|  |  |  | | | | | TE-RegenMed-StemCell feed | | | | | | | | | | | | | | | | | | | The role of ECM proteins and protein fragments in guiding cell behavior in regenerative medicine. Biomaterials. 2011 Jun;32(18):4211-4 Authors: Barker TH The promise of biomaterials design for regenerative medicine tissue engineering is predicated on the fundamental ability to direct or guide specific and highly coordinated cellular behaviors that culminate in the creation of physiologically functional tissues and organs. To date, our efforts have focused primarily on the grafting and presentation of short synthetic peptides with just cause. Short peptides are capable of high levels of control, can be manufactured relatively easily in a highly reproducible manner under GMP guidelines and are readily modified to enable their integration with numerous current and emerging chemistries for biomaterials grafting. However, while extracellular matrix (ECM)-derived peptides have demonstrated their initial purpose of promoting cell adhesion, their general lack of specificity and significantly decreased receptor binding affinities have proven detrimental in attempts to regulate highly specific and integrated processes necessary for tissue regeneration. Unlike adhesion peptides, the natural ECM displays a complex interplay with cells by supporting environmentally sensitive and cell dependent integrin specificity and binding affinity. Furthermore, the adhesion ligands on ECM proteins display a finely tuned and evolutionarily directed spatial periodicity, of which is dynamically controlled through both mechanical and chemical modifications. These and other emerging concepts from matrix biology require our attention if biomaterials design is to fulfill its promise. Here, we are charged with debating the statement 'The use of short synthetic adhesion peptides, like RGD, is the best approach in the design of biomaterials that guide cell behavior for regenerative medicine tissue engineering'. In this Leading Opinion Paper I will focus on aspects of natural ECM proteins and protein fragments that have proven difficult, if not impossible to date, to recapitulate in peptide-based systems. While this represents an argument against the use of peptides per se, it might also be viewed as outlining the challenges and opportunities for the biomaterials field. PMID: 21515169 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Distinct tissue formation by heterogeneous printing of osteo- and endothelial progenitor cells. Tissue Eng Part A. 2011 Apr 23; Authors: Fedorovich N, Wijnberg H, Dhert W, Alblas J The organ- or tissue printing (OP) approach, based on layered deposition of cell-laden hydrogels, is a new technique in regenerative medicine suitable to investigate whether mimicking the anatomical organization of cells, matrix and bioactive molecules is necessary for obtaining or improving functional engineered tissues. Currently data on performance of multicellular printed constructs in vivo is limited. In this study we illustrate the ability of the system to print intricate porous constructs containing two different cell types - endothelial progenitors (EPCs) and multipotent stromal cells (MSCs) - and show that these grafts retain heterogeneous cell organization after subcutaneous implantation in immunodeficient mice. We demonstrate that cell differentiation leading to the expected tissue formation occurs at the site of the deposited progenitor cell type. While perfused blood vessels are formed in the EPC-laden part of the constructs, bone formation is taking place in the MSC-laden part of the printed graft. PMID: 21513466 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | In vitro profiling of epigenetic modifications underlying heavy metal toxicity of tungsten-alloy and its components. Toxicol Appl Pharmacol. 2011 Apr 14; Authors: Verma R, Xu X, Jaiswal MK, Olsen C, Mears D, Caretti G, Galdzicki Z Tungsten-alloy has carcinogenic potential as demonstrated by cancer development in rats with intramuscular implanted tungsten-alloy pellets. This suggests a potential involvement of epigenetic events previously implicated as environmental triggers of cancer. Here, we tested metal induced cytotoxicity and epigenetic modifications including H3 acetylation, H3-Ser10 phosphorylation and H3-K4 trimethylation. We exposed human embryonic kidney (HEK293), human neuroepithelioma (SKNMC), and mouse myoblast (C2C12) cultures for 1-day and hippocampal primary neuronal cultures for 1-week to 50-200μg/ml of tungsten-alloy (91%tungsten/6%nickel/3%cobalt), tungsten, nickel, and cobalt. We also examined the potential role of intracellular calcium in metal mediated histone modifications by addition of calcium channel blockers/chelators to the metal solutions. Tungsten and its alloy showed cytotoxicity at concentrations >50μg/ml, while we found significant toxicity with cobalt and nickel for most tested concentrations. Diverse cell-specific toxic effects were observed, with C2C12 being relatively resistant to tungsten-alloy mediated toxic impact. Tungsten-alloy, but not tungsten, caused almost complete dephosphorylation of H3-Ser10 in C2C12 and hippocampal primary neuronal cultures with H3-hypoacetylation in C2C12. Dramatic H3-Ser10 dephosphorylation was found in all cobalt treated cultures with a decrease in H3 pan-acetylation in C2C12, SKNMC and HEK293. Trimethylation of H3-K4 was not affected. Tungsten-alloy and cobalt mediated H3-Ser10 dephosphorylation was reversed with BAPTA-AM, highlighting the role of intracellular calcium, confirmed with 2-photon calcium imaging. In summary, our results for the first time reveal epigenetic modifications triggered by tungsten-alloy exposure in C2C12 and hippocampal primary neuronal cultures suggesting the underlying synergistic effects of tungsten, nickel and cobalt mediated by changes in intracellular calcium homeostasis and buffering. PMID: 21513724 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Induced pluripotent stem cells for neural tissue engineering. Biomaterials. 2011 Apr 21; Authors: Wang A, Tang Z, Park IH, Zhu Y, Patel S, Daley GQ, Li S Induced pluripotent stem cells (iPSCs) hold great promise for cell therapies and tissue engineering. Neural crest stem cells (NCSCs) are multipotent and represent a valuable system to investigate iPSC differentiation and therapeutic potential. Here we derived NCSCs from human iPSCs and embryonic stem cells (ESCs), and investigated the potential of NCSCs for neural tissue engineering. The differentiation of iPSCs and the expansion of derived NCSCs varied in different cell lines, but all NCSC lines were capable of differentiating into mesodermal and ectodermal lineages, including neural cells. Tissue-engineered nerve conduits were fabricated by seeding NCSCs into nanofibrous tubular scaffolds, and used as a bridge for transected sciatic nerves in a rat model. Electrophysiological analysis showed that only NCSC-engrafted nerve conduits resulted in an accelerated regeneration of sciatic nerves at 1 month. Histological analysis demonstrated that NCSC transplantation promoted axonal myelination. Furthermore, NCSCs differentiated into Schwann cells and were integrated into the myelin sheath around axons. No teratoma formation was observed for up to 1 year after NCSC transplantation in vivo. This study demonstrates that iPSC-derived multipotent NCSCs can be directly used for tissue engineering and that the approach that combines stem cells and scaffolds has tremendous potential for regenerative medicine applications. PMID: 21514663 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Microglia activation during neuroregeneration in the adult vertebrate brain. Neurosci Lett. 2011 Apr 14; Authors: Kirkham M, Berg D, Simon A Brain injury and neuronal loss leads to an inflammatory response, which is initiated by the innate immune system. To what extent this immune response is beneficial or detrimental for neurogenesis and regeneration is unclear. We addressed this question during regeneration of dopamine neurons in the adult salamander brain. In contrast to mammals, ablation of dopamine neurons evokes robust neurogenesis leading to complete histological and functional regeneration within four weeks in salamanders. Here we show that similarly to mammals, ablation of dopamine neurons causes microglia activation and an increase in microglia numbers in the ablated areas. Furthermore, microglia numbers remains elevated compared to the uninjured brain at least six weeks after ablation. Suppression of the microglia response results in enhanced regeneration, concomitant with reduced death of dopamine neurons during the regeneration phase. Thus neuroregeneration is not dependent on the absence of an innate immune response, but the suppression of this response may be a means to promote neurogenesis in the adult vertebrate brain. PMID: 21515337 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Evolving the use of peptides as components of biomaterials. Biomaterials. 2011 Jun;32(18):4198-204 Authors: Collier JH, Segura T This manuscript is part of a debate on the statement that "the use of short synthetic adhesion peptides, like RGD, is the best approach in the design of biomaterials that guide cell behavior for regenerative medicine and tissue engineering". We take the position that although there are some acknowledged disadvantages of using short peptide ligands within biomaterials, it is not necessary to discard the notion of using peptides within biomaterials entirely, but rather to reinvent and evolve their use. Peptides possess advantageous chemical definition, access to non-native chemistries, amenability to de novo design, and applicability within parallel approaches. Biomaterials development programs that require such aspects may benefit from a peptide-based strategy. PMID: 21515167 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | The role of short synthetic adhesion peptides in regenerative medicine; The debate. Biomaterials. 2011 Jun;32(18):4195-7 Authors: Williams DF This paper constitutes the judgment on the four-paper debate in the same issue of this journal on the motion that ''the use of short synthetic adhesion peptides, like RGD, is the best approach in the design of biomaterials that guide cell behavior for regenerative medicine and tissue engineering". It is concluded that this motion is carried on the basis that the concept of installing multifunctionality and modularity in peptides is the more powerful concept, coupled with the potential attractiveness of retaining a chemically defined product, without any possibilities of disease transmission, and relative inexpensiveness and clearer regulatory pathways. PMID: 21515166 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Polymeric coating of surface modified nitinol stent with POSS-nanocomposite polymer. Colloids Surf B Biointerfaces. 2011 Apr 4; Authors: Bakhshi R, Darbyshire A, Evans JE, You Z, Lu J, Seifalian AM Stent angioplasty is a successful treatment for arterial occlusion, particularly in coronary artery disease. The clinical communities were enthusiastic about the use of drug-eluting stents; however, these stents have a tendency to be a contributory factor towards late stage thrombosis, leading to mortality in a significant number of patients per year. This work presents an innovative approach in self-expanding coronary stents preparation. We developed a new nanocomposite polymer based on polyhedral oligomeric silsesquioxanes (POSS) and poly(carbonate-urea)urethane (PCU), which is an antithrombogenic and a non-biodegradable polymer with in situ endothelialization properties. The aim of this work is to coat a NiTi stent alloy with POSS-PCU. In prolonged applications in the human body, the corrosion of the NiTi alloy can result in the release of deleterious ions which leads to unwanted biological reactions. Coating the nitinol (NiTi) surface with POSS-PCU can enhance surface resistance and improve biocompatibility. Electrohydrodynamic spraying was used as the polymer deposition process and thus a few experiments were carried out to compare this process with casting. Prior to deposition the NiTi has been surface modified. The peel strength of the deposit was studied before and after degradation of the coating. It is shown that the surface modification enhances the peel strength by 300%. It is also indicated how the adhesion strength of the POSS-PCU coating changes post-exposure to physiological solutions comprised of hydrolytic, oxidative, peroxidative and biological media. This part of the study shows that the modified NiTi presents far greater resistance to decay in peel strength compared to the non-modified NiTi. PMID: 21515031 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Hyaluronic acid-based clinical biomaterials derived for cell and molecule delivery in regenerative medicine. J Control Release. 2011 Apr 14; Authors: Prestwich GD The development of injectable and biocompatible vehicles for delivery, retention, growth, and differentiation of stem cells is of paramount importance for regenerative medicine. For cell therapy and the development of clinical combination products, we created a hyaluronan (HA)-based synthetic extracellular matrix (sECM) that provides highly reproducible, manufacturable, approvable, and affordable biomaterials. The composition of the sECM can be customized for use with progenitor and mature cell populations obtained from skin, fat, liver, heart, muscle, bone, cartilage, nerves, and other tissues. This overview describes the design criteria for "living" HA derivatives, and the many uses of this in situ crosslinkable HA-based sECM hydrogel for three-dimensional (3-D) culture of cells in vitro and translational use in vivo. Recent advances allow rapid expansion and recovery of cells in 3-D, and the bioprinting of engineered tissue constructs. The uses of HA-derived sECMs for cell and molecule delivery in vivo will be reviewed, including applications in cancer biology and tumor imaging. PMID: 21513749 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | New therapy for spinal cord injury shows positive results in rodent model. Regen Med. 2011 Mar;6(2):142-3 Authors: PMID: 21513086 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Chemical Strategies for Stem Cell Biology and Regenerative Medicine. Annu Rev Biomed Eng. 2010 Jul 21; Authors: Zhu S, Wei W, Ding S Stem cell technology holds great promises for the cures of devastating diseases, injuries, aging, and even cancers as it is applied in regenerative medicine. Recent breakthroughs in the development of induced pluripotent stem cell techniques and efficient differentiation strategies have generated tremendous enthusiasm and efforts to explore the therapeutic potential of stem cells. Small molecules, which target specific signaling pathways and/or proteins, have been demonstrated to be particularly valuable for manipulating cell fate, state, and function. Such small molecules not only are useful in generating desired cell types in vitro for various applications but also could be further developed as conventional therapeutics to stimulate patients' endogenous cells to repair and regenerate in vivo. Here, we focus on recent progress in the use of small molecules in stem cell biology and regenerative medicine. Expected final online publication date for the Annual Review of Biomedical Engineering Volume 13 is July 15, 2011. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates. PMID: 21513460 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | The effects of culture on genomic imprinting profiles in human embryonic and fetal mesenchymal stem cells. Epigenetics. 2011 Jan 1;6(1):52-62 Authors: Frost J, Monk D, Moschidou D, Guillot PV, Stanier P, Minger SL, Fisk NM, Moore HD, Moore GE Human embryonic stem (hES) cells and fetal mesenchymal stem cells (fMSC) offer great potential for regenerative therapy strategies. It is therefore important to characterise the properties of these cells in vitro. One major way the environment impacts on cellular physiology is through changes to epigenetic mechanisms. Genes subject to epigenetic regulation via genomic imprinting have been characterised extensively. The integrity of imprinted gene expression therefore provides a measurable index for epigenetic stability. Allelic expression of 26 imprinted genes and DNA methylation at associated differentially methylated regions (DMRs) was measured in fMSC and hES cell lines. Both cell types exhibited monoallelic expression of 13 imprinted genes, biallelic expression of six imprinted genes, and there were seven genes that differed in allelic expression between cell lines. fMSCs exhibited the differential DNA methylation patterns associated with imprinted expression. This was unexpected given that gene expression of several imprinted genes was biallelic. However, in hES cells, differential methylation was perturbed. These atypical methylation patterns did not correlate with allelic expression. Our results suggest that regardless of stem cell origin, in vitro culture affects the integrity of imprinted gene expression in human cells. We identify biallelic and variably expressed genes that may inform on overall epigenetic stability. As differential methylation did not correlate with imprinted expression changes we propose that other epigenetic effectors are adversely influenced by the in vitro environment. Since DMR integrity was maintained in fMSC but not hES cells, we postulate that specific hES cell derivation and culturing practices result in changes in methylation at DMRs. PMID: 20864803 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Difficulties in the translation of functionalized biomaterials into regenerative medicine clinical products. Biomaterials. 2011 Jun;32(18):4215-7 Authors: Ratcliffe A There are many ways to influence cell activities, and biomaterials with functional groups attached is an attractive method that clearly has the ability to modulate cell behavior. The evidence is clear that biomaterials, with or without growth factors and cells, have resulted in numerous products for the regenerative medicine field. In contrast the functionalized biomaterial products remain in the development phase. PMID: 21515170 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Characterization of Urine-Derived Stem Cells Obtained from Upper Urinary Tract for Use in Cell-Based Urological Tissue Engineering. Tissue Eng Part A. 2011 Apr 22; Authors: Bharadwaj S, Liu G, Shi Y, Markert CD, Andersson KE, Atala A, Zhang Y Abstract Background: The goals of this study were to characterize urine-derived stem cells obtained from the upper urinary tract (uUSC), induce these cells to differentiate into urothelial and smooth muscle cells and determine whether they could serve as a potential stem cell source for bladder tissue engineering. Materials: Urine samples were collected from 5 patients with normal upper urinary tracts during renal pyeloplasty. Cells were isolated from this urine and extensively expanded in vitro. Results: The mean population doubling of uUSC was 44.0 ± 8.5. The uUSC expressed surface markers associated with mesenchymal stem cells and pericytes. These cells could differentiate into smooth muscle-like cells that expressed smooth muscle-specific gene transcripts and proteins, including α-smooth muscle actin, desmin and myosin, when exposed to TGF-β1 and PDGF-BB. In a collagen lattice assay, these myogenic-differentiated uUSC displayed contractile function that was similar to that seen in native smooth muscle cells. Urothelial-differentiated uUSC expressed urothelial-specific genes and proteins such as uroplakin-Ia and -III, CK-7 and CK-13. Conclusions: uUSC possess expansion and differentiation (urothelial and myogenic) capabilities, and can potentially be used as an alternative cell source in bladder tissue engineering for patients needing cystoplasty. PMID: 21513463 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | From tissue engineering to regenerative medicine-the potential and the pitfalls. Adv Drug Deliv Rev. 2011 Apr 14; Authors: Schenke-Layland K PMID: 21515319 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Stereologic Analysis of Tibial-Plateau Cartilage and Femoral Cancellous Bone in Guinea Pigs With Spontaneous Osteoarthritis. Clin Orthop Relat Res. 2011 Apr 23; Authors: Wang SX, Arsenault L, Hunziker EB BACKGROUND: Two strains of guinea pig develop spontaneous osteoarthritis of the knee. Although the disease evolves at different rates in the two strains, it is not known whether these differences are reflected in the structure of the cartilage and cancellous bone. QUESTIONS/PURPOSES: We determined whether the three-dimensional structure of the tibial-plateau cartilage and femoral cancellous bone differed between the two strains. METHODS: Six Dunkin-Hartley and six GOHI/SPF guinea pigs were evaluated. The animals were sacrificed at 11 months of age. The 24 proximal tibias were used for a stereologic histomorphometric analysis of the tibial-plateau cartilage. The 24 femurs were used for a site-specific, three-dimensional quantitative analysis of the cancellous bone by micro-CT. RESULTS: Compared to the GOHI/SPF guinea pigs, the tibial-plateau cartilage of the Dunkin-Hartley strain had a larger lesion volume (3.8% versus 1.5%) and a thicker uncalcified cartilage layer (0.042 versus 0.035 mm), but a thinner calcified cartilage zone (0.008 versus 0.01 mm) and a thinner subchondral cortical bone plate (0.035 versus 0.039 mm). The femoral cancellous bone in the Dunkin-Hartley strain had a lower bone mineral density (477 versus 509 mg/cm(3)). However, the trabeculae were thicker (3.91 versus 3.53 pixels) and farther apart (7.8 versus 5.6 pixels). The osteoarthritic changes in the cartilage were topographically mirrored in the subchondral bone. They were most severe on the medial side of the joint, particularly in the anterior region. CONCLUSIONS: Spontaneous osteoarthritis in the guinea pig is associated with site-specific changes in the articular cartilage layer, which are topographically mirrored in the underlying subchondral bone. CLINICAL RELEVANCE: Three-dimensional structural information not revealed by two-dimensional radiography may help characterize the stages of osteoarthritis. PMID: 21516362 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Gene therapy vectors with enhanced transfection based on hydrogels modified with affinity peptides. Biomaterials. 2011 Apr 21; Authors: Shepard JA, Wesson PJ, Wang CE, Stevans AC, Holland SJ, Shikanov A, Grzybowski BA, Shea LD Regenerative strategies for damaged tissue aim to present biochemical cues that recruit and direct progenitor cell migration and differentiation. Hydrogels capable of localized gene delivery are being developed to provide a support for tissue growth, and as a versatile method to induce the expression of inductive proteins; however, the duration, level, and localization of expression is often insufficient for regeneration. We thus investigated the modification of hydrogels with affinity peptides to enhance vector retention and increase transfection within the matrix. PEG hydrogels were modified with lysine-based repeats (K4, K8), which retained approximately 25% more vector than control peptides. Transfection increased 5- to 15-fold with K8 and K4 respectively, over the RDG control peptide. K8- and K4-modified hydrogels bound similar quantities of vector, yet the vector dissociation rate was reduced for K8, suggesting excessive binding that limited transfection. These hydrogels were subsequently applied to an in vitro co-culture model to induce NGF expression and promote neurite outgrowth. K4-modified hydrogels promoted maximal neurite outgrowth, likely due to retention of both the vector and the NGF. Thus, hydrogels modified with affinity peptides enhanced vector retention and increased gene delivery, and these hydrogels may provide a versatile scaffold for numerous regenerative medicine applications. PMID: 21514659 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Junctions gone bad: Claudins and loss of the barrier in cancer. Biochim Biophys Acta. 2011 Apr 15; Authors: Turksen K, Troy TC The tight junctions (TJs) of epithelia are responsible for regulating the "fence and gate" function of polarized epithelial cells. It is now well-established that dysregulation of these functions contributes to initiation and progression of cancer. Recently, it has become clear that the Claudins, members of a large family of 23 closely related transmembrane proteins, play a crucial role in formation, integrity and function of TJs, the epithelial permeability barrier and epithelial polarization. A growing body of data indicates that Claudin expression is altered in numerous epithelial cancers in a stage- and tumor-specific manner. While a single universal mechanism is still lacking, accumulating evidence supports a role for epigenetic regulation of Claudin expression in tumorgenesis, with concomitant alterations in barrier function. We review here new insights and challenges in understanding Claudin function in normal physiology and cancer. PMID: 21515339 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Mitochondrial biogenesis drives tumor cell proliferation. Am J Pathol. 2011 May;178(5):1949-52 Authors: Martinez-Outschoorn UE, Pavlides S, Sotgia F, Lisanti MP PMID: 21514412 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Adipose tissue-derived stem cell in vitro differentiation in a three-dimensional dental bud structure. Am J Pathol. 2011 May;178(5):2299-310 Authors: Ferro F, Spelat R, Falini G, Gallelli A, D'Aurizio F, Puppato E, Pandolfi M, Beltrami AP, Cesselli D, Beltrami CA, Ambesi-Impiombato FS, Curcio F Tooth morphogenesis requires sequential and reciprocal interactions between the cranial neural crest-derived mesenchymal cells and the stomadial epithelium, which regulate tooth morphogenesis and differentiation. We show how mesenchyme-derived single stem cell populations can be induced to transdifferentiate in vitro in a structure similar to a dental bud. The presence of stem cells in the adipose tissue has been previously reported. We incubated primary cultures of human adipose tissue-derived stem cells in a dental-inducing medium and cultured the aggregates in three-dimensional conditions. Four weeks later, cells formed a three-dimensional organized structure similar to a dental bud. Expression of dental tissue-related markers was tested assaying lineage-specific mRNA and proteins by RT-PCR, immunoblot, IHC, and physical-chemical analysis. In the induction medium, cells were positive for ameloblastic and odontoblastic markers as both mRNAs and proteins. Also, cells expressed epithelial, mesenchymal, and basement membrane markers with a positional relationship similar to the physiologic dental morphogenesis. Physical-chemical analysis revealed 200-nm and 50-nm oriented hydroxyapatite crystals as displayed in vivo by enamel and dentin, respectively. In conclusion, we show that adipose tissue-derived stem cells in vitro can transdifferentiate to produce a specific three-dimensional organization and phenotype resembling a dental bud even in the absence of structural matrix or scaffold to guide the developmental process. PMID: 21514442 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | [Comparison of the efficiency of three techniques for labeling human adipose-derived stem cells.] Nan Fang Yi Ke Da Xue Xue Bao. 2011 Apr 20;31(4):582-586 Authors: Li KC, Chang Q, Lu F OBJECTIVE: To explore the optimal methods for labeling human adipose-derived stem cells (ASCs). METHODS: ASCs were isolated by collagenase digestion and density gradient centrifugation, and their cell surface markers and ability to differentiate into the adipogenic, chondrogenic, and osteogenic lineages were examined in vitro. Three different cell labeling methods, namely 5 µl DiI, 10 µg/ml BrdU and 50 MOI adenovirus carrying GFP, were used for ASC labeling, and the labeling efficiency were compared at different time points and in different passages using fluorescent microscope. RESULTS: The isolated ASCs were capable of differentiating into adipogenic, osteogenic, chondrogenic lineages with positive stem cell marker expression. At 48 h after DiI staining, 100% of the ASCs emitted red fluorescence in the cytoplasm with fluorescent-negative nuclei, but the fluorescence intensity declined quickly after cell passaging. With 10 µg/ml BrdU, 90% of the cells showed green fluorescence in the cell nuclei at 48 h after the labeling, but the positivity rate also decreased gradually after cell passaging. Cell labeling with GFP adenovirus showed more stable labeling efficiency, and green fluorescence was detected at 24 h after labeling, and even till 5 days later more than 90% of the ASCs remained positive without an obvious attenuation of the fluorescent intensity even after cell passaging. CONCLUSION: All the 3 techniques are applicable for labeling ASCs. Cell labeling with DiI and BrdU can be convenient and economic and well serve the purpose of short-term labeling. Adenovirous carrying GFP gene is the optimal choice for long-term ASC tracing. PMID: 21515448 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Real time responses of fibroblasts to plastically compressed fibrillar collagen hydrogels. Biomaterials. 2011 Apr 22; Authors: Ghezzi CE, Muja N, Marelli B, Nazhat SN In vitro reconstituted type I collagen hydrogels are widely utilized for tissue engineering studies. However, highly hydrated collagen (HHC) gels exhibit insufficient mechanical strength and unstable geometrical properties, thereby limiting their therapeutic application. Plastic compression (PC) is a simple and reproducible approach for the immediate production of dense fibrillar collagen (DC) scaffolds which demonstrate multiple improvements for tissue engineered constructs including extracellular matrix (ECM)-like meso scale characteristics, increased mechanical properties (modulus and strength), enhanced cell growth and differentiation, and reduced long-term scaffold deformation. In order to determine at which stage these benefits become apparent, and the underlying mechanisms involved, the immediate response of NIH/3T3 fibroblasts to PC as well as longer-term cell growth within DC scaffolds were examined herein. The real time three-dimensional (3D) distribution of fluorescently labelled cells during PC was sequentially monitored using confocal laser scanning microscopy (CLSM), observing excellent cell retention and negligible numbers of expelled cells. Relative to cells grown in HHC gels, a significant improvement in cell survival within DC scaffolds was evident as early as day 1. Cell growth and metabolic activity within DC gels were significantly increased over the course of one week. While cells within DC scaffolds reached confluency, an inhomogeneous distribution of cells was present in HHC gels, as detected using x-ray computed micro-tomography analysis of phosphotungstic acid labelled cells and CLSM, which both showed a significant cell loss within the HHC core. Therefore, PC generates a DC gel scaffold without detrimental effects towards seeded cells, surpassing HHC gels as a 3D scaffold for tissue engineering. PMID: 21514662 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Preparation and characterization of composite nanofibers of polycaprolactone and nanohydroxyapatite for osteogenic differentiation of mesenchymal stem cells. Colloids Surf B Biointerfaces. 2011 Apr 5; Authors: Chen JP, Chang YS Nanocomposites of nanohydroxyapatite (nHAP) dispersed in poly(ɛ-caprolactone) (PCL) were prepared by electrospinning (ES) to obtain PCL/nHAP nanofibers. Nanofibers with similar diameters (340±30nm) but different nHAP concentrations (0-50%) were fabricated and studied for growth and osteogenic differentiation of bone marrow mesenchymal stem cells (MSCs). The nanofibrous membranes were subjected to detailed analysis for its physicochemical properties by scanning electron microscopy (SEM), thermogravimetric analysis, X-ray diffraction, Fourier-transform infrared spectroscopy, and mechanical tensile testing. nHAP particles (∼30nm diameter) embedded in nanofibers increased the nanofibrous membrane's ultimate stress and the elastic modulus, while decreased the strain at failure. When cultured under an osteogenic stimulation condition on nanofibers, MSCs showed normal phenotypic cell morphology, and time-dependent mineralization and osteogenic differentiation from SEM observations and alkaline phosphatase activity assays. The nanofibers could support the growth of mesenchymal stem cells without compromising their osteogenic differentiation capability up to 21 days and the enhancement of cell differentiation by nHAP is positively correlated with its concentration in the nanofibers. Energy dispersive X-ray analysis of Ca and P elements indicated mineral deposits on the cell surface. The mineralization extent was significantly raised in nanofibers with 50% nHAP where a Ca/P ratio similar to that of bone was found. The present study indicated that electrospun composite PCL/nHAP nanofibrous membranes are suitable for mineralization of MSCs intended for bone tissue engineering. PMID: 21514800 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | From tissue engineering to regenerative medicine-the potential and the pitfalls. Adv Drug Deliv Rev. 2011 Apr 14; Authors: Schenke-Layland K PMID: 21515319 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | An Agent-Based Model for the Investigation of Neovascularization within Porous Scaffolds. Tissue Eng Part A. 2011 Apr 22; Authors: Artel A, Mehdizadeh H, Chiu YC, Brey EM, Cinar A The ability to control blood vessel assembly in polymer scaffolds is important for clinical success in tissue engineering. A mathematical and computational representation of the relationship between scaffold properties and neovascularization may provide a better understanding of the fundamental process itself and help guide the design of new therapeutic approaches. This paper proposes a multi-layered, multi-agent framework to model sprouting angiogenesis in porous scaffolds and examines the impact of pore structure on vessel invasion and network structure. We have defined the speed of vessel sprouting in the agent-based model based on in vivo results in the absence of a polymer scaffold. A number of cases were run to investigate the effect of scaffold pore size on angiogenesis. The simulation results indicate that the rate of scaffold vascularization increases with pore size. Pores of larger size (160-270 μm) support rapid and extensive angiogenesis throughout the scaffold. Model predictions were compared to experimental results of vascularization in porous poly(ethylene glycol) hydrogels in order to validate the results. This model can be used to provide insight into optimal scaffold properties that support vascularization of engineered tissues. PMID: 21513462 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Multiplexed, high-throughput analysis of 3D microtissue suspensions. Integr Biol (Camb). 2010 Oct 4;2(10):517-27 Authors: Chen AA, Underhill GH, Bhatia SN Three-dimensional (3D) tissue models have significantly improved our understanding of structure/function relationships and promise to lead to new advances in regenerative medicine. However, despite the expanding diversity of 3D tissue fabrication methods, approaches for functional assessment have been relatively limited. Here, we describe the fabrication of microtissue (μ-tissue) suspensions and their quantitative evaluation with techniques capable of analyzing large sample numbers and performing multiplexed parallel analysis. We applied this platform to 3D μ-tissues representing multiple stages of liver development and disease including: embryonic stem cells, bipotential hepatic progenitors, mature hepatocytes, and hepatoma cells photoencapsulated in polyethylene glycol hydrogels. Multiparametric μ-tissue cytometry enabled quantitation of fluorescent reporter expression within populations of intact μ-tissues (n≥ 10²-10³) and sorting-based enrichment of subsets for subsequent studies. Further, 3D μ-tissues could be implanted in vivo, respond to systemic stimuli, retrieved and quantitatively assessed. In order to facilitate multiplexed 'pooled' experimentation, fluorescent labeling strategies were developed and utilized to investigate the impact of μ-tissue composition and exposure to soluble factors. In particular, examination of drug/gene interactions on collections of 3D hepatoma μ-tissues indicated synergistic influence of doxorubicin and siRNA knockdown of the anti-apoptotic gene BCL-XL. Collectively, these studies highlight the broad utility of μ-tissue suspensions as an enabling approach for high n, populational analysis of 3D tissue biology in vitro and in vivo. PMID: 20820630 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | The role of ECM proteins and protein fragments in guiding cell behavior in regenerative medicine. Biomaterials. 2011 Jun;32(18):4211-4 Authors: Barker TH The promise of biomaterials design for regenerative medicine tissue engineering is predicated on the fundamental ability to direct or guide specific and highly coordinated cellular behaviors that culminate in the creation of physiologically functional tissues and organs. To date, our efforts have focused primarily on the grafting and presentation of short synthetic peptides with just cause. Short peptides are capable of high levels of control, can be manufactured relatively easily in a highly reproducible manner under GMP guidelines and are readily modified to enable their integration with numerous current and emerging chemistries for biomaterials grafting. However, while extracellular matrix (ECM)-derived peptides have demonstrated their initial purpose of promoting cell adhesion, their general lack of specificity and significantly decreased receptor binding affinities have proven detrimental in attempts to regulate highly specific and integrated processes necessary for tissue regeneration. Unlike adhesion peptides, the natural ECM displays a complex interplay with cells by supporting environmentally sensitive and cell dependent integrin specificity and binding affinity. Furthermore, the adhesion ligands on ECM proteins display a finely tuned and evolutionarily directed spatial periodicity, of which is dynamically controlled through both mechanical and chemical modifications. These and other emerging concepts from matrix biology require our attention if biomaterials design is to fulfill its promise. Here, we are charged with debating the statement 'The use of short synthetic adhesion peptides, like RGD, is the best approach in the design of biomaterials that guide cell behavior for regenerative medicine tissue engineering'. In this Leading Opinion Paper I will focus on aspects of natural ECM proteins and protein fragments that have proven difficult, if not impossible to date, to recapitulate in peptide-based systems. While this represents an argument against the use of peptides per se, it might also be viewed as outlining the challenges and opportunities for the biomaterials field. PMID: 21515169 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | A PEGylated fibrin-based wound dressing with antimicrobial and angiogenic activity. Acta Biomater. 2011 Apr 13; Authors: Seetharaman S, Natesan S, Stowers RS, Mullens C, Baer DG, Suggs LJ, Christy RJ Wounds sustained under battlefield conditions are considered to be contaminated and their initial treatment should focus on decreasing this contamination and thus reducing the possibility of infection. The early and aggressive administration of antimicrobial treatment starting with intervention on the battlefield has resulted in improved patient outcomes and is considered the standard of care. Chitosan microspheres (CSM) loaded with silver sulfadiazine (SSD) were developed via a novel water in oil emulsion technique to address this problem. The SSD-loaded spheres were porous with needle-like structures (attributed to SSD) that were evenly distributed over the spheres. The average particle size of the SSD-CSM was 125-180μm with 76.50±2.8% drug entrapment. As a potential new wound dressing with angiogenic activity SSD-CSM particles were impregnated in polyethylene glycol (PEGylated) fibrin gels. In vitro drug release studies showed that a burst release of 27.02% in 6h was achieved, with controlled release for 72h, with an equilibrium concentration of 27.7% (70μg). SSD-CSM-PEGylated fibrin gels were able to exhibit microbicidal activity at 125 and 100μgml(-1) against Staphylococcus aureus and Pseudomonas aeruginosa, respectively. The in vitro vasculogenic activity of this composite dressing was shown by seeding adipose-derived stem cells (ASC) in SSD-CSM-PEGylated fibrin gels. The ASC spontaneously formed microvascular tube-like structures without the addition of any exogenous factors. This provides a method for the extended release of an antimicrobial drug in a matrix that may provide an excellent cellular environment for revascularization of infected wounds. PMID: 21515420 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Biomechanical forces exert anabolic effects on osteoblasts by activation of SMAD 1/5/8 through type 1 BMP receptor. Biorheology. 2011 Jan 1;48(1):37-48 Authors: Rath B, Nam J, Deschner J, Schaumburger J, Tingart M, Grässel S, Grifka J, Agarwal S Osteoblasts are mechanosensitive cells, which respond to biomechanical stimuli to regulate the bone structure through anabolic and catabolic gene regulation. To examine the effects of mechanical forces on the osteogenic responses through the SMAD signaling in osteoblasts, the cells were cultured in well-characterized mechanoresponsive 3-D scaffolds and exposed to 10% dynamic compressive strain (Cmp) at 1 Hz. Subsequently, SMAD phosphorylation and osteogenic gene induction was examined. Osteoblasts cultured in 3-D scaffolds exhibited increased constitutive SMAD 1/5/8 phosphorylation, as compared to monolayers cultures. This SMAD 1/5/8 phosphorylation was further upregulated after 10, 30 and 60 min in response to Cmp, exhibiting a peak activation at 30 min. No significant changes in SMAD2 phosphorylation were observed, suggesting signals generated by Cmp may not activate the Transforming Growth Factor-β signaling cascade. Subsequently, biomechanical stimulation-induced SMAD 1/5/8 phosphorylation upregulated the expression of osteogenic genes such as Osteoprotegrin, Msx2 and Runx2. Dorsomorphin, a selective inhibitor of the bone morphogenetic protein (BMP) receptor type 1 (BMPR1), blocked Cmp-induced SMAD 1/5/8 phosphorylation, as well as Osteoprotegrin, Msx2 and Runx2 gene expression. Collectively, the present findings demonstrate that biomechanical stimulation of osteoblasts activates SMAD 1/5/8 in the BMP signaling pathway through BMPR1 and may enhance osteogenesis by upregulating SMAD-dependent osteogenic genes. PMID: 21515935 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | In vitro models for the evaluation of angiogenic potential in bone engineering. Acta Pharmacol Sin. 2011 Jan;32(1):21-30 Authors: Cenni E, Perut F, Baldini N Blood vessels have a fundamental role both in skeletal homeostasis and in bone repair. Angiogenesis is also important for a successful bone engineering. Therefore, scaffolds should be tested for their ability to favour endothelial cell adhesion, proliferation and functions. The type of endothelial cell to use for in vitro assays should be carefully considered, because the properties of these cells may depend on their source. Morphological and functional relationships between endothelial cells and osteoblasts are evaluated with co-cultures, but this model should still be standardized, particularly for distinguishing the two cell types. Platelet-rich plasma and recombinant growth factors may be useful for stimulating angiogenesis. PMID: 21042285 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Evaluation of in vitro spermatogenesis using poly(D,L-lactic-co-glycolic acid) (PLGA)-based macroporous biodegradable scaffolds. J Tissue Eng Regen Med. 2011 Feb;5(2):130-7 Authors: Lee JH, Oh JH, Lee JH, Kim MR, Min CK Successful in vitro differentiation of spermatogenic cells into spermatids appears to offer extremely attractive potential for the treatment of impaired spermatogenesis and male infertility. Experimental evidence indicates that biocompatible polymers may improve in vitro reconstitution and regeneration of tissues of various origins. Here, we fabricated highly porous biodegradable poly(D,L-lactic-co-glycolic acid) or PLGA co-polymer scaffolds by combining the gas-foaming and salt-leaching methods, using ammonium bicarbonate as a porogen, which allowed us to generate polymer scaffolds with a high density of interconnected pores of 400-500 µm in average diameter, concomitant with a high malleability to mould a wide range of temporal tissue scaffolds requiring a specific shape and geometry. The PLGA scaffolds were biocompatible and biodegradable, as evidenced by the fact that they survived almost 3 month long subcutaneous xenografting into immunodeficient host mice and became easily destroyable after recovery. Immature rat testicular cells that were seeded onto the surface of the scaffold exhibited about 65% seeding efficiency and up to 75% viability after 18 days in culture. Furthermore, our scaffolds enhanced the proliferation and differentiation of spermatogenic germ cells to a greater extent than conventional in vitro culture methods, such as monolayer or organ culture. Taken together, an implication of the present findings is that the PLGA-based macroporous scaffold may provide a novel means by which spermatocytes could be induced to differentiate into presumptive spermatids. PMID: 20603864 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Outstanding survival and regeneration process by the use of intelligent acellular dermal matrices and mesenchymal stem cells in a burn pig model. Transplant Proc. 2010 Dec;42(10):4275-8 Authors: Mansilla E, Spretz R, Larsen G, Nuñez L, Drago H, Sturla F, Marin GH, Roque G, Martire K, Díaz Aquino V, Bossi S, Gardiner C, Lamonega R, Lauzada N, Cordone J, Raimondi JC, Tau JM, Biasi NR, Marini JE, Patel AN, Ichim TE, Riordan N, Maceira A A pig model with a deep large burn was used to study the regeneration process induced by mesenchymal stem cells (MSCs) and acellular pig dermal matrices, made intelligent by the combination with biodegradable nanofibers loaded with growth factors (granulocyte-macrophage colony-stimulating factor and epidermal growth factor) and coated with the anti-CD44 monoclonal antibody (intelligent acellular dermal matrices, IADMs). These IADMs are specially designed to integrate in the wound bed as new biological scaffolds as well as to specifically recruit and attach circulating and/or externally applied MSCs through the anti-CD44 antibody while delivering precise amounts of growth factors. In this way, the reparative process as well as the aesthetic and functional results were enhanced in our burn model. The animal survived, the wound was completely closed, and total regeneration of the skin was obtained without much scarring. Surprisingly, hair follicles and other skin appendages developed despite the severity and deepness of the burn. Even burned muscles and ribs seemed to have undergone a regenerative process by the end of the study. Based on these findings, we have proposed the use of IADMs and autologous, allogeneic or xenogeneic MSCs, as a new paradigm for the future treatment of large burns and probably other dermatological and cosmetic human conditions. PMID: 21168681 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Characterization of Urine-Derived Stem Cells Obtained from Upper Urinary Tract for Use in Cell-Based Urological Tissue Engineering. Tissue Eng Part A. 2011 Apr 22; Authors: Bharadwaj S, Liu G, Shi Y, Markert CD, Andersson KE, Atala A, Zhang Y Abstract Background: The goals of this study were to characterize urine-derived stem cells obtained from the upper urinary tract (uUSC), induce these cells to differentiate into urothelial and smooth muscle cells and determine whether they could serve as a potential stem cell source for bladder tissue engineering. Materials: Urine samples were collected from 5 patients with normal upper urinary tracts during renal pyeloplasty. Cells were isolated from this urine and extensively expanded in vitro. Results: The mean population doubling of uUSC was 44.0 ± 8.5. The uUSC expressed surface markers associated with mesenchymal stem cells and pericytes. These cells could differentiate into smooth muscle-like cells that expressed smooth muscle-specific gene transcripts and proteins, including α-smooth muscle actin, desmin and myosin, when exposed to TGF-β1 and PDGF-BB. In a collagen lattice assay, these myogenic-differentiated uUSC displayed contractile function that was similar to that seen in native smooth muscle cells. Urothelial-differentiated uUSC expressed urothelial-specific genes and proteins such as uroplakin-Ia and -III, CK-7 and CK-13. Conclusions: uUSC possess expansion and differentiation (urothelial and myogenic) capabilities, and can potentially be used as an alternative cell source in bladder tissue engineering for patients needing cystoplasty. PMID: 21513463 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Artificial skin in perspective: concepts and applications. Pigment Cell Melanoma Res. 2011 Feb;24(1):35-50 Authors: Brohem CA, Cardeal LB, Tiago M, Soengas MS, Barros SB, Maria-Engler SS Skin, the largest organ of the human body, is organized into an elaborate layered structure consisting mainly of the outermost epidermis and the underlying dermis. A subcutaneous adipose-storing hypodermis layer and various appendages such as hair follicles, sweat glands, sebaceous glands, nerves, lymphatics, and blood vessels are also present in the skin. These multiple components of the skin ensure survival by carrying out critical functions such as protection, thermoregulation, excretion, absorption, metabolic functions, sensation, evaporation management, and aesthetics. The study of how these biological functions are performed is critical to our understanding of basic skin biology such as regulation of pigmentation and wound repair. Impairment of any of these functions may lead to pathogenic alterations, including skin cancers. Therefore, the development of genetically controlled and well characterized skin models can have important implications, not only for scientists and physicians, but also for manufacturers, consumers, governing regulatory boards and animal welfare organizations. As cells making up human skin tissue grow within an organized three-dimensional (3D) matrix surrounded by neighboring cells, standard monolayer (2D) cell cultures do not recapitulate the physiological architecture of the skin. Several types of human skin recombinants, also called artificial skin, that provide this critical 3D structure have now been reconstructed in vitro. This review contemplates the use of these organotypic skin models in different applications, including substitutes to animal testing. PMID: 21029393 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | The role of short synthetic adhesion peptides in regenerative medicine; The debate. Biomaterials. 2011 Jun;32(18):4195-7 Authors: Williams DF This paper constitutes the judgment on the four-paper debate in the same issue of this journal on the motion that ''the use of short synthetic adhesion peptides, like RGD, is the best approach in the design of biomaterials that guide cell behavior for regenerative medicine and tissue engineering". It is concluded that this motion is carried on the basis that the concept of installing multifunctionality and modularity in peptides is the more powerful concept, coupled with the potential attractiveness of retaining a chemically defined product, without any possibilities of disease transmission, and relative inexpensiveness and clearer regulatory pathways. PMID: 21515166 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Effective combination of aligned nanocomposite nanofibers and human unrestricted somatic stem cells for bone tissue engineering. Acta Pharmacol Sin. 2011 Apr 25; Authors: Bakhshandeh B, Soleimani M, Ghaemi N, Shabani I Aim:Bioartificial bone tissue engineering is an increasingly popular technique to solve bone defect challenges. This study aimed to investigate the interactions between matrix composition and appropriate cell type, focusing on hydroxyapatite (HA), to achieve a more effective combination for bone regeneration.Methods:Human unrestricted somatic stem cells (USSCs) were isolated from placental cord blood. The cellular and molecular events during the osteo-induction of USSCs were evaluated for 21 d under the following conditions: (1) in basal culture, (2) supplemented with hydroxyapatite nanoparticle (nHA) suspension, and (3) seeded on electrospun aligned nanofibrous poly-ɛ-caprolactone/poly-L-lactic acid/nHA (PCL/PLLA/nHA) scaffolds. The scaffolds were characterized using scanning electron microscope (SEM), fourier transform infrared spectroscopy (FTIR) and tensile test.Results:Maintenance of USSCs for 21 d in basal or osteogenic culture resulted in significant increase in osteoblast differentiation. With nHA suspension, even soluble osteo-inductive additives were ineffective, probably due to induced apoptosis of the cells. In contrast to the hindrance of proliferation by nHA suspension, the scaffolds improved cell growth. The scaffolds mimic the nanostructure of natural bone matrix with the combination of PLLA/PCL (organic phase) and HA (inorganic phase) offering a favorable surface topography, which was demonstrated to possess suitable properties for supporting USSCs. Quantitative measurement of osteogenic markers, enzymatic activity and mineralization indicated that the scaffolds did not disturb, but enhanced the osteogenic potential of USSCs. Moreover, the alignment of the fibers led to cell orientation during cell growth.Conclusion:The results demonstrated the synergism of PCL/PLLA/nHA nanofibrous scaffolds and USSCs in the augmentation of osteogenic differentiation. Thus, nHA grafted into PCL/PLLA scaffolds can be a suitable choice for bone tissue regeneration.Acta Pharmacologica Sinica advance online publication, 25 Apr 2011; doi: 10.1038/aps.2011.8. PMID: 21516135 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Autofluorescence assessment of extracellular matrices of a cartilage-like tissue construct using a fluorescent image analyser. J Tissue Eng Regen Med. 2011 Feb;5(2):163-8 Authors: Hagiwara Y, Hattori K, Aoki T, Ohgushi H, Ito H For the assessment of tissue-engineered cartilage, easy and precise evaluation methods are anticipated. We have investigated the use of a fluorescent image analyser and the application of fluorescence intensity measurements to describe and evaluate a tissue-engineered cartilage-like tissue construct. We evaluated the matrix production of a tissue-engineered cartilage-like tissue (TCL) construct in vitro using the image analyser. The autofluorescence intensity of the TCL construct was determined using the image analyser driven by a software program after 1, 2, 3 and 4 weeks of culture. The autofluorescence intensity of the TCL construct gradually increased with increasing culture time. However, there was very little change in the autofluorescence intensity of gels without articular chondrocytes as controls. The relationship between the autofluorescence intensity of the TCL construct and the hydroxyproline content, reflecting the collagen content, was investigated. There was a significant correlation between the two items. Our findings demonstrate the ability of a commercial fluorescent image analyser to evaluate the matrix production of the TCL construct in vitro on the basis of objective data, such as the autofluorescence intensity. Therefore, the image analyser can be used to judge the success or failure of the in vitro culture process of TCL construct without a biopsy. Copyright © 2010 John Wiley & Sons, Ltd. PMID: 20734346 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Evolving the use of peptides as components of biomaterials. Biomaterials. 2011 Jun;32(18):4198-204 Authors: Collier JH, Segura T This manuscript is part of a debate on the statement that "the use of short synthetic adhesion peptides, like RGD, is the best approach in the design of biomaterials that guide cell behavior for regenerative medicine and tissue engineering". We take the position that although there are some acknowledged disadvantages of using short peptide ligands within biomaterials, it is not necessary to discard the notion of using peptides within biomaterials entirely, but rather to reinvent and evolve their use. Peptides possess advantageous chemical definition, access to non-native chemistries, amenability to de novo design, and applicability within parallel approaches. Biomaterials development programs that require such aspects may benefit from a peptide-based strategy. PMID: 21515167 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Impact of pre-existing elastic matrix on TGFβ1 and HA oligomer-induced regenerative elastin repair by rat aortic smooth muscle cells. J Tissue Eng Regen Med. 2011 Feb;5(2):85-96 Authors: Gacchina CE, Ramamurthi A Regenerating elastic matrices lost to disease (e.g. in aneurysms) is vital to re-establishing vascular homeostasis but is challenged by the poor elastogenicity of post-neonatal cells. We previously showed that exogenous hyaluronan oligomers (HA-o) and TGFβ1 synergistically enhance tropo- and matrix elastin deposition by healthy adult rat aortic SMCs (RASMCs). Towards treating aortic aneurysms (AAs), which exhibit cause- and site-specific heterogeneity in matrix content/structure and contain proteolytically-injured SMCs, we investigated the impact of pre-existing elastic matrix degeneration on elastogenic induction of injured RASMCs. Elastin-rich RASMC layers at 21 days of culture were treated with 0.15 U/ml (PPE15) and 0.75 U/ml (PPE75) porcine pancreatic elastase to degrade the elastic matrix variably, or left uninjured (control). One set of cultures was harvested at 21 days, before and after injury, to quantify viable cell count, matrix elastin loss. Other injured cell layers were cultured to 42 days with or without factors (0.2 µg/ml HA oligomers, 1 ng/ml TGFβ1). We showed that: (a) the ability of cultures to self-repair and regenerate elastic matrices following proteolysis is limited when elastolysis is severe; (b) HA oligomers and TGFβ1 elastogenically stimulate RASMCs in mildly-injured (i.e. PPE15) cultures to restore both elastic matrix amounts and elastic fibre deposition to levels in healthy cultures; and (c) in severely injured (i.e. PPE75) cultures, the factors stimulate matrix elastin synthesis and crosslinking, although not to control levels. The outcomes underscore the need to enhance elastogenic factor doses based on the severity of elastin loss. This study will help in customizing therapies for elastin regeneration within AAs, based on cause and location. PMID: 20653044 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Induced pluripotent stem cells for neural tissue engineering. Biomaterials. 2011 Apr 21; Authors: Wang A, Tang Z, Park IH, Zhu Y, Patel S, Daley GQ, Li S Induced pluripotent stem cells (iPSCs) hold great promise for cell therapies and tissue engineering. Neural crest stem cells (NCSCs) are multipotent and represent a valuable system to investigate iPSC differentiation and therapeutic potential. Here we derived NCSCs from human iPSCs and embryonic stem cells (ESCs), and investigated the potential of NCSCs for neural tissue engineering. The differentiation of iPSCs and the expansion of derived NCSCs varied in different cell lines, but all NCSC lines were capable of differentiating into mesodermal and ectodermal lineages, including neural cells. Tissue-engineered nerve conduits were fabricated by seeding NCSCs into nanofibrous tubular scaffolds, and used as a bridge for transected sciatic nerves in a rat model. Electrophysiological analysis showed that only NCSC-engrafted nerve conduits resulted in an accelerated regeneration of sciatic nerves at 1 month. Histological analysis demonstrated that NCSC transplantation promoted axonal myelination. Furthermore, NCSCs differentiated into Schwann cells and were integrated into the myelin sheath around axons. No teratoma formation was observed for up to 1 year after NCSC transplantation in vivo. This study demonstrates that iPSC-derived multipotent NCSCs can be directly used for tissue engineering and that the approach that combines stem cells and scaffolds has tremendous potential for regenerative medicine applications. PMID: 21514663 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Continuum-level modelling of cellular adhesion and matrix production in aggregates. Comput Methods Biomech Biomed Engin. 2011 May;14(5):403-10 Authors: Geris L, Ashbourn JM, Clarke T Key regulators in tissue-engineering processes such as cell culture and cellular organisation are the cell-cell and cell-matrix interactions. As mathematical models are increasingly applied to investigate biological phenomena in the biomedical field, it is important, for some applications, that these models incorporate an adequate description of cell adhesion. This study describes the development of a continuum model that represents a cell-in-gel culture system used in bone-tissue engineering, namely that of a cell aggregate embedded in a hydrogel. Cell adhesion is modelled through the use of non-local (integral) terms in the partial differential equations. The simulation results demonstrate that the effects of cell-cell and cell-matrix adhesion are particularly important for the survival and growth of the cell population and the production of extracellular matrix by the cells, concurring with experimental observations in the literature. PMID: 21516526 [PubMed - in process] | | | | | | | | | |  | |  | |  |
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