| |  | | | | | | | | | | | | | | | |  | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | An imprinted signature helps isolate ESC-equivalent iPSCs. Cell Res. 2010 Aug 10; Authors: Lujan E, Wernig M PMID: 20697429 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Can HIV be cured with stem cell therapy? Nat Biotechnol. 2010 Aug;28(8):807-10 Authors: Deeks SG, McCune JM PMID: 20697404 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Prospects for Minocycline Neuroprotection. Arch Neurol. 2010 Aug 9; Authors: Plane JM, Shen Y, Pleasure DE, Deng W Minocycline is a clinically available antibiotic and anti-inflammatory drug that also demonstrates neuroprotective properties in a variety of experimental models of neurological diseases. There have thus far been more than 300 publications on minocycline neuroprotection including a growing number of human studies. Our objective is to critically review the biological basis and translational potential of this action of minocycline on the nervous system. PMID: 20697034 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | A model for the stretch-mediated enzymatic degradation of silk fibers. J Mech Behav Biomed Mater. 2010 Oct;3(7):538-547 Authors: Kluge JA, Thurber A, Leisk GG, Kaplan DL, Luis Dorfmann A To restore physiological function through regenerative medicine, biomaterials introduced into the body must degrade at a rate that matches new tissue formation. For effective therapies, it is essential that we understand the interaction between physiological factors, such as routine mechanical loading specific to sites of implantation, and the resultant rate of material degradation. These relationships are poorly characterized at this time. We hypothesize that mechanical forces alter the rates of remodeling of biomaterials, and this impact is modulated by the concentration of enzymes and the duration of the mechanical loads encountered in situ. To test this hypothesis we subjected silk fibroin fibers to repeated cyclic loading in the presence of enzymatic degradation (either alpha-chymotrypsin or Protease XIV) and recorded the stress-strain response. Data were collected daily for a duration of 2 weeks and compared to the control cases of stretched fibers in the presence of phosphate buffered saline or non-stretched samples in the presence of enzyme alone. We observed that incubation with proteases in the absence of mechanical loads causes a reduction of the ultimate tensile strength but no change in stiffness. However, cyclic loading caused the accumulation of residual strain and softening in the material's properties. We utilize these data to formulate a mathematical model to account for residual strain and reduction of mechanical properties during silk fiber degradation. Numerical predictions are in fair agreement with experimental data. The improved understanding of the degradation phenomenon will be significant in many clinical repair cases and may be synergistic to decrease silk's mechanical properties after in vivo implantation. PMID: 20696419 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Interview with Prof. Dr. Augustinus Bader. Rejuvenation Res. 2010 Aug 9; Authors: Editor's note: The interview series in Rejuvenation Research is a unique and, I believe, highly valuable feature of the journal, giving readers insights into the thinking and motivation of some of the most influential movers and shakers in the many disciplines-not only scientific( 1-7 ) but also political,( 8 , 9 ) sociological,( 10 ) ethical,( 11 ) and more-that impinge on the crusade to defeat aging. This issue's interview features a leading academic from the world of tissue engineering and stem cells, who has enthusiastically embraced the application of those technologies to the problem of aging. Moreover, Professor Bader is highly active in the effort to birng such groups together across both national and scientific borders, as exemplified by the recent adoption of Rejuvenation Research as the official journal of the World Federation and World Virtual Institute of Regenerative Medicine, which he leads. Such efforts constitute a powerful force in stimulating high-quality communication between the field of biomedical gerontology and the many constituencies that will be affected by progress against aging-a debate that, as I( 12-22 ) and others( 23-28 ) have noted recently, is essential if we are to develop effective interventions against aging with all possible speed. PMID: 20695819 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | de novo MYOCARDIAL REGENERATION: ADVANCES AND PITFALLS. Antioxid Redox Signal. 2010 Aug 9; Authors: Haider KH, Buccini S, Ahmed RP, Ashraf M The capability of adult tissue derived stem cells for cardiogenesis has been extensively studied in experimental animals and clinical studies for treatment of post-ischemic cardiomyopathy. The less than anticipated improvement in the heart function in most clinical studies with skeletal myoblasts and bone marrow cells has warranted a search for alternative sources of stem cells. Despite their multilineage differentiation potential, ethical issues, teratogenicity and tissue rejection are main obstacles in developing clinically feasible methods for embryonic stem (ES) cell transplantation into patients. A decade long research on ES cells has paved the way for discovery of alternative approaches for generating pluripotent stem cells. Genetic manipulation of somatic cells for pluripotency genes reprograms the cells to pluripotent status. Efforts are currently focused to make reprogramming protocols safer for clinical applications of the reprogrammed cells. We summarizes the advancements and complicating features of stem cell therapy and discuss the decade and a half long efforts made by stem cell researchers for moving the field from bench to the bedside as an adjunct therapy or as an alternative to the contemporary therapeutic modalities for routine clinical application. The review also provides a special focus on the advancements made in the field of somatic cell reprogramming. PMID: 20695792 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Analysis of the Soluble Human Tooth Proteome and Its Ability to Induce Dentin/Tooth Regeneration. Tissue Eng Part A. 2010 Aug 9; Authors: Chun SY, Lee HJ, Choi YA, Kim KM, Baek SH, Park HS, Kim JY, Ahn JM, Cho JY, Cho DW, Shin HI, Park EK While the soluble proteins of human teeth consist of various extracellular matrix (ECM) and bioactive proteins, they have not yet been characterized fully. Moreover, the role they play in tooth regeneration is not clear. Analysis of the soluble proteins in human teeth by liquid chromatography-mass spectrometry (LC-MS/MS) revealed 147 different EDTA-soluble tooth proteins (ESTPs). Of these, 29 had not been shown previously to be present in human teeth. To determine their effect on the in vitro responses of dental pulp stem cells (DPSCs), DPSCs were cultured in ESTP-coated culture plates and 3-dimensional scaffolds. The ESTPs significantly enhanced DPSC odontoblast differentiation and mineralization in vitro, but had only partial effect on bone marrow stem cells or adipose tissue stem cells. To test the effect of ESTPs on in vivo dentin and tooth formation, mouse embryonic tooth-forming primordia and xenogenic murine apical bud epithelium/human DPSC composites were treated with ESTPs prior to implantation under the renal capsule of ICR mice. ESTP treatment promoted the formation of morphologically normal teeth by the tooth-forming primordium regions and enhanced the development of a regular and large dentin structure by the composites. These observations suggest that human ESTPs contain dentinogenic proteins and can promote dentin and tooth formation. PMID: 20695775 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Mechanical signals activate vascular endothelial growth factor receptor-2 to upregulate endothelial cell proliferation during inflammation. J Immunol. 2010 Jul 15;185(2):1215-21 Authors: Liu J, Agarwal S Signals generated by the dynamic mechanical strain critically regulate endothelial cell proliferation and angiogenesis; however, the molecular basis remains unclear. We investigated the mechanisms by which human dermal microvascular endothelial cells (HDMECs) perceive mechanical signals and relay them intracellularly to regulate gene expression and endothelial cell proliferation. HDMECs were exposed to low/physiologic levels of dynamic strain and probed for the differential activation/inhibition of kinases in the mechanosignaling cascade associated with endothelial cell gene activation. Because angiogenesis is important at inflammatory sites, we also assessed the mechanisms of mechanosignaling in the presence of an proinflammatory cytokine IL-1beta. In this article, we demonstrate that the mechanosignaling cascade is initiated by vascular endothelial growth receptor-2 (VEGFR2) activation. Mechanoactivation of VEGFR2 results in its nuclear translocation and elevation of PI3K-dependent Ser473-Akt phosphorylation. Subsequently, activated Akt inactivates the kinase activity of the serine/threonine kinase, glycogen synthase kinase-3beta (GSK3beta), via its Ser9 phosphorylation. Thus, inactive GSK3beta fails to phosphorylate cyclin D1 and prevents its proteosomal degradation and, consequently, promotes endothelial cell survival and proliferation. In the presence of IL-1beta, cyclin D1 is phosphorylated and degraded, leading to inhibition of cell proliferation. However, mechanical signals repress cyclin D1 phosphorylation and upregulate cell proliferation, despite the presence of IL-1beta. The data indicate that the VEGFR2/Akt/GSK3beta signaling cascade plays a critical role in sensing and phospho-relaying mechanical stimuli in endothelial cells. Furthermore, mechanical forces control highly interconnected networks of proinflammatory and Akt signaling cascades to upregulate endothelial cell proliferation. PMID: 20548028 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Adjuvant engineering for cancer immunotherapy: Development of a synthetic TLR2 ligand with increased cell adhesion. Cancer Sci. 2010 Apr 5; Authors: Akazawa T, Inoue N, Shime H, Kodama K, Matsumoto M, Seya T The development of effective immunoadjuvants for tumor immunotherapy is of fundamental importance. The use of Mycobacterium bovis bacillus Calmette-Guérin cell wall skeleton (BCG-CWS) in tumor immunotherapy has been examined in various clinical applications. Because BCG-CWS is a macromolecule that cannot be chemically synthesized, the development of an alternative synthetic molecule is necessary to ensure a constant supply of adjuvant. In the present study, a new adjuvant was designed based on the structure of macrophage-activating lipopeptide (MALP)-2, which is a Toll-like receptor (TLR)-2 ligand similar to BCG-CWS. Macrophage-activating lipopeptide-2, [S-(2,3-bispalmitoyloxypropyl)Cys (P2C) - GNNDESNISFKEK], originally identified in a Mycoplasma species, is a lipopeptide that can be chemically synthesized. A MALP-2 peptide was substituted with a functional motif, RGDS, creating a novel molecule named P2C-RGDS. RGDS was selected because its sequence constitutes an integrin-binding motif and various integrins are expressed in immune cells including dendritic cells (DCs). Thus, this motif adds functionality to the ligand. P2C-RGDS activated DCs and splenocytes more efficiently than MALP-2 over short incubation times in vitro, and the RGDS motif contributed to their activation. Furthermore, P2C-RGDS showed higher activity than MALP-2 in inducing migration of DCs to draining lymph node, and in inhibiting tumor growth in vivo. This process of designing and developing synthetic adjuvants has been named "adjuvant engineering," and the evaluation and improvement of P2C-RGDS constitutes a first step in the development of stronger synthetic adjuvants in the future. (Cancer Sci 2010). PMID: 20507323 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | A novel bioengineering clinical device testing cellular homeostatic potentials to customize and monitor nutritional-related age-management interventional strategies. Rejuvenation Res. 2010 Apr-Jun;13(2-3):256-9 Authors: Mariani G, Dellaglio F, Marotta F Most devices assessing body composition harbor a number of drawbacks and hardly assess the phenomena taking place at a cellular membrane level. The present single-frequency bioelectrical potential homeostatic structure analysis (PHoSA) technology requires only a proper hands contact on fixed electrodes and determines the phase displacement between tested current and voltage by using a 50-KHz alternate sinusoidal current. This allows quick testing time with high degree of precision, sensitivity, and specificity of sectorial functional body compartments analysis. Such assessment may prove to be an integrated part of either a diagnostic workup or monitoring tool in tailoring nutritional/nutraceutical, pharmacological, and exercise activity, all being framed within a proactive, preventive, age-intervention management strategy. PMID: 20462382 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Demographic consequences of defeating aging. Rejuvenation Res. 2010 Apr-Jun;13(2-3):329-34 Authors: Gavrilov LA, Gavrilova NS A common objection against starting a large-scale biomedical war on aging is the fear of catastrophic population consequences (overpopulation). This fear is only exacerbated by the fact that no detailed demographic projections for radical life extension scenario have been conducted so far. This study explores different demographic scenarios and population projections, in order to clarify what could be the demographic consequences of a successful biomedical war on aging. A general conclusion of this study is that population changes are surprisingly slow in their response to a dramatic life extension. For example, we applied the cohort-component method of population projections to 2005 Swedish population for several scenarios of life extension and a fertility schedule observed in 2005. Even for very long 100-year projection horizon, with the most radical life extension scenario (assuming no aging at all after age 60), the total population increases by 22% only (from 9.1 to 11.0 million). Moreover, if some members of society reject to use new anti-aging technologies for some religious or any other reasons (inconvenience, non-compliance, fear of side effects, costs, etc.), then the total population size may even decrease over time. Thus, even in the case of the most radical life extension scenario, population growth could be relatively slow and may not necessarily lead to overpopulation. Therefore, the real concerns should be placed not on the threat of catastrophic population consequences (overpopulation), but rather on such potential obstacles to a success of biomedical war on aging, as scientific, organizational, and financial limitations. PMID: 20426616 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Characterization of the complete fiber network topology of planar fibrous tissues and scaffolds. Biomaterials. 2010 Jul;31(20):5345-54 Authors: D'Amore A, Stella JA, Wagner WR, Sacks MS Understanding how engineered tissue scaffold architecture affects cell morphology, metabolism, phenotypic expression, as well as predicting material mechanical behavior has recently received increased attention. In the present study, an image-based analysis approach that provides an automated tool to characterize engineered tissue fiber network topology is presented. Micro-architectural features that fully defined fiber network topology were detected and quantified, which include fiber orientation, connectivity, intersection spatial density, and diameter. Algorithm performance was tested using scanning electron microscopy (SEM) images of electrospun poly(ester urethane)urea (ES-PEUU) scaffolds. SEM images of rabbit mesenchymal stem cell (MSC) seeded collagen gel scaffolds and decellularized rat carotid arteries were also analyzed to further evaluate the ability of the algorithm to capture fiber network morphology regardless of scaffold type and the evaluated size scale. The image analysis procedure was validated qualitatively and quantitatively, comparing fiber network topology manually detected by human operators (n = 5) with that automatically detected by the algorithm. Correlation values between manual detected and algorithm detected results for the fiber angle distribution and for the fiber connectivity distribution were 0.86 and 0.93 respectively. Algorithm detected fiber intersections and fiber diameter values were comparable (within the mean +/- standard deviation) with those detected by human operators. This automated approach identifies and quantifies fiber network morphology as demonstrated for three relevant scaffold types and provides a means to: (1) guarantee objectivity, (2) significantly reduce analysis time, and (3) potentiate broader analysis of scaffold architecture effects on cell behavior and tissue development both in vitro and in vivo. PMID: 20398930 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | The role of simvastatin in the osteogenesis of injectable tissue-engineered bone based on human adipose-derived stromal cells and platelet-rich plasma. Biomaterials. 2010 Jul;31(20):5325-35 Authors: Zhou Y, Ni Y, Liu Y, Zeng B, Xu Y, Ge W An injectable tissue-engineered bone (ITB) composed of human adipose-derived stromal cells (hADSCs) and platelet-rich plasma (hPRP) was preliminarily constructed, but its osteogenic capability needs improving. This study aimed to evaluate if simvastatin can be applied as a bone anabolic agent for this ITB. We found 0.01 microm, 0.1 microm, and 1 microm simvastatin could induce hADSCs' osteoblastic differentiation in vitro that accompanied with non-inhibition on cell proliferation, high alkaline phosphatase activity, more mineralization deposition and more expression of osteoblast-related genes such as osteocalcin, core binding factor alpha1, bone morphogenetic protein-2, vascular endothelial growth factor, and basic fibroblast growth factor. Simvastatin at 1 mum seemed the most optimal concentration due to its high osteocalcin secretion in media (P < 0.01). Quantitative mineralization assay also showed 1 microm SIM had the most obvious synergistic effect on hPRP's induction for matrix mineralization of hADSCs (P < 0.01). When 1 microm Simvastatin was applied to this ITB to restore the critical-sized calvarial defects in mice, more bone formation was observed in defected regions, and the peripheries just outside the defect margins by X-ray analysis, and H&E staining. These findings indicate that simvastatin at optimal concentrations can be used to promote this ITB's osteogenesis. However, simvastatin's effects on this ITB await long-term investigation. PMID: 20381859 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Practice of postponing human aging. Rejuvenation Res. 2010 Apr-Jun;13(2-3):356-8 Authors: Kristjuhan U People want to be wealthy, attractive, young, and healthy, and they integrate these wishes into most of their activities. As a result, aging processes are somewhat postponed in the human organism. Both average life expectancy and expected life in good health increase by 0.15-0.2 years every year in most developed countries. Studies show that using contemporary knowledge enables an increase in life expectancy up to 90 years in these countries, and much more in the future. Increases in life expectancy depend on the interest of society in these problems and on the amount of scientific research directed at postponing aging and rejuvenation. PMID: 20370496 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Affordable rejuvenation: a prototype facility in action. Rejuvenation Res. 2010 Apr-Jun;13(2-3):350-2 Authors: Prokopov AF, Reinmuth J The Strategies for Engineered Negligible Senescence (SENS) agenda contemplates specialized centers that offer a periodic rejuvenation and biogerontological maintenance for their clients. Although high-tech interventions are still in an early research phase, well-proven natural techniques, such as various forms of caloric/nutritional restriction, physical training, and preconditioning treatments, are not unanimously embraced due to poor adherence of patients. The practicability of such interventions can be significantly improved by "engineering" them for higher efficiency and better user friendliness. We describe practical experience in developing and running a prototype facility that uses rejuvenative treatment protocols, derived from two natural life span-prolonging strategies: Intermittent calorie/nutritive restriction (ICR) and intermittent oxygen restriction (IOR). PMID: 20370490 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | The Drosophila planar polarity proteins inturned and multiple wing hairs interact physically and function together. Genetics. 2010 Jun;185(2):549-58 Authors: Lu Q, Yan J, Adler PN The conserved frizzled (fz) pathway regulates planar cell polarity in both vertebrate and invertebrate animals. This pathway has been most intensively studied in the wing of Drosophila, where the proteins encoded by pathway genes all accumulate asymmetrically. Upstream members of the pathway accumulate on the proximal, distal, or both cell edges in the vicinity of the adherens junction. More downstream components including Inturned and Multiple Wing Hairs accumulate on the proximal side of wing cells prior to hair initiation. The Mwh protein differs from other members of the pathway in also accumulating in growing hairs. Here we show that the two Mwh accumulation patterns are under different genetic control with the early proximal accumulation being regulated by the fz pathway and the latter hair accumulation being largely independent of the pathway. We also establish recruitment by proximally localized Inturned to be a putative mechanism for the localization of Mwh to the proximal side of wing cells. Genetically inturned (in) acts upstream of mwh (mwh) and is required for the proximal localization of Mwh. We show that Mwh can bind to and co-immunoprecipitate with Inturned. We also show that these two proteins can function in close juxtaposition in vivo. An InMwh fusion protein provided complete rescue activity for both in and mwh mutations. The fusion protein localized to the proximal side of wing cells prior to hair formation and in growing hairs as expected if protein localization is a key for the function of these proteins. PMID: 20351219 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Apical polarity in three-dimensional culture systems: where to now? J Biol. 2010;9(1):2 Authors: Inman JL, Bissell MJ Delineation of the mechanisms that establish and maintain the polarity of epithelial tissues is essential to understanding morphogenesis, tissue specificity and cancer. Three-dimensional culture assays provide a useful platform for dissecting these processes but, as discussed in a recent study in BMC Biology on the culture of mammary gland epithelial cells, multiple parameters that influence the model must be taken into account. PMID: 20092610 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Urodynamic evaluation of fesoterodine metabolite, doxazosin and their combination in a rat model of partial urethral obstruction. BJU Int. 2010 Jul;106(2):287-93 Authors: Füllhase C, Soler R, Gratzke C, Brodsky M, Christ GJ, Andersson KE OBJECTIVE: To evaluate the urodynamic effects of fesoterodine, a new antimuscarinic agent, alone and combined with doxazosin, in a rat model of partial urethral obstruction (PUO), as 35-83% of men with bladder outlet obstruction (BOO) secondary to benign prostatic hyperplasia (BPH) have overactive bladder (OAB) syndrome, and as the combination of alpha(1)-adrenoceptor- and muscarinic-receptor antagonists has been proposed to be beneficial for these patients. MATERIALS AND METHODS: Thirty-seven male Sprague-Dawley rats (250 g) had surgically induced PUO; 2 weeks later they were evaluated by cystometry with no anaesthesia or any restraint. After a 1-h period either 5-hydroxymethyl tolterodine (5-HMT, the active metabolite of fesoterodine, previously known as SPM 7605), doxazosin or a combination of both, was given intravenously (0.1 mg/kg body weight), and cystometry was continued for another 45 min. Fifteen healthy, age-matched rats served as a control. RESULTS: At 2 weeks after surgery the obstructed rats had an greater bladder weight, threshold pressure (TP) and micturition frequency (MF), and lower bladder capacity (BCap) and micturition volume (MV) than the controls. 5-HMT did not cause urinary retention in obstructed rats, but decreased TP, maximum pressure (MP), spontaneous bladder activity (SA) and, paradoxically, increased MF. Doxazosin alone decreased TP, MP, MF and increased BCap and MV. 5-HMT and doxazosin together did not depress the ability to empty the bladder, and showed decreased TP, MP and SA. CONCLUSIONS: 5-HMT, alone and in combination, did not impair the voiding ability in obstructed rats. Doxazosin counteracted some of the 'negative' effects of 5-HMT in this model (increase of MF) and did not attenuate the 'positive' effects (decrease of bladder SA). In this model, the combination of 5-HMT and doxazosin appeared to be urodynamically safe and well tolerated. PMID: 19888972 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [Clone screening and interference efficiency of seed cells with CCL20 gene knockdown for tissue-engineered skin] Zhonghua Wai Ke Za Zhi. 2009 Apr 15;47(8):621-4 Authors: Wang LH, Peng DZ, Liu J, Zhou X, Wang Y, He SD, He B, Zheng BX, Dong ZX, Zhou GQ OBJECTIVE: To screen stable cell clones of CCL20 gene knockdown and assess their interference effects, recombinant lentivirus vectors with CCL20 gene specific shRNA were applied to infect human immortal keratinocyte line (HaCaT). METHODS: The three pHSER-CCL20-shRNA-GFP vectors (pHCG-1 and pHCG-2 were CCL20 gene specific, and pHCG-3 was used as mismatch control) have been previously constructed. The virus packaging cell line 293FT was transfected with these vectors by using CaCl2 methods to produce lectiviral particles. After the viral titers of these three harvested cell supernatants were determined by flow cytometry, HaCaT cells were transfected by these viruses and screened under the pressure of G418. The CCL20 mRNA from HaCaT cell clones and the CCL20 protein levels in the supernatants of HaCaT cell clones were detected by Real-time RT-PCR and ELISA, respectively. RESULTS: The titers of three lentiviruses were 7.08 x 10(5) transduced units (TU)/ml, 1.88 x 10(5) TU /ml and 2.08 x 10(5) TU/ml, respectively. Two HaCaT cell clones from each lectiviral vectors were obtained after G418 screening for 5 - 8 weeks. Four CCL20 gene specific clones showed stable interference effect in both Real-time RT-PCR and ELISA. The mRNA expression and protein level of CCL20 gene specific clones were down regulated significantly. CONCLUSIONS: The four human immortal keratinocyte clones with long term CCL20 gene knockdown have been screened by recombinant lentivirus vectors with CCL20 gene specific shRNA. These clones might be served as seed cells for novel tissue-engineered skin with lower rejection. PMID: 19595046 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [Experimental study on spinal fusion induced by hBMP-4 gene modified tissue engineered bone] Zhonghua Wai Ke Za Zhi. 2009 Feb 1;47(3):197-201 Authors: Zheng ZM, Dong ZY, Kuang GM, Chen H, Lü Y, Zhang KB, Liu H, Li FB OBJECTIVE: To evaluate the efficacy of hBMP-4 gene modified tissue engineered bone graft in the enhancement of rabbit spinal fusion and find an ideal kind of substitute for the autograft bone. METHODS: Rabbit BMSCs were cultured and transfected with AAV-hBMP-4 using different MOI value. The optimal MOI value were determined by observing cell's morphology change. BMSCs were then transfected with AAV-hBMP4 and AAV-EGFP respectively, following which the transfected cells were evenly suspended in a collagen sponge I, and implanted to either side of the L5,6 intertransverse spaces posterolateral in the New Zealand rabbits to induce spinal fusion. Fourteen rabbits were randomly divided into 2 groups. Group 1: AAV-hBMP-4 transfected BMSCs in the right side (hBMP-4 side) and autograft bone in the left side. Group 2: AAV-hBMP-4 transfected BMSCs in the right side (hBMP-4 side) and AAV-EGFP transfected BMSCs in the left side (EGFP side). Radiographs and three-dimensional CT of the spine, manual palpation, gross and histological examination of the fusion masses for all the animals were performed subsequent to animals having been sacrificed at 12 weeks after surgery. RESULTS: Evaluation has been taken in 12 New Zealand rabbits delivered into 2 groups which meet the criterion after operation. Eleven in 12 implemented sides involved hBMP-4 achieved bony fusion, to which 5 in 6 autografted sides was similar. But only 2 in 6 sides in EGFP-group achieved bony fusion meanwhile. Three-dimensional CT scan and palpation also evidenced the results. Bone formation was observed obviously on specimen both in hBMP4 sides and autografted ones. EGFP-group also got bony integration, but the quantity was small. CONCLUSION: Tissue-engineered bone graft constructed from application of hBMP4 is a fine substitute for autograft. Effective enhancement of bony integration in spinal fusion surgery has been evidenced in vivo. PMID: 19563074 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Cryogenic prototyping of chitosan scaffolds with controlled micro and macro architecture and their effect on in vivo neo-vascularization and cellular infiltration. J Biomed Mater Res A. 2010 Sep 15;94(4):1303-11 Authors: Lim TC, Chian KS, Leong KF A major challenge in tissue engineering has been to develop scaffolds with controlled complex geometries, on both the macro- and micro-scale. One group of techniques, using rapid prototyping (RP) processes, has the capability to produce complex three-dimensional structures with good control over the size, geometry, and connectivity of the pores. In this article, a novel technique based on RP technology, that is, cryogenic prototyping (CP), that has the capability to fabricate scaffolds with controlled macro- and micro-structures, is presented. Our in vivo studies showed that the micro architecture (i.e., both pore size and pore orientation) and macro structures of the CP scaffolds affect both cellular infiltration and neo-vascularization. Full cellular infiltration and neo-vascularization were observed after 28 days in scaffolds with micropore sizes of 90 mum. In addition, it was observed that channels (300 mum) created in scaffolds were effective at enhancing cellular infiltration and vascularization. Our results have demonstrated that CP is a viable method for fabricating scaffolds for a wide range of tissue engineering applications. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694998 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Aligned poly(L-lactic-co-e-caprolactone) electrospun microfibers and knitted structure: A novel composite scaffold for ligament tissue engineering. J Biomed Mater Res A. 2010 Sep 15;94(4):1270-82 Authors: Vaquette C, Kahn C, Frochot C, Nouvel C, Six JL, De Isla N, Luo LH, Cooper-White J, Rahouadj R, Wang X We developed a novel technique involving knitting and electrospinning to fabricate a composite scaffold for ligament tissue engineering. Knitted structures were coated with poly(L-lactic-co-e-caprolactone) (PLCL) and then placed onto a rotating cylinder and a PLCL solution was electrospun onto the structure. Highly aligned 2-mum-diameter microfibers covered the space between the stitches and adhered to the knitted scaffolds. The stress-strain tensile curves exhibited an initial toe region similar to the tensile behavior of ligaments. Composite scaffolds had an elastic modulus (150 +/- 14 MPa) similar to the modulus of human ligaments. Biological evaluation showed that cells proliferated on the composite scaffolds and they spontaneously orientated along the direction of microfiber alignment. The microfiber architecture also induced a high level of extracellular matrix secretion, which was characterized by immunostaining. We found that cells produced collagen type I and type III, two main components found in ligaments. After 14 days of culture, collagen type III started to form a fibrous network. We fabricated a composite scaffold having the mechanical properties of the knitted structure and the morphological properties of the aligned microfibers. It is difficult to seed a highly macroporous structure with cells, however the technique we developed enabled an easy cell seeding due to presence of the microfiber layer. Therefore, these scaffolds presented attractive properties for a future use in bioreactors for ligament tissue engineering. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694995 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Primary human osteoblast culture on 3D porous collagen-hydroxyapatite scaffolds. J Biomed Mater Res A. 2010 Sep 15;94(4):1244-50 Authors: Jones GL, Walton R, Czernuszka J, Griffiths SL, El Haj AJ, Cartmell SH There is a need in tissue-engineering for 3D scaffolds that mimic the natural extracellular matrix of bone to enhance cell adhesion, proliferation, and differentiation. The scaffold is also required to be degradable. A highly porous scaffold has been developed to incorporate two of the extracellular components found in bone-collagen and hydroxyapatite (HA). The scaffold's collagen component is an afibrillar monomeric type I atelocollagen extracted from foetal calf's skin. This provided a novel environment for the inclusion of HA powder. Five hundred thousand primary human osteoblasts were seeded onto 4 mm cubed scaffolds that varied in ratio of HA to collagen. Weight ratios of 1:99, 25:75, 50:50, and 75:25 hydroxyapatite:collagen (HA:Collagen) were analysed. The scaffolds plus cells were cultured for 21 days. DNA assays and live/dead viability staining demonstrated that all of the scaffolds supported cell proliferation and viability. An alkaline phosphatase assay showed similar osteoblast phenotype maintenance on all of the 3D scaffolds analysed at 21 days. MicroCT analysis demonstrated an increase in total sample volume (correlating to increase in unmineralised matrix production). An even distribution of HA throughout the collagen matrix was observed using this technique. Also at 3 weeks, reductions in the percentage of the mineralised phase of the constructs were seen. These results indicate that each of the ratios of HA/collagen scaffolds have great potential for bone tissue engineering. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694991 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Bioactivation of porous polyurethane scaffolds using fluorinated RGD surface modifiers. J Biomed Mater Res A. 2010 Sep 15;94(4):1226-35 Authors: Blit PH, Shen YH, Ernsting MJ, Woodhouse KA, Santerre JP Biomaterial scaffolds for tissue engineering require appropriate cell adhesion, proliferation, and infiltration into their three-dimensional (3D) porous structures. Surface modification techniques have the potential to enhance cell infiltration into synthetic scaffolds while retaining bulk material properties intact. The objective of this work was to assess the potential of achieving a uniform surface modification in 3D porous constructs through the blending of surface-modifying additives known as bioactive fluorinated surface modifiers (BFSMs) with a base polyurethane material. By coupling RGD peptides to the fluorinated surface modifiers to form RGD-BFSMs, the BFSMs can act as a vehicle for the delivery of RGD moieties to the surface without direct covalent attachment to the polymer substrate. Fluorescent RGD-BFSMs were shown to migrate to the polymer-air interfaces within the porous scaffolds by two-photon confocal microscopy. A-10 rat aortic smooth muscle cells were cultured for 4 weeks on nonmodified and RGD-BFSM-modified porous scaffolds, and cell adhesion, proliferation, and viability were quantified at different depths. RGD-BFSM-modified scaffolds showed significantly greater cell numbers within deeper regions of the scaffolds, and this difference became more pronounced over time. This study demonstrates an effective approach to promote cell adhesion and infiltration within thick ( approximately 0.5 cm) porous synthetic scaffolds by providing a uniform distribution of adhesive peptide throughout the scaffolds without the use of covalent surface reaction chemistry. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694989 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Bladder tissue engineering: Tissue regeneration and neovascularization of HA-VEGF-incorporated bladder acellular constructs in mouse and porcine animal models. J Biomed Mater Res A. 2010 Sep 15;94(4):1205-15 Authors: Loai Y, Yeger H, Coz C, Antoon R, Islam SS, Moore K, Farhat WA Successful tissue engineering requires appropriate recellularization and vascularization. Herein, we assessed the regenerative and angiogenic effects of porcine bladder acellular matrix (ACM) incorporated with hyaluronic acid (HA) and vascular endothelial growth factor (VEGF) in mouse and porcine models. Prepared HA-ACMs were rehydrated in different concentrations of VEGF (1, 2, 3, 10, and 50 ng/g ACM). Grafts were implanted in mice peritoneum in situ for 1 week. Angiogenesis was quantified with CD31 and Factor VIII immunostaining using Simple PCI. Selected optimal VEGF concentration that induced maximum vascularization was then used in porcine bladder augmentation model. Implants were left in for 4 and 10 weeks. Three groups of six pigs each were implanted with ACM alone, HA-ACM, and HA-VEGF-ACM. Histological, immunohistochemical (Uroplakin III, alpha-SMA, Factor VIII), and immunofluorescence (CD31) analysis were performed to assess graft regenerative capacity and angiogenesis. In mouse model, statistically significant increase in microvascular density was demonstrated in the 2 ng/g ACM group. When this concentration was used in porcine model, recellularization increased significantly from weeks 4 to 10 in HA-VEGF-ACM, with progressive decrease in fibrosis. Significantly increased vascularization, coupled with increased urothelium and smooth muscle cell (SMC) regeneration, was observed in HA-VEGF grafts at week 10 in the center and periphery, compared with week 4. HA-VEGF grafts displayed highest in vivo epithelialization, neovascularization, and SMCs regeneration. A total of 2 ng/g tissue VEGF when incorporated with HA proved effective in stimulating robust graft recellularization and vascularization, coordinated with increased urothelial bladder development and SMC augmentation into bundles by week 10. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694987 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Role of mesenchymal stem cells in tissue engineering of meniscus. J Biomed Mater Res A. 2010 Sep 15;94(4):1150-61 Authors: Zellner J, Mueller M, Berner A, Dienstknecht T, Kujat R, Nerlich M, Hennemann B, Koller M, Prantl L, Angele M, Angele P Tissue engineering is a promising approach for the treatment of tissue defects. Mesenchymal stem cells are of potential use as a source of repair cells or of important growth factors for tissue engineering. The purpose of this study was to examine the role of mesenchymal stem cells in meniscal tissue repair. This was tested using several cell and biomaterial-based treatment options for repair of defects in the avascular zone of rabbit menisci. Circular meniscal punch defects (2 mm) were created in the avascular zone of rabbit menisci and left empty or filled with hyaluronan-collagen composite matrices without cells, loaded with platelet-rich plasma, autologous bone marrow, or autologous mesenchymal stem cells. In some experiments, matrices with stem cells were precultured in chondrogenic medium for 14 days before implantation. Rabbits were then allowed free cage movement after surgery for up to 12 weeks. Untreated defects and defects treated with cell-free implants had muted fibrous healing responses. Neither bone marrow nor platelet-rich plasma loaded in matrices produced improvement in healing compared with cell-free implants. The implantation of 14 days precultured chondrogenic stem cell-matrix constructs resulted in fibrocartilage-like repair tissue, which was only partially integrated with the native meniscus. Non-precultured mesenchymal stem cells in hyaluronan-collagen composite matrices stimulated the development of completely integrated meniscus-like repair tissue. The study shows the necessity of mesenchymal stem cells for the repair of meniscal defects in the avascular zone. Mesenchymal stem cells seem to fulfill additional repair qualities besides the delivery of growth factors. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694982 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Fabrication of cardiac patch with decellularized porcine myocardial scaffold and bone marrow mononuclear cells. J Biomed Mater Res A. 2010 Sep 15;94(4):1100-10 Authors: Wang B, Borazjani A, Tahai M, de Jongh Curry AL, Simionescu DT, Guan J, To F, Elder SH, Liao J Tissue engineered cardiac grafts are a promising therapeutic mode for ventricular wall reconstruction. Recently, it has been found that acellular tissue scaffolds provide natural ultrastructural, mechanical, and compositional cues for recellularization and tissue remodeling. We thus assess the potential of decellularized porcine myocardium as a scaffold for thick cardiac patch tissue engineering. Myocardial sections with 2-mm thickness were decellularized using 0.1% sodium dodecyl sulfate and then reseeded with differentiated bone marrow mononuclear cells. We found that thorough decellularization could be achieved after 2.5 weeks of treatment. Reseeded cells were found to infiltrate and proliferate in the tissue constructs. Immunohistological staining studies showed that the reseeded cells maintained cardiomyocyte-like phenotype and possible endothelialization was found in locations close to vasculature channels, indicating angiogenesis potential. Both biaxial and uniaxial mechanical testing showed a stiffer mechanical response of the acellular myocardial scaffolds; however, tissue extensibility and tensile modulus were found to recover in the constructs along with the culture time, as expected from increased cellular content. The cardiac patch that we envision for clinical application will benefit from the natural architecture of myocardial extracellular matrix, which has the potential to promote stem cell differentiation, cardiac regeneration, and angiogenesis. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694977 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Surface nanoscale patterning of bioactive glass to support cellular growth and differentiation. J Biomed Mater Res A. 2010 Sep 15;94(4):1091-9 Authors: Lei B, Chen X, Wang Y, Zhao N, Du C, Fang L Bioactive glasses (BGs) have been widely used for bone tissue regeneration as they are able to bond directly with bone. Clinical applications of these materials are likely to be in particulate form. Nanoscale materials can mimic the surface properties of natural tissues, which have exhibited superior cytocompatible property and improved tissue regeneration. The objective of this study is to prepare bioactive glass particles with nanoscale or non-nanoscale surface features and investigate their microstructure, apatite-forming bioactivity and cellular response. The microstructure and micro-nanoscale surface morphology were controlled by adding a hydroxyl-carboxyl acid (citric acid) in the sol-gel process. Results shown that the addition of citric acid induced the formation of nanoscale surface structure and increased the specific surface area, pore volume and pore size of bioactive glass particles. The citric acid with low-concentration-derived sol-gel bioactive glasses (CBGs) resulted in an enhanced apatite-formation ability in simulated body fluids (SBF) compared to normal bioactive glasses. The attachment and proliferation of rat marrow mesenchymal stem cells (RMSCs) on CBGs (low concentration) were higher than those of normal BGs, demonstrating that the CBGs had the excellent cytocompatibility. RMSCs on CBGs (low concentration) expressed the higher alkaline phosphatase activity (ALP) than normal BGs and tissue culture plastic, revealing that CBGs can induced differentiation of RMSCs to the osteogenic lineage. Such improved physical and biological properties of CBGs (low concentration) should be useful in developing new bioactive glass materials for stem cell-based bone regeneration or biomimic tissue engineering scaffolds. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694976 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Endothelialization of PVA/gelatin cryogels for vascular tissue engineering: Effect of disturbed shear stress conditions. J Biomed Mater Res A. 2010 Sep 15;94(4):1080-90 Authors: Vrana NE, Cahill PA, McGuinness GB Mechanically, poly(vinyl alcohol) (PVA)-based cryogels are extremely well suited for vascular tissue engineering applications. However, their surface properties lead to a slow rate of endothelialization, and the mode of cell attachment leaves the endothelium susceptible to removal under physiological shear stress conditions. In this study, abrupt and ramped disturbed shear stress conditions created by a turbulent orbital flow were used to examine endothelialization on PVA/gelatin cryogels. Cell proliferation rate and apoptosis were evaluated by fluorescent activated cell sorter (FACS) analysis, and the expression of cell-adhesion molecules was used to evaluate the response of cells on cryogels to static and shear conditions by real-time polymerase chain reaction (RT-PCR). Application of a ramped shear stress had a profound effect on endothelial cell proliferation (22.30 +/- 0.20-fold increase), necrosis (eliminated), apoptosis (1.04 +/- 0.18 increase), and overall facilitation of endothelialization while concomitantly increasing nitric oxide (NO) synthesis levels. Ramped shear stress was also effective in helping the retention of the endothelial cells on the cryogel surface, whereas abrupt application caused widespread removal. Under static conditions, Selectin-P expression decreased, whereas both inter-cellular adhesion molecule (ICAM) and platelet endothelial cell adhesion molecule (PECAM)-I expression increased on cryogels over a 10-day culture period. Under both shear stress conditions, Selectin-P expression was decreased both on cryogels and tissue culture polystyrene (TCPS). Controlled application of disturbed shear stress shortens endothelialization times on cryogel surfaces, in contrast to the established antiproliferative effect of shear stress caused by laminar flow, without compromising their functionality. This demonstrates how such mechanical stimuli can be exploited to alter cellular behavior and facilitate the required outcomes for tissue engineering applications. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694975 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Comparison of morphology, orientation, and migration of tendon derived fibroblasts and bone marrow stromal cells on electrochemically aligned collagen constructs. J Biomed Mater Res A. 2010 Sep 15;94(4):1070-9 Authors: Gurkan UA, Cheng X, Kishore V, Uquillas JA, Akkus O There are approximately 33 million injuries involving musculoskeletal tissues (including tendons and ligaments) every year in the United States. In certain cases the tendons and ligaments are damaged irreversibly and require replacements that possess the natural functional properties of these tissues. As a biomaterial, collagen has been a key ingredient in tissue engineering scaffolds. The application range of collagen in tissue engineering would be greatly broadened if the assembly process could be better controlled to facilitate the synthesis of dense, oriented tissue-like constructs. An electrochemical method has recently been developed in our laboratory to form highly oriented and densely packed collagen bundles with mechanical strength approaching that of tendons. However, there is limited information whether this electrochemically aligned collagen bundle (ELAC) presents advantages over randomly oriented bundles in terms of cell response. Therefore, the current study aimed to assess the biocompatibility of the collagen bundles in vitro, and compare tendon-derived fibroblasts (TDFs) and bone marrow stromal cells (MSCs) in terms of their ability to populate and migrate on the single and braided ELAC bundles. The results indicated that the ELAC was not cytotoxic; both cell types were able to populate and migrate on the ELAC bundles more efficiently than that observed for random collagen bundles. The braided ELAC constructs were efficiently populated by both TDFs and MSCs in vitro. Therefore, both TDFs and MSCs can be used with the ELAC bundles for tissue engineering purposes. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694974 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Combination of enzymes and flow perfusion conditions improves osteogenic differentiation of bone marrow stromal cells cultured upon starch/poly(epsilon-caprolactone) fiber meshes. J Biomed Mater Res A. 2010 Sep 15;94(4):1061-9 Authors: Martins AM, Saraf A, Sousa RA, Alves CM, Mikos AG, Kasper FK, Reis RL Previous studies have shown that alpha-amylase and lipase are capable of enhancing the degradation of fiber meshes blends of starch and poly(epsilon-caprolactone) (SPCL) under dynamic conditions, and consequently to promote the proliferation and osteogenic differentiation of bone marrow stromal cells (MSCs). This study investigated the effect of flow perfusion bioreactor culture in combination with enzymes on the osteogenic differentiation of MSCs. SPCL fiber meshes were seeded with MSCs and cultured with osteogenic medium supplemented with alpha-amylase, lipase, or a combination of the two for 8 or 16 days using static or flow conditions. Lipase and its combination with alpha-amylase enhanced cell proliferation after 16 days. In addition, the flow perfusion culture enhanced the infiltration of cells and facilitated greater distribution of extracellular matrix (ECM) throughout the scaffolds in the presence/absence of enzymes. A significant amount of calcium was detected after 16 days in all groups cultured in flow conditions compared with static cultures. Nevertheless, when alpha-amylase and lipase were included in the flow perfusion cultures, the calcium content was 379 +/- 30 mug/scaffold after as few as 8 days. The highest calcium content (1271 +/- 32 mug/scaffold) was obtained for SPCL/cell constructs cultured for 16 days in the presence of lipase and flow. Furthermore, von Kossa staining and tetracycline fluorescence of histological sections demonstrated mineral deposition within the scaffolds for all groups cultured for 16 days under flow. However, all the data corroborate that lipase coupled with flow perfusion conditions improve the osteogenic differentiation of MSCs and enhance ECM mineralization. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694973 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Concentrated collagen-chondroitin sulfate scaffolds for tissue engineering applications. J Biomed Mater Res A. 2010 Sep 15;94(4):1050-60 Authors: Liang WH, Kienitz BL, Penick KJ, Welter JF, Zawodzinski TA, Baskaran H Collagen-chondroitin sulfate biomaterial scaffolds have been used in a number of tissue-engineered products under development or in the clinics. In this article, we describe a new approach based on centrifugation for obtaining highly concentrated yet porous collagen scaffolds. Water uptake, chondroitin sulfate retention, morphology, mechanical properties, and tissue-engineering potential of the concentrated scaffolds were investigated. Our results show that the new approach can lead to scaffolds containing four times as much collagen as that in conventional unconcentrated scaffolds. Further, water uptake in the concentrated scaffolds was significantly greater while chondroitin sulfate retention in the concentrated scaffolds was unaffected. The value of mean pore diameter in the concentrated scaffolds was smaller than that in the unconcentrated scaffolds and the walls of the pores in the former comprised of a continuous sheet of collagen. The mechanical properties measured as moduli of elasticity in compression and tension were improved by as much as 30 times in the concentrated scaffolds. In addition, our tissue culture results with human mesenchymal stem cells and foreskin keratinocytes show that the new scaffolds can be used for cartilage and skin tissue-engineering applications. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20694972 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Molecular characterisation of post-bio-electrosprayed human brain astrocytoma cells. Analyst. 2010 Aug 6; Authors: Eddaoudi A, Townsend-Nicholson A, Timms JF, Schorge S, Jayasinghe SN Bio-electrospraying (BES) is a method for directly jetting living cells under conditions that allow their distribution in the x, y, and z axes. Previous work has been focused on achieving jetting in stable cone-jet mode, which is required for precision placement, and these studies have demonstrated that there are no significant effects of bio-electrospraying on cell morphology or viability. In this work, we examine the biological properties of bio-electrosprayed cells using assays of cellular function that range from the molecular level through to integrated cellular systems, and include proteomics, signal transduction, cell growth and proliferation, and the characterisation of apoptotic blebs. From these molecular methods, we have determined that bio-electrospraying, under the electric field conditions used to achieve stable cone-jet mode, causes no alterations to the biological properties and function of the cells being jetted. Bio-electrosprayed and control cells had similar viability, proliferation properties and virtually indistinguishable cell cycle profiles. The biophysical properties of large conducting (BK) potassium channels were unchanged, as were the pharmacological responses of the endogenous muscarinic and exogenous P2Y(11) receptors, both of which are cell surface receptors of the 7TM superfamily. Proteomic analyses revealed that although three proteins had subtle differences in expression level between bio-electrosprayed and control cells, none of these fold differences was above the 1.5-fold cut-off threshold required for further analyses. These findings support the further development of bio-electrosprays as a viable technology for a wide diversity of tissue engineering, regenerative biology, advanced cellular theraputics and medicinal applications, having significance in the clinic. PMID: 20694206 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | An engineered mammalian band-pass network. Nucleic Acids Res. 2010 Aug 6; Authors: Greber D, Fussenegger M Gene expression circuitries, which enable cells to detect precise levels within a morphogen concentration gradient, have a pivotal impact on biological processes such as embryonic pattern formation, paracrine and autocrine signalling, and cellular migration. We present the rational synthesis of a synthetic genetic circuit exhibiting band-pass detection characteristics. The components, involving multiply linked mammalian trans-activator and -repressor control systems, were selected and fine-tuned to enable the detection of 'low-threshold' morphogen (tetracycline) concentrations, in which target gene expression was triggered, and a 'high-threshold' concentration, in which expression was muted. In silico predictions and supporting experimental findings indicated that the key criterion for functional band-pass detection was the matching of componentry that enabled sufficient separation of the low and high threshold points. Using the circuitry together with a fluorescence-encoded target gene, mammalian cells were genetically engineered to be capable of forming a band-like pattern of differentiation in response to a tetracycline chemical gradient. Synthetic gene networks designed to emulate naturally occurring gene behaviours provide not only insight into biological processes, but may also foster progress in future tissue engineering, gene therapy and biosensing applications. PMID: 20693530 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | The generation of 3-D tissue models based on hyaluronan hydrogel-coated microcarriers within a rotating wall vessel bioreactor. Biomaterials. 2010 Aug 6; Authors: Skardal A, Sarker SF, Crabbé A, Nickerson CA, Prestwich GD With the increasing necessity for functional tissue- and organ equivalents in the clinic, the optimization of techniques for the in vitro generation of organotypic structures that closely resemble the native tissue is of paramount importance. The engineering of a variety of highly differentiated tissues has been achieved using the rotating wall vessel (RWV) bioreactor technology, which is an optimized suspension culture allowing cells to grow in three-dimensions (3-D). However, certain cell types require the use of scaffolds, such as collagen-coated microcarrier beads, for optimal growth and differentiation in the RWV. Removal of the 3-D structures from the microcarriers involves enzymatic treatment, which disrupts the delicate 3-D architecture and makes it inapplicable for potential implantation. Therefore, we designed a microcarrier bead coated with a synthetic extracellular matrix (ECM) composed of a disulfide-crosslinked hyaluronan and gelatin hydrogel for 3-D tissue engineering, that allows for enzyme-free cell detachment under mild reductive conditions (i.e. by a thiol-disulfide exchange reaction). The ECM-coated beads (ECB) served as scaffold to culture human intestinal epithelial cells (Int-407) in the RWV, which formed viable multi-layered cell aggregates and expressed epithelial differentiation markers. The cell aggregates remained viable following dissociation from the microcarriers, and could be returned to the RWV bioreactor for further culturing into bead-free tissue assemblies. The developed ECBs thus offer the potential to generate scaffold-free 3-D tissue assemblies, which could further be explored for tissue replacement and remodeling. PMID: 20692703 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Regeneration of the articular surface of the rabbit synovial joint by cell homing: a proof of concept study. Lancet. 2010 Aug 7;376(9739):440-448 Authors: Lee CH, Cook JL, Mendelson A, Moioli EK, Yao H, Mao JJ BACKGROUND: A common approach for tissue regeneration is cell delivery, for example by direct transplantation of stem or progenitor cells. An alternative, by recruitment of endogenous cells, needs experimental evidence. We tested the hypothesis that the articular surface of the synovial joint can regenerate with a biological cue spatially embedded in an anatomically correct bioscaffold. METHODS: In this proof of concept study, the surface morphology of a rabbit proximal humeral joint was captured with laser scanning and reconstructed by computer-aided design. We fabricated an anatomically correct bioscaffold using a composite of poly-varepsilon-caprolactone and hydroxyapatite. The entire articular surface of unilateral proximal humeral condyles of skeletally mature rabbits was surgically excised and replaced with bioscaffolds spatially infused with transforming growth factor beta3 (TGFbeta3)-adsorbed or TGFbeta3-free collagen hydrogel. Locomotion and weightbearing were assessed 1-2, 3-4, and 5-8 weeks after surgery. At 4 months, regenerated cartilage samples were retrieved from in vivo and assessed for surface fissure, thickness, density, chondrocyte numbers, collagen type II and aggrecan, and mechanical properties. FINDINGS: Ten rabbits received TGFbeta3-infused bioscaffolds, ten received TGFbeta3-free bioscaffolds, and three rabbits underwent humeral-head excision without bioscaffold replacement. All animals in the TGFbeta3-delivery group fully resumed weightbearing and locomotion 3-4 weeks after surgery, more consistently than those in the TGFbeta3-free group. Defect-only rabbits limped at all times. 4 months after surgery, TGFbeta3-infused bioscaffolds were fully covered with hyaline cartilage in the articular surface. TGFbeta3-free bioscaffolds had only isolated cartilage formation, and no cartilage formation occurred in defect-only rabbits. TGFbeta3 delivery yielded uniformly distributed chondrocytes in a matrix with collagen type II and aggrecan and had significantly greater thickness (p=0.044) and density (p<0.0001) than did cartilage formed without TGFbeta3. Compressive and shear properties of TGFbeta3-mediated articular cartilage did not differ from those of native articular cartilage, and were significantly greater than those of cartilage formed without TGFbeta3. Regenerated cartilage was avascular and integrated with regenerated subchondral bone that had well defined blood vessels. TGFbeta3 delivery recruited roughly 130% more cells in the regenerated articular cartilage than did spontaneous cell migration without TGFbeta3. INTERPRETATION: Our findings suggest that the entire articular surface of the synovial joint can regenerate without cell transplantation. Regeneration of complex tissues is probable by homing of endogenous cells, as exemplified by stratified avascular cartilage and vascularised bone. Whether cell homing acts as an adjunctive or alternative approach of cell delivery for regeneration of tissues with different organisational complexity warrants further investigation. FUNDING: New York State Stem Cell Science; US National Institutes of Health. PMID: 20692530 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | In-vivo tissue engineering of biological joint replacements. Lancet. 2010 Aug 7;376(9739):394-6 Authors: Warnke PH PMID: 20692514 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Emerging nanotechnology approaches in tissue engineering for peripheral nerve regeneration. Nanomedicine. 2010 Aug 5; Authors: Cunha C, Panseri S, Antonini S Effective nerve regeneration and functional recovery subsequent to peripheral nerve injury is still a clinical challenge. Autologous nerve graft transplantation is a feasible treatment in several clinical cases, but limited by donor site morbidity and insufficient donor tissue, impairing complete functional recovery. Tissue engineering has introduced innovative approaches to promote and guide peripheral nerve regeneration by using biomimetic conduits creating favorable microenvironments for nervous ingrowth, but despite a plethora of nerve prostheses developed, nowadays few approaches entered into the clinic. Promising strategies using nanotechnology have recently been proposed, such as the use of scaffolds with functionalized cell binding domains, the use of guidance channels with cell-scale internal oriented fibers and the possibility of sustained release of neurotrophic factors. This review addresses the fabrication, advantages, drawbacks and results achieved by the most recent nanotechnology approaches in view of future solutions for peripheral nerve repair. PMID: 20692373 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Allogeneic administration of fetal membrane-derived mesenchymal stem cells attenuates acute myocarditis in rats. J Mol Cell Cardiol. 2010 Aug 5; Authors: Ishikane S, Yamahara K, Sada M, Harada K, Kodama M, Ishibashi-Ueda H, Hayakawa K, Mishima K, Iwasaki K, Fujiwara M, Kangawa K, Ikeda T We reported previously that the autologous administration of bone marrow-derived mesenchymal stem cells (BM-MSC) significantly attenuated myocardial dysfunction and injury in a rat model of acute myocarditis by stimulating angiogenesis and reducing inflammation. Because BM aspiration procedures are invasive and can yield low numbers of MSC after processing, we focused on fetal membranes (FMs) as an alternative source of MSC to provide a large number of cells. We investigated whether the allogeneic administration of FM-derived MSC (FM-MSC) attenuates myocardial injury and dysfunction in a rat myocarditis model. Experimental autoimmune myocarditis (EAM) was induced in male Lewis rats by injecting porcine cardiac myosin. Allogeneic FM-MSC obtained from major histocompatibility complex-mismatched ACI rats (5x10(5) cells/animal) were injected intravenously into Lewis rats one week after myosin administration. At day 21, severe cardiac inflammation and deterioration of cardiac function were observed. The allogeneic administration of FM-MSC significantly attenuated inflammatory cell infiltration and monocyte chemoattractant protein 1 expression in the myocardium and improved cardiac function. In a T-lymphocyte proliferation assay, the proliferative response of splenic T lymphocytes was significantly lower in cells obtained from FM-MSC-treated EAM rats that reacted to myosin than in cells obtained from vehicle-treated rats with EAM. T-lymphocyte activation was significantly reduced by coculture with FM-MSC. The allogeneic administration of FM-MSC attenuated myocardial dysfunction and inflammation, and the host cell-mediated immune response was attenuated in a rat model of acute myocarditis. These results suggest that allogeneic administration of FM-MSC might provide a new therapeutic strategy for the treatment of acute myocarditis. PMID: 20692268 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Lessons for cardiac regeneration and repair through development. Trends Mol Med. 2010 Aug 2; Authors: Alexander JM, Bruneau BG Cell-based regenerative strategies have the potential to revolutionize the way cardiovascular injury is treated, but successful therapies will require a precise understanding of the mechanisms that dictate cell fate, survival and differentiation. Recent advances in the study of cardiac development hold promise for unlocking the keys for successful therapies. Using mouse models and embryonic stem cells, researchers are uncovering cardiac progenitor cells in both embryonic and adult contexts. Furthermore, the signaling molecules and transcriptional regulators that govern these cells and their behavior are being revealed. Here, we focus on the recent advances in these areas of cardiac developmental research and their impact on the expanding field of regenerative medicine. PMID: 20692205 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Studies on neuronal differentiation and signalling processes with a novel impedimetric biosensor. Biosens Bioelectron. 2010 Jul 23; Authors: Valero T, Moschopoulou G, Kintzios S, Hauptmann P, Naumann M, Jacobs T The differentiation of neural cells is an important process during the development of the central nervous system. Studies on the mechanisms of the differentiation process is of special importance, e.g. in the field of regenerative medicine. In this contribution the cellular differentiation of gel matrix embedded neuronal cells was studied. The three-dimensional organization of neuronal cells represents a new cellular model system closer to the physiology than conventional two-dimensional cell cultures. Neuro2a (N2a) neuroblastoma cells were immobilized in different gel matrices and the grade of differentiation was compared. Furthermore, the impact of the cell number and selected differentiation factors were analyzed. Experimental results revealed that gel matrices based on collagen-laminin mixtures in contact with serum free medium enable neural differentiation. Therefore, collagen-laminin gels appear as a suitable three-dimensional model for drug screening in developmental neurobiology. Following optimization of the immobilization process, a novel impedimetric sensor and electrical impedance spectroscopy technique was applied to on-line monitor the differentiation process by means of changes in the dielectric and conductive properties. Experimental results showed an increase in the impedance magnitude that can be mainly attributed to differentiating cells accompanied by an increase in the specific resistivity of the bare gel mixture. PMID: 20692153 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Stem Cells and DNA Damage: Persist or Perish? Cell. 2010 Aug 6;142(3):360-362 Authors: Lane AA, Scadden DT Stem cells repopulate tissues after injury while also renewing themselves, but this makes them vulnerable to genotoxic damage. Mohrin et al. (2010) and Milyavsky et al. (2010) now show that mouse and human hematopoietic stem cells make opposing decisions about whether to die or to persist in response to DNA damage. PMID: 20691895 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Lung Organogenesis. Curr Top Dev Biol. 2010;90C:73-158 Authors: Warburton D, El-Hashash A, Carraro G, Tiozzo C, Sala F, Rogers O, Langhe SD, Kemp PJ, Riccardi D, Torday J, Bellusci S, Shi W, Lubkin SR, Jesudason E Developmental lung biology is a field that has the potential for significant human impact: lung disease at the extremes of age continues to cause major morbidity and mortality worldwide. Understanding how the lung develops holds the promise that investigators can use this knowledge to aid lung repair and regeneration. In the decade since the "molecular embryology" of the lung was first comprehensively reviewed, new challenges have emerged-and it is on these that we focus the current review. Firstly, there is a critical need to understand the progenitor cell biology of the lung in order to exploit the potential of stem cells for the treatment of lung disease. Secondly, the current familiar descriptions of lung morphogenesis governed by growth and transcription factors need to be elaborated upon with the reinclusion and reconsideration of other factors, such as mechanics, in lung growth. Thirdly, efforts to parse the finer detail of lung bud signaling may need to be combined with broader consideration of overarching mechanisms that may be therapeutically easier to target: in this arena, we advance the proposal that looking at the lung in general (and branching in particular) in terms of clocks may yield unexpected benefits. PMID: 20691848 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Preparation of three-dimensional interconnected macroporous cellulosic hydrogels for soft tissue engineering. Biomaterials. 2010 Aug 4; Authors: Yue Z, Wen F, Gao S, Ang MY, Pallathadka PK, Liu L, Yu H This work exploits the thermal responsive phase behavior of hydroxypropylcellulose to produce 3D interconnected macroporous hydrogels in aqueous environment. Hydroxypropylcellulose was modified with allyl isocyanate, and their temperature mediated phase behavior was studied as a function of degree of modification (DS). A derivative with a DS of 1.5 was selected for scaffold preparation. Its aqueous solutions were warmed up to trigger the formation of biphasic systems. Such state was then immobilized efficiently by gamma-ray irradiated crosslinking. Lyophilization of the crosslinked hydrogels yielded 3D macroporous sponges. The re-hydrated gels demonstrate a combination of interconnected macroporosity, high water content and mechanical integrity to soft tissues. Cytocompatibility was demonstrated among various cell types, and in vivo biocompatibility test showed minimal inflammatory response within 12 weeks' subcutaneous implantation in mice. The potential applications of these macroporous hydrogels in tissue engineering are discussed. PMID: 20691470 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Biodegradable and synthetic membranes for the expansion and functional differentiation of rat embryonic liver cells. Acta Biomater. 2010 Aug 3; Authors: Piscioneri A, Campana C, Salerno S, Morelli S, Bader A, Giordano F, Drioli E, Bartolo LD Insufficient donor organs for orthotopic liver transplantation worldwide have urgently increased the requirement for new therapies for acute and chronic liver disease. The creation of unlimited source of donor cells for hepatocyte transplantation therapy and pharmaceutical applications may be the isolation and expansion of liver progenitor cells or stem cells. Here we report the expansion and the functional differentiation of rat embryonic liver cells on biodegradable and synthetic polymeric membranes in comparison with traditional substrates such as collagen and polystyrene culture dishes. Membranes prepared from chitosan and modified polyetheretherketone were used for the culture of liver progenitor cells derived from rat embryonic liver. Cells proliferated generating a significant increase in their number within 8-11 days. On all investigated substrates cells gained a functional differentiation showing the urea synthesis, albumin production and diazepam biotransformation. In particular on chitosan membrane the liver specific functions were expressed at significant higher levels for prolonged time compared to other synthetic membrane, utilizing the traditional substrates (collagen and PSCD) as references. These results demonstrate that chitosan membrane offered to cells favourable conditions to promote the expansion and functional differentiation of embryonic liver cells that could be effectively used in liver tissue engineering and in pharmaceutical applications. PMID: 20691288 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Highly efficient osteogenic differentiation of human mesenchymal stem cells by eradication of STAT3 signaling. Int J Biochem Cell Biol. 2010 Aug 3; Authors: Levy O, Ruvinov E, Reem T, Granot Y, Cohen S Human bone marrow-derived mesenchymal stem cells (hMSCs) are promising candidates for cellular therapy owing to their multipotency to differentiate into several cell lineages. Elucidating the signaling events involved in the response of hMSCs to diverse stimulants affecting their differentiation may considerably promote their clinical use. In this study, we attempted to illuminate the molecular signaling networks involved in bone morphogenetic protein (BMP)-stimulated hMSC osteogenic differentiation. We demonstrate that eradication of signal transducers and activators of transcription (STAT) signaling considerably enhances BMP-induced osteogenic differentiation of hMSCs. BMP 2 and 4 are shown for the first time to activate the Janus-activated kinase (JAK)-STAT pathway in hMSC. Specifically, we reveal that JAK2 mediates STAT3 tyrosine phosphorylation in response to the two BMPs, whereas BMP2- and BMP4-induced STAT3 serine phosphorylation involves two divergent cascades, namely the mTOR and ERK1/2 cascades, respectively. Furthermore, elimination of the STAT3 signaling pathway by the inhibitors, AG490 or STAT3 siRNA, results in the acceleration and augmentation of BMPs-induced osteogenic differentiation, thus proposing a role for JAK-STAT signaling as a negative regulator of this process in MSCs. We believe that the findings presented in this study may be the basis for the development of a useful strategy to better control stem cell fate through intervention in molecular signaling networks. Hopefully, such a strategy will include the development of more efficient and controllable protocols for hMSC differentiation and facilitate their use in regenerative medicine. PMID: 20691278 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | An economical single-sided antibody incubation method for Western blotting. J Virol Methods. 2010 Aug 3; Authors: Pan C, Lan X, Chen H, Bishop CE A simple, single-sided antibody method for incubating primary and secondary antibodies in Western blotting was developed, which generates significant savings on the use of antibodies. Compared with the conventional immersion technique for antibody incubation, the present economical single-sided antibody incubation method resulted in a saving of 80% of antibody use. Besides, the present incubation method did not compromise the Western blot results and was not affected by the expression levels of target proteins. PMID: 20691216 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Leptin gene therapy in the fight against diabetes. Expert Opin Biol Ther. 2010 Aug 9; Authors: Wang Y, Asakawa A, Inui A, Kosai KI Importance of the field: The incidence of diabetes is increasing worldwide, yet current treatments are not always effective for all patient or disease types. Areas covered in this review: Here, we summarize the biologic and clinical roles of leptin in diabetes, and discuss candidate viral vectors that may be employed in the clinical use of central leptin gene therapy for diabetes. What the reader will gain: We discuss how studies on leptin, a regulator of the insulin-glucose axis, have significantly advanced our understanding of the roles of energy homeostasis and insulin resistance in the pathogeneses of metabolic syndrome and diabetes. Recent studies have demonstrated the long-term therapeutic effects of central leptin gene therapy in obesity and diabetes via decreased insulin resistance and increased glucose metabolism. Many of these studies have employed viral vectors, which afford high in vivo gene transduction efficiencies compared with non-viral vectors. Take home message: Adeno-associated viral vectors are particularly well suited for central leptin gene therapy owing to their low toxicity and ability to drive transgene expression for extended periods. PMID: 20690892 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Biodegradability and biocompatibility of thermoreversible hydrogels formed from mixing a sol and a precipitate of block copolymers in water. Biomacromolecules. 2010 Aug 9;11(8):2169-78 Authors: Yu L, Zhang Z, Zhang H, Ding J This study examines in vitro and in vivo biodegradation and biocompatibility of a thermogelling polymeric material, which we call a mixture hydrogel. The mixture contains two ABA-type triblock copolymers poly(d,l-lactic acid-co-glycolic acid)-b-poly(ethylene glycol)-b-poly(d,l-lactic acid-co-glycolic acid) (PLGA-PEG-PLGA) with different block ratios, and one polymer is soluble in water, but the other is not. The aqueous solutions of some mixtures with appropriate mix ratios form hydrogels at the body temperature. The degradation of mixture hydrogels proceeded by hydrolysis of ester bonds followed by the erosion of gel in phosphate saline buffer solution at 37 degrees C for nearly one month. The mass loss and reduction of molecular weight were detected. The mix ratio was found to significantly influence the degradation profiles. The rapid in vivo gel formation was confirmed after subcutaneous injection of the thermogelling copolymer mixtures into Sprague-Dawley rats. The in vivo degradation was a bit accelerated than in vitro hydrolysis, and the persistence time of injected hydrogels in vivo was found to be tuned by mix ratio. MTT assay and histological observations were used to examine the copolymer mixtures. Both in vitro and in vivo results illustrate acceptable biocompatibility of our materials. As such, the thermosensitive hydrogel of copolymer mixture is confirmed to be a promising candidate of an injectable biomaterial for drug delivery and tissue engineering. PMID: 20690723 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Templated mineralization of synthetic hydrogels for bone-like composite materials: role of matrix hydrophobicity. Biomacromolecules. 2010 Aug 9;11(8):2060-8 Authors: Phadke A, Zhang C, Hwang Y, Vecchio K, Varghese S Bone-mimetic mineral-polymer composite materials have several applications ranging from artificial bone grafts to scaffolds for bone tissue engineering; templated mineralization is an effective approach to fabricate such composites. In this study, we synthesized bone-like composites using synthetic hydrogels having pendant side chains terminating with carboxyl groups as a template for mineralization. The role of matrix hydrophobicity on mineralization was examined using poly(ethylene glycol) hydrogels modified with varying lengths of anionic pendant side chains (CH(2) horizontal lineCHCONH(CH(2))(n)COOH, where n = 1, 3, 5, and 7). The ability of these hydrogels to undergo templated mineralization was found to be strongly dependent upon the length of the pendant side chain as is evident from the extent of calcification and morphology of the minerals. Moreover, mineralized phases formed on the hydrogels were confirmed to resemble apatite-like structures. In addition to demonstrating the importance of material hydrophobicity as a design parameter for the development of bone-like synthetic materials, our study also provides a potential explanation for the in vitro differences between the apatite-nucleating capacity of aspartate-rich osteopontin and glutamate-rich bone sialoprotein. PMID: 20690714 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Characterization of tunable fgf-2 releasing polyelectrolyte multilayers. Biomacromolecules. 2010 Aug 9;11(8):2053-9 Authors: Macdonald ML, Rodriguez NM, Shah NJ, Hammond PT Fibroblast growth factor 2 (FGF-2) is a potent mediator of stem cell differentiation and proliferation. Although FGF-2 has a well-established role in promoting bone tissue formation, flaws in its delivery have limited its clinical utility. Polyelectrolyte multilayer films represent a novel system for FGF-2 delivery that has promise for local, precisely controlled, and sustained release of FGF-2 from surfaces of interest, including medical implants and tissue engineering scaffolds. In this work, the loading and release of FGF-2 from synthetic hydrolytically degradable multilayer thin films of various architectures is explored; drug loading was tunable using at least three parameters (number of nanolayers, counterpolyanion, and type of degradable polycation) and yielded values of 7-45 ng/cm(2) of FGF-2. Release time varied between 24 h and approximately five days. FGF-2 released from these films retained in vitro activity, promoting the proliferation of MC3T3 preosteoblast cells. The use of biologically derived counterpolyanions heparin sulfate and chondroitin sulfate in the multilayer structures enhanced FGF-2 activity. The control over drug loading and release kinetics inform future in vivo bone and tissue regeneration models for the exploration of clinical relevance of LbL growth factor delivery films. PMID: 20690713 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Immobilization of Human Mesenchymal Stem Cells within RGD-Grafted Alginate Microspheres and Assessment of Their Angiogenic Potential. Biomacromolecules. 2010 Aug 9;11(8):1956-64 Authors: Bidarra SJ, Barrias CC, Barbosa MA, Soares R, Granja PL In this work, human mesenchymal stem cells (hMSC) immobilized in RGD-coupled alginate microspheres, with a binary composition of high and low molecular weight alginate, were investigated. Cells immobilized within RGD-alginate microspheres (during 21 days) showed metabolic activity, with an overall viability higher than 90%, short cell extensions, and, when induced, they were able to differentiate into the osteogenic lineage. In osteogenic conditions (comparing to basal conditions), immobilized cells presented alkaline phosphatase (ALP) activity and an upregulation of ALP, collagen type I, and Runx 2 expression. Moreover, mineralization was also detected in immobilized cells under osteogenic stimulus. In addition, it was demonstrated for the first time that MSCs immobilized in this 3D matrix were able to enhance the ability of neighboring endothelial cells to form tubelike structures. Overall, these findings represent a step forward in the development of injectable stem cell carriers for bone tissue engineering. PMID: 20690708 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Investigation of pore size effect on chondrogenic differentiation of adipose stem cells using a pore size gradient scaffold. Biomacromolecules. 2010 Aug 9;11(8):1948-55 Authors: Oh SH, Kim TH, Im GI, Lee JH In this study, polycaprolactone (PCL) cylindrical scaffolds with gradually increasing pore size along the longitudinal direction were fabricated by the centrifugation and thermal fibril-bonding process. The fabricated PCL scaffold showed a gradual increasing pore size (from approximately 90 to approximately 400 mum) and porosity (from approximately 80 to approximately 97%) along the cylindrical axis. The pore size gradient PCL scaffold was used to investigate the effect of pore size on the chondrogenic differentiation of adipose stem cells (ASCs). From the in vitro culture of ASCs, it was observed that the scaffold section having a pore size range of 370-400 mum provided a more favorable environment for chondrogenic differentiation than other pore size groups. The pore size gradient scaffolds can be a good tool for the systematic study of determining optimum pore size ranges for a variety of stem cell differentiation to a specific cell type. PMID: 20690707 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Influence of Gelatin Cues in PCL Electrospun Membranes on Nerve Outgrowth. Biomacromolecules. 2010 Aug 6; Authors: Alvarez-Perez MA, Guarino V, Cirillo V, Ambrosio L The design of functionalized polymers that can elicit specific biological responses and the development of methods to fabricate new devices that incorporate biological cues are of great interest to the biomedical community. The realization of nanostructured matrices that exhibit biological properties and that comprise fibers with diameters of similar scale to those of the natural extracellular matrix (ECM) would enable the provision of tailored materials for tissue engineering. Accordingly, the goal of this work is to create a biologically active functionalized electrospun matrix capable of guiding neurite growth for the regeneration of nerve tissue. In this study, nanoscale electrospun membranes made of poly epsilon-caprolactone enhanced with gelatin from calf skin were investigated to validate their biological response under in vitro culture of PC-12 nerve cells. Preliminary observations from SEM studies supported by image analysis highlighted the nanoscale texture of the scaffold with fiber diameters equal to 0.548 +/- 0.140 mum. In addition, contact angle measurements confirmed the hydrophilic behavior of the membranes, ascribable to the gelatin content. We demonstrate that the balance of morphological and biochemical properties improves all the fundamental biological events of nerve regeneration, enhancing cell adhesion, proliferation, and differentiation in comparison with PCL nanofibrous scaffolds, as well as supporting the neurite outgrowth. PMID: 20690634 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Directing the Morphology and Differentiation of Skeletal Muscle Cells Using Oriented Cellulose Nanowhiskers. Biomacromolecules. 2010 Aug 6; Authors: Dugan JM, Gough JE, Eichhorn SJ Radially oriented submonolayer surfaces of 10-15 nm diameter cellulose nanowhiskers (CNWs) were prepared by spin-coating. The response of myoblasts (muscle cells) to the surfaces was assessed using atomic force microscopy (AFM), immunocytochemistry, and image analysis. Despite the small size of the CNWs, the myoblasts oriented along the CNW surfaces. Upon differentiation, the myoblasts produced striking radial patterns of myotubes, following the radial pattern of the CNWs. This facile method of nanopatterning surfaces may be applied where the directed growth of tissue is required and shows for the first time the potential of CNWs for tissue engineering applications. PMID: 20690614 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | State of the art of Micro-CT applications in dental research. Int J Oral Sci. 2009 Dec;1(4):177-88 Authors: Swain MV, Xue J This review highlights the recent advances in X-ray microcomputed tomography (Micro-CT) applied in dental research. It summarizes Micro-CT applications in measurement of enamel thickness, root canal morphology, evaluation of root canal preparation, craniofacial skeletal structure, micro finite element modeling, dental tissue engineering, mineral density of dental hard tissues and about dental implants. Details of studies in each of these areas are highlighted along with the advantages of Micro-CT, and finally a summary of the future applications of Micro-CT in dental research is given. PMID: 20690421 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Multilineage differentiation of dental pulp stem cells from green fluorescent protein transgenic mice. Int J Oral Sci. 2010 Mar;2(1):21-7 Authors: Grottkau BE, Purudappa PP, Lin YF AIM: The aim of this study was to confirm the multilineage differentiation ability of dental pulp stem cells (DPSCs) from green fluorescent protein (GFP) transgenic mice. The expression of GFP in DPSCs was also observed during differentiation. METHODOLOGY: DPSCs were harvested from the dental pulp tissue of transgenic nude mice, and then transferred to osteogenic, adipogenic, and chondrogenic media. The morphological characterization of induced cells was observed by microscopy and histological staining. The expression of marker genes was measured by RT-PCR. RESULTS: The endogenous GFP and multilineage potential of transgenic DPSCs had no influence on each other. Moreover, the results of fluorescence microscopic imaging suggest that there was no significant decline of GFP expression during DPSCs differentiation. CONCLUSION: As the population of GFP labeled DPSCs can be easily identified, this will be a promising method for tracking DPSCs in vivo. PMID: 20690415 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | An in vitro model of chronic wound biofilms to test wound dressings and assess antimicrobial susceptibilities. J Antimicrob Chemother. 2010 Jun;65(6):1195-206 Authors: Hill KE, Malic S, McKee R, Rennison T, Harding KG, Williams DW, Thomas DW OBJECTIVES: The targeted disruption of biofilms in chronic wounds is an important treatment strategy and the subject of intense research. In the present study, an in vitro model of chronic wound biofilms was developed to assess the efficacy of antimicrobial treatments for use in the wound environment. METHODS: Using chronic wound isolates, assays of bacterial coaggregation established that aerobic and anaerobic wound bacteria were able to coaggregate and form biofilms. A constant depth film fermenter (CDFF) was used to develop wound biofilms in vitro, which were analysed using light microscopy and scanning electron microscopy. The susceptibility of bacteria within these biofilms was examined in response to the most frequently prescribed 'chronic wound' antibiotics and a series of iodine- and silver-containing commercial antimicrobial products and lactoferrin. RESULTS: Defined biofilms were rapidly established within 1-2 days. Antibiotic treatment demonstrated that mixed Pseudomonas and Staphylococcus biofilms were not affected by ciprofloxacin (5 mg/L) or flucloxacillin (15 mg/L), even at concentrations equivalent to twice the observed peak serum levels. The results contrasted with the ability of povidone-iodine (1%) to disrupt the wound biofilm; an effect that was particularly pronounced in the dressing testing where iodine-based dressings completely disrupted established 7 day biofilms. In contrast, only two of six silver-containing dressings exhibited any effect on 3 day biofilms, with no effect on 7 day biofilms. CONCLUSIONS: This wound model emphasizes the potential role of the biofilm phenotype in the observed resistance to antibiotic therapies that may occur in chronic wounds in vivo. PMID: 20378671 [PubMed - indexed for MEDLINE] | | | | | | | | | | |  |
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