|  |  |  | | | | | TE-RegenMed-StemCell feed | | | | | | | | | | | | | | | | | | Nature magazine today weighed in with its story on the nominations for the new chairman of the $3 billion California stem cell agency.
Writing on the publication's Spoonful of Medicine blog, Elie Dolgin said that the CIRM board of directors faces "a stark choice over who will lead the San Francisco-based institute as it enters into its next phase" – a cardiologist or bond financier.
The piece | | | | | | | | | | | | | | | | | | | | | | | | The latest news coverage of selection of a new leader for California's $3 billion stem cell agency is meager, to say the least.
Absent from the game this morning is most of the major media in the state, including the Los Angeles Times, the San Francisco Chronicle and the San Jose Mercury News. The Chronicle's and Mercury's backyards include the headquarters of CIRM, not to mention the major | | | | | | | | | | | | | | | | | | | | | | | | | Rapid and highly efficient generation of induced pluripotent stem cells from human umbilical vein endothelial cells. PLoS One. 2011;6(5):e19743 Authors: Panopoulos AD, Ruiz S, Yi F, Herrerías A, Batchelder EM, Izpisua Belmonte JC The ability to induce somatic cells to pluripotency by ectopic expression of defined transcription factors (e.g. KLF-4, OCT4, SOX2, c-MYC, or KOSM) has transformed the future of regenerative medicine. Here we report somatic cell reprogramming of human umbilical vein endothelial cells (HUVECs), yielding induced pluripotent stem (iPS) cells with the fastest kinetics, and one of the highest reprogramming efficiencies for a human somatic cell to date. HUVEC-derived iPS (Huv-iPS) cell colonies appeared as early as 6 days after a single KOSM infection, and were generated with a 2.5-3% reprogramming efficiency. Furthermore, when HUVEC reprogramming was performed under hypoxic conditions in the presence of a TGF-beta family signaling inhibitor, colony formation increased an additional ∼2.5-fold over standard conditions. Huv-iPS cells were indistinguishable from human embryonic stem (ES) cells with regards to morphology, pluripotent marker expression, and their ability to generate all embryonic germ layers in vitro and in vivo. The high efficiency and rapid kinetics of Huv-iPS cell formation, coupled with the ease by which HUVECs can be collected, expanded and stored, make these cells an attractive somatic source for therapeutic application, and for studying the reprogramming process. PMID: 21603572 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Cardiac differentiation of pluripotent stem cells. Stem Cells Int. 2011;2011:383709 Authors: Rajala K, Pekkanen-Mattila M, Aalto-Setälä K The ability of human pluripotent stem cells to differentiate towards the cardiac lineage has attracted significant interest, initially with a strong focus on regenerative medicine. The ultimate goal to repair the heart by cardiomyocyte replacement has, however, proven challenging. Human cardiac differentiation has been difficult to control, but methods are improving, and the process, to a certain extent, can be manipulated and directed. The stem cell-derived cardiomyocytes described to date exhibit rather immature functional and structural characteristics compared to adult cardiomyocytes. Thus, a future challenge will be to develop strategies to reach a higher degree of cardiomyocyte maturation in vitro, to isolate cardiomyocytes from the heterogeneous pool of differentiating cells, as well as to guide the differentiation into the desired subtype, that is, ventricular, atrial, and pacemaker cells. In this paper, we will discuss the strategies for the generation of cardiomyocytes from pluripotent stem cells and their characteristics, as well as highlight some applications for the cells. PMID: 21603143 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | "Humanized" stem cell culture techniques: the animal serum controversy. Stem Cells Int. 2011;2011:504723 Authors: Tekkatte C, Gunasingh GP, Cherian KM, Sankaranarayanan K Cellular therapy is reaching a pinnacle with an understanding of the potential of human mesenchymal stem cells (hMSCs) to regenerate damaged tissue in the body. The limited numbers of these hMSCs in currently identified sources, like bone marrow, adipose tissue, and so forth, bring forth the need for their in vitro culture/expansion. However, the extensive usage of supplements containing xenogeneic components in the expansion-media might pose a risk to the post-transplantation safety of patients. This warrants the necessity to identify and develop chemically defined or "humanized" supplements which would make in vitro cultured/processed cells relatively safer for transplantation in regenerative medicine. In this paper, we outline the various caveats associated with conventionally used supplements of xenogenic origin and also portray the possible alternatives/additives which could one day herald the dawn of a new era in the translation of in vitro cultured cells to therapeutic interventions. PMID: 21603148 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Robust fat suppression at 3T in high-resolution diffusion-weighted single-shot echo-planar imaging of human brain. Magn Reson Med. 2011 May 20; Authors: Sarlls JE, Pierpaoli C, Lalith Talagala S, Luh WM Single-shot echo-planar imaging is the most common acquisition technique for whole-brain diffusion tensor imaging (DTI) studies in vivo. Higher field MRI systems are readily available and advantageous for acquiring DTI due to increased signal. One of the practical issues for DTI with single-shot echo-planar imaging at high-field is incomplete fat suppression resulting in a chemically shifted fat artifact within the brain image. Unsuppressed fat is especially detrimental in DTI because the diffusion coefficient of fat is two orders of magnitude lower than that of parenchyma, producing brighter appearing fat artifacts with greater diffusion weighting. In this work, several fat suppression techniques were tested alone and in combination with the goal of finding a method that provides robust fat suppression and can be used in high-resolution single-shot echo-planar imaging DTI studies. Combination of chemical shift saturation with slice-select gradient reversal within a dual-spin-echo diffusion preparation period was found to provide robust fat suppression at 3 T. Magn Reson Med, 2011. © 2011 Wiley-Liss, Inc. PMID: 21604298 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | [Effect of allogenic adipose-derived stem cell transplantation on bone mass in rats with glucocorticoid-induced osteoporosis.] Nan Fang Yi Ke Da Xue Xue Bao. 2011 May 20;31(5):817-821 Authors: Tao H, Yu MC, Yang HY, Yang C, Qu RM, Dai JX, Yu L, Yuan L OBJECTIVE: To investigate the effects of systemic transplantation of allogenic adipose-derived stem cells (ADSCs), the main undifferentiated cells in the supporting-storing system based on the fasciology hypothesis, on the bone mineral density (BMD) and histomorphometry in rats with glucocorticoid-induced osteoporosis (GIOP) rats, and explore a new therapeutic approach for osteoporosis. METHODS: Forty female adult Wistar rats were randomized equally into blank control group (A), model group (B), control treatment group (C) and treatment group (D). In groups B, C, and D, osteoporosis was induced by injection of prednisolone (8 mg/kg) via the tail vein 3 times a week for 12 consecutive weeks. After successful establishment of osteoporosis, allogenic ADSCs (3×10(6)/ml) were transplanted via the tail vein. The BMD at the L3-L5 levels and of the right femurs were detected, and the histomorphometry of the right tibias was analyzed in all the rats. RESULTS: After prednisolone injection, the BMD of L3-L5 vertebrae and the right femurs, the percent trabecular area, trabecular thickness and trabecular number of the right tibias were all obviously lowered while the trabecular separation and osteoclast number increased in group B as compared to those in group A (P<0.05). Four weeks after ADSC transplantation, all these indices were significantly improved in group D, showing significant differences from those in group B (P<0.05) and also group C. CONCLUSION: Transplantation of allogenic ADSCs can restore the BMD and bone histomorphometric properties of rats with GIOP, and may serve as a potential treatment for GIOP. These results also provide partial xperimental evidence supporting the hypothesis of fasciaology. PMID: 21602132 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Scaffold porosity and oxygenation of printed hydrogel constructs affect functionality of embedded osteogenic progenitors. Tissue Eng Part A. 2011 May 20; Authors: Fedorovich N, Kuipers E, Gawlitta D, Dhert W, Alblas J Insufficient supply of oxygen and nutrients throughout the graft is considered one of the principal limitations in development of large, tissue-engineered bone grafts. Organ- or tissue printing (OP) by means of three-dimensional (3D) fiber deposition is a novel modality in regenerative medicine that combines pore formation and defined cell placement, and is used here for development of cell-laden hydrogel structures with reproducible internal architecture to sustain oxygen supply and to support adequate tissue development. In this study we tested the effect of porosity on multipotent stromal cells (MSCs) embedded in hydrogel constructs printed with a 3D fiber deposition (3DF) machine. For this, porous and solid alginate hydrogel scaffolds, with MSCs homogeneously dispersed throughout the construct, were printed and analyzed in vitro for the presence of hypoxia markers, metabolism, survival and osteogenic differentiation. We demonstrated that porosity promotes oxygenation of MSCs in printed hydrogel scaffolds and supported the viability and osteogenic differentiation of embedded cells. Porous and solid printed constructs were subsequently implanted subcutaneously in immunodeficient mice to analyze tissue formation in relation to hypoxia responses of embedded cells. Implantation of printed grafts resulted in ingrowth of vascularized tissue and significantly enhanced oxygenation of embedded MSCs. In conclusion, the introduction of pores significantly enhances the conductive properties of printed hydrogel constructs and contributes to the functionality of embedded osteogenic progenitors. PMID: 21599540 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | LSD1 regulates the balance between self-renewal and differentiation in human embryonic stem cells. Nat Cell Biol. 2011 May 22; Authors: Adamo A, Sesé B, Boue S, Castaño J, Paramonov I, Barrero MJ, Belmonte JC We identify LSD1 (lysine-specific demethylase 1; also known as KDM1A and AOF2) as a key histone modifier that participates in the maintenance of pluripotency through the regulation of bivalent domains, a chromatin environment present at the regulatory regions of developmental genes that contains both H3K4 di/trimethylation and H3K27 trimethylation marks. LSD1 occupies the promoters of a subset of developmental genes that contain bivalent domains and are co-occupied by OCT4 and NANOG in human embryonic stem cells, where it controls the levels of H3K4 methylation through its demethylase activity. Thus, LSD1 has a role in maintaining the silencing of several developmental genes in human embryonic stem cells by regulating the critical balance between H3K4 and H3K27 methylation at their regulatory regions. PMID: 21602794 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Caveolin-1 overexpression enhances androgen-dependent growth and proliferation in the mouse prostate. Int J Biochem Cell Biol. 2011 May 12; Authors: Bryant KG, Camacho J, Jasmin JF, Wang C, Addya S, Casimiro MC, Fortina P, Balasubramaniam S, Knudsen KE, Schwarting R, Lisanti MP, Mercier I Prostate cancer (PCa) continues to be one of the leading causes of cancer- related deaths among American men. The prostate relies upon the androgen receptor (AR) to mediate the effects of androgens on normal growth, a reliance that is maintained during malignant prostate growth. Caveolin-1 (Cav-1), the main structural component of caveolae, has been shown to promote malignant growth and invasion of prostate tumors. In vitro work has shown that Cav-1 can act as an AR coactivator by enhancing its transciptional activity. However, it is unknown how Cav-1 affects androgen-dependent growth and signaling in vivo. To explore this role, a novel mouse model of Cav-1 overexpression was developed with a hormone-insensitive promoter. Cav-1 transgenic (Tg) mice subjected to castration and androgen stimulation display enlarged prostate weights and increased DNA synthesis. Through gene transcript and proteomic profiling, we demonstrate that Cav-1 overexpression favors androgen-regulated responses and enhances processes involved in transcription, cell cycle and protein synthesis. Interestingly, Cav-1 overexpression was associated with an increase in the phosphorylation of AR on serine 210, a post-translational modification linked to its activity in androgen-stimulated conditions. In addition, these mice exhibited an increase in the phosphorylation of ribosomal S6 protein on serine 235/236 (pS6), a marker of protein synthesis and downstream component of the mTOR pathway. Thus, Cav-1 Tg mice could serve as a novel model for studying AR-regulated pathways involved in prostate growth and proliferation. PMID: 21601007 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | In vitro human tissue models - moving towards personalized regenerative medicine. Adv Drug Deliv Rev. 2011 May 17; Authors: Schenke-Layland K, Nerem RM PMID: 21600252 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Wnt/β-catenin Signaling in Embryonic Stem Cell Self-renewal and Somatic Cell Reprogramming. Stem Cell Rev. 2011 May 21; Authors: Miki T, Yasuda SY, Kahn M Embryonic stem cells and induced pluripotent stem (iPS) cells are characterized by their ability to self-renew and to generate differentiated cells of all three germ layers. This potential makes them an attractive source to address question of developmental and also for use in clinical regenerative medicine. Although the culture conditions to maintain pluripotency and reprogramming technologies have been established, the underlying molecular mechanisms are incompletely understood. Accumulating evidence indicates that the Wnt/β-catenin signaling pathway plays a pivotal role in the maintenance of pluripotency as well as in the process of somatic cell reprogramming. Reciprocally, Wnt/β-catenin signaling also plays a critical role in the lineage decision/commitment process. These dramatically different outcomes upon activation of the Wnt signaling cascade has fueled enormous controversy concerning the role of Wnt signaling in the maintenance of potency and induction of differentiation in stem cells. Here, we discuss and explore the divergent roles of the Wnt signaling pathways based on findings from our lab. Accumulated results from our lab indicate the usage of a critical switching mechanism that regulates the divergent Wnt/catenin transcriptional programs associated with either maintenance of potency or initiation of differentiation. PMID: 21603945 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Gellan gum-based hydrogels for intervertebral disc tissue-engineering applications. J Tissue Eng Regen Med. 2011 Jun;5(6):e97-e107 Authors: Silva-Correia J, Oliveira JM, Caridade SG, Oliveira JT, Sousa RA, Mano JF, Reis RL Intervertebral disc (IVD) degeneration is a challenging clinical problem that urgently demands viable nucleus pulposus (NP) implant materials. The best suited biomaterial for NP regeneration has yet to be identified, but it is believed that biodegradable hydrogel-based materials are promising candidates. In this work, we have developed ionic- and photo-crosslinked methacrylated gellan gum (GG-MA) hydrogels to be used in acellular and cellular tissue-engineering strategies for the regeneration of IVDs. The physicochemical properties of the developed hydrogels were investigated by Fourier-transform infrared spectroscopy, (1) H nuclear magnetic resonance and differential scanning calorimetry. The swelling ability and degradation rate of hydrogels were also analysed in phosphate-buffered saline solution at physiological pH for a period of 30 days. Additionally, the morphology and mechanical properties of the hydrogels were assessed under a scanning electron microscope and dynamic compression, respectively. An in vitro study was carried out to screen possible cytotoxicity of the gellan gum-based hydrogels by culturing rat lung fibroblasts (L929 cells) with hydrogel leachables up to 7 days. The results demonstrated that gellan gum was successfully methacrylated. We observed that the produced GG-MA hydrogels possess improved mechanical properties and lower water uptake ability and degradation rate as compared to gellan gum. This work also revealed that GG-MA hydrogels are non-cytotoxic in vitro, thus being promising biomaterials to be used in IVD tissue-engineering strategies. Copyright © 2010 John Wiley & Sons, Ltd. PMID: 21604382 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Microchannel-patterned and heparin micro-contact-printed biodegradable composite membranes for tissue-engineering applications. J Tissue Eng Regen Med. 2011 Jun;5(6):e108-14 Authors: Baran ET, Tuzlakoğlu K, Salgado A, Reis RL Microchannel-patterned starch-poly(capro-lactone)/hydydroxyapatite (SPCL-HA) and starch-poly(lactic acid) (SPLA) composite membranes were produced for use as a laminated tissue-engineering scaffold that incorporates both physical and biochemical patterns. For this purpose, SPCL (30% starch) blended with inorganic hydroxyl apatite (50%) and SPLA (50% starch) membranes were made with compressive moulding. Consequently, the microchannel structures (width 102 µm, 174 µm intervals) were developed on the composite membranes by means of micro-patterned metal mould(s) and hydraulic pressing. An elastomer poly(dimetylsiloxane) stamp was used to transfer heparin as a biochemical cue over the microchannel surfaces by micro-contact printing (µCP). Toluidine blue staining of developed capillaries and heparin µCP-coated membranes showed that heparin was transferred predominantly over the microchannel surfaces. Fibroblast cell culture over the microchannel-formed and heparin µCP-modified SPCL-HA and SPLA membranes showed distinct growth patterns. In contrast to the uniform cell layer formed on unmodified microchannels, the cells were bridging across the grooves of heparin-printed microchannels. At extended culture periods, the heparin-printed microchannels were covered with a layer of fibroblast cells without cellular ingrowths inside. This study indicated that the topographical pattern could induce an organization of fibroblasts only with the biochemical cue and the cells' functions can be controlled spatially over the microchannels by using both cues. Copyright © 2010 John Wiley & Sons, Ltd. PMID: 21604378 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | "Humanized" stem cell culture techniques: the animal serum controversy. Stem Cells Int. 2011;2011:504723 Authors: Tekkatte C, Gunasingh GP, Cherian KM, Sankaranarayanan K Cellular therapy is reaching a pinnacle with an understanding of the potential of human mesenchymal stem cells (hMSCs) to regenerate damaged tissue in the body. The limited numbers of these hMSCs in currently identified sources, like bone marrow, adipose tissue, and so forth, bring forth the need for their in vitro culture/expansion. However, the extensive usage of supplements containing xenogeneic components in the expansion-media might pose a risk to the post-transplantation safety of patients. This warrants the necessity to identify and develop chemically defined or "humanized" supplements which would make in vitro cultured/processed cells relatively safer for transplantation in regenerative medicine. In this paper, we outline the various caveats associated with conventionally used supplements of xenogenic origin and also portray the possible alternatives/additives which could one day herald the dawn of a new era in the translation of in vitro cultured cells to therapeutic interventions. PMID: 21603148 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Toward Transgene-Free Induced Pluripotent Stem Cells: Lessons from Transdifferentiation Studies. Cell Reprogram. 2011 May 20; Authors: Chua SJ, Casper RF, Rogers IM Abstract Regenerative medicine has received much attention over the years due to its clinical and commercial potential. The excitement around regenerative medicine waxes and wanes as new discoveries add to its foundation but are not immediately clinically applicable. The recent discovery of induced pluripotent stem cells has lead to a sustained effort from many research groups to develop clinically relevant regenerative medicine therapies. A major focus of cellular reprogramming is to generate safe cellular products through the use of proteins or small molecules instead of transgenes. The successful reprogramming of somatic nuclei to generate pluripotential cells capable of embryo development was pioneered over 50 years ago by Briggs and King and followed by Gurdon in the early 1960s. The success of these studies, the cloning of Dolly, and more current studies involving adult stem cells and transdifferentiation provide us with a large repository of potential candidate molecules and experimental systems that will assist in the generation of safe, transgene-free pluripotential cells. PMID: 21599518 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Metabolic profiling based quantitative evaluation of hepatocellular metabolism in presence of adipocyte derived extracellular matrix. PLoS One. 2011;6(5):e20137 Authors: Sharma NS, Nagrath D, Yarmush ML The elucidation of the effect of extracellular matrices on hepatocellular metabolism is critical to understand the mechanism of functional upregulation. We have developed a system using natural extracellular matrices [Adipogel] for enhanced albumin synthesis of rat hepatocyte cultures for a period of 10 days as compared to collagen sandwich cultures. Primary rat hepatocytes isolated from livers of female Lewis rats recover within 4 days of culture from isolation induced injury while function is stabilized at 7 days post-isolation. Thus, the culture period can be classified into three distinct stages viz. recovery stage [day 0-4], pre-stable stage [day 5-7] and the stable stage [day 8-10]. A Metabolic Flux Analysis of primary rat hepatocytes cultured in Adipogel was performed to identify the key metabolic pathways modulated as compared to collagen sandwich cultures. In the recovery stage [day 4], the collagen-soluble Adipogel cultures shows an increase in TriCarboxylic Acid [TCA] cycle fluxes; in the pre-stable stage [day 7], there is an increase in PPP and TCA cycle fluxes while in the stable stage [day 10], there is a significant increase in TCA cycle, urea cycle fluxes and amino acid uptake rates concomitant with increased albumin synthesis rate as compared to collagen sandwich cultures throughout the culture period. Metabolic analysis of the collagen-soluble Adipogel condition reveals significantly higher transamination reaction fluxes, amino acid uptake and albumin synthesis rates for the stable vs. recovery stages of culture. The identification of metabolic pathways modulated for hepatocyte cultures in presence of Adipogel will be a useful step to develop an optimization algorithm to further improve hepatocyte function for Bioartificial Liver Devices. The development of this framework for upregulating hepatocyte function in Bioartificial Liver Devices will facilitate the utilization of an integrated experimental and computational approach for broader applications of Adipogel in tissue e engineering and regenerative medicine. PMID: 21603575 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Use of the 2A Peptide for Generation of Multi-Transgenic Pigs through a Single Round of Nuclear Transfer. PLoS One. 2011;6(5):e19986 Authors: Deng W, Yang D, Zhao B, Ouyang Z, Song J, Fan N, Liu Z, Zhao Y, Wu Q, Nashun B, Tang J, Wu Z, Gu W, Lai L Multiple genetic modifications in pigs can essentially benefit research on agriculture, human disease and xenotransplantation. Most multi-transgenic pigs have been produced by complex and time-consuming breeding programs using multiple single-transgenic pigs. This study explored the feasibility of producing multi-transgenic pigs using the viral 2A peptide in the light of previous research indicating that it can be utilized for multi-gene transfer in gene therapy and somatic cell reprogramming. A 2A peptide-based double-promoter expression vector that mediated the expression of four fluorescent proteins was constructed and transfected into primary porcine fetal fibroblasts. Cell colonies (54.3%) formed under G418 selection co-expressed the four fluorescent proteins at uniformly high levels. The reconstructed embryos, which were obtained by somatic cell nuclear transfer and confirmed to express the four fluorescent proteins evenly, were transplanted into seven recipient gilts. Eleven piglets were delivered by two gilts, and seven of them co-expressed the four fluorescent proteins at equivalently high levels in various tissues. The fluorescence intensities were directly observed at the nose, hoof and tongue using goggles. The results suggest that the strategy of combining the 2A peptide and double promoters efficiently mediates the co-expression of the four fluorescent proteins in pigs and is hence a promising methodology to generate multi-transgenic pigs by a single nuclear transfer. PMID: 21603633 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | In vivo monitoring of adult neurogenesis in health and disease. Front Neurosci. 2011;5:67 Authors: Couillard-Despres S, Vreys R, Aigner L, Van der Linden A Adult neurogenesis, i.e., the generation of new neurons in the adult brain, presents an enormous potential for regenerative therapies of the central nervous system. While 5-bromo-2'-deoxyuridine labeling and subsequent histology or immunohistochemistry for cell-type-specific markers is still the gold standard in studies of neurogenesis, novel techniques, and tools for in vivo imaging of neurogenesis have been recently developed and successfully applied. Here, we review the latest progress on these developments, in particular in the area of magnetic resonance imaging (MRI) and optical imaging. In vivo in situ labeling of neural progenitor cells (NPCs) with micron-sized iron oxide particles enables longitudinal visualization of endogenous progenitor cell migration by MRI. The possibility of genetic labeling for cellular MRI was demonstrated by using the iron storage protein ferritin as the MR reporter-gene. However, reliable and consistent results using ferritin imaging for monitoring endogenous progenitor cell migration have not yet been reported. In contrast, genetic labeling of NPCs with a fluorescent or bioluminescent reporter has led to the development of some powerful tools for in vivo imaging of neurogenesis. Here, two strategies, i.e., viral labeling of stem/progenitor cells and transgenic approaches, have been used. In addition, the use of specific promoters for neuronal progenitor cells such as doublecortin increases the neurogenesis-specificity of the labeling. Naturally, the ultimate challenge will be to develop neurogenesis imaging methods applicable in humans. Therefore, we certainly need to consider other modalities such as positron emission tomography and proton magnetic resonance spectroscopy ((1)H-MRS), which have already been implemented for both animals and humans. Further improvements of sensitivity and neurogenesis-specificity are nevertheless required for all imaging techniques currently available. PMID: 21603226 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | On-chip characterization of cryoprotective agent mixtures using an EWOD-based digital microfluidic device. Lab Chip. 2011 May 20; Authors: Park S, Wijethunga PA, Moon H, Han B For tissue engineering and regenerative medicine, cryopreservation, a technique for preserving biomaterials in the frozen state with cryoprotective agents (CPAs), is critically important for preserving engineered tissues (ETs) as well as cells necessary to create ETs. As more diverse ETs are produced using various cell types, CPAs and corresponding freeze/thaw (F/T) protocols need to be developed cell/tissue-type specifically. This is because CPAs and F/T protocols that have been successful for one cell/tissue type have proven to be difficult to adapt to other cell/tissue types. The most critical barrier to address this challenge is the inability to screen and identify CPA or CPA mixtures efficiently. In this paper, we developed an "electro-wetting-on-dielectic" (EWOD) based digital microfluidic platform to characterize and screen CPA mixtures cell-type specifically. The feasibility of the EWOD platform was demonstrated by characterizing and optimizing a mixture of dimethlysulfoxide (DMSO) and PBS for human breast cancer cell line as model CPA mixture and cell line. The developed platform multiplexed droplets of DMSO and PBS to create an array of DMSO-PBS mixtures, and mapped the phase change diagram of the mixture. After loading cell suspensions on the platform, the mixture was further screened on-chip for toxicity and cryoprotection. The results were discussed to illustrate the capabilities and limitations of the EWOD platform for cell and tissue-type specific optimization of CPA mixtures and F/T protocols. PMID: 21603697 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Rapid vascularization of starch-poly(caprolactone) in vivo by outgrowth endothelial cells in co-culture with primary osteoblasts. J Tissue Eng Regen Med. 2011 Jun;5(6):e136-43 Authors: Ghanaati S, Fuchs S, Webber MJ, Orth C, Barbeck M, Gomes ME, Reis RL, James Kirkpatrick C The successful integration of in vitro-generated tissues is dependent on adequate vascularization in vivo. Human outgrowth endothelial cells (OECs) isolated from the mononuclear cell fraction of peripheral blood represent a potent population of circulating endothelial progenitors that could provide a cell source for rapid anastomosis and scaffold vascularization. Our previous work with these cells in co-culture with primary human osteoblasts has demonstrated their potential to form perfused vascular structures within a starch-poly(caprolactone) biomaterial in vivo. In the present study, we demonstrate the ability of OECs to form perfused vascular structures as early as 48 h following subcutaneous implantation of the biomaterial in vivo. The number of OEC-derived vessels increased throughout the study, an effect that was independent of the OEC donor. This finding of rapid and thorough OEC-mediated scaffold vascularization demonstrates the great potential for OEC-based strategies to promote vascularization in tissue engineering. OECs have the potential to contribute to host-derived scaffold vascularization, and formed vascular structures at a similar density as those arising from the host. Additionally, immunohistochemical evidence demonstrated the close interaction between OECs and the co-cultured osteoblasts. In addition to the known paracrine activity osteoblasts have in modulating angiogenesis of co-cultured OECs, we demonstrate the potential of osteoblasts to provide additional structural support for OEC-derived vessels, perhaps acting in a pericyte-like role. Copyright © 2010 John Wiley & Sons, Ltd. PMID: 21604380 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Transforming growth factor-β3 regulates assembly of a non-fibrotic matrix in a 3D corneal model. J Tissue Eng Regen Med. 2011 May 23; Authors: Karamichos D, Hutcheon AE, Zieske JD Corneal tissue engineering has attracted the attention of many researchers over the years, in part due to the cornea's avascularity and relatively straightforward structure. However, the highly organized and structured nature of this optically clear tissue has presented a great challenge. We have previously developed a model in which human corneal fibroblasts (HCFs) are stimulated by a stable vitamin C (VitC) derivative to self-assemble an extracellular matrix (ECM). Addition of TGFβ1 enhanced the assembly of ECM; however, it was accompanied by the upregulation of specific fibrotic markers. In this study, we tested the effects of all three TGFβ isoforms (-β1, -β2 and -β3) on ECM production, as well as expression of fibrotic markers. HCFs were grown in four media conditions for 4 weeks: control, VitC only; T1, VitC + TGFβ1; T2, VitC + TGFβ2; and T3, VitC + TGFβ3. The cultures were analysed with western blots, TEM and indirect immunofluorescence (IF). Compared to controls, all TGFβ isoforms stimulated matrix production by about three-fold. IF showed the presence of type III collagen and smooth muscle actin (SMA) in T1 and T2; however, T3 showed little to no expression. In western blots, T3 stimulated a lower type III:type I collagen ratio when compared to the other conditions. In addition, TEM indicated that T3 stimulated a higher level of matrix alignment and organization. HCFs stimulated by VitC and TGFβ3 appear to generate a matrix that mimics the normal adult or developing human cornea, whereas TGF-β1 and -β2 drive the constructs towards a more fibrotic path. Copyright © 2011 John Wiley & Sons, Ltd. PMID: 21604386 [PubMed - as supplied by publisher] | | | | | | | | | |  | |  | |  |
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