| |  | | | | | MAINTENANCE Feed My Inbox will be undergoing maintenance on Saturday January 29th from approx 8-11 am CST. Emails will be delayed during this time period. We appreciate your patience. More details | | | | | | | | | | | | |  | |  | | | | | TE-RegenMed-StemCell feed | | | | | | | | | | | | | | | | | | | Abstracts of bone-tec 2009, International Bone-Tissue-Engineering Congress. October 8-11, 2009. Hannover, Germany. Tissue Eng Part A. 2010 Aug;16(8):A2-29 Authors: PMID: 20687812 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Fabrication of micro-tissues using modules of collagen gel containing cells. J Vis Exp. 2010;(46): Authors: Chamberlain MD, Butler MJ, Ciucurel EC, Fitzpatrick LE, Khan OF, Leung BM, Lo C, Patel R, Velchinskaya A, Voice DN, Sefton MV This protocol describes the fabrication of a type of micro-tissues called modules. The module approach generates uniform, scalable and vascularized tissues. The modules can be made of collagen as well as other gelable or crosslinkable materials. They are approximately 2 mm in length and 0.7 mm in diameter upon fabrication but shrink in size with embedded cells or when the modules are coated with endothelial cells. The modules individually are small enough that the embedded cells are within the diffusion limit of oxygen and other nutrients but modules can be packed together to form larger tissues that are perfusable. These tissues are modular in construction because different cell types can be embedded in or coated on the modules before they are packed together to form complex tissues. There are three main steps to making the modules: neutralizing the collagen and embedding cells in it, gelling the collagen in the tube and cutting the modules and coating the modules with endothelial cells. PMID: 21178971 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Scaffold stiffness affects the contractile function of three-dimensional engineered cardiac constructs. Biotechnol Prog. 2010 Sep;26(5):1382-90 Authors: Marsano A, Maidhof R, Wan LQ, Wang Y, Gao J, Tandon N, Vunjak-Novakovic G We investigated the effects of the initial stiffness of a three-dimensional elastomer scaffold--highly porous poly(glycerol sebacate)--on functional assembly of cardiomyocytes cultured with perfusion for 8 days. The polymer elasticity varied with the extent of polymer cross-links, resulting in three different stiffness groups, with compressive modulus of 2.35 ± 0.03 (low), 5.28 ± 0.36 (medium), and 5.99 ± 0.40 (high) kPa. Laminin coating improved the efficiency of cell seeding (from 59 ± 15 to 90 ± 21%), resulting in markedly increased final cell density, construct contractility, and matrix deposition, likely because of enhanced cell interaction and spreading on scaffold surfaces. Compact tissue was formed in the low and medium stiffness groups, but not in the high stiffness group. In particular, the low stiffness group exhibited the greatest contraction amplitude in response to electric field pacing, and had the highest compressive modulus at the end of culture. A mathematical model was developed to establish a correlation between the contractile amplitude and the cell distribution within the scaffold. Taken together, our findings suggest that the contractile function of engineered cardiac constructs positively correlates with low compressive stiffness of the scaffold. PMID: 20945492 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Keratinocytes drive melanoma invasion in a reconstructed skin model. Melanoma Res. 2010 Oct;20(5):372-80 Authors: Van Kilsdonk JW, Bergers M, Van Kempen LC, Schalkwijk J, Swart GW Melanoma progression is a multistep progression from a common melanocytic nevus through the radial growth phase, the invasive vertical growth phase finally leading to metastatic spread into distant organs. Migration and invasion of tumor cells requires secretion of proteases to facilitate remodeling of the extracellular matrix including basement membranes. Here we used a reconstructed skin model to investigate melanoma growth and invasion in vitro. Using this model we show that the dermoepidermal basement membrane prevents the invasion of metastatic melanoma BLM and MV3 cells in the absence of a stratified epidermis. In the reconstructed skin model, matrix metalloproteinase-9, a protease activated early in melanoma development, is secreted by the keratinocytes and subsequently activated by an unknown soluble factor secreted by the melanoma cells. The dynamic interplay between keratinocytes and melanoma cells is further shown by an altered growth pattern of melanoma cells and the finding that a reconstructed epidermis induces invasion. Overall, our findings show that the invasive behavior of melanoma cells is determined by the melanoma cells themselves, but that the interplay between surrounding keratinocytes and the melanoma cells plays an important role in melanoma invasion. PMID: 20700063 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | A novel dermal tissue construct: development and in vitro characterization. Biotechnol Prog. 2010 Sep;26(5):1424-30 Authors: Deshpande MS, Kuchroo PV A dermal tissue construct composed of human dermal fibroblasts and a chitosan sponge has been developed, targeted towards the treatment of diabetic nonhealing ulcers. The construct has been designed in a way that the dermal fibroblasts are arranged as a three-dimensional sheet adhered entirely on one side of the chitosan sponge. This design would allow maximal diffusion of growth factors from the cells to the wound bed when the construct is applied on the wound with the cellular sheet side making contact with the wound bed. The diffusion of secreted growth factors would take place directly from cells to the wound bed without being impeded by a matrix. The cells are present at a high density in the dermal construct, which would aid in accelerated wound healing. The construct has a porous chitosan sponge base, which would allow gas exchange, and renders the dermal construct very flexible so that it would take the shape of the wound contours well, while having mechanical integrity. The viability of cells in the construct is greater than 90%. The dermal construct produces a high amount of vascular endothelial growth factor, from 42 ng to 31 ng in 24 h. The construct also produces high amounts of Interleukin-8 (IL-8), from 375 ng to 1065 ng in the first 24 h. Both VEGF and IL-8 have important roles in the healing of chronic diabetic ulcers. PMID: 20549681 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Influence of acellular natural lung matrix on murine embryonic stem cell differentiation and tissue formation. Tissue Eng Part A. 2010 Aug;16(8):2565-80 Authors: Cortiella J, Niles J, Cantu A, Brettler A, Pham A, Vargas G, Winston S, Wang J, Walls S, Nichols JE We report here the first attempt to produce and use whole acellular (AC) lung as a matrix to support development of engineered lung tissue from murine embryonic stem cells (mESCs). We compared the influence of AC lung, Gelfoam, Matrigel, and a collagen I hydrogel matrix on the mESC attachment, differentiation, and subsequent formation of complex tissue. We found that AC lung allowed for better retention of cells with more differentiation of mESCs into epithelial and endothelial lineages. In constructs produced on whole AC lung, we saw indications of organization of differentiating ESC into three-dimensional structures reminiscent of complex tissues. We also saw expression of thyroid transcription factor-1, an immature lung epithelial cell marker; pro-surfactant protein C, a type II pneumocyte marker; PECAM-1/CD31, an endothelial cell marker; cytokeratin 18; alpha-actin, a smooth muscle marker; CD140a or platelet-derived growth factor receptor-alpha; and Clara cell protein 10. There was also evidence of site-specific differentiation in the trachea with the formation of sheets of cytokeratin-positive cells and Clara cell protein 10-expressing Clara cells. Our findings support the utility of AC lung as a matrix for engineering lung tissue and highlight the critical role played by matrix or scaffold-associated cues in guiding ESC differentiation toward lung-specific lineages. PMID: 20408765 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | In vitro regulation of neural differentiation and axon growth by growth factors and bioactive nanofibers. Tissue Eng Part A. 2010 Aug;16(8):2641-8 Authors: Lam HJ, Patel S, Wang A, Chu J, Li S Human embryonic stem cell (ESC)-derived neural cells are a potential cell source for neural tissue regeneration. Understanding the biochemical and biophysical regulation of neural differentiation and axon growth will help us develop cell therapies and bioactive scaffolds. We demonstrated that basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) had different effects on human ESC differentiation into neural cells. EGF was more effective in inducing expression of neuron and glial markers and cell extensions. In addition to biochemical cues, poly(l-lactic acid) scaffolds with aligned nanofibers increased axon growth from ESC-derived neural cells, demonstrating the significant effects of biophysical guidance at nanoscale. To combine the biochemical and biophysical cues, bFGF and EGF were either adsorbed or bound to heparin on nanofibrous scaffolds. EGF, but not bFGF, was effectively adsorbed onto nanofibers. However, adsorbed EGF and bFGF did not effectively enhance axon growth. In contrast, immobilization of bFGF or EGF onto nanofibers using heparin as the adapter molecule significantly promoted axon growth. This study elucidated the effect of bFGF and EGF in neural differentiation and axon growth, and demonstrated a method to immobilize active bFGF and EGF onto aligned nanofibers to promote neural tissue regeneration. PMID: 20367289 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Development of a decellularized lung bioreactor system for bioengineering the lung: the matrix reloaded. Tissue Eng Part A. 2010 Aug;16(8):2581-91 Authors: Price AP, England KA, Matson AM, Blazar BR, Panoskaltsis-Mortari A We developed a decellularized murine lung matrix bioreactor system that could be used to evaluate the potential of stem cells to regenerate lung tissue. Lungs from 2-3-month-old C57BL/6 female mice were excised en bloc with the trachea and heart, and decellularized with sequential solutions of distilled water, detergents, NaCl, and porcine pancreatic DNase. The remaining matrix was cannulated and suspended in small airway growth medium, attached to a ventilator to simulate normal, murine breathing-induced stretch. After 7 days in an incubator, lung matrices were analyzed histologically. Scanning electron microscopy and histochemical staining demonstrated that the pulmonary matrix was intact and that the geographic placement of the proximal and distal airways, alveoli and vessels, and the basement membrane of these structures all remained intact. Decellularization was confirmed by the absence of nuclear 4',6-diamidino-2-phenylindole staining and negative polymerase chain reaction for genomic DNA. Collagen content was maintained at normal levels. Elastin, laminin, and glycosaminglycans were also present, although at lower levels compared to nondecellularized lungs. The decellularized lung matrix bioreactor was capable of supporting growth of fetal alveolar type II cells. Analysis of day 7 cryosections of fetal-cell-injected lung matrices showed pro-Sp-C, cytokeratin 18, and 4',6-diamidino-2-phenylindole-positive cells lining alveolar areas that appeared to be attached to the matrix. These data illustrate the potential of using decellularized lungs as a natural three-dimensional bioengineering matrix as well as provide a model for the study of lung regeneration from pulmonary stem cells. PMID: 20297903 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Inhibition of gamma-secretase activity promotes differentiation of embryonic pancreatic precursor cells into functional islet-like clusters in poly(ethylene glycol) hydrogel culture. Tissue Eng Part A. 2010 Aug;16(8):2593-603 Authors: Mason MN, Mahoney MJ We assessed the ability of a gamma-secretase inhibitor to promote the in vitro differentiation of induced embryonic pancreatic precursor cell aggregates into functional islet-like clusters when encapsulated within a three-dimensional hydrogel. Undifferentiated pancreatic precursor cells were isolated from E.15 rat embryos, dissociated into single cells, and aggregated in suspension-rotation culture. Aggregates were photoencapsulated into poly(ethylene glycol) hydrogels with entrapped collagen type 1 and cultured for 14 days with or without a gamma-secretase inhibitor. Gene expression, proinsulin content, and C-peptide release were measured to determine differentiation and maturation of encapsulated precursor cell aggregates. In the control medium, scattered breakthrough beta cell differentiation was observed; however, cells remained largely insulin negative. Upon addition of a gamma-secretase inhibitor the majority of cells in clusters became insulin positive, and insulin per DNA and glucose-stimulated insulin release measurements for these cultures were comparable with those for adult rat islets. Cluster counts after culture day 14 were 88% of those initially encapsulated, demonstrating excellent cluster survival in hydrogel culture. These results indicate that concerted differentiation of pancreatic precursor cell aggregates into functionally mature islet-like clusters can be achieved in poly(ethylene glycol)-based hydrogel cultures by blocking cell contact-mediated Notch signaling with a gamma-secretase inhibitor. PMID: 20236034 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Derivation of functional smooth muscle cells from multipotent human hair follicle mesenchymal stem cells. Tissue Eng Part A. 2010 Aug;16(8):2553-64 Authors: Liu JY, Peng HF, Gopinath S, Tian J, Andreadis ST We investigated the potential of human hair follicle cells for multilineage differentiation and as a source of functional smooth muscle cells (SMCs). We report that human hair follicle stem cells (HFCs) isolated from individual follicles expressed surface markers that are characteristic of mesenchymal stem cells such as CD44, CD49b, CD73, CD90, and CD105 but lacked hematopoietic markers CD45 and CD34. In addition, HFCs differentiated toward adipocytes, chondrocytes, osteoblasts, or SMCs in the appropriate induction medium. Treatment with basic fibroblast growth factor increased proliferation and prevented myogenic differentiation, suggesting that basic fibroblast growth factor can be used to expand the population of undifferentiated HFCs to the large numbers needed for therapeutic applications. SMCs were isolated from HFCs using tissue-specific promoters and flow cytometry sorting. Cylindrical vascular constructs engineered with HF-SMCs showed remarkable contractility in response to receptor and nonreceptor agonists such KCl, endothelin-1, and the thromboxane mimetic, U46619, as well as superior mechanical properties compared to their counterparts with human vascular SMCs. Our results suggest that HF is a rich source of mesenchymal stem cells with great potential for myogenic differentiation providing functional SMCs for tissue regeneration and cell therapies. PMID: 20236033 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Microtopographical cues in 3D attenuate fibrotic phenotype and extracellular matrix deposition: implications for tissue regeneration. Tissue Eng Part A. 2010 Aug;16(8):2519-27 Authors: Ayala P, Lopez JI, Desai TA Recent studies have highlighted the role of external biophysical cues on cell morphology and function. In particular, substrate geometry and rigidity in two dimensions has been shown to impact cell growth, death, differentiation, and motility. Knowledge of how these physical cues affect cell function in three dimensions is critical for successful development of novel regenerative therapies. In this work, the effect of discrete micromechanical cues in three-dimensional (3D) system on cell proliferation, gene expression, and extracellular matrix synthesis was investigated. Poly(ethylene glycol) dimethacrylate hydrogel microrods were fabricated using photolithography and suspended in gel to create a 3D culture with microscale cues of defined mechanical properties in the physiological range (2-50 kPa). These microrods significantly affected fibroblast proliferation, matrix production, and gene expression. Cultures with stiff microrods reduced fibroblast proliferation and downregulated expression of key extracellular matrix proteins involved in scar tissue formation. In addition, the contractility marker alpha smooth muscle actin and adhesion molecule integrin alpha3 were also significantly downregulated. Cultures with soft microrods had no significant difference on fibroblast proliferation and expression of Cyclin D1, alpha smooth muscle actin, and integrin alpha3 compared to cultures with no microrods. Here, we present a new platform of potentially injectable microrods with tunable elasticity; in addition, we show that cell proliferation and gene expression are influenced by discrete physical cues in 3D. PMID: 20235832 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Spontaneous redifferentiation of dedifferentiated human articular chondrocytes on hydrogel surfaces. Tissue Eng Part A. 2010 Aug;16(8):2529-40 Authors: Yang JJ, Chen YM, Liu JF, Kurokawa T, Gong JP Chondrocytes rapidly dedifferentiate into a more fibroblastic phenotype on a two-dimensional polystyrene substratum. This impedes fundamental research on these cells as well as their clinical application. This study investigated the redifferentiation behavior of dedifferentiated chondrocytes on a hydrogel substratum. Dedifferentiated normal human articular chondrocyte-knee (NHAC-kn) cells were released from the sixth-passage monolayer cultured on a polystyrene surface. These cells were then subcultured on a chemically crosslinked copolymer hydrogel, that is, poly(NaAMPS-co-DMAAm), and the cells thus obtained were used as the seventh-passage cultivation. Copolymer gels were synthesized from a negatively charged monomer, the sodium salt of 2-acrylamido-2-methyl-1-propanesulfonic acid (NaAMPS), and a neutral monomer, N,N-dimethylacrylamide (DMAAm). These gels were of different compositions because the molar fraction (F) of NaAMPS was varied (F = 0, 0.2, 0.4, 0.6, 0.8, and 1.0). The dedifferentiated NHAC-kn cells spontaneously redifferentiated to normal NHAC-kn cells on neutral (F = 0) and poly(NaAMPS-co-DMAAm) hydrogels of low charge density (F = 0.2). This was deduced from the cell morphology and expression of cartilage-specific genes and proteins. These results should enable us to establish a simple and efficient method for preparing large amounts of chondrocytes by cultivation on the surfaces of neutral and low-charge-density hydrogels. PMID: 20233009 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Host myeloid cells are necessary for creating bioengineered human vascular networks in vivo. Tissue Eng Part A. 2010 Aug;16(8):2457-66 Authors: Melero-Martin JM, De Obaldia ME, Allen P, Dudley AC, Klagsbrun M, Bischoff J The recruitment of myeloid cells has been consistently associated with the formation of new blood vessels during pathological angiogenesis. However, the participation of myeloid cells in bioengineered vascular networks remains unclear. Therefore, we tested whether host myeloid cells play a role in the formation of bioengineered vascular networks that occurs in vivo upon coimplantation of blood-derived endothelial progenitor cells and bone-marrow-derived mesenchymal progenitor cells, suspended as single cells in Matrigel, into immune-deficient mice. We observed an influx of spatially organized host CD11b(+) myeloid cells into the Matrigel implant 1 to 3 days after implantation, which was shown to be cell mediated rather than a nonspecific response. Myeloid cells were significantly reduced once the implants were fully vascularized at days 6 and 7, suggesting an active role during steps that precede formation of functional anastomoses and perfused vessels. Importantly, depletion of circulating myeloid cells resulted in a significant reduction in microvessel density in the implants. In summary, the recruitment of myeloid cells occurs rapidly after coimplantation of endothelial and mesenchymal progenitor cells and is necessary for full vascularization in this model. This is the first demonstration of a role for recruited myeloid cells in the formation of bioengineered vascular networks. PMID: 20218762 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Comparison of ectopic bone formation of embryonic stem cells and cord blood stem cells in vivo. Tissue Eng Part A. 2010 Aug;16(8):2475-83 Authors: Handschel J, Naujoks C, Langenbach F, Berr K, Depprich RA, Ommerborn MA, Kübler NR, Brinkmann M, Kögler G, Meyer U Cell-based reconstruction therapies promise new therapeutic opportunities for bone regeneration. Unrestricted somatic stem cells (USSC) from cord blood and embryonic stem cells (ESCs) can be differentiated into osteogenic cells. The purpose of this in vivo study was to compare their ability to induce ectopic bone formation in vivo. Human USSCs and murine ESCs were cultured as both monolayer cultures and micromasses and seeded on insoluble collagenous bone matrix (ICBM). One week and 1, 2, and 3 months after implanting the constructs in immune-deficient rats, computed tomography scans were performed to detect any calcification. Subsequently, the implanted constructs were examined histologically. The radiological examination showed a steep increase in the mineralized bone-like tissue in the USSC groups. This increase can be considered as statistically significant compared to the basic value. Moreover, the volume and the calcium portion measured by computed tomography scans were about 10 times higher than in the ESC group. The volume of mineralization in the ESC group increased to a much smaller extent over the course of time, and the control group (ICBM without cells) showed almost no alterations during the study. The histological examinations parallel the radiological findings. Cord blood stem cells in combination with ICBM-induced ectopic bone formation in vivo are stronger than ESCs. PMID: 20214449 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | A survey on cellular and engineered tissue therapies in europe in 2008. Tissue Eng Part A. 2010 Aug;16(8):2419-27 Authors: Martin I, Baldomero H, Tyndall A, Niederwieser D, Gratwohl A Cellular therapy is an evolving investigational treatment modality in regenerative medicine, but little published information is available on its current use. Starting from the established European group for Blood and Marrow Transplantation activity survey on hematopoietic stem cell transplantation, a joint committee of four major scientific organizations made a coordinated attempt to collect detailed information in Europe for the year 2008. Thirty-three teams from 16 countries reported data on 656 patients to a "novel cellular therapy" survey, which were combined to additional 384 records reported to the standard European group for Blood and Marrow Transplantation survey. Indications were cardiovascular (29%; 100% autologous), musculoskeletal (18%; 97% autologous), neurological (9%; 39% autologous), epithelial/parenchymal (9%; 18% autologous), autoimmune diseases (12%; 77% autologous), or graft-versus-host disease (23%; 13% autologous). Reported cell types were hematopoietic stem cells (39%), mesenchymal stromal cells (47%), chondrocytes (5%), keratinocytes (7%), myoblasts (2%), and others (1%). In 51% of the grafts, cells were delivered after expansion; in 4% of the cases, cells were transduced. Cells were delivered intravenously (31%), intraorgan (45%), on a membrane or gel (14%), or using three-dimensional scaffolds (10%). This data collection platform is expected to capture and foresee trends for novel cellular therapies in Europe, and warrants further consolidation and extension. PMID: 20184422 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | 50 Years Later: Remembering the Paper. Radiat Res. 2011 Feb;175(2):143-144 Authors: Weissman IL PMID: 21268706 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Immunofluorescent detection of two thymidine analogues (CldU and IdU) in primary tissue. J Vis Exp. 2010;(46): Authors: Tuttle AH, Rankin MM, Teta M, Sartori DJ, Stein GM, Kim GJ, Virgilio C, Granger A, Zhou D, Long SH, Schiffman AB, Kushner JA Accurate measurement of cell division is a fundamental challenge in experimental biology that becomes increasingly complex when slowly dividing cells are analyzed. Established methods to detect cell division include direct visualization by continuous microscopy in cell culture, dilution of vital dyes such as carboxyfluorescein di-aetate succinimidyl ester (CFSE), immuno-detection of mitogenic antigens such as ki67 or PCNA, and thymidine analogues. Thymidine analogues can be detected by a variety of methods including radio-detection for tritiated thymidine, immuno-detection for bromo-deoxyuridine (BrdU), chloro-deoxyuridine (CldU) and iodo-deoxyuridine (IdU), and chemical detection for ethinyl-deoxyuridine (EdU). We have derived a strategy to detect sequential incorporation of different thymidine analogues (CldU and IdU) into tissues of adult mice. Our method allows investigators to accurately quantify two successive rounds of cell division. By optimizing immunostaining protocols our approach can detect very low dose thymidine analogues administered via the drinking water, safe to administer to mice for prolonged periods of time. Consequently, our technique can be used to detect cell turnover in very long-lived tissues. Optimal immunofluoresent staining results can be achieved in multiple tissue types, including pancreas, skin, gut, liver, adrenal, testis, ovary, thyroid, lymph node, and brain. We have also applied this technique to identify oncogenic transformation within tissues. We have further applied this technique to determine if transit-amplifying cells contribute to growth or renewal of tissues. In this sense, sequential administration of thymidine analogues represents a novel approach for studying the origins and survival of cells involved in tissue homeostasis. PMID: 21178965 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | FGF10 controls the patterning of the tracheal cartilage rings via Shh. Development. 2011 Jan;138(2):273-82 Authors: Sala FG, Del Moral PM, Tiozzo C, Alam DA, Warburton D, Grikscheit T, Veltmaat JM, Bellusci S During embryonic development, appropriate dorsoventral patterning of the trachea leads to the formation of periodic cartilage rings from the ventral mesenchyme and continuous smooth muscle from the dorsal mesenchyme. In this work, we have investigated the role of two crucial morphogens, fibroblast growth factor 10 and sonic hedgehog, in the formation of periodically alternating cartilaginous and non-cartilaginous domains in the ventral mesenchyme. Using a combination of gain- and loss-of-function approaches for FGF10 and SHH, we demonstrate that precise spatio-temporal patterns and appropriate levels of expression of these two signaling molecules in the ventral area are crucial between embryonic day 11.5 and 13.5 for the proper patterning of the cartilage rings. We conclude that the expression level of FGF10 in the mesenchyme has to be within a critical range to allow for periodic expression of Shh in the ventral epithelium, and consequently for the correct patterning of the cartilage rings. We propose that disturbed balances of Fgf10 and Shh may explain a subset of human tracheomalacia without tracheo-esophageal fistula or tracheal atresia. PMID: 21148187 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Sonic Hedgehog influences the balance of osteogenesis and adipogenesis in mouse adipose-derived stromal cells. Tissue Eng Part A. 2010 Aug;16(8):2605-16 Authors: James AW, Leucht P, Levi B, Carre AL, Xu Y, Helms JA, Longaker MT Adipose-derived stromal cells (ASCs) present a great potential for tissue engineering, as they are capable of differentiating into osteogenic and adipogenic cell types, among others. In this study, we examined the role of Hedgehog signaling in the balance of osteogenic and adipogenic differentiation in mouse ASCs. Results showed that Hedgehog signaling increased during early osteogenic differentiation (Shh, Ptc1, and Gli1), but decreased during adipogenic differentiation. N-terminal Sonic Hedgehog (Shh-N) significantly increased in vitro osteogenic differentiation in mouse ASCs, by all markers examined (*p < 0.01). Concomitantly, Shh-N abrogated adipogenic differentiation, by all markers examined (*p < 0.01). Conversely, blockade of endogenous Hedgehog signaling, with the Hedgehog antagonist cyclopamine, enhanced adipogenesis at the expense of osteogenesis. We next translated these results to a mouse model of appendicular skeletal regeneration. Using quantitative real-time polymerase chain reaction and in situ hybridization, we found that skeletal injury (a monocortical 1 mm defect in the tibia) results in a localized increase in Hedgehog signaling. Moreover, grafting of ASCs treated with Shh-N resulted in significantly increased bone regeneration within the defect site. In conclusion, Hedgehog signaling enhances the osteogenic differentiation of mouse ASCs, at the expense of adipogenesis. These data suggest that Hedgehog signaling directs the lineage differentiation of mesodermal stem cells and represents a promising strategy for skeletal tissue regeneration. PMID: 20367246 [PubMed - indexed for MEDLINE] | | | | | | | | | |  | |  | |  |
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