Jérôme Guicheux

Cartilage engineering: a crucial combination of cells, biomaterials and biofactors

Injuries to articular cartilage are one of the most challenging issues of musculoskeletal medicine due to the poor intrinsic ability of this tissue for repair. The lack of efficient modalities of treatment has prompted research into tissue engineering combining chondrogenic cells, scaffold materials and environmental factors. The aim of this review is to focus on the recent advances made in exploiting the potential of biomaterial-assisted cell therapy for cartilage engineering. We discuss the requirements for identifying additional specific growth factors and evaluating the optimal combination of cells, growth factors and scaffolds that is able to respond to the functional demand placed upon cartilage tissue replacement in clinics. Finally, some of the major obstacles encountered in cartilage engineering are discussed, as well as future trends in clinical applications.

Activation of p38 Mitogen‐Activated Protein Kinase and c‐Jun‐NH2‐Terminal Kinase by BMP‐2 and Their Implication in the Stimulation of Osteoblastic Cell Differentiation

Signaling involved in osteoblastic cell differentiation remains largely unknown. This study further investigates mechanisms involved in BMP‐2‐induced osteoblastic cell differentiation. We report that BMP‐2 can activate JNK and p38 in osteoblastic cells and provide evidences that these MAP kinases have distinct roles in regulating alkaline phosphatase and osteocalcin expression.

Evidence for a role of p38 MAP kinase in expression of alkaline phosphatase during osteoblastic cell differentiation

In the present study, we investigate the implication of the mitogen-activated protein kinases (MAPKs) Erk, p38, and JNK in mediating the effect of fetal calf serum (FCS) on the differentiation of MC3T3-E1 osteoblast-like cells. Erk is stimulated by FCS in proliferating, early-differentiating, as well as in mature cells. Activation of p38 by FCS is not detected in proliferating cells but is observed as the cells differentiate. JNK is activated in response to FCS throughout the entire differentiation process, but a maximal stimulation is observed in early differentiating cells. The roles of Erk and p38 pathways in mediating MC3T3-E1 cell differentiation was determined using specific inhibitors such as U0126 and SB203580, respectively. These experiments confirmed that the Erk pathway is essential for mediating cell proliferation in response to FCS, but indicated that this MAP kinase has little effect in regulating the differentiation of MC3T3-E1 cells. In contrast, p38 only marginally influenced proliferation, but appeared to be critical for the control of alkaline phosphatase (ALP) expression in differentiating cells. Finally, results obtained with high doses of SB203580, which also affected JNK activity, suggest that p38 and/or JNK are probably also involved in the control of type 1 collagen and osteocalcin expression in differentiating cells. The data indicate that MAPKs regulate different stages of MC3T3-E1 cell development in response to FCS. Distinct MAPK pathways seem to independently modulate osteoblastic cell proliferation and differentiation, with Erk playing an essential role in cell replication, whereas p38 is involved in the regulation of ALP expression during osteoblastic cell differentiation. JNK is also probably involved in the regulation of osteoblastic cell differentiation, but its precise role requires further investigation.

An injectable vehicle for nucleus pulposus cell-based therapy

An injectable hydrogel, acting as a reservoir for cell delivery and mimicking the native environment, offers promise for nucleus pulposus (NP) repair and regeneration. Herein, the potential of a stabilised type II collagen hydrogel using poly(ethylene glycol) ether tetrasuccinimidyl glutarate (4S-StarPEG) cross-linker, enriched with hyaluronic acid (HA) was investigated. The optimally stabilised type II collagen hydrogel was determined by assessing free amine groups, resistance to enzymatic degradation, gel point. The potential toxicity of the cross-linker was initially assessed against adipose-derived stem cells (ADSCs). After addition of HA (molar ratio type II collagen:HA 9:0, 9:1, 9:4.5, 9:9) within the hydrogel, the behaviour of the encapsulated NP cells was evaluated using cell proliferation assay, gene expression analysis, cell distribution and cell morphology. A significant decrease (p < 0.05) in the free amine groups of collagen was observed, confirming successful cross-linking. Gelation was independent of the concentration of 4S-StarPEG (8 min at 37 °C). The 1 mm cross-linked hydrogel yielded the most stable after enzymatic degradation (p < 0.05). No toxicity of the 4S-StarPEG was noted for the ADSCs. NP cell viability was high regardless of the concentration of HA (>80%). A cell proliferation was not seen after 14 days in its presence. At a gene expression level, HA did not influence NP cells phenotype after seven days in culture. After seven days in culture, the type I collagen mRNA expression was maintained (p > 0.05). The optimally stabilised and functionalised type II collagen/HA hydrogel system developed in this study shows promise as an injectable reservoir system for intervertebral disc regeneration.

Calcium phosphate biomaterials as bone drug delivery systems: a review.

A short review is proposed on the existing literature for the research performed in calcium phosphate (CaP) biomaterials used as drug delivery systems. In the first part, a brief update is given on the performance of both CaP ceramics and CaP cements. Second, a review of the research and clinical situation is developed for CaP materials already used as drug delivery systems. Experimental works performed for local delivery are reported. In particular, a description is given of the in vitro and in vivo studies in which these materials are loaded with various proteins and drugs.

Enhanced Expression of the Inorganic Phosphate Transporter Pit-1 Is Involved in BMP-2–Induced Matrix Mineralization in Osteoblast-Like Cells†

Pi handling by osteogenic cells is important for bone mineralization. The role of Pi transport in BMP‐2–induced matrix calcification was studied. BMP‐2 enhances Pit‐1 Pi transporters in osteogenic cells. Experimental analysis suggest that this response is required for bone matrix calcification.

Phosphate is a specific signal for ATDC5 chondrocyte maturation and apoptosis-associated mineralization: possible implication of apoptosis in the regulation of endochondral ossification

Involvement of Pi and Ca in chondrocyte maturation was studied because their levels increase in cartilage growth plate. In vitro results showed that Pi increases type X collagen expression, and together with Ca, induces apoptosis‐associated mineralization, which is similar to that analyzed in vivo, thus suggesting a role for both ions and apoptosis during endochondral ossification.

Articular cartilage calcification in osteoarthritis: Insights into crystal-induced stress

Osteoarthritis (OA) is the most common form of rheumatic disease, leading ultimately to chronic pain, restriction of joint mobility, and disability. OA affects millions of people worldwide, and its prevalence is expected to increase with the concomitant prevalence of obesity and aging (1). OA is a disease affecting the whole joint and is characterized by articular cartilage destruction along with changes occurring in other joint components including bone, menisci, synovium, ligaments, capsule, and muscles (2). Multiple parameters contribute to the pathogenesis and progression of OA, including hereditary factors, mechanical overload, intraarticular calcium-containing microcrystal (Ca crystals) deposition, and aging. Ca crystals, which encompass calcium pyrophosphate dihydrate (CPPD) and basic calcium phosphate (BCP) crystals, have been associated with severe OA for many years. However, the role of CPPD and BCP crystals in OA remains controversial (3,4). BCP crystals, a heterogeneous group of apatite crystals including carbonated apatite, octacalcium phosphate, immature amorphous apatite, tricalcium phosphate (5), and magnesium-substituted apatite (whitlockite) crystals (6), are considered by some investigators to be “innocent bystanders” and/or markers of end-stage disease, because they can be released from bone as apatite mineral (4). Similarly, CPPD crystal deposition is considered to be a common aging-related process (4,7). However, numerous clinical and experimental data provide strong evidence that cartilage calcification occurs as an active process, and that calcification clearly plays a pathogenic role in OA, a phenomenon referred to as “microcrystal-induced stress.” This is different from the previously described and well-known mechanical, oxidative, or cytokine stresses in OA. The aim of this review is to highlight this old but forgotten and neglected concept. Ultimately, cartilage calcification could be considered as a potential therapeutic target in OA. As such, several therapeutic strategies will be discussed.

From osteoarthritis treatments to future regenerative therapies for cartilage

Osteoarthritis (OA) is associated with cartilage degeneration and an accompanying inflammatory syndrome of the synovium in addition to alteration of the subchondral bone. The molecular and cellular events involved in OA have only partially been elucidated. This review provides a global view of the physiopathology of OA, as well as non-pharmacological and pharmacological treatments for the disorder. An update on surgical treatments and their indications is given with an orientation towards the management of OA and cartilage repair by cell-based regenerative therapies. These promising biological technologies will, potentially, play a major role in the treatment of cartilage-associated diseases.

Human osteosarcoma cells express functional receptor activator of nuclear factor-kappa B.

RANK, RANK ligand (RANKL) and osteoprotegerin (OPG) are the key regulators of bone metabolism, both in normal and pathological conditions. Previous data have demonstrated that human osteosarcoma biopsies express RANKL as well as OPG, and functional RANK is expressed in a murine osteosarcoma cell line. As RANK expression in human osteosarcoma remains controversial, the aim of the present study was to analyse its expression in vitro in human osteosarcoma cell lines, ex vivo using pathological tissues, and then to determine its functionality in terms of signal transduction pathways modulated by RANKL. RT‐PCR analysis and immunohistochemistry experiments revealed that RANK is expressed at both transcriptional and protein levels in MNNG/HOS, Saos‐2 and MG‐63 human osteosarcoma cell lines, in contrast to the U‐2 OS osteosarcoma cell line and human osteoblasts, which were negative. RANK was also expressed in 57% of osteosarcoma biopsies. Furthermore, western blot experiments clearly demonstrated the functionality of RANK. Thus, RANKL significantly induced the phosphorylation of ERK1/2, p38 and IκB in RANK‐positive osteosarcoma cells. This study is the first report of functional RANK expression in human osteosarcoma cells: this strengthens the involvement of the RANK–RANKL–OPG axis in primary bone tumour biology and identifies novel therapeutic approaches targeting RANK‐positive osteosarcoma. Copyright