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Adult bone-marrow-derived mesenchymal stroma cells (BMSC) seem to be a potential cell source for tissue engineering of the ligament. The objective of this work was to study the time-related changes in mRNA expression and protein levels of collagen types I and III and of tenascin-C in BMSC under co-culture with fibroblasts or under a uniaxial cyclic condition. Rat BMSC harvested from the femur and tibial bone marrow were co-cultured with ligament fibroblasts or stimulated by cyclic 10% uniaxial stretching at 1 Hz. Image analysis showed significant cell loss in stretched BMSC, particularly in the directions close to the stretching direction. However, these BMSC displayed an equivalent growth rate to that of non-stretched cells. Real-time reverse transcription/polymerase chain reaction revealed that the mRNA expression of collagen types I and III and of tenascin-C by BMSC was significantly up-regulated by co-culture and cyclic stretching. Radioimmunoassay results confirmed the effects of these stimulations, showing increases in the level of these proteins. Thus, BMSC might be useful as a cell source for the tissue engineering of ligament.

Bone marrow mesenchymal stem cells (BMSC) have the potential to differentiate into a variety of cell types like osteoblasts, chondroblasts, adipocytes, etc. It is well known that mechanical forces regulate the biological function of cells. The aim of this study was to investigate the effect of uniaxial stretching on the orientation and biological functions of BMSC. Rat BMSCs were harvested from femoral and tibial bone marrow by density gradient centrifugation. Cells from passages 1-6 were characterized by flow cytometry using monoclonal antibodies. The recovered cells were stably positive for the markers CD90 and CD44 and negative for CD34 and CD45. A cyclic 10% uniaxial stretching at 1Hz was applied on rat BMSC for different time-courses. The length, width, and orientation of the cells were subsequently determined. Expression of collagen types I and III and tenascin-C mRNAs was measured by real-time RT-PCR, and the synthesis of these receptors was determined by radioimmunoassay. Results showed that uniaxial stretching lengthened and rearranged the cells. Compared with control groups, expression of collagen types I and III mRNAs was up-regulated after 12-h of stretching, while significant increase in synthesis of the two collagen protein types was not observed until after 24-h stretching. The expression of tenascin-C mRNA was significantly increased after a 24-h stretching. These data suggest that cyclic stretching promotes the synthesis of collagen types I and III and tenascin-C by the rat BMSC.

Bioreactors are defined as devices in which biological and/or biochemical processes develop under closely monitored and tightly controlled environmental and operating conditions (e.g. pH, temperature, mechanical conditions, nutrient supply and waste removal). In functional tissue engineering of musculoskeletal tissues, a bioreactor capable of controlling dynamic loading plays a determinant role. It has been shown that mechanical stretching promotes the expression of type I and III collagens, fibronectin, tenascin-C in cultured ligament fibroblasts (J.C.-H. Goh et al., Tissue Eng. 9 (2003), S31) and that human bone marrow mesenchymal stem cells (hBMMSC) – even in the absence of biochemical regulators – could be induced to differentiate into ligament-like fibroblast by the application of physiologically relevant cyclic strains (G. Vunjak-Novakovic et al., Ann. Rev. Biomed. Eng. 6 (2004), 131; H.A. Awad et al., Tissue Eng. 5 (1999), 267; R.G. Young et al., J. Orthop. Res. 16 (1998), 406). Different bioreactors are commercially available but they are too generic to be used for a given tissue, each tissue showing specific mechanical loading properties. In the case of ligament tissue engineering, the design of a bioreactor is still an open question. Our group proposes a bioreactor allowing cyclic traction–torsion on a scaffold seeded with stem cells. [ABSTRACT FROM AUTHOR]