients. Current treatments, such as partial meniscectomy, alleviate these symptoms in the short term but lead to premature osteoarthritis as a result of compromised stability and changes in knee biomechanics. Thus, tissue engineering of the meniscus may provide an alternative treatment modality to overcome this problem. In this experiment, a scaffold-based tissue-engineering approach was utilized to regenerate the meniscus. Meniscus cells were cultured on poly-L-lactic acid scaffolds in normoxic ( approximately 21% oxygen) or hypoxic ( approximately 2% oxygen) conditions in the presence or absence of the growth factor, basic fibroblast growth factor (bFGF). At t = 4 weeks, histological sections of constructs showed presence of collagen and glycosaminoglycan (GAG) in all groups. Immunohistochemical staining showed the presence of collagen I in all groups and collagen II in groups cultured under hypoxic conditions. bFGF in the culture medium significantly increased cell number/construct by 25%, regardless of culture conditions. For GAG/construct, synergistic increases were observed in constructs cultured in hypoxic conditions and bFGF (two-fold) when compared to constructs cultured in normoxic conditions. Compressive tests showed synergistic increases in the relaxation modulus and coefficient of viscosity and additive increases in the instantaneous modulus for constructs cultured under hypoxic conditions and bFGF, when compared to constructs cultured under normoxic conditions. Overall, these results demonstrate that bFGF and hypoxia can significantly enhance the ability of meniscus cells to produce GAGs and improve the compressive properties of tissue-engineered meniscus constructs in vitro. Copyright (c) 2009 John Wiley & Sons, Ltd. PMID: 19937913 [PubMed - as supplied by publisher] |
| Enamel matrix derivative enhances tissue formation around scaffolds used for tissue engineering of ligaments.
November 26, 2009 at 6:24 am
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| | Enamel matrix derivative enhances tissue formation around scaffolds used for tissue engineering of ligaments. J Tissue Eng Regen Med. 2009 Nov 20; Authors: Messenger MP, Raïf EM, Seedhom BB, Brookes SJ The following in vitro translational study investigated whether enamel matrix derivative (EMD), an approved biomimetic treatment for periodontal disease (Emdogain((R))) and hard-to-heal wounds (Xelma((R))), enhanced synovial cell colonization and protein synthesis around a scaffold used clinically for in situ tissue engineering of the torn anterior cruciate ligament (ACL). Synovial cells were enzymatically extracted from bovine synovium and dynamically seeded onto polyethylene terephthalate (PET) scaffolds. The cells were cultured in low-serum medium (0.5% FBS) for 4 weeks with either a single administration of EMD at the start of the 4 week period or multiple administrations of EMD at regular intervals throughout the 4 weeks. Samples were harvested and evaluated using the Hoechst DNA assay, BCA protein assay, cresolphthalein complexone calcium assay, SDS-PAGE, ELISA and electron microscopy. A significant increase in cell number (DNA) (p < 0.01), protein content (p < 0.01) and TGFbeta1 synthesis (p < 0.01) was observed with multiple administrations of EMD. Additionally, SDS-PAGE showed an increase in high molecular weight proteins, characteristic of the fibril-forming collagens. Electron microscopy supported these findings, showing that scaffolds treated with multiple administrations of EMD were heavily coated with cells and extracellular matrix (ECM) that enveloped the fibres. Multiple administrations of EMD to synovial cell-seeded scaffolds enhanced the formation of tissue in vitro. Additionally, it was shown that EMD enhanced TGFbeta1 synthesis of synovial cells, suggesting a potential mode of action for EMD's capacity to stimulate tissue regeneration. Copyright (c) 2009 John Wiley & Sons, Ltd. PMID: 19937644 [PubMed - as supplied by publisher] |
| Investigating the importance of flow when utilizing hyaluronan scaffolds for tissue engineering.
November 26, 2009 at 6:24 am
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| | Investigating the importance of flow when utilizing hyaluronan scaffolds for tissue engineering. J Tissue Eng Regen Med. 2009 Nov 20; Authors: Donegan GC, Hunt JA, Rhodes N Esterified hyaluronan scaffolds offer significant advantages for tissue engineering. They are recognized by cellular receptors, interact with many other extracellular matrix proteins and their metabolism is mediated by intrinsic cellular pathways. In this study differences in the viability and structural integrity of vascular tissue models cultured on hyaluronan scaffolds under laminar flow conditions highlighted potential differences in the biodegradation kinetics, processes and end-products, depending on the culture environment. Critical factors are likely to include seeding densities and the duration and magnitude of applied biomechanical stress. Proteomic evaluation of the timing and amount of remodelling protein expression, the resulting biomechanical changes arising from this response and metabolic cell viability assay, together with examination of tissue morphology, were conducted in vascular tissue models cultured on esterified hyaluronan felt and PTFE mesh scaffolds. The vascular tissue models were derived using complete cell sheets derived from harvested and expanded umbilical cord vein cells. This seeding method utilizes high-density cell populations from the outset, while the cells are already supported by their own abundant extracellular matrix. Type I and type IV collagen expression in parallel with MMP-1 and MMP-2 expression were monitored in the tissue models over a 10 day culture period under laminar flow regimes using protein immobilization technologies. Uniaxial tensile testing and scanning electron microscopy were used to compare the resulting effects of hydrodynamic stimulation upon structural integrity, while viability assays were conducted to evaluate the effects of shear on metabolic function. The proteomic results showed that the hyaluronan felt-supported tissues expressed higher levels of all remodelling proteins than those cultured on PTFE mesh. Overall, a 21% greater expression of type I collagen, 24% higher levels of type IV collagen, 24% higher levels of MMP-1 and 34% more MMP-2 were observed during hydrodynamic stress. This was coupled with a loss of structural integrity in these models after the introduction of laminar flow, as compared to the increases in all mechanical properties observed in the PTFE mesh-supported tissues. However, under flow conditions, the hyaluronan-supported tissues showed some recovery of the viability originally lost during static culture conditions, in contrast to PTFE mesh-based models, where initial gains were followed by a decline in metabolic viability after applied shear stress. Proteomic, cell viability and mechanical testing data emphasized the need for extended in vitro evaluations to enable better understanding of multi-stage remodelling and reparative processes in tissues cultured on biodegradable scaffolds. This study also highlighted the possibility that in high-density tissue culture with a biodegradable component, dynamic conditions may be more conducive to optimal tissue development than the static environment because they facilitate the efficient removal of high concentrations of degradation end-products accumulating in the pericellular space. Copyright (c) 2009 John Wiley & Sons, Ltd. PMID: 19937643 [PubMed - as supplied by publisher] |
| Crosslinking effect of Nordihydroguaiaretic acid (NDGA) on decellularized heart valve scaffold for tissue engineering.
November 26, 2009 at 6:24 am
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| | Crosslinking effect of Nordihydroguaiaretic acid (NDGA) on decellularized heart valve scaffold for tissue engineering. J Mater Sci Mater Med. 2009 Nov 20; Authors: Lü X, Zhai W, Zhou Y, Zhou Y, Zhang H, Chang J Decellularized heart valve scaffolds possess many desirable properties in valvular tissue engineering. However, their current applications were limited by short durability, easily structural dysfunction and immunological competence. Although crosslinking with chemical reagents, such as glutaraldehyde (GA), will enhance the mechanical properties, the low long-term stability and cytotoxicity of the scaffolds remains potential problem. Nordihydroguaiaretic acid (NDGA) is a bioactive natural product which is able to crosslink collagen and was proven to be effective in preparation of scaffold for tendon tissue engineering. In this paper, NDGA crosslinked decellularized heart valve scaffolds demonstrated higher tensile strength, enzymatic hydrolysis resistance and store stability than the non-crosslinked ones. Its mechanical properties and cytocompability were superior to that of GA-crosslinked heart valve matrix. Below the concentration of 10 mug/ml, NDGA has no visible cytotoxic effect on both endothelial cells (EC) and valvular interstitial cells (VIC) and its cytotoxicity is much less than that of GA. The LC50 (50% lethal concentration) of NDGA on ECs and VICs are 32.6 mug/ml and 47.5 mug/ml, respectively, while those of GA are almost 30 times higher than NDGA (P < 0.05). ECs can attach to and maintain normal morphology on the surface of NDGA-crosslinked valvular scaffolds but not GA-crosslinked ones. This study demonstrated that NDGA-crosslinking of decellularized valvular matrix is a promising approach for preparation of heart valve tissue engineering scaffolds. PMID: 19936890 [PubMed - as supplied by publisher] | | | 
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