|  |  |  | | | | | TE-RegenMed-StemCell feed | | | | | | | | | | | | | | | | | | | Phases I-III Clinical Trials Using Adult Stem Cells. Stem Cells Int. 2010;2010:579142 Authors: Sanz-Ruiz R, Gutiérrez Ibañes E, Arranz AV, Fernández Santos ME, Fernández PL, Fernández-Avilés F First randomized clinical trials have demonstrated that stem cell therapy can improve cardiac recovery after the acute phase of myocardial ischemia and in patients with chronic ischemic heart disease. Nevertheless, some trials have shown that conflicting results and uncertainties remain in the case of mechanisms of action and possible ways to improve clinical impact of stem cells in cardiac repair. In this paper we will examine the evidence available, analyze the main phase I and II randomized clinical trials and their limitations, discuss the key points in the design of future trials, and depict new directions of research in this fascinating field. PMID: 21076533 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Monoclonal antibody therapy directed against human acute myeloid leukemia stem cells. Oncogene. 2010 Nov 15; Authors: Majeti R Accumulating evidence indicates that many human cancers are organized as a cellular hierarchy initiated and maintained by self-renewing cancer stem cells. This cancer stem cell model has been most conclusively established for human acute myeloid leukemia (AML), although controversies still exist regarding the identity of human AML stem cells (leukemia stem cell (LSC)). A major implication of this model is that, in order to eradicate the cancer and cure the patient, the cancer stem cells must be eliminated. Monoclonal antibodies have emerged as effective targeted therapies for the treatment of a number of human malignancies and, given their target antigen specificity and generally minimal toxicity, are well positioned as cancer stem cell-targeting therapies. One strategy for the development of monoclonal antibodies targeting human AML stem cells involves first identifying cell surface antigens preferentially expressed on AML LSC compared with normal hematopoietic stem cells. In recent years, a number of such antigens have been identified, including CD123, CD44, CLL-1, CD96, CD47, CD32, and CD25. Moreover, monoclonal antibodies targeting CD44, CD123, and CD47 have demonstrated efficacy against AML LSC in xenotransplantation models. Hopefully, these antibodies will ultimately prove to be effective in the treatment of human AML.Oncogene advance online publication, 15 November 2010; doi:10.1038/onc.2010.511. PMID: 21076471 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | A defined glycosaminoglycan-binding substratum for human pluripotent stem cells. Nat Methods. 2010 Nov 14; Authors: Klim JR, Li L, Wrighton PJ, Piekarczyk MS, Kiessling LL To exploit the full potential of human pluripotent stem cells for regenerative medicine, developmental biology and drug discovery, defined culture conditions are needed. Media of known composition that maintain human embryonic stem (hES) cells have been developed, but finding chemically defined, robust substrata has proven difficult. We used an array of self-assembled monolayers to identify peptide surfaces that sustain pluripotent stem cell self-renewal. The effective substrates displayed heparin-binding peptides, which can interact with cell-surface glycosaminoglycans and could be used with a defined medium to culture hES cells for more than 3 months. The resulting cells maintained a normal karyotype and had high levels of pluripotency markers. The peptides supported growth of eight pluripotent cell lines on a variety of scaffolds. Our results indicate that synthetic substrates that recognize cell-surface glycans can facilitate the long-term culture of pluripotent stem cells. PMID: 21076418 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | The orphan nuclear receptor Nurr1 restricts the proliferation of haematopoietic stem cells. Nat Cell Biol. 2010 Nov 14; Authors: Sirin O, Lukov GL, Mao R, Conneely OM, Goodell MA Successful haematopoiesis requires long-term retention of haematopoietic stem cells (HSCs) in a quiescent state. The transcriptional regulation of stem cell quiescence, especially by factors with specific functions in HSCs, is only beginning to be understood. Here, we demonstrate that Nurr1, a nuclear receptor transcription factor, has such a regulatory role. Overexpression of Nurr1 drives early haematopoietic progenitors into quiescence. When stem cells overexpressing Nurr1 are transplanted into lethally irradiated mice, they localize to the bone marrow, but do not contribute to regeneration of the blood system. Furthermore, the loss of only one allele of Nurr1 is sufficient to induce HSCs to enter the cell cycle and proliferate. Molecular analysis revealed an association between Nurr1 overexpression and upregulation of the cell-cycle inhibitor p18 (also known as INK4C), suggesting a mechanism by which Nurr1 could regulate HSC quiescence. Our findings provide critical insight into the transcriptional control mechanisms that determine whether HSCs remain dormant or enter the cell cycle and begin to proliferate. PMID: 21076412 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Induction of Kidney Allograft Tolerance by Soluble CD83 Associated With Prevalence of Tolerogenic Dendritic Cells and Indoleamine 2,3-Dioxygenase. Transplantation. 2010 Nov 11; Authors: Lan Z, Ge W, Arp J, Jiang J, Liu W, Gordon D, Healey D, Debenedette M, Nicolette C, Garcia B, Wang H BACKGROUND.: Tolerogenic dendritic cells (Tol-DCs) play a critical role in inducing and maintaining tolerance. Recognizing that both T-cell inactivation and activation are contingent on signals provided by DCs and that graft-specific activated T cells are major mediators of transplant rejection, we aimed to create an environment favoring Tol-DCs with a novel reagent, human soluble CD83 (hsCD83). METHODS.: Life-supporting orthotopic kidney transplantation was performed in a C57BL/6-to-BALB/c mouse model. The study group was treated with hsCD83 (100 μg/mouse/day, postoperative days -1 to +7, intravenously) and compared with untreated controls. RESULTS.: Treatment with hsCD83 achieved kidney allograft tolerance (>100 days), with negligible antidonor antibody detected. In contrast, kidney grafts in untreated recipients demonstrated severe rejection after 35 days, characterized by cellular infiltration, interstitial hemorrhage and edema, and glomerular and tubular necrosis, as well as high antidonor antibody titers. In addition, splenic DCs of tolerant recipients exhibited significantly decreased levels of surface major histocompatibility complex class II, CD40, CD80, and intracellular interleukin-12, as well as reduced allogeneic stimulatory capacity. Adoptive transfer of CD11cDCs from tolerant hsCD83-treated animals induced kidney allograft tolerance in syngeneic recipients. Blocking indoleamine 2,3-dioxygenase with 1-methyl-tryptophan (15 mg/mouse/day; gavage) prevented the immunosuppressive effect of hsCD83, abrogating hsCD83-induced Tol-DCs and graft tolerance, and leading to acute kidney graft rejection in 22 days. CONCLUSION.: hsCD83 alone was capable of inducing kidney allograft tolerance through a mechanism involving Tol-DC generation and, at least in part, indoleamine 2,3-dioxygenase activity. Because sCD83 is of human origin, the therapeutic approach used in our mouse transplant model holds significant promise for clinical transplantation. PMID: 21076370 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Core transcription factors, Oct4, Sox2 and Nanog, individually form complexes with nucleophosmin (Npm1) to control embryonic stem (ES) cell fate determination. Aging (Albany NY). 2010 Nov 12; Authors: Johansson H, Simonsson S Embryonic stem (ES) cells have therapeutic potential in regenerative medicine, although the molecular mechanism controlling their pluripotency is not completely understood. Depending on interaction partners most proteins can be involved in several different cellular mechanisms. We screened for novel protein-protein interactions usingin situ proximity ligation assays together with specific antibodies directed against known important ES cell proteins. We found that all three core transcription factors, namely Oct4, Sox2 and Nanog, individually formed complexes with nucleophosmin (Npm1). We showed that the Npm1/Sox2 complex was sustained when cells were induced to differentiate by retinoic acid, while decreased in the other differentiation pathways. Moreover, Oct4 also formed individual complexes with translationally controlled tumor protein (Tpt1). Downregulation of Npm1 or Tpt1 increased mRNA levels for genes involved in mesoderm and ectoderm differentiation pathways, respectively, indicative of their involvement in ES cell maintenance. We have here described four novel protein-protein interactions in ES cell involving all three core transcription factors. Our findings improve the current knowledge about ES cell-specific protein networks and indicate the importance of Npm1 and Tpt1 to maintain the ES cell phenotype. PMID: 21076177 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Pharmacologic modulation of the calcium-sensing receptor enhances hematopoietic stem cell lodgment in the adult bone marrow. Blood. 2010 Nov 12; Authors: Lam BS, Cunningham C, Adams GB The ability of hematopoietic stem cells (HSCs) to undergo self-renewal is partly regulated by external signals originating from the stem cell niche. Our previous studies using HSCs obtained from the fetal liver of mice deficient for the calcium-sensing receptor (CaR) have shown the crucial role of this receptor in HSC lodgment and engraftment in the bone marrow (BM) endosteal niche. Using a CaR agonist, Cinacalcet, we assessed the effects of stimulating the CaR on the function of murine HSCs. Our results demonstrate that CaR stimulation increases primitive hematopoietic cell activity in vitro, including growth in stromal cell co-cultures, adhesion to extracellular matrix molecules such as collagen I and fibronectin, and migration towards the chemotactic stimulus, SDF-1α. Receptor stimulation also led to augmented in vivo homing, CXCR4-mediated lodgment at the endosteal niche, and engraftment capabilities. These mechanisms by which stimulating the CaR dictates preferential localization of HSCs in the BM endosteal niche provide additional insights into the fundamental interrelationship between the stem cell and its niche. These studies also have implications in the area of clinical stem cell transplantation, where ex vivo modulation of the CaR may be envisioned as a strategy to enhance HSC engraftment in the BM. PMID: 21076044 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | [Vascularized periosteum and bone regeneration.] Chir Main. 2010 Oct 8; Authors: Moukoko D The osteogenic potential of periosteum is widely recognized. During development, it plays a prominent role in the radial growth of long bones. Similarly, it has a key role in the consolidation of fractures. The physiological function of periosteum in the healthy, mature skeleton remains relatively subtle; however, its detachment from the bone surface reactivates its potential for fibrogenic and osteochondrogenic regeneration. This discreet anatomical structure is actually a reservoir of mesenchymal progenitor cells capable of proliferating and differentiating, by reinitializing cellular and molecular cascades of embryogenesis in mesenchymal tissues. However, given the hitherto limited knowledge of the quantitative potential of periosteum and of the pathways regulating tissue differentiation during regeneration, human applications have remained anecdotal. The findings of several in vivo and in vitro experiments indicate that the maintenance of the periosteum's vascularization stimulates its quantitative potential. The structural organization of the regenerated material in vivo is governed by locoregional biological and mechanical regulatory mechanisms that serve to make it capable of performing its new functions. The increasing awareness of periosteum's potential is stimulating active research in the fields of cellular biology and tissue engineering. The demonstration of its regenerative potential in animals gives reason to believe that strips of vascularized periosteum could become part of the developing armamentarium of regenerative medicine. PMID: 21075660 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Carbohydrate-mediated inhibition of ice recrystallization in cryopreserved human umbilical cord blood. Carbohydr Res. 2010 Oct 26; Authors: Wu LK, Tokarew JM, Chaytor JL, Moos EV, Li Y, Palii C, Ben RN, Allan DS Cryopreservation of human umbilical cord blood (UCB) typically involves the cryoprotectant dimethylsulfoxide (DMSO), however, infusional toxicity and reductions in cell viability remain a concern. Ice recrystallization (IR) is an important source of cryopreservation-induced cellular injury and limits the stem cell dose in UCB units. Carbohydrates have wide-ranging intrinsic IR inhibition (IRI) activity related to structural properties. We investigated the impact of carbohydrate IRI on cell viability, induction of apoptosis and hematopoietic progenitor function in cryopreserved UCB. Mononuclear cells (MNCs) from UCB were cryopreserved in storage media containing specific carbohydrates (200mM) and compared to 5% DMSO. Samples were analyzed under conditions of high IR ('slow' thaw) and low IR ('fast' thaw). Thawed samples were analyzed for viability and apoptosis by flow cytometry and hematopoietic function using colony-forming unit (CFU) assays. IRI of carbohydrate solutions was determined using the 'splat cooling' assay. Greater IRI capacity of carbohydrates correlated with increased yield of viable MNCs (r(2)=0.92, p=0.004) and CD34(+) cells (r(2)=0.96, p=0.019) after thawing under conditions of high IR. The correlations were less apparent under conditions of low IR. Carbohydrates with greater IRI modulate the induction of early apoptosis during thawing, especially in CD34+ cells (r(2)=0.96, p=0.0001) as compared to total mononuclear cells (p=0.006), and preserve CFU capacity in vitro (r(2)=0.92, p=<0.0001). Our results suggest that carbohydrates with potent IRI increase the yield of non-apoptotic and functional hematopoietic progenitors and provide a foundation for the development of novel synthetic carbohydrates with enhanced IRI properties to improve cryopreservation of UCB. PMID: 21075361 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Vision from the right stem. Trends Mol Med. 2010 Nov 11; Authors: Pellegrini G, Rama P, De Luca M Cultures of limbal cells are a safe and effective treatment for the destruction of the human cornea owing to chemical burns. The essential feature of the graft is the presence of an adequate number of stem cells, which can be determined by the expression of the p63 transcription factor. Here, we will discuss the general principles defining the rigorous criteria for graftable limbal cultures in light of their clinical performances. Such criteria might prove relevant to the future therapeutic use of any cultured cell type. PMID: 21075055 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | High-density collagen gel tubes as a matrix for primary human bladder smooth muscle cells. Biomaterials. 2010 Nov 11; Authors: Micol LA, Ananta M, Engelhardt EM, Mudera VC, Brown RA, Hubbell JA, Frey P Tissue-engineered grafts for the urinary tract are being investigated for the potential treatment of several urologic diseases. These grafts, predominantly tubular-shaped, usually require in vitro culture prior to implantation to allow cell engraftment on initially cell-free scaffolds. We have developed a method to produce tubular-shaped collagen scaffolds based on plastic compression. Our approach produces a ready cell-seeded graft that does not need further in vitro culture prior to implantation. The tubular collagen scaffolds were in particular investigated for their structural, mechanical and biological properties. The resulting construct showed an especially high collagen density, and was characterized by favorable mechanical properties assessed by axial extension and radial dilation. Young modulus in particular was greater than non-compressed collagen tubes. Seeding densities affected proliferation rate of primary human bladder smooth muscle cells. An optimal seeding density of 10(6) cells per construct resulted in a 25-fold increase in Alamar blue-based fluorescence after 2 wk in culture. These high-density collagen gel tubes, ready seeded with smooth muscle cells could be further seeded with urothelial cells, drastically shortening the production time of graft for urinary tract regeneration. PMID: 21074843 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Modeling rett syndrome with stem cells. Cell. 2010 Nov 12;143(4):499-500 Authors: Walsh RM, Hochedlinger K The discovery that somatic cells can be reprogrammed into induced pluripotent stem cells (iPSCs) raised the exciting possibility of modeling diseases with patient-specific cells. Marchetto et al. (2010) now use iPSC technology to generate, characterize, and treat an in vitro model for the autism spectrum disorder Rett syndrome. PMID: 21074040 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Multipotent stem cells in the biliary tree. Ital J Anat Embryol. 2010;115(1-2):85-90 Authors: Cardinale V, Wang Y, Carpino G, Alvaro D, Reid L, Gaudio E The biliary tree system consists of two divisions: intrahepatic bile ducts and extrahepatic bile ducts. The development of the biliary tree, and secondarily the liver, shares a common origin with ventral pancreas. A common progenitor for liver, biliary duct system, and ventral pancreas exists at early stages of development, when the anterior definitive endoderm is forming the foregut. Several studies indicate that the biliary tree contains stem cell compartments for liver, pancreas and the bile duct system and persisting into adulthood. These stem cell compartments are present in the peribiliary glands and possibly give rise to committed progenitors in gallbladder that does not have peribiliary glands. The biliary tree stem/progenitors represent a new source of cells that can be used as tools for regenerative medicine of liver, bile duct and pancreas. PMID: 21072995 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Nanotechnology in ophthalmology. Can J Ophthalmol. 2010 Oct;45(5):457-76 Authors: Zarbin MA, Montemagno C, Leary JF, Ritch R Nanotechnology involves the creation and use of materials and devices at the size scale of intracellular structures and molecules, and involves systems and constructs in the order of <100 nm. The aim of nanomedicine is the comprehensive monitoring, control, construction, repair, defence, and improvement of human biological systems at the molecular level, using engineered nanodevices and nanostructures that operate massively in parallel at the single-cell level, ultimately to achieve medical benefit. In this review we consider general principles of nanotechnology as applied to nanomedicine (e.g., biomimicry and pseudointelligence). Some applications of nanotechnology to ophthalmology are described (including treatment of oxidative stress; measurement of intraocular pressure; theragnostics; use of nanoparticles to treat choroidal new vessels, prevent scarring after glaucoma surgery, and treat retinal degenerative disease with gene therapy; prosthetics; and regenerative nanomedicine). Nanotechnology will revolutionize our approach to current therapeutic challenges (e.g., drug delivery, postoperative scarring) and will enable us to address currently unsolvable problems (e.g., sight-restoring therapy for patients with retinal degenerative disease). Obstacles to the incorporation of nanotechnology remain, such as safe manufacturing techniques and unintended biological consequences of nanomaterial use. These obstacles are not insurmountable, and revolutionary treatments for ophthalmic diseases are expected to result from this burgeoning field. PMID: 20871642 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Effect of a Scaffold Fabricated Thermally from Acetylated PLGA on the Formation of Engineered Cartilage. Macromol Biosci. 2010 Nov 15; Authors: Kang SW, Lee SJ, Kim JS, Choi EH, Cha BH, Shim JH, Cho DW, Lee SH A MHDS has been employed to fabricate 3D scaffolds from PLGA with acetyl endgroups to achieve in vivo regeneration of cartilage tissue. The fabricated acetylated-PLGA scaffold showed open pores and interconnected structures. Rabbit chondrocytes were seeded on the PLGA scaffolds and transplanted immediately into subcutaneous sites of athymic mice. Chondrocytes transplantation with untreated PLGA scaffolds served as a control. Histological analysis of the implants at 4 weeks with H&E staining and alcian blue staining revealed higher extracellular matrix and GAG expression at the neocartilage in the PLGA-6Ac scaffolds than that of the PLGA-6OH scaffold group. This endgroup-modified scaffold may be useful for successful cartilage tissue engineering in orthopedic applications. PMID: 21077228 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Multi-Featured Macroporous Agarose-Alginate Cryogel: Synthesis and Characterization for Bioengineering Applications. Macromol Biosci. 2010 Nov 15; Authors: Tripathi A, Kumar A In this study agarose-alginate scaffolds are synthesized using cryogelation technology in different formats like monolith, sheet, discs, and beads, and show amiable mechanical strength like soft tissue properties and high interconnected macroporous degradable architecture. In cell-material interactions, fibroblast (NIH-3T3) cells showed good adherence and proliferation on these scaffolds presenting its potential application in soft tissue engineering. The application of cryogel beads and monoliths was also examined by the efficient immobilization of bacterial cells (BL21) on these matrices revealing their use for recovery of product from continuous fermentation systems without cell leakage. These scaffolds also showed potential as a filter for repeated recovery of heavy metal binding, such as copper and nickel from the waste water. The cryogels prepared herein do have a number of unique features that make them an important class of soft materials for developing multi-featured scaffolds as a novel carrier for bioengineering applications. PMID: 21077225 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Synthesis of Water Soluble, Biodegradable, and Electroactive Polysaccharide Crosslinker with Aldehyde and Carboxylic Groups for Biomedical Applications. Macromol Biosci. 2010 Nov 15; Authors: Wang Q, He W, Huang J, Liu S, Wu G, Teng W, Wang Q, Dong Y We report the synthesis and characterization of a polysaccharide crosslinker of tetraaniline grafting oxidized sodium alginate with large aldehyde and carboxylic groups. We demonstrate that this copolymer has the following properties: it is water soluble under any pH, biodegradable, electroactive, and noncytotoxic; it can self-assemble into nanoparticles with large active functional groups on the outer surface; it can crosslink materials with amino and aminoderivative groups like gelatin to form hydrogels, and thus the electroactivity is readily introduced to the materials. This copolymer has potential applications in biomedical fields such as tissue engineering, drug delivery, and nerve probes where electroactivity is required. PMID: 21077224 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Self-Assembled Gels for Biomedical Applications. Chem Asian J. 2010 Nov 12; Authors: Truong WT, Su Y, Meijer JT, Thordarson P, Braet F Natural and synthetic gel-like materials have featured heavily in the development of biomaterials for wound healing and other tissue-engineering purposes. More recently, molecular gels have been designed and tailored for the same purpose. When mixed with, or conjugated to therapeutic drugs or bioactive molecules, these materials hold great promise for treating/curing life-threatening and degenerative diseases, such as cancer, osteoarthritis, and neural injuries. This focus review explores the latest advances in this field and concentrates on self-assembled gels formed under aqueous conditions (i.e., self-assembled hydrogels), and critically compares their performance within different biomedical applications, including three-dimensional cell-culture studies, drug delivery, and tissue engineering. Although stability and toxicity issues still need to be addressed in more detail, it is clear from the work reviewed here that self-assembled gels have a bright future as novel biomaterials. PMID: 21077096 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Hydrolyzed fish collagen induced chondrogenic differentiation of equine adipose tissue-derived stromal cells. Histochem Cell Biol. 2010 Nov 14; Authors: Raabe O, Reich C, Wenisch S, Hild A, Burg-Roderfeld M, Siebert HC, Arnhold S Adipose-derived stromal cells (ADSCs) are multipotent cells which, in the presence of appropriate stimuli, can differentiate into various lineages such as the osteogenic, adipogenic and chondrogenic. In this study, we investigated the effect of transforming growth factor beta 1 (TGF-β1) in comparison to hydrolyzed fish collagen in terms of the chondrogenic differentiation potential of ADSCs. ADSCs were isolated from subcutaneous fat of horses by liposuction. Chondrogenesis was investigated using a pellet culture system. The differentiation medium was either supplemented with TGF-β1 (5 ng/ml) or fish collagen (0.5 mg/ml) for a 3 week period. After the 3 weeks in vitro differentiation, RT-PCR and histological staining for proteoglycan synthesis and type II collagen were performed to evaluate the degree of chondrogenic differentiation and the formation of cartilaginous extracellular matrix (ECM). The differentiation of ADSCs induced by TGF-β1 showed a high expression of glycosaminoglycan (GAG). Histological analysis of cultures stimulated by hydrolyzed fish collagen demonstrated an even higher GAG expression than cultures stimulated under standard conditions by TGF-β1. The expression of cartilage-specific type II collagen and Sox9 was about the same in both stimulated cultures. In this study, chondrogenesis was as effectively induced by hydrolyzed fish collagen as it was successfully induced by TGF-β1. These findings demonstrated that hydrolyzed fish collagen alone has the potential to induce and maintain ADSCs-derived chondrogenesis. These results support the application of ADSCs in equine veterinary tissue engineering, especially for cartilage repair. PMID: 21076963 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Functional stability of endothelial cells on a novel hybrid scaffold for vascular tissue engineering. Biofabrication. 2010 Sep 24;2(4):041001 Authors: Pankajakshan D, Krishnan V K, Krishnan LK Porous and pliable conduits made of biodegradable polymeric scaffolds offer great potential for the development of blood vessel substitutes but they generally lack signals for cell proliferation, survival and maintenance of a normal phenotype. In this study we have prepared and evaluated porous poly(ε-caprolactone) (PCL) integrated with fibrin composite (FC) to get a biomimetic hybrid scaffold (FC PCL) with the biological properties of fibrin, fibronectin (FN), gelatin, growth factors and glycosaminoglycans. Reduced platelet adhesion on a human umbilical vein endothelial cell-seeded hybrid scaffold as compared to bare PCL or FC PCL was observed, which suggests the non-thrombogenic nature of the tissue-engineered scaffold. Analysis of real-time polymerase chain reaction (RT-PCR) after 5 days of endothelial cell (EC) culture on a hybrid scaffold indicated that the prothrombotic von Willebrand factor and plasminogen activator inhibitor (PAI) were quiescent and stable. Meanwhile, dynamic expressions of tissue plasminogen activator (tPA) and endothelial nitric oxide synthase indicated the desired cell phenotype on the scaffold. On the hybrid scaffold, shear stress could induce enhanced nitric oxide release, which implicates vaso-responsiveness of EC grown on the tissue-engineered construct. Significant upregulation of mRNA for extracellular matrix (ECM) proteins, collagen IV and elastin, in EC was detected by RT-PCR after growing them on the hybrid scaffold and FC-coated tissue culture polystyrene (FC TCPS) but not on FN-coated TCPS. The results indicate that the FC PCL hybrid scaffold can accomplish a remodeled ECM and non-thrombogenic EC phenotype, and can be further investigated as a scaffold for cardiovascular tissue engineering. PMID: 21076184 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Collagen-induced expression of collagenase-3 by primary chondrocytes is mediated by integrin {alpha}1 and discoidin domain receptor 2: a protein kinase C-dependent pathway. Rheumatology (Oxford). 2010 Nov 12; Authors: Vonk LA, Doulabi BZ, Huang C, Helder MN, Everts V, Bank RA Objectives. To investigate whether maintaining the chondrocyte's native pericellular matrix prevents collagen-induced up-regulation of collagenase-3 (MMP-13) and whether integrin α1 (ITGα1) and/or discoidin domain receptor 2 (DDR2) modulate MMP-13 expression and which signalling pathway plays a role in collagen-stimulated MMP-13 expression. Methods. Goat articular chondrocytes and chondrons were cultured on collagen coatings. Small interfering RNA (siRNA) oligonucleotides targeted against ITGα1 and DDR2 were transfected into primary chondrocytes. Chemical inhibitors for mitogen-activated protein kinase kinase (MEK1) (PD98059), focal adhesion kinase (FAK) (FAK inhibitor 14), mitogen-activated protein kinase 8 (JNK) (SP600125) and protein kinase C (PKC) (PKC412), and a calcium chelator (BAPTA-AM) were used in cell cultures. Real-time PCR was performed to examine gene expression levels of MMP-13, ITGα1 and DDR2 and collagenolytic activity was determined by measuring the amount of hydroxyproline released in the culture medium. Results. Maintaining the chondrocyte's native pericellular matrix prevented MMP-13 up-regulation and collagenolytic activity when the cells were cultured on a collagen coating. Silencing of ITGα1 and DDR2 reduced MMP-13 gene expression and collagenolytic activity by primary chondrocytes cultured on collagen. Incubation with the PKC inhibitor strongly reduced MMP-13 gene expression levels. Gene expression levels of MMP-13 were also decreased by chondrocytes incubated with the MEK, FAK or JNK inhibitor. Conclusion. Maintaining the native pericellular matrix of chondrocytes prevents collagen-induced up-regulation of MMP-13. Both ITGα1 and DDR2 modulate MMP-13 expression after direct contact between chondrocytes and collagen. PKC, FAK, MEK and JNK are involved in collagen-stimulated expression of MMP-13. PMID: 21075784 [PubMed - as supplied by publisher] | | | | | | | | | |  | |  | |  |
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