Differential cartilaginous tissue formation by human synovial membrane, fat pad, meniscus cells and articular chondrocytes

Objective: To identify an appropriate cell source for the generation of meniscus substitutes, among those which would be available by arthroscopy of injured knee joints. Methods: Human inner meniscus cells, fat pad cells (FPC), synovial membrane cells (SMC) and articular chondrocytes (AC) were expanded with or without specific growth factors (Transforming growth factor-betal, Fibroblast growth factor-2 and Plate let-derived growth factor bb, TFP) and then induced to form three-dimensional cartilaginous tissues in pellet cultures, or using a hyaluronan-based scaffold (Hyaff(R)-11), in culture or in nude mice. Human native menisci were assessed as reference. Results: Cell expansion with TFP enhanced glycosaminoglycan (GAG) deposition by all cell types (up to 4.1-fold) and messenger RNA expression of collagen type II by FPC and SMC (up to 472-fold) following pellet culture. In all models, tissues generated by AC contained the highest fractions of GAG (up to 1.9 were positively stained for collagen type II (specific of the inner avascular region of meniscus), type IV (mainly present in the outer vascularized region of meniscus) and types I, III and VI (common to both meniscus regions). Instead, inner meniscus, FPC and SMC developed tissues containing negligible GAG and no detectable collagen type II protein. Tissues generated by AC remained biochemically and phenotypically stable upon ectopic implantation. Conclusions: Under our experimental conditions, only AC generated tissues containing relevant amounts of GAG and with cell phenotypes compatible with those of the inner and outer meniscus regions. Instead, the other investigated cell sources formed tissues resembling only the outer region of meniscus. It remains to be determined whether grafts based on AC will have the ability to reach the complex structural and functional organization typical of meniscus tissue. (C) 2006 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights rese

The molecular biology of bacterial xylanases

Available from British Library Document Supply Centre-DSC:DXN007333 / BLDSC - British Library Document Supply CentreSIGLEGBUnited Kingdo

Evidence for a general role for high affinity non-catalytic cellulose binding domains in microbial plant cell wall hydrolases

International audienc

Roles for the interleukin-4 receptor and associated JAK/STAT proteins in human articular chondrocyte mechanotransduction

SummaryObjectiveTo identify functional interleukin-4 (IL4) receptor (IL4R) subtypes and associated Janus kinase/signal transducers and activators of transcription (JAK/STAT) molecules in human articular chondrocytes and assess the role of JAK/STAT proteins in chondrocyte mechanotransduction.MethodsExpression of IL4R subunits and associated molecules was assessed by immunohistochemistry and western blotting. Functional IL4R were identified by chemical crosslinking of IL4-stimulated chondrocytes and western blotting. JAK and STAT phosphorylation was assessed by western blotting.ResultsChondrocytes from normal and osteoarthritic (OA) cartilage express IL4Rα, γc and IL13Rα1 subunits (components of the Type I and Type II IL4R). In the presence of IL4 only functional Type II IL4Rs were identified in normal or OA chondrocytes. With the exception of STAT2, no differences in JAK/STAT expression were detected between normal and OA cartilage. STAT2 was expressed in OA but not normal chondrocytes. Mechanical stimulation (MS) resulted in an IL4R-dependent increase in phosphorylated Tyk2 in normal chondrocytes, which could be abolished by IL1β preincubation. No phosphorylation of STAT5 or STAT6 was detected in either normal or OA chondrocytes following mechanical stimulation (MS) IL4 stimulation resulted in a decrease in Tyk2 phosphorylation and an increase in phosphorylation of STAT6 in both normal and OA chondrocytes.ConclusionChondrocytes from normal and OA cartilage signal through a Type II IL4R. This signalling is via a STAT6-independent pathway. Differences in IL4 signalling are likely due to crosstalk between integrin and cytokine signalling pathways, and not differences in IL4R expression

Evidence for JNK-dependent up-regulation of proteoglycan synthesis and for activation of JNK1 following cyclical mechanical stimulation in a human chondrocyte culture model

SummaryObjectiveTo examine the expression of mitogen-activated protein kinases (MAPKs) in human chondrocytes, to investigate whether selective activation of MAPKs is involved in up-regulation of proteoglycan (PG) synthesis following cyclical mechanical stimulation (MS), and to examine whether MS is associated with integrin-dependent or independent activation of MAPKs.MethodsThe C-28/I2 and C-20/A4 human chondrocyte cell lines were mechanically stimulated in monolayer cell culture. PG synthesis was assessed by [35S]-sulphate incorporation in the presence and absence of the p38 inhibitor SB203580, and the extracellular-regulated kinase (ERK1/2) inhibitor PD98059. Kinase expression and activation were assessed by Western blotting using phosphorylation status-dependent and independent antibodies, and by kinase assays. The Jun N-terminal kinase (JNK) inhibitor SP600125 and the anti-β1 integrin (CD29) function-blocking antibody were used to assess JNK activation and integrin dependence, respectively.ResultsIncreased PG synthesis following 3h of cyclic MS was abolished by pretreatment with 10μM SB203580, but was not affected by 50μM PD98059. The kinases p38, ERK1/ERK2 and JNKs were expressed in both stimulated and unstimulated cells. Phosphorylated p38 was detected at various time points following 0.5, 1, 2 and 3h MS in C-28/I2, but not detected in C-20/A4 cell lines. Phosphorylation of ERK1 and ERK2 was not significantly affected by MS. Phosphorylation of the 54 and 46kDa JNKs increased following 0.5, 1, 2 and 3h of MS, and following CO2 deprivation. MS-induced JNK phosphorylation was inhibited by SB203580 at concentrations ≥5μM and activation of JNK1 following MS was blocked by SP600125 and partially inhibited by anti-CD29.ConclusionsThe data suggest JNK, rather than p38 or ERK dependent increases in PG synthesis, and selective, partially integrin-dependent, activation of JNK kinases in human chondrocyte cell lines following cyclical MS. JNK activation is also very sensitive to changes in CO2/pH in this chondrocyte culture model