Plasticity of fetal cartilaginous cells.

Tissue-specific stem cells found in adult tissues can participate in the repair process following injury. However, adult tissues, such as articular cartilage and intervertebral disc, have low regeneration capacity, whereas fetal tissues, such as articular cartilage, show high regeneration ability. The presence of fetal stem cells in fetal cartilaginous tissues and their involvement in the regeneration of fetal cartilage is unknown. The aim of the study was to assess the chondrogenic differentiation and the plasticity of fetal cartilaginous cells. We compared the TGF-β3-induced chondrogenic differentiation of human fetal cells isolated from spine and cartilage tissues to that of human bone marrow stromal cells (BMSC). Stem cell surface markers and adipogenic and osteogenic plasticity of the two fetal cell types were also assessed. TGF-β3 stimulation of fetal cells cultured in high cell density led to the production of aggrecan, type I and II collagens, and variable levels of type X collagen. Although fetal cells showed the same pattern of surface stem cell markers as BMSCs, both type of fetal cells had lower adipogenic and osteogenic differentiation capacity than BMSCs. Fetal cells from femoral head showed higher adipogenic differentiation than fetal cells from spine. These results show that fetal cells are already differentiated cells and may be a good compromise between stem cells and adult tissue cells for a cell-based therapy

Efficient decellularization of equine tendon with preserved biomechanical properties and cytocompatibility for human tendon surgery indications.

Chronic and acute tendon injuries are frequent afflictions, for which treatment is often long and unsatisfactory. When facing extended injuries, matrices and scaffolds with sufficient biomechanical properties are required for surgical repair and could additionally serve as supports for cellular therapies to improve healing. In this study, protocols of either commonly used detergents only (SDS 1%, Triton 1%, TBP 1%, and Tween-20 1%) or a combination of freeze/thaw (F/T) cycles with decellularization agents (NaCl 1M, ddH <sub>2</sub> O) were evaluated for the decellularization of horse equine superficial digital flexor tendon (SDFT) for hand flexor or extensor tendon reconstruction. Decellularization efficiency was assessed microscopically by histological staining (HE, DAPI) and DNA quantification. Macroscopical structure and biomechanical integrity of the tendon matrices were further assessed by gross observation, histological staining (SR), and mechanical testing (ultimate strain and stress, Young's modulus, energy to failure) for select protocols. Decellularization with hypertonic NaCl 1M in association with F/T cycles produced the most robust tendon matrices, which were nontoxic after 10 days for subsequent recellularization with human fetal progenitor tendon cells (hFPTs). This standardized protocol uses a less aggressive decellularization agent than current practice, which allows subsequent reseeding with allogenic cells, therefore making them very suitable and bioengineered tendon matrices for human tendon reconstruction in the clinic

Modeling of failure mode in knee ligaments depending on the strain rate

BACKGROUND: The failure mechanism of the knee ligament (bone-ligament-bone complex) at different strain rates is an important subject in the biomechanics of the knee. This study reviews and summarizes the literature describing ligament injury as a function of stain rate, which has been published during the last 30 years. METHODS: Three modes of injury are presented as a function of strain rate, and they are used to analyze the published cases. The number of avulsions is larger than that of ligament tearing in mode I. There is no significant difference between the number of avulsions and ligament tearing in mode II. Ligament tearing happens more frequently than avulsion in mode III. RESULTS: When the strain rate increases, the order of mode is mode I, II, III, I, and II. Analytical models of ligament behavior as a function of strain rate are also presented and used to provide an integrated framework for describing all of the failure regimes. In addition, this study showed the failure mechanisms with different specimens, ages, and strain rates. CONCLUSION: There have been several a numbers of studies of ligament failure under various conditions including widely varying strain rates. One issue in these studies is whether ligament failure occurs mid-ligament or at the bone attachment point, with assertions that this is a function of the strain rate. However, over the range of strain rates and other conditions reported, there has appeared to be discrepancies in the conclusions on the effect of strain rate. The analysis and model presented here provides a unifying assessment of the previous disparities, emphasizing the differential effect of strain rate on the relative strengths of the ligament and the attachment

Synthetic nano-fibrillar extracellular matrices with predesigned macroporous architectures

Scaffolding plays a pivotal role in tissue engineering. To mimic the architecture of a natural extracellular matrix component—collagen, nona-fibrous matrices have been created with synthetic biodegradable polymers in our laboratory using a phase-separation technique. To improve the cell seeding, distribution, mass transport, and new tissue organization, three-dimensional macroporous architectures are built in the nano-fibrous matrices. Water-soluble porogen materials are first fabricated into three-dimensional negative replicas of the desired macroporous architectures. Polymer solutions are then cast over the porogen assemblies in a mold, and are thermally phase-separated to form nano-fibrous matrices. The porogen materials are leached out with water to finally form the synthetic nano-fibrous extracellular matrices with predesigned macroporous architectures. In this way, synthetic polymer matrices are created with architectural features at several levels, including the anatomical shape of the matrix, macroporous elements (100 Μm to millimeters), interfiber distance (microns), and the diameter of the fibers (50–500 nm). These scaffolding materials circumvent the concerns of pathogen transmission and immuno-rejection associated with natural collagen. With the flexibility in the design of chemical structure, molecular weight, architecture, degradation rate, and mechanical properties, these novel synthetic matrices may serve as superior scaffolding for tissue engineering. © 2000 John Wiley & Sons, Inc. J Biomed Mater Res, 52, 430–438, 2000.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/34418/1/25_ftp.pd

Effect of temporal onsets of mechanical loading on bone formation inside a tissue engineering scaffold combined with cell therapy.

Several approaches to combine bone substitutes with biomolecules, cells or mechanical loading have been explored as an alternative to the limitation and risk-related bone auto- and allo-grafts. In particular, human bone progenitor cells seeded in porous poly(L-lactic acid)/tricalcium phosphate scaffolds have shown promising results. Furthermore, the application of mechanical loading has long been known to be a key player in the regulation of bone architecture and mechanical properties. Several in vivo studies have pointed out the importance of its temporal offset. When an early mechanical loading was applied a few days after scaffold implantation, it was ineffective on bone formation, whereas a delayed mechanical loading of several weeks was beneficial for bone tissue regeneration. No information is reported to date on the effectiveness of applying a mechanical loading in vivo on cell-seeded scaffold with respect to bone formation in a bone site. In our study, we were interested in human bone progenitor cells due to their low immunogenicity, sensitivity to mechanical loading and capacity to differentiate into osteogenic human bone progenitor cells. The latest capacity allowed us to test two different bone cell fates originating from the same cell type. Therefore, the general aim of this study was to assess the outcome on bone formation when human bone progenitor cells or pre-differentiated osteogenic human bone progenitor cells are combined with early and delayed mechanical loading inside bone tissue engineering scaffolds. Scaffolds without cells, named cell-free scaffold, were used as control. Surprisingly, we found that (1) the optimal solution for bone formation is the combination of cell-free scaffolds and delayed mechanical loading and that (2) the timing of the mechanical application is crucial and dependent on the cell type inside the implanted scaffolds

The cytotoxic effect of titanium particles phagocytosed by osteoblasts

The cytotoxic effect of different concentrations of titanium particles on osteoblasts was studied in vitro. It was found that the viability of the osteoblasts was inversely proportional to the particle concentration. Phagocytosis of particles by the osteoblasts was evident and was demonstrated to be responsible for cell necrosis. Moreover, during and after phagocytosis, the osteoblasts released products that were cytotoxic for other osteoblasts, as established with a conditioned medium assay. The titanium particles thus had both a direct and an indirect effect on osteoblast viability. It also was observed that the titanium particles induced a process of programmed cell death (apoptosis) when co-cultured with osteoblasts. The results of this study suggest that not only is the amount of wear debris generated important, but the local accumulation of the debris also may have a significant impact on bone cell function

Development of Standardized Fetal Progenitor Cell Therapy for Cartilage Regenerative Medicine: Industrial Transposition and Preliminary Safety in Xenogeneic Transplantation

Diverse cell therapy approaches constitute prime developmental prospects for managing acute or degenerative cartilaginous tissue affections, synergistically complementing specific surgical solutions. Bone marrow stimulation (i.e., microfracture) remains a standard technique for cartilage repair promotion, despite incurring the adverse generation of fibrocartilagenous scar tissue, while matrix-induced autologous chondrocyte implantation (MACI) and alternative autologous cell-based approaches may partly circumvent this effect. Autologous chondrocytes remain standard cell sources, yet arrays of alternative therapeutic biologicals present great potential for regenerative medicine. Cultured human epiphyseal chondro-progenitors (hECP) were proposed as sustainable, safe, and stable candidates for chaperoning cartilage repair or regeneration. This study describes the development and industrial transposition of hECP multi-tiered cell banking following a single organ donation, as well as preliminary preclinical hECP safety. Optimized cell banking workflows were proposed, potentially generating millions of safe and sustainable therapeutic products. Furthermore, clinical hECP doses were characterized as non-toxic in a standardized chorioallantoic membrane model. Lastly, a MACI-like protocol, including hECPs, was applied in a three-month GLP pilot safety evaluation in a caprine model of full-thickness articular cartilage defect. The safety of hECP transplantation was highlighted in xenogeneic settings, along with confirmed needs for optimal cell delivery vehicles and implantation techniques favoring effective cartilage repair or regeneration

Polylactic acid-phosphate glass composite foams as scaffolds for tissue engineering

Phosphate glass (PG) of the composition 0.46(CaO)-0.04(Na(2)O)- 0.5(P(2)O(5)) was used as filler in poly-L-lactic acid (PLA) foams developed as degradable scaffolds for bone tissue engineering. The effect of PG on PLA was assessed both in bulk and porous composite foams. Composites with various PG content (0, 5, 10, and 20 wt %) were melt-extruded, and either compression-molded or foamed through supercritical CO(2). Dynamic mechanical analysis on the bulk composites showed that incorporating 20 wt % PG resulted in a significant increase in storage modulus. Aging studies in deionized water in terms of weight loss, pH change, and ion release inferred that the degradation was due to PG dissolution, and dependent on the amount of glass in the composites. Foaming was only possible for composites containing 5 and 10 wt % PG, as an increase in PG increased the foam densities; however, the level of porosity was maintained above 75%. PLA-T(g) in the foams was higher than those obtained for the bulk. Compressive moduli showed no significant reinforcement with glass incorporation in either expansion direction, indicating no anisotropy. Biocompatibility showed that proliferation of human fetal bone cells was more rapid for PLA compared to PLA-PG foams. However, the proliferation rate of PLA-PG foams were similar to those obtained for foams of PLA with either hydroxyapatite or beta-tricalcium phosphate

The effects of calcium phosphate cement particles on osteoblast functions

Calcium phosphate cements (CPC) are increasingly used in the orthopedic field. This kind of cement has potential applications in bone defect replacements, osteosynthetic screw reinforcements or drug delivery. In vivo studies have demonstrated a good osteointegration of CPC. However, it was also observed that the resorption of CPC could create particles. It is known from orthopedic implant studies that particles can be responsible for the peri-implant osteolysis. Biocompatibility assessment of CPC should then be performed with particles. In this study, we quantified the functions of osteoblasts in the presence of beta-TCP, brushite and cement particles. Two particle sizes were prepared. The first one corresponded to the critical diameter range 1-10 microm and the second one had a diameter larger than 10 microm. We found that CPC particles could adversely affect the osteoblast functions. A decrease in viability, proliferation and production of extracellular matrix was measured. A dose effect was also observed. A ratio of 50 CPC particles per osteoblast could be considered as the maximum number of particles supported by an osteoblast. The smaller particles had stronger negative effects on osteoblast functions than the larger ones. Future CPC development should minimize the generation of particles smaller than 10 microm

Colloquium: Mechanical formalisms for tissue dynamics

The understanding of morphogenesis in living organisms has been renewed by tremendous progressin experimental techniques that provide access to cell-scale, quantitative information both on theshapes of cells within tissues and on the genes being expressed. This information suggests that ourunderstanding of the respective contributions of gene expression and mechanics, and of their crucialentanglement, will soon leap forward. Biomechanics increasingly benefits from models, which assistthe design and interpretation of experiments, point out the main ingredients and assumptions, andultimately lead to predictions. The newly accessible local information thus calls for a reflectionon how to select suitable classes of mechanical models. We review both mechanical ingredientssuggested by the current knowledge of tissue behaviour, and modelling methods that can helpgenerate a rheological diagram or a constitutive equation. We distinguish cell scale ("intra-cell")and tissue scale ("inter-cell") contributions. We recall the mathematical framework developpedfor continuum materials and explain how to transform a constitutive equation into a set of partialdifferential equations amenable to numerical resolution. We show that when plastic behaviour isrelevant, the dissipation function formalism appears appropriate to generate constitutive equations;its variational nature facilitates numerical implementation, and we discuss adaptations needed in thecase of large deformations. The present article gathers theoretical methods that can readily enhancethe significance of the data to be extracted from recent or future high throughput biomechanicalexperiments.Comment: 33 pages, 20 figures. This version (26 Sept. 2015) contains a few corrections to the published version, all in Appendix D.2 devoted to large deformation