| |  | | | | | | | | | | | | | | | |  | | | | | | | | | | | | | | | | | | | | | | | | Proangiogenic scaffolds as functional templates for cardiac tissue engineering. Proc Natl Acad Sci U S A. 2010 Aug 9; Authors: Madden LR, Mortisen DJ, Sussman EM, Dupras SK, Fugate JA, Cuy JL, Hauch KD, Laflamme MA, Murry CE, Ratner BD We demonstrate here a cardiac tissue-engineering strategy addressing multicellular organization, integration into host myocardium, and directional cues to reconstruct the functional architecture of heart muscle. Microtemplating is used to shape poly(2-hydroxyethyl methacrylate-co-methacrylic acid) hydrogel into a tissue-engineering scaffold with architectures driving heart tissue integration. The construct contains parallel channels to organize cardiomyocyte bundles, supported by micrometer-sized, spherical, interconnected pores that enhance angiogenesis while reducing scarring. Surface-modified scaffolds were seeded with human ES cell-derived cardiomyocytes and cultured in vitro. Cardiomyocytes survived and proliferated for 2 wk in scaffolds, reaching adult heart densities. Cardiac implantation of acellular scaffolds with pore diameters of 30-40 mum showed angiogenesis and reduced fibrotic response, coinciding with a shift in macrophage phenotype toward the M2 state. This work establishes a foundation for spatially controlled cardiac tissue engineering by providing discrete compartments for cardiomyocytes and stroma in a scaffold that enhances vascularization and integration while controlling the inflammatory response. PMID: 20696917 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Surgical anatomy of the swine face. Lab Anim. 2010 Aug 9; Authors: Sasaki R, Watanabe Y, Yamato M, Aoki S, Okano T, Ando T In order to develop tissue engineering applications for oral and maxillofacial surgery, the surgical anatomy of the miniature pig's face was investigated in three miniature pig cadavers and three anaesthetized miniature pigs using identical procedures that were previously described for humans. A preauricular incision with a retromandibular and a submandibular extension was initially made through the facial skin and subcutaneous tissues. The underlying tissues were then carefully dissected in order to progressively expose the platysma muscle, the superficial layer of deep cervical fascia, the marginal mandibular branch and buccal branch of the facial nerve, the mental nerve and the mandibular skeleton. The marginal mandibular branch of the facial nerve has an upper and lower division. Stimulation of the facial nerve and its branches showed that the upper division of the marginal mandibular branch innervates muscles and tissues in the upper lip and nose region, and the lower division innervates muscles and tissues in the lower lip region. The gross anatomy of the maxillofacial region in the pigs was found to be similar to that of humans. Although the distributions of the marginal mandibular branch of the facial nerve and the mental nerve are different from that of humans, we concluded that miniature pigs are a suitable experimental model for the preclinical development of tissue engineering applications in oral and maxillofacial surgery. PMID: 20696789 [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] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Improvement of mechanical properties and biocompatibility of forsterite bioceramic addressed to bone tissue engineering materials. J Mech Behav Biomed Mater. 2010 Oct;3(7):530-537 Authors: Kharaziha M, Fathi MH This work deals with the fabrication and characterization of nanostructured forsterite bulk. This material may have better biocompatibility and mechanical properties than coarse grain forsterite for the development of bone tissue engineering materials. Nanostructured forsterite bulks were prepared by two step sintering of sol-gel derived forsterite nanopowder. Their sinterability and mechanical properties were then studied. Biocompatibility of the nanostructured forsterite bulk was also evaluated by cell attachment and proliferation experiments. In addition, the effects of ionic products from forsterite nanopowder dissolution on osteoblasts were studied. Results show that dense nanostructured forsterite bulk was prepared with hardness and fracture toughness of about 1102 Hv and 4.3 MPa m(1/2), respectively. Nanostructured forsterite was biocompatible and the MTT test confirmed that the products from forsterite nanopowder dissolution significantly promoted osteoblast proliferation within a certain concentration range. In addition, cells attached to and spread on the surface of nanostructured forsterite bulks. Mechanical properties of the nanostructured forsterite were much higher than that of hydroxyapatite. It was concluded that nanostructured forsterite is a bioactive ceramic with good biocompatibility that can be used as a bone tissue engineering material. PMID: 20696418 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | An ultrasound-responsive nano delivery system of tissue-type plasminogen activator for thrombolytic therapy. J Control Release. 2010 Aug 6; Authors: Uesugi Y, Kawata H, Jo JI, Saito Y, Tabata Y This study is undertaken to design a novel nano-sized delivery system of tissue-type plasminogen activator (t-PA) which has a suppressed thrombolytic activity of t-PA, but recovered the activity only when exposed to ultrasound. Various amounts of ethylenediamine were chemically introduced into gelatin (cationized gelatins) to complex with t-PA. To modify the surface of complexes with polyethylene glycol (PEG), PEG was chemically grafted to the anionic gelatin (PEG-gelatin). The simple mixing with the PEG-gelatin enabled the t-PA-cationized gelatin complex to form a nano-sized delivery complex with PEG chains on the surface. The t-PA activity of PEG-modified complexes was significantly suppressed to be 45% of original t-PA. However, when exposed to ultrasound in vitro, the t-PA activity was fully recovered. A cell culture experiment demonstrated no cytotoxicity of PEG-modified complexes. The body distribution study indicated that the half-life of t-PA in the blood circulation was prolonged about 3 times. In a rabbit thrombosis model, the intravenous administration of PEG-modified complexes followed by ultrasound irradiation resulted in complete recanalization, in remarked contrast to the complex administration alone. It is concluded that the PEG-modified complex is a promising t-PA delivery system to enhance the biological activity at the site necessary only by a local ultrasound irradiation. PMID: 20696194 [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] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Chimeric vessel tissue engineering driven by endothelialized modules in immunosuppressed Sprague-Dawley rats. Tissue Eng Part A. 2010 Aug 9; Authors: Chamberlain MD, Gupta R, Sefton MV Modular tissue engineering is a means of building functional, vascularized tissues using small (~1 mm long x 0.5 mm diameter) components. While this approach is being explored for its utility in adipose and cardiac tissue engineering and in islet transplantation, the initial question in this study was to assess the fate of the endothelial cells after transplantation delivered on the surface of modules, without an embedded cell. Rat aortic endothelial cell (RAEC) covered collagen gel modules were transplanted into the omental pouch of allogeneic (outbred) Sprague-Dawley rats with and without immunosuppressive drug treatment (atorvastatin and tacrolimus) for 3 to 60 days. There was a significant increase in vessel density at all time points in the drug treated rats as compared to untreated rats. Green fluorescent protein (GFP) positive donor RAEC migrated from the surface of the modules and formed primitive vessels by day 7. In the untreated rats, the GFP positive cells were not seen after day 7. In drug treated rats, GFP positive vessels matured over time, accumulated erythrocytes, were supported by host smooth muscle cells and formed chimeric vessels that survived until day 60. This resulted in the formation of a densely vascularised, perfusable network by day 60. To our knowledge, this is the first study that demonstrates that primary unmodified endothelial cells, without the addition of supporting cells, form a chimeric and stable vascular bed in allogeneic, albeit drug treated, animals. PMID: 20695789 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | CELL-DERIVED MATRIX ENHANCES OSTEOGENIC PROPERTIES OF HYDROXYAPATITE. Tissue Eng Part A. 2010 Aug 9; Authors: Tour G, Wendel M, Tcacencu I The study aimed to evaluate osteogenic properties of hydroxyapatite (HA) scaffold combined with extracellular matrix (ECM) derived in vitro from rat primary calvarial osteoblasts or dermal fibroblasts. The cellular viability, and the ECM deposited onto synthetic HA microparticles were assessed by MTT, Glycosaminoglycan, and Hydroxyproline assays as well as immunohistochemistry and scanning electron microscopy after 21 days of culture. The decellularized HA-ECM constructs were implanted in critical-sized calvarial defects of Sprague-Dowley rats, followed by bone repair and local inflammatory response assessments by histomorphometry and immunohistochemistry at 12 weeks postoperatively. We demonstrated that HA supported cellular adhesion, growth and ECM production in vitro, and the HA-ECM constructs significantly enhanced calvarial bone repair (p<0.05, Mann-Whitney U test), compared to HA alone, despite the significantly increased number of CD68+ macrophages, and foreign body giant cells (p<0.05, Mann-Whitney U test). Selective accumulation of BSP, OPN and periostin was observed at the tissue-HA interfaces. In conclusion, in vitro-derived ECM mimics the native bone matrix, enhances the osteogenic properties of the HA microparticles, and might modulate the local inflammatory response in a bone repair-favorable way. Our findings highlight the ability to produce functional HA-ECM constructs for bone tissue engineering applications. PMID: 20695777 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Guiding epithelial cell phenotypes with engineered integrin specific recombinant fibronectin fragments. Tissue Eng Part A. 2010 Aug 9; Authors: Barker TH, Brown AC, Rowe J The extracellular matrix (ECM) provides important cues for directing cell phenotypes. Cells interact with underlying ECM through cell-surface receptors known as integrins, which bind to specific sequences on their ligand(s). During tissue development, repair, and regeneration of epithelial tissues, cells must interact with an interstitial fibronectin (Fn)-rich matrix which has been shown to direct a more migratory/repair phenotype, presumably through interaction with Fn's cell binding domain comprised of both synergy (PHSRN) and RGD sequences. We hypothesized that the Fn synergy site is critical to the regulation of epithelial cell phenotype by directing integrin specificity. Epithelial cells were cultured on Fn fragments displaying stabilized synergy and RGD (FnIII9'10), or RGD alone (FnIII10) and cell phenotype analyzed by cytoskeleton changes, epithelial cell-cell contacts, changes in gene expression of epithelial and mesenchymal markers, and wound healing assay. Data indicate that epithelial cells engage RGD only with alphav integrins and display a significant shift toward a mesenchymal phenotype due, in part, to enhanced TGFbeta activation and/or signaling compared to cells on the synergy containing FnIII9'10. These studies demonstrate the importance of synergy in regulating epithelial cell phenotype relevant to tissue engineering as well as the utility of engineered integrin-specific ECM fragments in guiding cell phenotype. PMID: 20695776 [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] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Osteogenic potential of mesenchymal stem cells on expanded polytetrafluoroethylene coated with both a poly-amino-acid urethane copolymer and collagen. Tissue Eng Part A. 2010 Aug 9; Authors: Matsumoto T, Hattori K, Matsushima A, Tadokoro M, Yagyuu T, Kodama M, Sato J, Ohgushi H We have developed an expanded polytetrafluoroethylene polymer (e-P) coated with both poly-amino-acid urethane copolymer and collagen (e-PPC) to be used for cell culture substrata. Rat mesenchymal stem cells (MSC) were cultured on the e-P and e-PPC polymers in the presence of dexamethasone. After 24 hours, the MSC well contacted to the e-PPC surface and after 14 days, the MSC showed high levels of alkaline phosphatase activity, calcium and bone specific osteocalcin protein deposition. The MSC cultured on these polymers for one week were then implanted at rat subcutaneous sites and harvested after 4 weeks. Micro-computed tomography as well as histological analyses showed that any hard tissue could not be seen in implants of the MSC/e-P composites, whereas new bone formation could be detected in the MSC/e-PPC composites. These in vitro as well as in vivo results confirmed the importance of polymer surface to support the osteogenic differentiation, which resulted in new bone formation. The surface modification using poly-amino-acid urethane copolymer and collagen together with tissue engineering technology might facilitate bone anchoring to the polymers for dental and orthopedic applications. PMID: 20695773 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Investigation of the regenerative capacity of an acellular porcine medial meniscus for tissue engineering applications. Tissue Eng Part A. 2010 Aug 9; Authors: Stapleton TW, Ingram J, Fisher J, Ingham E Previously, we have described the development of an acellular porcine meniscal scaffold. The aims of this study were to determine the immunocompatibility of the scaffold and capacity for cellular attachment and infiltration in order to gain insight into its potential for meniscal repair and replacement. Porcine menisci were decellularized by exposing the tissue to freeze thaw cycles, incubation in hypotonic tris buffer, 0.1 % (w/v) sodium dodecyl sulphate in hypotonic buffer plus protease inhibitors, nucleases, hypertonic buffer followed by disinfection using 0.1% (v/v) peracetic and final washing in phosphate-buffered saline. <i>In vivo</i> immunocompatibility was assessed following implantation of the acellular meniscal scaffold subcutaneously into GTKO mice for 3 months in comparison to fresh and acellular tissue treated with alpha-galactosidase (negative control). The cellular infiltrates in the explants were assessed by histology and characterized using monoclonal antibodies against: CD3, CD4, CD34, F4/80 and C3c. Static culture was used to assess the potential of acellular porcine meniscal scaffold to support the attachment and infiltration of primary human dermal fibroblasts and primary porcine meniscal cells in vitro. The explants were surrounded by capsules which were more pronounced for the fresh meniscal tissue compared to the acellular tissues. Cellular infiltrates compromised mononuclear phagocytes, CD34 positive cells and non-labelled fibroblastic cells. T-lymphocytes were sparse in all explanted tissue types and there was no evidence of C3c deposition. The analysis revealed an absence of a specific immune response to all of the implanted tissues. Acellular porcine meniscus was shown to be capable of supporting the attachment and infiltration of primary human fibroblasts and primary porcine meniscal cells. In conclusion, acellular porcine meniscal tissue exhibits excellent immunocompatibility and potential for cellular regeneration in the longer term. PMID: 20695759 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [Research progress of tissue engineered nerve grafts] Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2010 Jul;24(7):860-3 Authors: Gu X OBJECTIVE: To summarize the recent advance in the research of tissue engineered nerve grafts. METHODS: The clinical and experimental research papers about tissue engineered nerve grafts were extensively reviewed and analyzed. RESULTS: The porosity, mechanical property and surface topography of a nerve scaffold, which was either made up of natural biodegradable polymers or synthetic polyesters, were pivotal factors that influenced the capacity of the scaffold in supporting nerve regeneration. Of various candidate supporting cells for nerve tissue engineering, the bone marrow mesenchymal stem cells had been paid more attention because of their advantages. Several model designs of drug delivery systems for controlled release of growth factors had been attempted. In clinical settings, short nerve gaps were demonstrated to be treatable with several nerve conduits which were commercially available, with functional recovery approximating to nerve autografting. CONCLUSION: The field of nerve tissue engineering has witnessed a rapid development not only in experimental research but also in clinical application. PMID: 20695386 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [Research and application of functional tissue engineered tendons] Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2010 Jul;24(7):850-3 Authors: Ning L, Qin T OBJECTIVE: To review the research and application of functional tissue engineered tendons (FTETs). METHODS: Recent literature concerning the research of FTETs was reviewed and analyzed. RESULTS: Functional tissue engineering (FTE) was a new approach that placed an emphasis on the importance of mechanical stress in determining the success of tissue engineered constructs and the effect of tissue remodeling in vivo. The concept of FTE was introduecd into the research of tissue engineered tendons; by measuring in vivo loads of normal tendon and using the information as a guideline, the appropriate tissue engineered tendon which can withstand in vivo loads was designed. It would be possible to solve the problems that the biomechanical function of the tissue engineered tendons could not meet the requirements of the loading environment in vivo. CONCLUSION: FTETs have a more promising future for the treatment of tendon defects. PMID: 20695384 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [Differentiation of human embryonic stem cells into hepatocyte-like cells in an adherent culture system with single-step induction] Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2010 Jul;24(7):838-42 Authors: Hu Z, Luo M, Liang D OBJECTIVE: To establish a new induction method from human embryonic stem cells (hESCs) differentiating into hepatocyte-like cells using an adherent culture system with single-step induction. METHODS: Undifferentiated hESCs were cultured on Matrigel-coated culture plates for 4 days, hepatic differentiation was initiated at 60%-70% confluence by adding Activin A for 5 days. Then the induction medium was replaced by hepatocyte induction medium (HIM) supplemented with fibroblast growth factor 1 (FGF-1) and bone morphogenetic protein 4 (BMP-4) for another 6 days. Finally, the cells were treated with HIM adding hepatocyte growth factor (HGF) and Oncostatin M (OSM) for 5-7 days. The characteristics of differentiated cells were determined by morphology, immunofluorescence staining, RT-PCR, and Periodic acid-Schiff (PAS) test. RESULTS: Differentiated cells treated with Activin A, FGF-1, BMP-4, HGF, and OSM sequentially were morphologically larger and became spherical, oval or polygon. Some cells had 2 or 3 nuclei, suggesting that the cells have a hepatocyte-like morphology. Differentiated cells at first induction stage could be stained positive by SOX17 and Forkhead (FOX)A2 after induction by Activin A. Then they turned to be a fetoprotein (AFP) and al antitripsin positive cells at second induction stage after induction by FGF-1 and BMP-4. Finally, the differentiated cells treated with HGF and OSM showed PAS positive for glycogen detection. The differentiated cells at various stages also expressed at early (SOX17, FOXA2, and GATA-4), middle (AFP, albumin, and cytokeratin 18), and mature (alcohol dehydrogenase 1C and Cytochrome P4501B1) stage hepatic genes, respectively. CONCLUSION: Using a simple-step induction method and by supplied with cytokines consequently, hESCs can be induced to differentiate into hepatocyte-like cells. The differentiation method can provide seed cells for hepatic tissue engineering or cell-therapy. PMID: 20695382 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [An experimental study on rabbit bone marrow mesenchymal stem cells double-labeled by PKH26 and 5-bromo-2'-deoxyuriding in vitro and application in cardiac patch] Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2010 Jul;24(7):828-33 Authors: Zhang J, Zhi W, Tan M, Chen X, Li X, Deng L OBJECTIVE: To study the biological characteristic of rabbit bone marrow mesenchymal stem cells (BMSCs) double-labeled by PKH26 and BrdU in vitro, and to construct tissue engineered cardiac patch in vitro. METHODS: The BMSCs were harvested from 6-month-old New Zealand rabbits and labeled with PKH26 and BrdU. The growth and fluorescent intensity were observed by inverted phase contrast microscope, fluorescent microscope, flow cytometry, and MTT detection. The characteristics of double-labeled BMSCs differentiating into osteoblasts and adipocytes, respectively, in vitro were identified by alkaline phosphatase (ALP) staining, Alizarin red staining, Oil red O staining, immunocytochemical technique of collagen type I, and osteocalcin expression. The labeled BMSCs were seeded on the small intestinal submucosa (SIS) and co-cultured for 5-7 days to construct tissue engineered cardiac patch. The patches were tested by inverted phase contrast microscope, fluorescent microscope, scanning electron microscope, and HE staining to observe the cell proliferation. RESULTS: The double-labeled cells grew well and showed red fluorescence. There was no significant difference in the growth characteristic between the labeled and unlabeled cells. There was no significant difference in the expression of stem cell specific surface antigen between before labeling and after labeling. After osteogenic induction of labeled BMSCs, ALP staining and Alizarin red staining were positive, and the cells expressed collagen type I and osteocalcin. After adipocytes induction, lipid droplets could be observed in cytoplasm by Oil red O staining. After the co-culture in vitro for 5-7 days, the double-labeled cells grew well, showing a multi-layer cellular structure on the surface of SIS. CONCLUSION: Rabbit BMSCs can be double-labeled with PKH26 and BrdU stably. The labeled cells still have the potential of self-renewal ability and multi potent differentiation ability; tissue engineered cardiac patch can be constructed by co-culturing labeled BMSCs and SIS in vitro. PMID: 20695380 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [Strain-induced tenogenic differentiation of bone marrow mesenchymal stem cells] Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2010 Jul;24(7):817-21 Authors: Liao M, Ning L, Chen X, Li X, Luo J, Qin T OBJECTIVE: To study the possibility of bone marrow mesenchymal stem cells (BMSCs) differentiation into tenocytes (TCs) under strain stimulation by co-culture of BMSCs-small intestinal submucosa (SIS) composites in vitro. METHODS: BMSCs were isolated by adherent culture from the bone marrow of 1-week-old SD rats. Inducing method of multiple differentiation and flow cytometry were applied to identify the cells. The stress-strain curve of SIS was measured with Instron machine. Purified BMSCs (2nd passage, 2.5 x 10(5) cells/cm2) were seeded on SIS (3 cm x 1 cm at size) and cultured for 2 days and then continued for another 5 days under strain stimulation (stretching frequency was 0.02 Hz, action time was 15 minutes/hour and 12 hours/day, strain amplitude was 5%) as experimental group, while the BMSCs-SIS composites were sustained static culture as control group. TCs were isolated from tail of 1-week-old SD rats. TCs-SIS composites were cultured under non-strained as positive control group. Scanning electron microscope (SEM) was used to examine the morphological changes of BMSCs after strain stimulation. The contents of Scleraxis and Tenomodulin in supernatant were tested by ELISA kit. Results The BMSCs could be induced to differentiate into osteoblasts and lipocytes, and showed the results of CD34-, CD45-, and CD90+, which were accorded with the biological characteristics of BMSCs. The failure test of SIS showed that the average elastic strain was 39.5%. SEM observation showed that the strain-stimulated BMSCs had the TCs-like morphological characteristics. The contents of Scleraxis and Tenomodulin in supernatant of experimental group, control group, and positive control group were (3.56 +/- 0.91) micromol/L and (4.27 +/- 1.10) micromol/L, (0.23 +/- 0.14) micromol/L and (0.16 +/- 0.10) micromol/L, and (14.73 +/- 2.30) micromol/L and (10.65 +/- 1.51) micromol/L, respectively. There were significant differences among 3 groups (P < 0.05). CONCLUSION: Appropriate strain stimulation could induce BMSCs differentiate into TCs, and the best conditions of strain stimulation need more experiments. PMID: 20695378 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [Osteogenic activity of porous calcium phosphate ceramics fabricated by rapid prototyping] Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2010 Jul;24(7):792-6 Authors: He C, Zhao L, Lin L, Gu H, Zhou H, Cui L OBJECTIVE: Calcium phosphate bioceramics has a broad application prospect because of good biocompatibility, but porous scaffolds with complex shape can not be prepared by the traditional methods. To fabricate porous calcium phosphate ceramics by rapid prototyping and to investigate the in vitro osteogenic activities. METHODS: The porous calcium phosphate ceramics was fabricated by rapid prototyping. The bone marrow mesenchymal stem cells (BMSCs) were isolated from bone marrow of Beagle canine, and the 3rd passage BMSCs were seeded onto the porous ceramics. The cell/ceramics composite cultured in osteogenic medium were taken as the experimental group (group A) and the cell/ceramics composite cultured in growth medium were taken as the control group (group B). Meanwhile, the cells seeded on the culture plate were cultured in osteogenic medium or growth medium respectively as positive control (group C) or negative control (group D). After 1, 3, and 7 days of culture, the cell proliferation and osteogenic differentiation on the porous ceramics were evaluated by DNA quantitative analysis, histochemical staining and alkaline phosphatase (ALP) activity. After DiO fluorescent dye, the cell adhesion, growth, and proliferation on the porous ceramics were also observed by confocal laser scanning microscope (CLSM). RESULTS: DNA quantitative analysis results showed that the number of BMSCs in all groups increased continuously with time. Plateau phase was not obvious in groups A and B, but it was clearly observed in groups C and D. The CLSM observation indicated that the activity of BMSCs was good and the cells spread extensively, showing good adhesion and proliferation on the porous calcium phosphate ceramics prepared by rapid prototyping. ALP quantitative analysis results showed that the stain of cells on the ceramics became deeper and deeper with time in groups A and B, the staining degree in group A were stronger than that in group B. There was no significant difference in the change of the ALP activity among 4 groups at the first 3 days (P > 0.05); the ALP activity increased obviously in 4 groups at 7 days, group A was significantly higher than other groups (P < 0.05) and groups C, D were significantly higher than group D (P < 0.05). CONCLUSION: The porous calcium phosphate ceramics has good cytocompatibility and the designed pores are favorable for cell ingrowth. The porous ceramics fabricated by rapid prototyping has prominent osteogenic differentiation activity and can be used as a choice of scaffolds for bone tissue engineering. PMID: 20695373 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [Experimental comparative study on osteogenic activity between freeze-dried tissue engineered bone and tissue engineered bone] Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2010 Jul;24(7):779-84 Authors: Hou T, Luo F, Liu J, Bian B, Xu J OBJECTIVE: Tissue engineered bone (TEB) lacks of an effective and feasible method of storage and transportation. To evaluate the activity of osteogenesis and capability of ectopic osteogenesis for TEB after freeze-dried treatment in vitro and in vivo and to explore a new method of preserving and transporting TEB. METHODS: Human bone marrow mesenchymal stem cells (hBMSCs) and decalcified bone matrix (DBM) were harvested from bone marrow and bone tissue of the healthy donators. TEB was fabricated with the 3rd passage hBMSCs and DBM, and they were frozen and dried at extremely low temperatures after 3, 5, 7, 9, 12, and 15 days of culture in vitro to obtain freeze-dried tissue engineered bone (FTEB). TEB and FTEB were observed by gross view and scanning electron microscope (SEM). Western blot was used to detect the changes of relative osteogenic cytokines, including bone morphogenetic protein 2 (BMP-2), transforming growth factor beta1 (TGF-beta1), and insulin-like growth factor 1 (IGF-1) between TEB and FTEB. The ectopic osteogenesis was evaluated by the methods of X-ray, CT score, and HE staining after TEB and FTEB were transplanted into hypodermatic space in athymic mouse. RESULTS: SEM showed that the cells had normal shape in TEB, and secretion of extracellular matrix increased with culture time; in FTEB, seeding cells were killed by the freeze-dried process, and considerable extracellular matrix were formed in the pore of DBM scaffold. The osteogenic cytokines (BMP-2, TGF-beta1, and IGF-1) in TEB were not decreased after freeze-dried procedure, showing no significant difference between TEB and FTEB (P > 0.05) except TGF-beta1 15 days after culture (P < 0.05). The ectopic osteogenesis was observed in TEB and FTEB groups 8 and 12 weeks after transplantation, there was no significant difference in the calcified level of grafts between TEB and FTEB groups by the analysis of X-ray and CT score. On the contrary, there was no ectopic osteogenesis in group DBM 12 weeks after operation. HE staining showed that DBM scaffold degraded and disappeared 12 weeks after operation. CONCLUSION: The osteogenic activity of TEB and FTEB is similar, which provides a new strategy to preserve and transport TEB. PMID: 20695371 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [Combined application of green fluorescent protein labeling and confocal laser scanning microscope three-dimensional reconstruction to monitor construction and in vivo transplantation of tissue engineered bone] Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2010 Jul;24(7):774-8 Authors: Hou T, Bian B, Luo F, Wu X, Jin H, Jiang H, Xu J OBJECTIVE: The combined application of green fluorescent protein (GFP) and confocal laser scanning microscope three-dimensional reconstruction (CLSM-3DR) were used to monitor the construction and in vivo transplantation of tissue engineered bone (TEB), to provide for technology in selection of scaffolds and three-dimensional constructional methods. METHODS: After bone marrow mesenchymal stem cells (BMSCs) were isolated from a 2-year-old green goat by a combination method of density gradient centrifugation and adherent culture, and the expressions of CD29, CD60L, CD45, and CD44 in BMSCs were detected by flow cytometry. Plasmid of pLEGFP-N1 was amplified, digested by enzymes (Hind III, BamH I, Sal I, and Bgl II), and identified. Transfection of pLEGFP-N1 into PT67 cells was performed under the help of liposome. Positive PT67 cells were picked out with G418, and proliferated for harvesting virus. Based on the titre of virus, after BMSCs were infected by virus containing pLEGFP-N1, GFP positive BMSCs were collected and proliferated for seeding cells. TEB was fabricated by GFP positive BMSCs and decalcified bone matrix (DBM) and observed by CLSM-3DR for the evaluation of the distribution and proliferation of seeding cells. After TEB was transplanted in the defect of goat femur, CLSM was used for observing the survival and distribution of GFP positive cells in the grafts. RESULTS: The isolated cells were fibroblast-like morphous, with the positive expression of CD29 and CD44, and negative expression of CD60L and CD45. The digested production of pLEGFP-N1 was collected for ionophoresis, whose results showed the correct fragment length (6 900 bp). The virus of pLEGFP-N 1 was harvested by transfection of pLEGFP-N1 into PT67 cells and used for further infection to obtain GFP positive BMSCs. The proliferated GFP positive BMSCs and DBM were used for fabrication of TEB. The distribution, proliferation, and migration of BMSCs in TEB were observed by CLSM-3DR. GFP positive cells also were observed in images of TEB graft in goat femur 28 days after transplantation. CONCLUSION: The BMSCs labeled by GFP in three-dimensional scaffold in vivo were monitored well by CLSM-3DR. It suggests a wide use potency in monitoring of three-dimensional cultured TEB. PMID: 20695370 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [New progress of related research of bone tissue engineering] Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2010 Jul;24(7):769-73 Authors: Xu J OBJECTIVE: To review new progress of related research of bone tissue engineering in recent years. METHODS: Domestic and international literature concerning bone tissue engineering was reviewed and analyzed. RESULTS: In the recent years, great progression had been made in the research and development of bone tissue engineering, it had been used in more and more hospitals, and relevant national regulations and protocols had been set up. As to seed cells of bone tissue engineering, autologous and allogeneic stem cells had been widely used, while recently embryonic stem cells and induced pluri potent stem cells had attracted most attentions. In the field of scaffolds materials, significant improvements had been made, from natural extractions to artificial polymers; from single construction to multiple compounds with surface modifications. As to the methods of construction, the static seeding approach had been widely accepted, and the applications of bioreactor had provided a stable and various micro-enviroment for the vitro-culture of different stem cells, which had been regarded as an alternative way of vitro-culture and construction for bone tissue engineering. CONCLUSION: With the tremendous help of the techniques and approaches above, we shall expect a promising future of a new generation bone tissue engineering based medical products in the years to come. PMID: 20695369 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | 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 - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | Adjuvant engineering for cancer immunotherapy: Development of a synthetic TLR2 ligand with increased cell adhesion. Cancer Sci. 2010 Jul;101(7):1596-603 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. PMID: 20507323 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | 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 - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | 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 - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | 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 - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | 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 lentiviral 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 lentiviral 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 - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | [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 - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | 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] | | | | | | | | | | |  |
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