|  |  |  | | | | | TE-RegenMed-StemCell feed | | | | | | | | | | | | | | | | | | | Mesenchymal stem cell therapy regenerates the native bone-tendon junction after surgical repair in a degenerative rat model. PLoS One. 2010;5(8): Authors: Nourissat G, Diop A, Maurel N, Salvat C, Dumont S, Pigenet A, Gosset M, Houard X, Berenbaum F BACKGROUND: The enthesis, which attaches the tendon to the bone, naturally disappears with aging, thus limiting joint mobility. Surgery is frequently needed but the clinical outcome is often poor due to the decreased natural healing capacity of the elderly. This study explored the benefits of a treatment based on injecting chondrocyte and mesenchymal stem cells (MSC) in a new rat model of degenerative enthesis repair. METHODOLOGY: The Achilles' tendon was cut and the enthesis destroyed. The damage was repaired by classical surgery without cell injection (group G1, n = 52) and with chondrocyte (group G2, n = 51) or MSC injection (group G3, n = 39). The healing rate was determined macroscopically 15, 30 and 45 days later. The production and organization of a new enthesis was assessed by histological scoring of collagen II immunostaining, glycoaminoglycan production and the presence of columnar chondrocytes. The biomechanical load required to rupture the bone-tendon junction was determined. PRINCIPAL FINDINGS: The spontaneous healing rate in the G1 control group was 40%, close to those observed in humans. Cell injection significantly improved healing (69%, p = 0.0028 for G2 and p = 0.006 for G3) and the load-to-failure after 45 days (p<0.05) over controls. A new enthesis was clearly produced in cell-injected G2 and G3 rats, but not in the controls. Only the MSC-injected G3 rats had an organized enthesis with columnar chondrocytes as in a native enthesis 45 days after surgery. CONCLUSIONS: Cell therapy is an efficient procedure for reconstructing degenerative entheses. MSC treatment produced better organ regeneration than chondrocyte treatment. The morphological and biomechanical properties were similar to those of a native enthesis. PMID: 20805884 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Review article: stem cell therapies for inflammatory bowel disease - efficacy and safety. Aliment Pharmacol Ther. 2010 Aug 19; Authors: GarcÃa-Bosch O, Ricart E, Panés J Background Drugs available for the treatment of inflammatory bowel disease fail to induce and maintain remission in a significant number of patients. Aim To assess the value of stem cell therapies for treatment of inflammatory bowel disease based on published studies. Methods Publications were identified through a MEDLINE search using the Medical Subject Heading terms: inflammatory bowel diseases, or Crohn's disease, or ulcerative colitis, and stem cell, or stromal cell or transplant. Results Haematopoietic stem cell therapy as a primary treatment for inflammatory bowel disease was originally supported by animal experiments, and by remissions in patients undergoing transplant for haematological disorders. Later, transplantation specifically performed for patients with refractory Crohn's disease showed long-lasting clinical remission and healing of inflammatory intestinal lesions. Use of autologous nonmyeloablative regimens and concentration of the procedures in centres with large experience are key in reducing treatment-related mortality. Initial trials of mesenchymal stem cell therapy with local injection in Crohn's perianal fistulas had positive results. Conclusions Autologous haematopoietic stem cell transplant changes the natural course of Crohn's disease, and may be a therapeutic option in patients with refractory disease if surgery is not feasible due to disease location or extension. PMID: 20804451 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Micropatterning and characterization of electrospun poly(epsilon-caprolactone)/gelatin nanofiber tissue scaffolds by femtosecond laser ablation for tissue engineering applications. Biotechnol Bioeng. 2010 Sep 1; Authors: Lim YC, Johnson J, Fei Z, Wu Y, Farson DF, Lannutti JJ, Choi HW, Lee LJ Experimental investigations aimed at assessing the effectiveness of femtosecond laser ablation for creating microscale features on electrospun poly(epsilon-caprolactone) (PCL)/gelatin nanofiber tissue scaffold capable of controlling cell distribution are described. Statistical comparisons of the fiber diameter and surface porosity on laser-machined and as-spun surface were made and results showed that laser ablation did not change the fiber surface morphology. The minimum feature size that could be created on electrospun nanofiber surfaces by direct-write ablation was measured over a range of laser pulse energies. The minimum feature size that could be created was limited only by the pore size of the scaffold surface. The chemical states of PCL/gelatin nanofiber surfaces were measured before and after femtosecond laser machining by Attenuated Total Reflectance Fourier Transform Infrared (ATR- FTIR) spectroscopy and x-ray photoelectron spectroscopy (XPS) and showed that laser machining produced no changes in the chemistry of the surface. In vitro, mES cells were cultured on as-spun surfaces and in laser-machined microwells. Cell densities were found to be statistically indistinguishable after one and two days of growth. Additionally, confocal microscope imaging confirmed that spreading of mES cells cultured within laser-machined microwells was constrained by the cavity walls, the expected and desired function of these cavities. The geometric constraint caused statistically significant smaller density of cells in microwells after three days of growth. It was concluded that femtosecond laser ablation is an effective process for microscale structuring of these electrospun nanofiber tissue scaffold surfaces. (c) 2010 Wiley Periodicals, Inc. PMID: 20812254 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Tissue engineering: Vision restored. Nature. 2010 Sep 2;467(7311):8 Authors: PMID: 20811413 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Tissue engineering in plastic surgery: a review. Plast Reconstr Surg. 2010 Sep;126(3):858-68 Authors: Wong VW, Rustad KC, Longaker MT, Gurtner GC Novel tissue- and organ-engineering strategies are needed to address the growing need for replacement biological parts. Collective progress in stem cell technology, biomaterials, engineering, and molecular medicine has advanced the state of regenerative medicine, yet many hurdles to clinical translation remain. Plastic surgeons are in an ideal position to capitalize on emerging technologies and will be at the forefront of transitioning basic science research into the clinical reconstructive arena. This review highlights fundamental principles of bioengineering, recent progress in tissue-specific engineering, and future directions for this exciting and rapidly evolving area of medicine. PMID: 20811219 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Depot-specific variation in the osteogenic and adipogenic potential of human adipose-derived stromal cells. Plast Reconstr Surg. 2010 Sep;126(3):822-34 Authors: Levi B, James AW, Glotzbach JP, Wan DC, Commons GW, Longaker MT BACKGROUND:: Adipose-derived stromal cells hold promise for use in tissue regeneration. However, multiple facets of their biology remain unclear. The authors examined the variations in osteogenesis and adipogenesis in adipose-derived stromal cells between subcutaneous fat depots and potential molecular causes. METHODS:: Adipose-derived stromal cells were isolated from human patients from subcutaneous fat depots, including arm, flank, thigh, and abdomen (n = 5 patients). Osteogenic and adipogenic differentiation was performed (alkaline phosphatase, alizarin red, and oil red O staining, and quantitative real-time polymerase chain reaction). Co-cultures were established to assess the paracrine effect of human adipose-derived stromal cells on mouse osteoblasts. Finally, HOX gene expression was analyzed by quantitative real-time polymerase chain reaction. RESULTS:: Subcutaneous fat depots retain markedly different osteogenic and adipogenic potentials. Osteogenesis was most robust in adipose-derived stromal cells from the flank and thigh, as compared with those from the arm and abdomen (p < 0.05 by all markers examined). This was accompanied by elevations of BMP4 and BMPR1B (p < 0.05 by all markers examined). The osteogenic advantage of cells from the flank and thigh was again observed when analyzing the paracrine effects of these cells. Conversely, those cells isolated from the flank had a lesser ability to undergo adipogenic differentiation. Adipose-associated HOX genes were less expressed in flank-derived adipose-derived stromal cells. CONCLUSIONS:: Variations exist between fat depots in terms of adipose-derived stromal cell osteogenic and adipogenic differentiation. Differences in HOX expression and bone morphogenetic protein signaling may underlie these observations. This study indicates that the choice of fat depot derivation of adipose-derived stromal cells may be an important one for future efforts in tissue engineering. PMID: 20811215 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Effects of surfactant and gentle agitation on inkjet dispensing of living cells. Biofabrication. 2010 Jun;2(2):025003 Authors: Parsa S, Gupta M, Loizeau F, Cheung KC Inkjet dispensing is a promising method for patterning cells and biomaterials for tissue engineering applications. In a novel approach, this work uses a biocompatible surfactant to improve the reliability of droplet formation in piezoelectric drop-on-demand inkjet printing of Hep G2 hepatocytes onto hydrogels. During a long printing process, cell aggregation and sedimentation within the inkjet reservoir can lead to inconsistent printing results. In order to improve repeatability, the effects of gentle agitation on cell sedimentation and aggregation within the inkjet reservoir were also investigated. Cell viability and proliferation when printed onto prepared collagen substrates were assessed using live/dead staining and the Alamar Blue metabolic assay. The addition of 0.05% Pluronic as a surfactant did not reduce cell viability, which remained above 95% 2 days after printing. The surfactant improved the reliability of droplet formation. Although gentle stirring of the inkjet reservoir was sufficient to maintain a cell suspension and reduce sedimentation, aggregation within the suspension continued to affect printing performance over a 180 min printing period. PMID: 20811131 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Fabricating a pearl/PLGA composite scaffold by the low-temperature deposition manufacturing technique for bone tissue engineering. Biofabrication. 2010 Jun;2(2):025002 Authors: Xu M, Li Y, Suo H, Yan Y, Liu L, Wang Q, Ge Y, Xu Y Here we developed a composite scaffold of pearl/poly(lactic-co-glycolic acid) (pearl/PLGA) utilizing the low-temperature deposition manufacturing (LDM). LDM makes it possible to fabricate scaffolds with designed microstructure and macrostructure, while keeping the bioactivity of biomaterials by working at a low temperature. Process optimization was carried out to fabricate a mixture of pearl powder, PLGA and 1,4-dioxane with the designed hierarchical structures, and freeze-dried at a temperature of -40 degrees C. Scaffolds with square and designated bone shape were fabricated by following the 3D model. Marrow stem cells (MSCs) were seeded on the pearl/PLGA scaffold and then cultured in a rotating cell culture system. The adhesion, proliferation and differentiation of MSCs into osteoblasts were determined using scanning electronic microscopy, WST-1 assay, alkaline phosphatase activity assay, immunofluorescence staining and real-time reverse transcription polymerase chain reaction. The results showed that the composite scaffold had high porosity (81.98 +/- 3.75%), proper pore size (micropores: <10 microm; macropore: 495 +/- 54 microm) and mechanical property (compressive strength: 0.81 +/- 0.04 MPa; elastic modulus: 23.14 +/- 0.75 MPa). The pearl/PLGA scaffolds exhibited better biocompatibility and osteoconductivity compared with the tricalcium phosphate/PLGA scaffold. All these results indicate that the pearl/PLGA scaffolds fulfill the basic requirements of bone tissue engineering scaffold. PMID: 20811130 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Tissue engineering by self-assembly and bio-printing of living cells. Biofabrication. 2010 Jun;2(2):022001 Authors: Jakab K, Norotte C, Marga F, Murphy K, Vunjak-Novakovic G, Forgacs G Biofabrication of living structures with desired topology and functionality requires the interdisciplinary effort of practitioners of the physical, life and engineering sciences. Such efforts are being undertaken in many laboratories around the world. Numerous approaches are pursued, such as those based on the use of natural or artificial scaffolds, decellularized cadaveric extracellular matrices and, most lately, bioprinting. To be successful in this endeavor, it is crucial to provide in vitro micro-environmental clues for the cells resembling those in the organism. Therefore, scaffolds, populated with differentiated cells or stem cells, of increasing complexity and sophistication are being fabricated. However, no matter how sophisticated scaffolds are, they can cause problems stemming from their degradation, eliciting immunogenic reactions and other a priori unforeseen complications. It is also being realized that ultimately the best approach might be to rely on the self-assembly and self-organizing properties of cells and tissues and the innate regenerative capability of the organism itself, not just simply prepare tissue and organ structures in vitro followed by their implantation. Here we briefly review the different strategies for the fabrication of three-dimensional biological structures, in particular bioprinting. We detail a fully biological, scaffoldless, print-based engineering approach that uses self-assembling multicellular units as bio-ink particles and employs early developmental morphogenetic principles, such as cell sorting and tissue fusion. PMID: 20811127 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Biomatrices and biomaterials for future developments of bioprinting and biofabrication. Biofabrication. 2010 Mar;2(1):014110 Authors: Nakamura M, Iwanaga S, Henmi C, Arai K, Nishiyama Y The next step beyond conventional scaffold-based tissue engineering is cell-based direct biofabrication techniques. In industrial processes, various three-dimensional (3D) prototype models have been fabricated using several different rapid prototyping methods, such as stereo-lithography, 3D printing and laser sintering, as well as others, in which a variety of chemical materials are utilized. However, with direct cell-based biofabrication, only biocompatible materials can be used, and the manufacturing process must be performed under biocompatible and physiological conditions. We have developed a direct 3D cell printing system using inkjet and gelation techniques with inkjet droplets, and found that it had good potential to construct 3D structures with multiple types of cells. With this system, we have used alginate and fibrin hydrogel materials, each of which has advantages and disadvantages. Herein, we discuss the roles of hydrogel for biofabrication and show that further developments in biofabrication technology with biomatrices will play a major part, as will developments in manufacturing technology. It is important to explore suitable biomatrices as the next key step in biofabrication techniques. PMID: 20811125 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Bio rapid prototyping by extruding/aspirating/refilling thermoreversible hydrogel. Biofabrication. 2010 Mar;2(1):014108 Authors: Iwami K, Noda T, Ishida K, Morishima K, Nakamura M, Umeda N This paper reports a method for rapid prototyping of cell tissues, which is based on a system that extrudes, aspirates and refills a mixture of cells and thermoreversible hydrogel as a scaffold. In the extruding mode, a cell-mixed scaffold solution in the sol state is extruded from a cooled micronozzle into a temperature-controlled substrate, which keeps the scaffold in the gel state. In the aspiration mode, the opposite process is performed by Bernoulli suction. In the refilling mode, the solution is extruded into a groove created in the aspiration mode. The minimum width of extruded hydrogel pattern is 114 +/- 15 microm by employing a nozzle of diameter 100 microm, and that of aspirated groove was 355 +/- 10 microm using a 500 microm-diameter nozzle. Gum arabic is mixed with the scaffold solution to avoid peeling-off of the gel pattern from the substrate. Patterning of Sf-9 cell tissue is demonstrated, and the stability of the patterned cell is investigated. This system offers a procedure for rapid prototyping and local modification of cell scaffolds for tissue engineering. PMID: 20811123 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | A droplet-based building block approach for bladder smooth muscle cell (SMC) proliferation. Biofabrication. 2010 Mar;2(1):014105 Authors: Xu F, Moon SJ, Emre AE, Turali ES, Song YS, Hacking SA, Nagatomi J, Demirci U Tissue engineering based on building blocks is an emerging method to fabricate 3D tissue constructs. This method requires depositing and assembling building blocks (cell-laden microgels) at high throughput. The current technologies (e.g., molding and photolithography) to fabricate microgels have throughput challenges and provide limited control over building block properties (e.g., cell density). The cell-encapsulating droplet generation technique has potential to address these challenges. In this study, we monitored individual building blocks for viability, proliferation and cell density. The results showed that (i) SMCs can be encapsulated in collagen droplets with high viability (>94.2 +/- 3.2%) for four cases of initial number of cells per building block (i.e. 7 +/- 2, 16 +/- 2, 26 +/- 3 and 37 +/- 3 cells/building block). (ii) Encapsulated SMCs can proliferate in building blocks at rates that are consistent (1.49 +/- 0.29) across all four cases, compared to that of the controls. (iii) By assembling these building blocks, we created an SMC patch (5 mm x 5 mm x 20 microm), which was cultured for 51 days forming a 3D tissue-like construct. The histology of the cultured patch was compared to that of a native rat bladder. These results indicate the potential of creating 3D tissue models at high throughput in vitro using building blocks. PMID: 20811120 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Laser printing of cells into 3D scaffolds. Biofabrication. 2010 Mar;2(1):014104 Authors: Ovsianikov A, Gruene M, Pflaum M, Koch L, Maiorana F, Wilhelmi M, Haverich A, Chichkov B One of the most promising approaches in tissue engineering is the application of 3D scaffolds, which provide cell support and guidance in the initial tissue formation stage. The porosity of the scaffold and internal pore organization influence cell migration and play a major role in its biodegradation dynamics, nutrient diffusion and mechanical stability. In order to control cell migration and cellular interactions within the scaffold, novel technologies capable of producing 3D structures in accordance with predefined design are required. The two-photon polymerization (2PP) technique, used in this report for the fabrication of scaffolds, allows the realization of arbitrary 3D structures with submicron spatial resolution. Highly porous 3D scaffolds, produced by 2PP of acrylated poly(ethylene glycol), are seeded with cells by means of laser-induced forward transfer (LIFT). In this laser printing approach, a propulsive force, resulting from laser-induced shock wave, is used to propel individual cells or cell groups from a donor substrate towards the receiver substrate. We demonstrate that with this technique printing of multiple cell types into 3D scaffolds is possible. Combination of LIFT and 2PP provides a route for the realization of 3D multicellular tissue constructs and artificial ECM engineered on the microscale. PMID: 20811119 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Bioprinting by laser-induced forward transfer for tissue engineering applications: jet formation modeling. Biofabrication. 2010 Mar;2(1):014103 Authors: Mézel C, Souquet A, Hallo L, Guillemot F In this paper, a nanosecond LIFT process is analyzed both from experimental and modeling points of view. Experimental results are first presented and compared to simple estimates obtained from physical analysis, i.e. energy balance, jump relations and analytical pocket dynamics. Then a self-consistent 2D axisymmetric modeling strategy is presented. It is shown that data accessible from experiments, i.e. jet diameter and velocity, can be reproduced. Moreover, some specific mechanisms involved in the rear-surface deformation and jet formation may be described by some scales of hydrodynamic process, i.e. shock waves propagation and expansion waves, as a consequence of the laser heating. It shows that the LIFT process is essentially driven by hydrodynamics and thermal transfer, and that a coupled approach including self-consistent laser energy deposition, heating by thermal conduction and specific models for matter is required. PMID: 20811118 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Combining electrospinning and fused deposition modeling for the fabrication of a hybrid vascular graft. Biofabrication. 2010 Mar;2(1):014102 Authors: Centola M, Rainer A, Spadaccio C, De Porcellinis S, Genovese JA, Trombetta M Tissue engineering of blood vessels is a promising strategy in regenerative medicine with a broad spectrum of potential applications. However, many hurdles for tissue-engineered vascular grafts, such as poor mechanical properties, thrombogenicity and cell over-growth inside the construct, need to be overcome prior to the clinical application. To surmount these shortcomings, we developed a poly-l-lactide (PLLA)/poly-epsilon-caprolactone (PCL) scaffold releasing heparin by a combination of electrospinning and fused deposition modeling technique. PLLA/heparin scaffolds were produced by electrospinning in tubular shape and then fused deposition modeling was used to armor the tube with a single coil of PCL on the outer layer to improve mechanical properties. Scaffolds were then seeded with human mesenchymal stem cells (hMSCs) and assayed in terms of morphology, mechanical tensile strength, cell viability and differentiation. This particular scaffold design allowed the generation of both a drug delivery system amenable to surmount thrombogenic issues and a microenvironment able to induce endothelial differentiation. At the same time, the PCL external coiling improved mechanical resistance of the microfibrous scaffold. By the combination of two notable techniques in biofabrication-electrospinning and FDM-and exploiting the biological effects of heparin, we developed an ad hoc differentiating device for hMSCs seeding, able to induce differentiation into vascular endothelium. PMID: 20811117 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Bioprinting is coming of age: report from the International Conference on Bioprinting and Biofabrication in Bordeaux (3B'09). Biofabrication. 2010 Mar;2(1):010201 Authors: Guillemot F, Mironov V, Nakamura M Abstract The International Conference on Bioprinting and Biofabrication in Bordeaux (3B'09) demonstrated that the field of bioprinting and biofabrication continues to evolve. The increasing number and broadening geography of participants, the emergence of new exciting bioprinting technologies, and the attraction of young investigators indicates the strong growth potential of this emerging field. Bioprinting can be defined as the use of computer-aided transfer processes for patterning and assembling living and non-living materials with a prescribed 2D or 3D organization in order to produce bio-engineered structures serving in regenerative medicine, pharmacokinetic and basic cell biology studies. The use of bioprinting technology for biofabrication of in vitro assay has been shown to be a realistic short-term application. At the same time, the principal feasibility of bioprinting vascularized human organs as well as in vivo bioprinting has been demonstrated. The bioprinting of complex 3D human tissues and constructs in vitro and especially in vivo are exciting, but long-term, applications. It was decided that the 5th International Conference on Bioprinting and Biofabrication would be held in Philadelphia, USA in October 2010. The specially appointed 'Eploratory Committee' will consider the possibility of turning the growing bioprinting community into a more organized entity by creating a new bioprinting and biofabrication society. The new journal Biofabrication was also presented at 3B'09. This is an important milestone per se which provides additional objective evidence that the bioprinting and biofabrication field is consolidating and maturing. Thus, it is safe to state that bioprinting technology is coming of age. 1. Back to Europe The International Conference on Bioprinting and Biofabrication in Bordeaux (3B'09), France, 6-8 July 2009, was held after several international meetings which had been organized previously. The First International Workshop on Bioprinting and Biopatterning [1] was held at the University of Manchester (UK) in September 2004 and was organized by Professor Brian Derby (University of Manchester), Douglas B Chrisey (Naval Research Laboratory, Washington, USA), Richard K Everett (ONR Global, London) and Nuno Reis (Universidade de Beira Interior, Covilha, Portugal). The Second International Workshop on Bioprinting, Biopatterning and Bioassembly was chaired by Vladimir Mironov in 2005 in Charleston (USA) [2]. The Third International Symposium on Bioprinting and Biofabrication was held in Kawasaki (Japan) in November 2006 and was organized by Professor Makoto Nakamura (University of Toyama, Japan). After three years without an international meeting, Fabien Guillemot (INSERM, University of Bordeaux, France) and Professor Makoto Nakamura decided to organize the International Conference on Bioprinting and Biofabrication (3B'09) [3] in Bordeaux, the wine capital of the world. This 4th international meeting on bioprinting was endorsed by TERMIS (Tissue Engineering and Regenerative Medicine International Society) and sponsored by the clusters Advanced Materials in Aquitaine, Materials in Bordeaux and Route des Lasers, INSERM and the Aquitaine Regional Council. It took place at the Burdigala Hotel in a very pleasant and friendly atmosphere where new players, especially the young, were welcome and were given the opportunity to discuss their work. The balanced geographical representation was another important feature of the 3B'09 conference. Indeed, more than 65 scientists and engineers from 11 countries (Belgium, France, Germany, Italy, Japan, Poland, Portugal, Romania, The Netherlands, UK, USA) attended this conference, which included five oral presentation sessions, one poster session and social events organized at the Bordeaux City Hall and at the Château Giscours. 2. Scientific program The scientific program was established to highlight the latest developments associated with bioprinting technologies and biofabrication approaches. Indeed, since the early pioneering works of Thomas Boland (Clemson University, USA) and Vladimir Mironov [4], the bioprinting scientific community has been evolving, bringing together physicists, biologists and physicians [5, 6]. Consequently, the oral presentation sessions covered: (1) the latest developments in bioprinting technologies; (2) the potential to combine the bioprinting process with other biofabrication and rapid prototyping methods; (3) matrices and biomaterials for bioprinting and biofabrication; (4) methods for designing, modeling and biomechanically evaluating 3D constructs; (5) developmental biology and tissue engineering. This scientific schedule emphasized some of the main areas that have to be connected prior to envisaging real applications in regenerative medicine, pharmacokinetic and toxicological studies, where high throughput and high-resolution bioprinting technologies are required. Thus, it is now obvious that, in addition to core bioprinting technologies, biomaterial and bioink properties, rapid prototyping approaches and basic cell biology have to be taken into account within a single perspective. Consequently, while the initial definition of our burgeoning field was formulated at the First International Workshop on Bioprinting and Biopatterning in Manchester (UK) as 'the use of material transfer processes for patterning and assembling biologically relevant materials, molecules, cells, tissues, and biodegradable biomaterials with a prescribed organization to accomplish one or more biological functions', we propose to enlarge this definition to 'the use of computer-aided transfer processes for patterning and assembling living and non-living materials with a prescribed 2D or 3D organization in order to produce bio-engineered structures serving in regenerative medicine, pharmacokinetic and basic cell biology studies'. Regarding the poster session, three young scientists were awarded a bottle of Château Giscours wine for their impressive work. Laureates were Virginie Kériquel in vivo high-throughput biological laser printing of nano-hydroxyapatite in mice calvaria critical sized defects: preliminary results) [7], Kayo Sakaue (Integration of 3D-micro tissue models for creation of tissue chip) [8] and Alberto Rainer (Regeneration of osteochondral segment via mesenchymal stem cells culturing on 3D rapid prototyped scaffolds) [9]. 3. On bioprinting technologies The 3B'09 conference was an excellent opportunity to perform an in-depth review of bioprinting technologies (jet-based and extrusion methods) through a number of exciting talks given by both academics and industrialists. Recent advances in ink-jet bioprinting technologies were reviewed by Professor Thomas Boland and supplemented mainly by the presentations given by Professor Makoto Nakamura [10, 11] and Professor Brian Derby. Regarding laser-based technologies, a series of talks dealing with experimental and modeling approaches of laser-assisted bioprinting (LAB) [12] were given by members of Fabien Guillemot [13] and Boris Chichkov's groups [14] from The University of Bordeaux and Lazer Zentrum in Hannover (Germany), respectively. Utkan Demirci (Harvard University, USA) introduced a new ultrasonic wave-based bioprinting technology, the so-called layer-by-layer 3D tissue epitaxy by cell-laden hydrogel droplets [15] while Hedges and Wirth from Germany presented an aerosol jet bioprinting technology and its applications in surface biofunctionalization [16]. Besides the above-mentioned jet-based bioprinting techniques, many presentations were related to 3D plotting by extrusion processes. Giovani Vozzi's group (University of Pisa, Italy) presented several talks dealing with the micro-fabrication of two- and three-dimensional structures by pressure-assisted micro-syringe (PAM) [17]. Moreover, processing considerations for the 3D plotting of thermoplastic scaffolds were presented by Kim Ragaert (Ghent University, Belgium) [18] and Hendrik John (Sys-Eng, Germany) while Kentaro Iwami (Tokyo University, Japan) described for the first time rapid prototyping by extruding/aspirating/refilling thermoreversible hydrogel [19]. Finally, additional 3D biofabrication methods were presented. Matsusaki et al (Japan) introduced possible, layer-by-layer, short-term applications of bioprinting for biofabrication of tissue chips. Frasca et al (University of Paris-Diderot, France) demonstrated how nanotechnology (magnetic nanoparticles) can be employed in magnetic force-driven tissue engineering. In summary, while each of these technologies displays specific properties such as high resolution, high throughput, low price, bio-safety, etc, it seems obvious that achieving more advanced applications in tissue engineering will require a combination of these tools, and thus a combination of their performances. Moreover, to print human organs, highly integrative approaches should be set up; these should include the development of new biomaterials, the improvement of reverse-engineering and rapid prototyping methods, intensification of scientific gateways between developmental biology and tissue engineering [20] (as introduced by Gabor Forgacs, University of Missouri, USA) [21], in particular with the help of numerical modeling. All these perspectives were discussed during the conference and are reported in the following sections. 4. Scaffold or not scaffold, that is the question! One of the controversial topics of discussion during the conference was the definition of 'scaffold'. Synthetic biodegradable scaffold is considered to be a fundamental principle of the traditional scaffold-based or 'top down' tissue engineering approach [22]. The emerging modular or 'bottom-up' approach is sometimes called 'scaffold-free' [23] or 'scaffold-less'. Some participants insisted that scaffold according to definition is a temporal and removable support. (ABSTRACT TRUNCATED) PMID: 20811115 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | The role of printing parameters and scaffold biopolymer properties in the efficacy of a new hybrid nano-bioprinting system. Biofabrication. 2009 Sep;1(3):035003 Authors: Buyukhatipoglu K, Jo W, Sun W, Clyne AM We created a hybrid nano-bioprinting system, which combines the initial patterning capabilities of direct cell writing with the active patterning capabilities of superparamagnetic nanoparticles. Biofabrication conditions, including printing parameters and scaffold biopolymer properties, may affect cell viability, nanoparticle manipulation and patterning capabilities. Nanoparticles were printed under varied conditions either in the biopolymer or loaded inside cells. Cell viability, alginate viscosity, nanoparticle movement and printing resolution were measured. We now show that while nanoparticles decreased cell viability, nozzle size had no significant effect. High printing pressure decreased cell viability, but viability loss was not accentuated by nanoparticles. High nanoparticle concentrations increased alginate viscosity at higher alginate concentrations. Nanoparticle velocity in response to a magnetic field was a function of nanoparticle diameter and scaffold viscosity, which agreed with a mathematical model of nanoparticle movement. Finally, the nano-bioprinting system resolution and patterning precision were not affected by nanoparticles in the prepolymer solution. These data suggest that nanoparticle incorporation in solid freeform fabrication does not change biofabrication parameters unless high nanoparticle concentrations are used. Future work includes developing vascularized tissue engineering constructs using the nano-bioprinting system. PMID: 20811107 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | A brief review of dispensing-based rapid prototyping techniques in tissue scaffold fabrication: role of modeling on scaffold properties prediction. Biofabrication. 2009 Sep;1(3):032001 Authors: Li MG, Tian XY, Chen XB Artificial scaffolds play vital roles in tissue engineering as they provide a supportive environment for cell attachment, proliferation and differentiation during tissue formation. Fabrication of tissue scaffolds is thus of fundamental importance for tissue engineering. Of the variety of scaffold fabrication techniques available, rapid prototyping (RP) methods have attracted a great deal of attention in recent years. This method can improve conventional scaffold fabrication by controlling scaffold microstructure, incorporating cells into scaffolds and regulating cell distribution. All of these contribute towards the ultimate goal of tissue engineering: functional tissues or organs. Dispensing is typically used in different RP techniques to implement the layer-by-layer fabrication process. This article reviews RP methods in tissue scaffold fabrication, with emphasis on dispensing-based techniques, and analyzes the effects of different process factors on fabrication performance, including flow rate, pore size and porosity, and mechanical cell damage that can occur in the bio-manufacturing process. PMID: 20811104 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Suppression of NF-{kappa}B Increases Bone Formation and Ameliorates Osteopenia in Ovariectomized Mice. Endocrinology. 2010 Sep 1; Authors: Alles N, Soysa NS, Hayashi J, Khan M, Shimoda A, Shimokawa H, Ritzeler O, Akiyoshi K, Aoki K, Ohya K Bone degenerative diseases, including osteoporosis, impair the fine balance between osteoclast bone resorption and osteoblast bone formation. Single-agent therapy for anabolic and anticatabolic effects is attractive as a drug target to ameliorate such conditions. Inhibition of nuclear factor (NF)-kappaB reduces the osteoclast bone resorption. The role of NF-kappaB inhibitors on osteoblasts and bone formation, however, is minimal and not well investigated. Using an established NF-kappaB inhibitor named S1627, we demonstrated that inhibition of NF-kappaB increases osteoblast differentiation and bone formation in vitro by up-regulating the mRNAs of osteoblast-specific genes like type I collagen, alkaline phosphatase, and osteopontin. In addition, S1627 was able to increase bone formation and repair bone defect in a murine calvarial defect model. To determine the effect of NF-kappaB on a model of osteoporosis, we injected two doses of inhibitor (25 and 50 mg/kg . d) twice a day in sham-operated or ovariectomized 12-wk-old mice and killed them after 4 wk. The anabolic effect of S1627 on trabecular bone was determined by micro focal computed tomography and histomorphometry. Bone mineral density of inhibitor-treated ovariectomized animals was significantly increased compared with sham-operated mice. Osteoblast-related indices like osteoblast surface, mineral apposition rate, and bone formation rate were increased in S1627-treated animals in a dose-dependent manner. NF-kappaB inhibition by S1627 increased the trabecular bone volume in ovariectomized mice. Furthermore, S1627 could inhibit the osteoclast number, and osteoclast surface to bone surface. In vitro osteoclastogenesis and bone resorbing activity were dose-dependently reduced by NF-kappaB inhibitor S1627. Taken collectively, our results suggest that NF-kappaB inhibitors are effective in treating bone-related diseases due to their dual anabolic and antiresorptive activities. PMID: 20810563 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | The primordium of a biological joint replacement: Coupling of two stem cell pathways in biphasic ultrarapid compressed gel niches. J Craniomaxillofac Surg. 2010 Aug 30; Authors: Brady MA, Sivananthan S, Mudera V, Liu Q, Wiltfang J, Warnke PH The impaired temporomandibular joint might be the first to benefit from applied tissue engineering techniques because it is small and tissue growth in larger amounts is challenging. Bone and cartilage require different competing environmental conditions to be cultivated in vitro. But coupling both the osteogenic and cartilaginous pathways of mesenchymal stem cell differentiation in homeostasis will be a key essential to grow osteochondral constructs or even the first biological joint replacement in the future. The aim of this study was to test a single source biomaterial and a single source cell type to engineer a biphasic osteochondral construct in vitro for future in vivo implantation. Ultrarapid tissue engineering techniques were used to create the biphasic matrix and primary human mesenchymal stem cells (MSCs) preconditioned in osteogenic and chondrogenic media were then seeded in opposite portions of the hyper-hydrated collagen gel in order to further substantiate the respective bone-like and cartilage-like layers thus potentially customising the collagen scaffold according to patient needs in regards to future biological joint replacements. After incubation for 7 days to allow cell growth and differentiation, mineralization of the bone-like layer was demonstrated using von Kossa staining and biochemical bone markers. The cartilage-like layer was demonstrated using alcian blue staining and biochemical cartilage markers. Integration of the bone-like and cartilage-like layers to simulate a tidemark layer was achieved through partial setting of the gels. Cell tracking was used to further confirm the establishment of distinct cartilage-like and bone-like areas within the single construct. This is the first report of one homogeneous human MSC population differentiating into dissimilar "bone-like" and "cartilage-like" zones hosted in a biphasic ultrarapid compressed gel phase niche and mimicking a primordial joint-like structure. PMID: 20810288 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Expansion of human articular chondrocytes and formation of tissue-engineered cartilage: A step towards exploring a potential use of matrix-induced cell therapy. Tissue Cell. 2010 Aug 30; Authors: Munirah S, Samsudin OC, Aminuddin BS, Ruszymah BH Monolayer culture expansion remains as a fundamental step to acquire sufficient number of cells for 3D constructs formation. It has been well-documented that cell expansion is however accompanied by cellular dedifferentiation. In order to promote cell growth and circumvent cellular dedifferentiation, we evaluated the effects of Transforming Growth Factor Beta-2 (TGF-beta2), Insulin-like Growth Factor-I (IGF-I) and basic Fibroblast Growth Factor (bFGF) combination on articular chondrocytes culture and 'chondrocytes-fibrin' construct formation. Chondrocytes were serially cultured in: (1) F12:DMEM+10% Foetal Bovine Serum (FBS) with growth factors (FD10GFs), (2) F12:DMEM+2%FBS with the growth factors (FD2GFs) and, (3) F12:DMEM+10%FBS without growth factors (FD) as control. Cultured chondrocytes were evaluated by means of growth kinetics parameters, cell cycle analysis, quantitative phenotypic expression of collagen type II, aggrecan core protein sox-9 and collagen type I and, immunochemistry technique. Harvested chondrocytes were incorporated with plasma-derived fibrin and were polymerized to form the 3D constructs and implanted subcutaneously at the dorsum of athymic nude mice for eight (8) weeks. Resulted constructs were assigned for gross inspections and microscopic evaluation using standard histochemicals staining, immunochemistry technique and, quantitative phenotypic expression of cartilage markers to reassure cartilaginous tissue formation. Growth kinetics performance of chondrocytes cultured in three (3) types of culture media from the most to least was in the following order: FD10GFs>FD2GFs>FD. Following growth kinetics analysis, we decided to use FD10GFs and FD (control) for further evaluation and 'chondrocytes-fibrin' constructs formation. Chondrocytes cultured in FD10GFs preserved the normal diploid state (2c) with no evidence of aneuploidy, haploidy or tetraploidy. Expression of cartilage-specific markers namely collagen type II, aggrecan core protein and sox-9 were significantly higher in FD10GFs when compared to control. After implantation, 'chondrocytes-fibrin' constructs exhibited firm, white, smooth and glistening cartilage-like properties. FD10GFs constructs formed better quality cartilage-like tissue than FD constructs in term of overall cartilaginous tissue formation, cells organization and extracellular matrix distribution in the specimens. Cartilaginous tissue formation was confirmed by the presence of lacunae and cartilage-isolated cells embedded within basophilic ground substance. Presence of proteoglycan was confirmed by positive Safranin O staining. Collagen type II exhibited immunopositivity at the pericellular and inter-territorial matrix area. Chondrogenic properties of the construct were further confirmed by the expression of genes encoding collagen type II, aggrecan core protein and sox9. In conclusion, FD10GFs promotes the proliferation of chondrocytes and formation of good quality 'chondrocytes-fibrin' constructs which may have potential use of matrix-induced cell implantation. PMID: 20810142 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Mechanical Strain Stabilizes Reconstituted Collagen Fibrils against Enzymatic Degradation by Mammalian Collagenase Matrix Metalloproteinase 8 (MMP-8). PLoS One. 2010;5(8): Authors: Flynn BP, Bhole AP, Saeidi N, Liles M, Dimarzio CA, Ruberti JW BACKGROUND: Collagen, a triple-helical, self-organizing protein, is the predominant structural protein in mammals. It is found in bone, ligament, tendon, cartilage, intervertebral disc, skin, blood vessel, and cornea. We have recently postulated that fibrillar collagens (and their complementary enzymes) comprise the basis of a smart structural system which appears to support the retention of molecules in fibrils which are under tensile mechanical strain. The theory suggests that the mechanisms which drive the preferential accumulation of collagen in loaded tissue operate at the molecular level and are not solely cell-driven. The concept reduces control of matrix morphology to an interaction between molecules and the most relevant, physical, and persistent signal: mechanical strain. METHODOLOGY/PRINCIPAL FINDINGS: The investigation was carried out in an environmentally-controlled microbioreactor in which reconstituted type I collagen micronetworks were gently strained between micropipettes. The strained micronetworks were exposed to active matrix metalloproteinase 8 (MMP-8) and relative degradation rates for loaded and unloaded fibrils were tracked simultaneously using label-free differential interference contrast (DIC) imaging. It was found that applied tensile mechanical strain significantly increased degradation time of loaded fibrils compared to unloaded, paired controls. In many cases, strained fibrils were detectable long after unstrained fibrils were degraded. CONCLUSIONS/SIGNIFICANCE: In this investigation we demonstrate for the first time that applied mechanical strain preferentially preserves collagen fibrils in the presence of a physiologically-important mammalian enzyme: MMP-8. These results have the potential to contribute to our understanding of many collagen matrix phenomena including development, adaptation, remodeling and disease. Additionally, tissue engineering could benefit from the ability to sculpt desired structures from physiologically compatible and mutable collagen. PMID: 20808784 [PubMed - in process] | | | | | | | | | |  | |  | |  |
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