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| Micro/nano-fabrication technologies for cell biology.
May 22, 2010 at 6:37 AM
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| | Micro/nano-fabrication technologies for cell biology. Med Biol Eng Comput. 2010 May 21; Authors: Qian T, Wang Y Micro/nano-fabrication techniques, such as soft lithography and electrospinning, have been well-developed and widely applied in many research fields in the past decade. Due to the low costs and simple procedures, these techniques have become important and popular for biological studies. In this review, we focus on the studies integrating micro/nano-fabrication work to elucidate the molecular mechanism of signaling transduction in cell biology. We first describe different micro/nano-fabrication technologies, including techniques generating three-dimensional scaffolds for tissue engineering. We then introduce the application of these technologies in manipulating the physical or chemical micro/nano-environment to regulate the cellular behavior and response, such as cell life and death, differentiation, proliferation, and cell migration. Recent advancement in integrating the micro/nano-technologies and live cell imaging are also discussed. Finally, potential schemes i! n cell biology involving micro/nano-fabrication technologies are proposed to provide perspectives on the future research activities. PMID: 20490938 [PubMed - as supplied by publisher] | | |
| Immunomodulatory effect of mesenchymal stem cells.
May 22, 2010 at 6:37 AM
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| | Immunomodulatory effect of mesenchymal stem cells. Braz J Med Biol Res. 2010 May;43(5):425-30 Authors: Herrero C, Pérez-Simón JA Mesenchymal stem cells (MSC) are multipotential nonhematopoietic progenitor cells capable of differentiating into multiple mesenchymal tissues. MSC are able to reconstitute the functional human hematopoietic microenvironment and promote engraftment of hematopoietic stem cells. MSC constitutively express low levels of major histocompatibility complex-I molecules and do not express costimulatory molecules such as CD80, CD86 or CD40, thus lacking immunogenicity. Furthermore, they are able to suppress T- and B-lymphocyte activation and proliferation and may also affect dendritic cell maturation. Based on these properties, MSC are being used in regenerative medicine and also for the treatment of autoimmune diseases and graft-versus-host disease. On the other hand, MSC from patients diagnosed with myelodysplastic syndromes or multiple myeloma display abnormalities, which could play a role in the physiopathology of the disease. Finally, in patients with immune thrombocyt! openic purpura, MSC have a reduced proliferative capacity and a lower inhibitory effect on T-cell proliferation compared with MSC from healthy donors. PMID: 20490429 [PubMed - in process] | | |
| Differentiation of embryonic stem cells into oligodendrocyte precursors.
May 22, 2010 at 6:37 AM
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| | Differentiation of embryonic stem cells into oligodendrocyte precursors. J Vis Exp. 2010;(39): Authors: Jiang P, Selvaraj V, Deng W Oligodendrocytes are the myelinating cells of the central nervous system. For regenerative cell therapy in demyelinating diseases, there is significant interest in deriving a pure population of lineage-committed oligodendrocyte precursor cells (OPCs) for transplantation. OPCs are characterized by the activity of the transcription factor Olig2 and surface expression of a proteoglycan NG2. Using the GFP-Olig2 (G-Olig2) mouse embryonic stem cell (mESC) reporter line, we optimized conditions for the differentiation of mESCs into GFP+Olig2+NG2+ OPCs. In our protocol, we first describe the generation of embryoid bodies (EBs) from mESCs. Second, we describe treatment of mESC-derived EBs with small molecules: (1) retinoic acid (RA) and (2) a sonic hedgehog (Shh) agonist purmorphamine (Pur) under defined culture conditions to direct EB differentiation into the oligodendroglial lineage. By this approach, OPCs can be obtained with high efficiency (>80%) in a time period o! f 30 days. Cells derived from mESCs in this protocol are phenotypically similar to OPCs derived from primary tissue culture. The mESC-derived OPCs do not show the spiking property described for a subpopulation of brain OPCs in situ. To study this electrophysiological property, we describe the generation of spiking mESC-derived OPCs by ectopically expressing Na(V;)1.2 subunit. The spiking and nonspiking cells obtained from this protocol will help advance functional studies on the two subpopulations of OPCs. PMID: 20489683 [PubMed - in process] | | |
| Bone tissue engineering using porous carbonate apatite and bone marrow cells.
May 22, 2010 at 6:37 AM
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| | Bone tissue engineering using porous carbonate apatite and bone marrow cells. J Craniofac Surg. 2010 Mar;21(2):473-8 Authors: Kasai T, Sato K, Kanematsu Y, Shikimori M, Kanematsu N, Doi Y Porous blocks of carbonate apatite (CA) were prepared by holding together CA particles ranging in size from 300 to 500 mum through sintering at 750 degrees C for 2 hours. Bone marrow cells taken from Fischer rats were seeded onto and inside the CA blocks and cultured for 14 days to allow stem cells to proliferate to osteoblasts capable of inducing bone formation. Hybrids made of CA blocks and cultured bone marrow cells were then implanted into the back of syngeneic rats. Microfocus x-ray computed tomographic images of tissues containing CA blocks before decalcification suggested that new bone was formed in this extraosseous site 4 and 8 weeks after implantation. These data indicate that the hybrid made of CA and bone marrow cells is capable of inducing heterotopic bone formation in vivo. PMID: 20489453 [PubMed - in process] | | |
| Production of l-lactic acid by a thermophilic Bacillus mutant using sodium hydroxide as neutralizing agent.
May 22, 2010 at 6:37 AM
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| | Production of l-lactic acid by a thermophilic Bacillus mutant using sodium hydroxide as neutralizing agent. Bioresour Technol. 2010 May 18; Authors: Qin J, Wang X, Zheng Z, Ma C, Tang H, Xu P A sodium lactate tolerant mutant strain named Bacillus sp. Na-2 was obtained and applied to sodium hydroxide-based l-lactic acid (LA) production process. The influences of aeration and pH were investigated to further improve the resistance of strain Na-2 against sodium lactate stress and to obtain the most efficient l-LA production process. Although mild aeration was favorable for cell growth and l-LA production, vigorous aeration resulted in a metabolic shift from homolactic to mixed-acid/acetoin fermentation. Therefore, a two-stage aeration control strategy was employed. Optimum pH was found to be 6.0. A total of 106.0g/l l-LA was produced in 30h by Bacillus sp. Na-2 using sodium hydroxide as neutralizing agent. Productivity, conversion rate and optical purity were 3.53g/l/h, 94% and 99.5%, respectively. The remarkable fermentation traits of Bacillus sp. Na-2 and the environment-friendly characteristics of NaOH-based process represent new insight for industrial ! scale production of l-LA. PMID: 20488697 [PubMed - as supplied by publisher] | | |
| SiRNA-loaded multi-shell nanoparticles incorporated into a multilayered film as a reservoir for gene silencing.
May 22, 2010 at 6:37 AM
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| | SiRNA-loaded multi-shell nanoparticles incorporated into a multilayered film as a reservoir for gene silencing. Biomaterials. 2010 May 18; Authors: Zhang X, Kovtun A, Mendoza-Palomares C, Oulad-Abdelghani M, Fioretti F, Rinckenbach S, Mainard D, Epple M, Benkirane-Jessel N In this study, we presented a new type of coating based on polyelectrolyte multilayers containing sequentially adsorbed active shRNA calcium phosphate nanoparticles for locally defined and temporarily variable gene silencing. Therefore, we investigated multi-shell calcium phosphate-shRNA nanoparticles embedded into a polyelectrolyte multilayer for gene silencing. As model system, we synthesized triple-shell calcium phosphate-shRNA nanoparticles (NP) and prepared polyelectrolyte multilayers films made of nanoparticles and poly-(l-lysine) (PLL). The biological activities of these polyelectrolyte multilayers films were tested by the production of osteopontin and osteocalcin in the human osteoblasts (HOb) which were cultivated on the PEM films. This new strategy can be used to efficiently control the bone formation and could be applicable in tissue engineering. PMID: 20488536 [PubMed - as supplied by publisher] | | |
| Impaired Bladder Function in Aging Male Rats.
May 22, 2010 at 6:37 AM
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| | Impaired Bladder Function in Aging Male Rats. J Urol. 2010 May 18; Authors: Zhao W, Aboushwareb T, Turner C, Mathis C, Bennett C, Sonntag WE, Andersson KE, Christ G PURPOSE: The prevalence of bladder dysfunctions increases with age. In humans it is difficult to separate changes related to exogenous factors from those directly related to the aging process. Some confounding variables can be avoided by studying age related changes in an animal model. We evaluated the impact of age on bladder function in vivo and in vitro, and characterized the corresponding morphological changes. MATERIALS AND METHODS: Young (4 to 6 months old) and old (older than 28 to 30 months) male Fischer/Brown Norway rats were used in the study. Cystometric studies were done in conscious, freely moving rats. After cystometry tissue strips from the bladder body were used in in vitro studies of muscarinic receptor activation and electrical field stimulation, and histological examination. RESULTS: Old rats had higher bladder weight than young rats but the bladder-to-body weight ratio did not change. We noted significant age related differences in 8 of 10 cyst! ometric parameters. Old rats had increased bladder capacity, post-void residual volume, micturition volume and frequency, baseline and intermicturition pressure, and spontaneous activity but decreased micturition pressure. Bladder strip responses to carbachol and electrical field stimulation were significantly lower in old than in young rats. Histological examination revealed urothelial thinning, lower muscle mass and higher collagen content in the bladders of old vs young rats. CONCLUSIONS: Physiological aging alters bladder function in male rats even when external factors remain constant. Thus, in old rats bladder capacity, post-void residual urine and spontaneous activity are higher, and responses to muscarinic receptor stimulation and electrical field stimulation are lower than in young rats. Such changes correspond to findings in aging human bladders, supporting the view that the Fischer/Brown Norway rat is a useful model in which to study age related bladder function ! changes. PMID: 20488483 [PubMed - as supplied by publisher] | | |
| Sirt1 plays an important role in mediating greater functionality of human ES/iPS-derived vascular endothelial cells.
May 22, 2010 at 6:37 AM
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| | Sirt1 plays an important role in mediating greater functionality of human ES/iPS-derived vascular endothelial cells. Atherosclerosis. 2010 Apr 22; Authors: Homma K, Sone M, Taura D, Yamahara K, Suzuki Y, Takahashi K, Sonoyama T, Inuzuka M, Fukunaga Y, Tamura N, Itoh H, Yamanaka S, Nakao K OBJECTIVE: We previously succeeded in inducing and isolating vascular endothelial cells (ECs) from both human embryonic stem (ES) and induced pluripotent stem (iPS) cells. Here, we compared the functionality of human adult ECs (HAECs), human ES-derived ECs (ESECs) and human iPS-derived ECs (iPSECs). METHODS AND RESULTS: We compared the cell proliferative potential, potential for migration, and tolerance to oxidative stress. ESECs were significantly superior to HAECs in all of these cell functions. The cell functions of iPSECs were comparable to those of ESECSs and also superior to HAECs. We then analyzed the gene expressions of HAECs, ESECs and iPSECs, and observed that the expression level of Sirt1, a nicotinamide adenine dinucleotide (NAD+)-dependent histone deacetylase, is higher in ESECs and iPSECs than in HAECs. The inhibition of Sirt1 with a Sirt1-specific inhibitor and siRNA antagonized these differences between the three types of cells. CONCLUSIONS: Sirt1 ! plays a key role in the high cellular function of ESECs and iPSECs. Although further in vivo investigations are required, this study initially demonstrated the potential of ESECs and iPSECs as the cell source for regenerative medicine, and also showed the potential of ES cells as a useful tool for elucidating the molecular mechanism of cell aging. PMID: 20488443 [PubMed - as supplied by publisher] | | |
| Rationale and design of the JUVENTAS trial for repeated intra-arterial infusion of autologous bone marrow-derived mononuclear cells in patients with critical limb ischemia.
May 22, 2010 at 6:37 AM
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| | Rationale and design of the JUVENTAS trial for repeated intra-arterial infusion of autologous bone marrow-derived mononuclear cells in patients with critical limb ischemia. J Vasc Surg. 2010 Jun;51(6):1564-1568 Authors: Sprengers RW, Moll FL, Teraa M, Verhaar MC, Critical limb ischemia (CLI) continues to form a substantial burden on Western healthcare. Many patients still face amputation as a last treatment option. Autologous bone marrow (BM)-derived cell administration has emerged as a potential new treatment, but proof for sustainable clinical effects of BM-derived cell therapy in CLI is still lacking. The JUVENTAS (reJUVenating ENdothelial progenitor cells via Transcutaneous intra-Arterial Supplementation) trial is the first randomized, placebo-controlled, double-blinded clinical trial on repeated intra-arterial BM mononuclear cell (MNC) infusion in 110 to 160 CLI patients, designed to provide definite proof for the efficacy of stem cell therapy. Primary outcome is the incidence of major amputation at 6 months. Inclusion of patients is well underway. If BM-MNC cells therapy is beneficial, it could become a novel treatment to prevent amputation in patients with CLI. PMID: 20488328 [PubMed - as supplied by publisher] | | |
| Corpus Cavernosal Smooth Muscle Relaxation Effect of a Novel AMPK Activator, Beta-Lapachone.
May 22, 2010 at 6:37 AM
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| | Corpus Cavernosal Smooth Muscle Relaxation Effect of a Novel AMPK Activator, Beta-Lapachone. J Sex Med. 2010 May 11; Authors: Bae JH, Kim JW, Kweon GR, Park MG, Jeong KH, Kim JJ, Moon DG ABSTRACT Introduction. Adenosine monophosphate-activated protein kinase (AMPK) activation is suggested to relax smooth muscle by endothelial nitric oxide synthase (eNOS) phosphorylation. Aim. To assess the mechanism and effect of a novel AMPK activator, beta-lapachone, upon cavernosal smooth muscle relaxation and the therapeutic potential for erectile dysfunction. Methods. Human umbilical vein endothelial cells (HUVECs) were treated with beta-lapachone. The lysates were blotted with specific antibodies for phosphorylated AMPK (p-AMPK) or phosphorylated eNOS (p-eNOS). The membranes were re-blotted for total AMP total eNOS, or beta-actin. The eNOS activity was measured by the conversion of L-14C-arginine to L-14C-citrulline in HUVECs lysates. In a separated experiment, cavernosal strips from New Zealand white rabbits were harvested for organ bath study and the relaxation effect of beta-lapachone on phenylephrine-induced contracted strips was evaluated and compared w! ith sodium nitroprusside, zaprinast, metformin, and aminoimidazole carboxamide ribonucleotide (AICAR). Methylene blue and L-NAME were used to assess the inhibition of cyclic guanosine monophosphate/nitric oxide pathway. Zinc-protoporphyrin-IX (ZnPP) was also used to investigate the contribution of mevalonate pathway. Main Outcome Measures. The expression of p-AMPK, p-eNOS, AMPK and eNOS induced by beta-lapachone in HUVECs study and the percent relaxation of cavernosal tissue in organ bath study. Results. Beta-lapachone clearly induced AMPK phosphorylation and, as a consequence, eNOS phosphorylation in HUVECs. Beta-lapachone-induced upregulation of eNOS activity was also observed in HUVECs and steadily increased up to 1 hour. In organ bath study, beta-lapachone significantly relaxed the phenylephrine pretreated strips in a dose-dependent manner. This relaxation effect was not totally blocked by methylene blue or L-NAME. After removing endothelium, the relaxation was totally ! blocked by ZnPP. Conclusions. A novel AMPK activator, beta-lap! achone h as a strong relaxation effect on precontracted cavernosal smooth muscle strips in the rabbit. And phosphorylation of AMPK and eNOS strongly related to the action of beta-lapachone. Mevalonate pathway also might be considered as a suggestive mechanism. Bae JH, Kim JW, Kweon GR, Park MG, Jeong K-H, Kim JJ, and Moon DG. Corpus cavernosal smooth muscle relaxation effect of a novel AMPK activator, beta-lapachone. J Sex Med **;**:**-**. PMID: 20487243 [PubMed - as supplied by publisher] | | |
| Regeneration of Dental Pulp-Like Tissue by Chemotaxis-induced Cell Homing.
May 22, 2010 at 6:37 AM
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| | Regeneration of Dental Pulp-Like Tissue by Chemotaxis-induced Cell Homing. Tissue Eng Part A. 2010 May 20; Authors: Kim J, Xin X, Moioli EK, Chung J, Lee CH, Chen M, Fu SY, Koch PD, Mao J Tooth infections or injuries involving dental pulp are treated routinely by root canal therapy. Endodontically treated teeth are devitalized, susceptible to re-infections, fractures and subsequent tooth loss. Here, we report regeneration of dental pulp-like tissue by cell homing and without cell transplantation. Upon in vivo implantation of endodontically treated real-size human teeth in mouse dorsum for the tested 3 wks, delivery of basic fibroblast growth factor and/or vascular endothelial growth factor (bFGF and/or VEGF) yielded re-cellularized and revascularized connective tissue that integrated to native dentinal wall in root canals. Furthermore, combined delivery of bFGF, VEGF or platelet derived growth factor (PDGF) with a basal set of nerve growth factor (NGF) and bone morphogenetic protein-7 (BMP7) generated cellularized and vascularized tissues positive of VEGF antibody staining and apparent neo-dentin formation over the surface of native dentinal wall i! n some, but not all, endodontically treated teeth. Newly formed dental-pulp tissue appeared dense with disconnected cells surrounded by extracellular matrix. Erythrocyte-filled blood vessels were present with endothelial-like cell lining. Reconstructed, multiple microscopic images showed complete fill of dental pulp-like tissue in the entire root canal from root apex to pulp chamber with tissue integration to dentinal wall upon delivery of bFGF, VEGF or PDGF with a basal set of NGF and BMP-7. Quantitative ELISA showed that combinatory delivery of bFGF, VEGF or PDGF with basal NGF and BMP7 elaborated von Willerbrand factor, dentin sialoprotein (DSP) and NGF. These findings represent the first demonstration of regenerated dental pulp-like tissue in endodontically treated root canals of real-size human teeth. The present chemotaxis-based approach has potent cell homing effects for re-cellularization and revascularization in endodontically treated root canals in vivo, albeit in! an ectopic model. Regeneration of dental pulp by cell homing,! rather than cell delivery, may accelerate clinical translation. PMID: 20486799 [PubMed - as supplied by publisher] | | |
| Biodegradable, photocrosslinked alginate hydrogels with independently tailorable physical properties and cell adhesivity.
May 22, 2010 at 6:37 AM
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| | Biodegradable, photocrosslinked alginate hydrogels with independently tailorable physical properties and cell adhesivity. Tissue Eng Part A. 2010 May 20; Authors: Jeon O, Powell C, Ahmed SM, Alsberg E Biocompatible polymers capable of photopolymerization are of immense interest for tissue engineering applications as they can be injected in a minimally invasive manner into a defect site, and then upon application of ultraviolet light, rapidly form hydrogels in situ. Cell adhesion interactions with a biomaterial are known to be important in regulating cell behaviors such as proliferation and differentiation. Therefore, we have covalently modified photocrosslinkable alginate with cell adhesion ligands containing the Arg-Gly-Asp (RGD) amino acid sequence to form biodegradable, photocrosslinked alginate hydrogels with controlled cell adhesivity. This unique polymer system allows for independent modulation of the physical and biochemical signaling environment presented to cells. The physical properties of the hydrogels such as elastic moduli, swelling ratios, and degradation profiles were similar at the same crosslinking density regardless of the presence of adhesion! ligands. Chondrocytes seeded on the surface of the adhesion ligand-modified hydrogels were able to attach and spread, whereas those seeded on unmodified hydrogels exhibited minimal adherence. Importantly, the adhesion ligand-modified hydrogels enhanced the proliferation and chondrogenic differentiated function of encapsulated chondrocytes as demonstrated by increased DNA content and production of glycosaminoglycans compared to unmodified control hydrogels. This new photocrosslinkable, biodegradable biomaterial system in which the soluble (e.g., growth factors) and insoluble (e.g., cell adhesion signals) biochemical signaling environment and the biomaterial physical properties (e.g., the elastic moduli) can be independently controlled may be a powerful tool for elucidating the individual and combined effects of these parameters on cell function for cartilage tissue engineering and other regenerative medicine applications. Keywords; Biomaterial, Drug delivery, Hydrogels, Tis! sue engineering, Chondrocytes. PMID: 20486798 [PubMed - as supplied by publisher] | | |
| In vivo evaluation of polysialic acid as part of tissue engineered nerve transplants.
May 22, 2010 at 6:37 AM
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| | In vivo evaluation of polysialic acid as part of tissue engineered nerve transplants. Tissue Eng Part A. 2010 May 20; Authors: Haastert K, Schaper-Rinkel J, Schmitte R, Rode B, Mühlenhoff M, Schwarzer D, Dräger G, Su Y, Scheper T, Gerardy-Schahn R, Grothe C With the aim to develop new biomaterials for peripheral nerve grafts, the current study used bioidentical polysialic acid as complement in synthetic conduits. Polysialic acid provides an important guidance cue during nervous system development and regeneration. First in vivo results on the use of cell-free and Schwann cell-containing synthetic peripheral nerve grafts complemented with soluble exogenous polysialic acid (K1-polySia) are presented. Reconstructing 10 mm rat sciatic nerve gaps, K1-polySia complementation significantly improved structural nerve regeneration in comparison to cell-free and K1-polySia-free grafts. Subsequently, long nerve gaps (13mm) were reconstructed by Schwann cell transplants plus K1-polySia and compared to nerve autotransplantation. Structural but also functional regeneration could be observed using K1-polySia-transplants, however, autotransplantation was still significantly more successful. Overall the current study demonstrates that! exogenous K1 polySia has no negative but rather regeneration promoting effects. This is important novel data on the applicability of exogenous polysialic acid in vivo. Further studies are required to develop solid 3D polysialic acid based scaffolds for nerve tissue engineering. Biocompatible and assessable biodegrading materials will ensure long lasting presence of polysialic acid to allow its applicability and prolonged efficacy in the slow regenerating scenario of human peripheral nerve reconstruction. PMID: 20486797 [PubMed - as supplied by publisher] | | |
| Restoration of the transepithelial potential within tissue-engineered human skin in vitro and during the wound healing process in vivo.
May 22, 2010 at 6:37 AM
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| | Restoration of the transepithelial potential within tissue-engineered human skin in vitro and during the wound healing process in vivo. Tissue Eng Part A. 2010 May 20; Authors: Dube J, Rochette O, Levesque P, Gauvin R, Roberge C, Auger FA, Goulet D, Bourdages M, Plante M, Germain L, Moulin VJ Normal human epidermis possesses a trans-epithelial potential (TEP) that varies in different parts of the body (10 to 60 mV). The role of TEP in normal epidermis is not yet identified but after skin injury, TEP disruption induces an endogenous direct current electric field (100 to 200 mV/mm) directed toward the middle of the wound. This endogenous electric field could be implicated in the wound healing process by attracting cells thus facilitating reepithelialization. However, little is known on the restoration of the TEP during human skin formation and wound healing. In the present study, the variations in TEP and Na+/K+ ATPase pump expression during the formation of the epithelium were investigated in vitro using human tissue-engineered skin (TES) reconstituted by tissue engineering and in vivo with a porcine wound healing model. Results showed that TEP undergoes ascending and decreasing phases during epithelium formation in TES as well as during wound repair wi! thin TES. Similar results were observed during in vivo reepithelialization of wounds. The ascending and decreasing TEP values were correlated with changes in the expression of Na+/K+ ATPase pump. The distribution of Na+/K+ ATPase pumps also varied according to epidermal differentiation. Taken together, these results suggest that the variations in the expression of Na+/K+ ATPase pump over time and across epidermis would be a determinant parameter of the TEP, dictating a cationic transport during the formation and the restoration of the epidermis. Therefore, this study brings a new perspective to understand the formation and restoration of TEP during the cutaneous wound healing process. This might have important future medical applications regarding the treatment of chronic wound healing. PMID: 20486795 [PubMed - as supplied by publisher] | | |
| Repair of full-thickness tendon injury using connective tissue progenitors efficiently derived from human embryonic stem cells and fetal tissues.
May 22, 2010 at 6:37 AM
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| | Repair of full-thickness tendon injury using connective tissue progenitors efficiently derived from human embryonic stem cells and fetal tissues. Tissue Eng Part A. 2010 May 20; Authors: Cohen S, Leschansky L, Zussman E, Burman M, Srouji S, Abramov N, Itskovitz-Eldor J The use of stem cells for tissue engineering (TE) encourages scientists to design new platforms in the field of regenerative and reconstructive medicine. Human embryonic stem cells (hESCs) have been proposed to be an important cell source for cell-based TE applications as well as an exciting tool for investigating the fundamentals of human development. Here, we describe the efficient derivation of connective tissue progenitors (CTPs) from hESC lines and fetal tissues. CTPs were significantly expanded and induced to generate tendon tissues in-vitro, with ultrastructural characteristics and biomechanical properties typical of mature tendons. We describe a simple method for engineering tendon grafts which can successfully repair injured Achilles tendons and restore the ankle joint extension movement in mice. We also show the CTP's ability to differentiate into bone, cartilage and fat both in vitro and in vivo. This study provides an evidence for the possibility of us! ing stem cell-derived engineered grafts to replace missing tissues, and sets a basic platform for future cell-based tissue engineering applications in the fields of orthopedics and reconstructive surgery. PMID: 20486794 [PubMed - as supplied by publisher] | | |
| Reduced thrombocyte adhesion to endothelialized poly 4-methyl-1-pentene (PMP) gas exchange membranes - A first step towards bioartificial lung development.
May 22, 2010 at 6:37 AM
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| | Reduced thrombocyte adhesion to endothelialized poly 4-methyl-1-pentene (PMP) gas exchange membranes - A first step towards bioartificial lung development. Tissue Eng Part A. 2010 May 20; Authors: Hess C, Wiegmann B, Maurer AN, Fischer P, Moeller L, Martin U, Hilfiker A, Haverich A, Fischer S Polymeric materials used in biomedical devices, bioartificial organs or for the fabrication of tissue engineering scaffolds should completely prevent the activation of the coagulation system and subsequent clot formation. Surface endothelialization is considered an important tool to optimize the blood compatibility of synthetic materials as a functional endothelial cell layer on an artificial material may help control hemostasis and therefore provide a solution to improve the biocompatibility of these materials. Here we report on the endothelialization of poly 4-methyl-1-pentene (PMP) gas exchange membranes using human cord blood derived late outgrowth endothelial cells (CB-ECFCs). We achieved complete endothelialization of PMP membranes and when seeded and cultivated on the membrane, CB-ECFCs maintained both endothelial characteristics and functionality. Endothelialization resulted in significantly lower platelet adhesion and activation compared to unseeded membr! anes. Of importance, the endothelial layer had no mayor impact on gas permeability of PMP membranes. This study is a first promising step towards the development of a biofunctionalized surface for the use in gas exchange devices with blood contacting surfaces and a straightforward approach towards a long-term bio-hybrid lung replacement system. PMID: 20486793 [PubMed - as supplied by publisher] | | |
| Interconnected macroporous poly(ethylene glycol) (PEG) cryogels as a cell scaffold for cartilage tissue engineering.
May 22, 2010 at 6:37 AM
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| | Interconnected macroporous poly(ethylene glycol) (PEG) cryogels as a cell scaffold for cartilage tissue engineering. Tissue Eng Part A. 2010 May 20; Authors: Hwang Y, Sangaj N, Varghese S Macroporous networks of poly(ethylene glycol) (PEG) with interconnected pores can be created by cryogelation techniques. In this study, we describe the potential application of such PEG cryogels as scaffolds for cartilage tissue engineering. Three dimensional macroporous cryogels were evaluated for chondrocyte growth and production of cartilage-specific extracellular matrix (ECM). Seeded primary bovine chondrocytes showed homogeneous distribution throughout the cryogels. DNA content suggests continuous cell proliferation over 4 weeks of in vitro culture. Analysis of the composition of cell-secreted ECM showed a culture time dependent increase in the amount of glycosaminoglycan (GAG) and collagen. The production of ECM by chondrocytes was confirmed using SEM analysis. Further histological and immunohistological analysis of the cell-laden scaffold confirmed the presence of accumulated cartilage-specific ECM within the scaffold. The interconnected macroporous network! promoted diffusion of cell secreted matrix within the cryogels. Our results indicated that interconnected macroporous PEG cryogels successfully supported attachment, viability, proliferation, and biosynthetic activity of seeded chondrocytes. PMID: 20486791 [PubMed - as supplied by publisher] | | |
| Modulation of MMP-2 levels by mechanical loading of three-dimensional mesenchymal stem cell constructs: impact on in vitro tube formation.
May 22, 2010 at 6:37 AM
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| | Modulation of MMP-2 levels by mechanical loading of three-dimensional mesenchymal stem cell constructs: impact on in vitro tube formation. Tissue Eng Part A. 2010 May 20; Authors: Glaeser JD, Geissler S, Ode A, Schipp CJ, Matziolis G, Taylor WR, Knaus P, Perka C, Duda G, Kasper G Angiogenesis is essential to tissue reconstitution, is sensitive to mechanical stresses and currently represents one of the major challenges in tissue engineering. The pro-angiogenic matrix metalloprotease-2 (MMP-2) is up-regulated in mechanically loaded mesenchymal stem cells (MSCs). Therefore, MMP-2 may provide a regulating link between angiogenesis and the surrounding mechanical conditions. This study aimed to modulate MMP-2 levels by mechanical loading of MSCs embedded in a three-dimensional matrix as well as to investigate the mechanism of MMP-2 regulation along with its contribution to angiogenesis stimulation. MMP-2-inducing conditions (30% compression, 1Hz, 72h) were defined after varying loading parameters. Addition of the Golgi-disturbing agent Brefeldin A suppressed this mechanical up-regulation of MMP-2. Analysis of enzymatic activities demonstrated an enhancement of pro-MMP-2, mature MMP-2 and tissue inhibitor of metalloproteases-2 (TIMP-2). Furthermo! re, mechano-regulation of MMP-14 and mature MMP-2 was dependent upon the activity of furin, a proprotein processing endoprotease. Angiogenesis was stimulated by conditioned media from MSCs loaded at inducing conditions. This augmentation of angiogenesis was hindered by inhibition of pro-MMP-2 and mature MMP-2. In conclusion, mechanical stimulation of MSCs in a 3D matrix induces pro-MMP-2 secretion and MMP-2 activation, potentially via the activation complex consisting of MMP-2/-14/TIMP-2. Mechano-regulated pro- and mature MMP-2 seem to contribute to angiogenesis stimulation. Thus, an application of these loading parameters could augment vascularization of tissue engineered constructs based on the described MMP-2-dependent mechanism. PMID: 20486790 [PubMed - as supplied by publisher] | | |
| Development of functional fibrous matrices for the controlled release of basic fibroblast growth factor (bFGF) to improve therapeutic angiogenesis.
May 22, 2010 at 6:37 AM
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| | Development of functional fibrous matrices for the controlled release of basic fibroblast growth factor (bFGF) to improve therapeutic angiogenesis. Tissue Eng Part A. 2010 May 20; Authors: Kim MS, Bhang SH, Yang HS, Rim NG, Kim SI, Kim BS, Shin H In this study, novel fibrous matrices were developed as a depot to store and liberate growth factors in a controlled manner. Specifically, heparin was covalently conjugated onto the surface of fibrous matrices (composites of polycaprolactone and gelatin crosslinked with genipin), and bFGF was then reversibly immobilized. The immobilization of bFGF was controlled as a function of the amount of conjugated heparin. The sustained release of bFGF from the fibrous matrices was successfully achieved over 4 weeks while physical adsorption of bFGF released quickly. The bFGF released from the fibrous matrices significantly enhanced in vitro proliferation of human umbilical vein endothelial cells (HUVECs). From the in vivo study, the group implanted with a higher amount of immobilized bFGF significantly facilitated neo-blood vessel formation as compared to other implantation groups. These results indicate that the sustained release of bFGF is important for the formation of b! lood vessels, and that our fibrous matrices could be useful for regulation of tissue damage requiring angiogenesis. Furthermore, our system can be combined with other growth factors with heparin binding domains, representing a facile depot for spatio-temporal control over the delivery of bioactive molecules in regenerative medicine. PMID: 20486788 [PubMed - as supplied by publisher] | | |
| The interactions between rASCs, rhBMP-2 and beta-TCP play an important role in bone tissue engineering.
May 22, 2010 at 6:37 AM
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| | The interactions between rASCs, rhBMP-2 and beta-TCP play an important role in bone tissue engineering. Tissue Eng Part A. 2010 May 20; Authors: E LL, Xu L, Wu X, Wang D, Lv Y, Wang J, Liu H ABSTRACT Cells, scaffolds, and growth factors are the three main factors for creating a stem cell-based tissue engineered construct, but the interactions between three factors are not very clear. We hereby explored the interactions between rat adipose-derived stromal cells (rASCs), bone morphogenetic protein-2 (rhBMP-2) and beta-tricalcium phosphate (ss-TCP) to provide evidence for their application in bone tissue engineering by evaluating the protein adsorption of ss-TCP, the cell attachment, alkaline phosphatase (ALP) activity/protein, osteocalcin content, mineral formation, calcium content, phosphonium content, cell vitality, gene expression and implantation in the backs of severe combined immunodeficient (SCID) mice of rhBMP-2 pre-inducing rASCs seeded onto ss-TCP. The attachment, proliferation and osteogenic properties of rASCs were supported by ss-TCP using scanning electron microscope (SEM); Compared with rASCs cultured on the culture plate, rASCs cultlured! on ss-TCP had significantly higher ALP activity/protein, osteocalcin content and mineral formation, the values for rASCs cultured on ss-TCP with rhBMP-2 increased most significantly; The rhBMP-2 significantly increased calcium content, phosphonium content and ALP, I type collagen, osteocalcin mRNAs levels of rASCs cultured on ss-TCP; After 8 and 12 weeks of implantation, each group displayed increased bone formation over the 12-week time period. The value for rASCs/ss-TCP construct was significantly higher than that for ss-TCP group. But the maximal and robust bone formation was presented in rASCs/ss-TCP with rhBMP-2. The results implied that stem cells existed in adult rat adipose tissue. ss-TCP could adsorb rhBMP-2 from the media and had osteoinductivity when alone implanted in the back of SCID mice. ss-TCP was also sufficient to trigger the differentiation of rASCs toward an osteoblastic phenotype without the addition of osteogenic factor. The rhBMP-2 could better suffi! ciently induce osteogenic differentiation of rASCs seeded onto! ss-TCP. The rASCs and rhBMP-2 could promote the dissolution of ss-TCP to provide Ca2+, PO3-4 needed by bone formation. The interactions between three factors could provide an optimizing microenvironment for osteogenic differentiation of rASCs, this might be essential for sufficient and timely bone formation in vivo. This study may provide insight into the clinical repair of bone defect with ASCs+ss-TCP+rhBMP-2 construct. PMID: 20486786 [PubMed - as supplied by publisher] | | |
| Comparative characterization of porcine mesenchymal stem cells derived from bone marrow extract and skin tissues.
May 22, 2010 at 6:37 AM
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| | Comparative characterization of porcine mesenchymal stem cells derived from bone marrow extract and skin tissues. Tissue Eng Part C Methods. 2010 May 20; Authors: Ock SA, Jeon BG, Rho GJ Mesenchymal stem cells (MSCs) offer a great promise for regenerative medicine. Present study compared the characterization of porcine MSCs (pMSCs) derived from bone marrow extract with adult ear and fetal skin derived cells on morphology, cell growth, alkaline phosphatase (AP) activity, proliferation ability, expression of cell surface (CD) markers (CD 29, 45 and 90), cell cycle, protein and mRNA levels of Oct-4, Sox-2 and Nanog and lineage differentiation ability. Skin derived cells exhibited AP activity and differentiation ability like pMSCs. pMSCs possessed a longer doubling time than skin derived cells and there was no difference in the ratio of G0/G1 phase between pMSCs and skin derived cells. Except for CD 29 and 90, all cells were found negative for CD 45. Protein and mRNA expression of Oct-4, Sox-2 and Nanog were observed with similar intensity in all cells. Taken together, pMSCs and skin derived cells revealed similar characteristics, and suggested the po! ssible supportive role of skin derived cells with MSCs for the regeneration of damaged tissues in cell based therapies. PMID: 20486783 [PubMed - as supplied by publisher] | | |
| Non expanded mesenchymal stem cells for regenerative medicine: yield in stromal vascular fraction from adipose tissues.
May 22, 2010 at 6:37 AM
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| | Non expanded mesenchymal stem cells for regenerative medicine: yield in stromal vascular fraction from adipose tissues. Tissue Eng Part C Methods. 2010 May 20; Authors: Faustini M, Bucco M, Chlapanidas T, Lucconi G, Marazzi M, Tosca M, Gaetani P, Klinger M, Villani S, Ferretti VV, Vigo D, Torre ML The adipose-derived stromal vascular fraction (SVF) represents a rich source of mesenchymal cells, potentially able to differentiate into adipocytes, chondrocytes, osteoblasts, myocytes, cardiomyocytes, hepatocytes, and neuronal, epithelial and endothelial cells. These cells are ideal candidates for use in regenerative medicine, tissue engineering, including gene therapyand cell replacement cancer therapies. In this work, aiming to the optimization of the adipose stem cell-based therapy, the effect of the collection site, surgical procedure and tissue processing techniques on SVF yield was evaluated in terms of cell recovery and live cells, taking into account the effect of gender, age and BMI. Adipose tissue samples were recovered from 125 informed subjects (37 males and 88 females, mean age: 51.31 y, range: 15-87 y), and digested in different condition with collagenase. A multivariate linear model put in evidence that the gender of the donor should be taken into! account, as well as the collection site and the tissue digestion conditions; the collection technique, the age and the body mass index of donor seem not to influence the cell yield. PMID: 20486782 [PubMed - as supplied by publisher] | | |
| Tissue Engineering by Molecular Disassembly and Reassembly: Biomimetic Retention of Mechanically Functional Aggrecan in Hydrogel.
May 22, 2010 at 6:37 AM
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| | Tissue Engineering by Molecular Disassembly and Reassembly: Biomimetic Retention of Mechanically Functional Aggrecan in Hydrogel. Tissue Eng Part C Methods. 2010 May 20; Authors: Han E, Wilensky LM, Schumacher BL, Chen AC, Masuda K, Sah RL In vitro assembly of key functional extracellular matrix constituents for tissue-engineered constructs may provide a tool to modulate the retention of proteoglycan (PG) aggregates, which are crucial to compressive biomechanical properties of connective tissues. This study tested the hypotheses that (1) biomimetic molecular reassembly of PG aggregates (native aggrecan (AGC) with hyaluronan (HA) +/- link proteins (LP)) affects AGC retention kinetics in hydrogel constructs, (2) the compressive properties of such hydrogel constructs are related to the content of retained AGC, and (3) the reassembly method is compatible with chondrocytes. Addition of HA to AGC in hydrogel constructs increased AGC retention in a dose-dependent manner, and the addition of LP to AGC+HA further enhanced AGC retention. The level of AGC retention, in turn, was associated with increased equilibrium compressive stress of the constructs. Chondrocytes could be included in the process, and mainta! ined expression of the chondrogenic phenotype, secreting type II collagen but little type I collagen. Thus, by altering the assembly of PG aggregates with HA +/- LP, which affects AGC retention, it may be possible to achieve the targeted levels of PG components in order to modulate the mechanical properties of the engineered construct for cartilage as well as other tissues containing PG and PG aggregates. PMID: 20486781 [PubMed - as supplied by publisher] | | |
| Telomerase reverse transcriptase (TERT) Related with Telomerase Activity Regulates Tumorigenic Potential of Mouse Embryonic Stem Cells.
May 22, 2010 at 6:37 AM
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| | Telomerase reverse transcriptase (TERT) Related with Telomerase Activity Regulates Tumorigenic Potential of Mouse Embryonic Stem Cells. Stem Cells Dev. 2010 May 20; Authors: Park KD, Seong SK, Park YM, Choi Y, Park JH, Lee SH, Baek DH, Kang JW, Choi KS, Park SN, Kim DS, Kim SH, Kim HS Embryonic stem cell (ESC) research gave rise to the possibility that stem cell therapy could be used in the treatment of incurable diseases such as neurodegenerative disorders. However, problems related to the tumorigenicity of undifferentiated ESCs must be resolved before such cells can be used in the application of cell replacement therapies. In the present study, we attempted to determine biomarkers that predicted tumor formation of undifferentiated ESCs in vivo. We differentiated mouse ESCs (R1 cell line) into neural lineage using a five-step method, and evaluated the expression of oncogenes (p53, Bax, c-myc, Bcl2, K-ras), telomerase related genes (TERT, TRF), as well as telomerase activity and telomere length during differentiation of ESCs. The expression of oncogenes did not show a significant change during differentiation steps but the expression of tert and telomerase activity correlated with mESCs differentiation. To investigate the possibility of mTERT a! s biomarker of tumorigenicity of undifferentiated ESCs, we established mTERT knockdown ESCs using the shRNA lentivirus vector and evaluated its tumorigenicity in vivo using nude mice. Tumor volumes significantly decreased and appearances of tumor formation in mice were delayed in the TERT-knockdown ESC treated group compared to undifferentiated ESCs treated group. Altogether, these results suggested that mTERT might be potentially beneficial as a biomarker, rather than oncogenes of somatic cells, for the assessment of ESCs tumorigenicity. PMID: 20486780 [PubMed - as supplied by publisher] | | |
| MicroRNA hsa-miR-138 inhibits adipogenic differentiation of human adipose tissue-derived mesenchymal stem cells through EID-1.
May 22, 2010 at 6:37 AM
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| | MicroRNA hsa-miR-138 inhibits adipogenic differentiation of human adipose tissue-derived mesenchymal stem cells through EID-1. Stem Cells Dev. 2010 May 20; Authors: Yang Z, Bian C, Zhou H, Huang S, Wang S, Liao L, Zhao RC A better understanding of the molecular mechanisms underlying the differentiation of human adipose tissue-derived mesenchymal stem cells (hAD-MSCs) could provide new insights into the pathogenesis of a number of diseases, such as obesity and diabetes, and broaden the spectrum of potential hAD-MSCs-based cell therapy. In this study, we reported that a microRNA, miR-138, could inhibit the adipogenic differentiation of hAD-MSCs. Our results showed that miR-138 was significantly down-regulated during adipogenic differentiation. Overexpression of miR-138 in hAD-MSCs could effectively reduce lipid droplets accumulation, inhibit expression of key adipogenic transcription factors CCAAT enhancer binding protein alpha (C/EBPalpha) and peroxisome proliferator-activated receptor gamma 2 (PPARgamma2) as well as several other adipogenic marker genes, such as fatty acid binding protein 4 (FABP4) and lipoprotein lipase (LPL). Further studies showed that the expression of E1A-like! inhibitor of differentiation 1 (EID-1), a nuclear receptor coregulator, was inversely correlated with that of miR-138 when hAD-MSCs were differentiated into adipocytes. Knockdown of EID-1 by RNA interference (RNAi) inhibited adipocyte differentiation of hAD-MSCs. In addition, luciferase reporter assays demonstrated that miR-138 directly targeted the 3'UTR region of EID-1, implying that the negative role of miR-138 in the adipocyte differentiation of hAD-MSCs is at least partially mediated via repressing EID-1. Taken together, this study shows that miR-138 plays a negative role in adipogenic differentiation and sheds light on the role of miRNAs during differentiation of hAD-MSCs toward adipocytes. PMID: 20486779 [PubMed - as supplied by publisher] | | |
| Combined Omics Analysis Identifies Transmembrane 4 L6 family member 1 as a Surface Protein Marker Specific to Human Mesenchymal Stem Cells.
May 22, 2010 at 6:37 AM
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| | Combined Omics Analysis Identifies Transmembrane 4 L6 family member 1 as a Surface Protein Marker Specific to Human Mesenchymal Stem Cells. Stem Cells Dev. 2010 May 20; Authors: Bae S, Shim SH, Park CW, Son HK, Lee HJ, Son JY, Jeon C, Kim H Mesenchymal stem cells (MSCs) are promising for cell therapy and regenerative medicine, but their lack of specific markers renders the cell culture at potential contamination risk with other cell types, in particular, fibroblasts. In this study, we mapped two differential transcriptome data of MSCs compared, one to mononuclear cells and the other to fibroblasts, onto the membrane proteome data, the analysis of which led to an identification of transmembrane 4 L6 family member 1 (TM4SF1) as a surface protein marker candidate that could discriminate MSCs simultaneously from blood cells and fibroblasts. Our analyses confirmed that TM4SF1 was abundantly expressed on MSCs, but neither on other blood/tissue cells nor on fibroblasts. TM4SF1 immunoselection from bone marrow and adipose tissues yielded homogeneous cell populations that were highly similar to MSCs, in terms of morphology, immunophenotype, and differentiation potential. These findings indicate that TM4SF1 ca! n serve as a surface protein marker that singly identifies MSCs from diverse cell sources, in particular, fibroblast-rich connective tissues. PMID: 20486778 [PubMed - as supplied by publisher] | | |
| Human Bone-Marrow And Adipose Tissue Mesenchymal Stem Cells: A User's Guide.
May 22, 2010 at 6:37 AM
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| | Human Bone-Marrow And Adipose Tissue Mesenchymal Stem Cells: A User's Guide. Stem Cells Dev. 2010 May 20; Authors: Mosna F, Sensebé L, Krampera M Mesenchymal Stem Cells (MSCs) are adult stem cells which hold great promise in the field of Regenerative Medicine. They can be isolated from almost any tissue of the body and display, after expansion, very similar properties and minor differences, probably due to their microenvironment of origin. Expansion in vitro can be obtained in cytokine-free, serum-enriched media, as well as in serum-free, basic Fibroblast Growth Factor-enriched media. A detailed immunophenotypic analysis is required to test the purity of the preparation, but no unique distinguishing marker has been described as yet. Functional assays, i.e. differentiation studies in vitro, are needed to prove multilineage differentiation of expanded cells, and demonstration of pluripotency is necessary to identify most immature precursors. MSCs show powerful immunomodulative properties towards most of the cells of the immune system: this strengthens the theoretical rationale for their use also in an allogen! eic setting across the MHC immunological barriers. Systemic intravenous injection, as well as local use have been tried: following systemic injection, MSCs show a high degree of chemotaxis based on pro-inflammatory cytokines, and localize at inflamed and neoplastic tissues; local regeneration has been improved using synthetic, as well as organic scaffolds. On the other hand, inadequate heterotopic in vivo differentiation and neoplastic transformation are potential risks of this form of cell therapy, even if evidence of this sort have been collected only from studies in mice, and generally after prolonged in vitro expansion. This review tries to provide a detailed technical overview of the methods used for human bone marrow-derived and adipose tissue-derived MSCs isolation, in vitro expansion and characterization for tissue repair. We chose to use BM-MSCs as a model to describe techniques that have been used for MSCs isolation and expansion from very different sources, and A! T-MSCs as an example of a reliable and increasingly common alt! ernative source. PMID: 20486777 [PubMed - as supplied by publisher] | | |
| Effect of losartan and benzbromarone on the level of human urate transporter 1 mRNA.
May 22, 2010 at 6:37 AM
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| | Effect of losartan and benzbromarone on the level of human urate transporter 1 mRNA. Arzneimittelforschung. 2010;60(4):186-8 Authors: Nindita Y, Hamada T, Bahrudin U, Hosoyamada M, Ichida K, Iwai C, Urashima S, Kuwabara N, Utami SB, Mizuta E, Yamada K, Igawa O, Shigemasa C, Ninomiya H, Tsuchihashi T, Hisatome I Both an angiotensin II receptor blocker, losartan (CAS 124750-99-8) and a serum urate lowering agent, benzbromarone (CAS 3562-84-3) exert a uricosuric action by inhibiting urate transporter 1 (URAT1). A recent clinical trial indicated that losartan could reduce the level of serum urate in hypertensive patients treated with urate lowering agents, suggesting the different mode of action of losartan from benzbromarone. In the present study, the effect of losartan and benzbromarone on the level of URAT1 mRNA was determined in transfected HEK293 cells. Losartan caused a significant reduction of its mRNA level, whereas it was not affected by benzbromarone. These results indicate that losartan decreases the level of human URAT1 mRNA, which may underlie the uricosuric action of losartan in hypertensive patients treated with serum urate lowering agents. PMID: 20486468 [PubMed - in process] | | |
| Adipocytes constitutively release factors that accelerate keratinocyte proliferation in vitro.
May 22, 2010 at 6:37 AM
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| | Adipocytes constitutively release factors that accelerate keratinocyte proliferation in vitro. Ann Plast Surg. 2010 Mar;64(3):327-32 Authors: Campbell CA, Cairns BA, Meyer AA, Hultman CS Keratinocytes grown directly on adipose tissue have greater proliferation rates than keratinocytes grown alone. It is unknown if factors released by adipose tissue into culture media could increase keratinocyte proliferation without requiring incorporation of adipose tissue into skin graft models, or serve as a substitute for the fibroblast feeder layer.Human keratinocytes were grown with and without NIH 3T3 fibroblast feeder layer in the following conditions (12 cultures per group) adipose tissue coculture (AT), cultures supplemented with medium from whole adipose tissue referred to as adipose-conditioned medium (ACM), and control. Proliferation was measured with a colorimetric proliferation assay and digital calculations of percent confluence over time. Culture morphology was assessed by light microscopy.ACM cultures without 3T3s, AT cultures with and without 3T3s, and 3T3 control cultures demonstrated a similarly significant keratinocyte proliferation increase ! over non-3T3 control (P < 0.05) corresponding with a 2-fold increase in percent confluence by day 7. ACM cultures with 3T3s proliferated significantly faster than all other treatment groups (P < 0.05) resulting in complete confluence by day 5. ACM cultures with and without 3T3s produced a thick keratinized layer by day 7 whereas all other cultures including AT cultures did not.Engineered tissue replacement can be accelerated and simplified by ACM without requiring the addition of adipose tissue or a fibroblast feeder layer to keratinocyte culture systems. ACM supplementation provides an additive proliferation benefit when combined with a feeder layer producing mature grafts in approximately half the time as keratinocytes alone by accelerating proliferation and increasing keratinization. PMID: 20179485 [PubMed - indexed for MEDLINE] | | | | 
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