In vivo inhibition of osteoblastic metalloproteinases leads to increased trabecular bone mass.

UNLABELLED: Mice specifically overexpressing TIMP-1 in osteoblasts have been generated to investigate the role of MMPs in bone in vivo. These mice displayed increased trabecular bone volume and decreased bone turnover. This model provides evidence of the role played by the MMPs in bone remodeling and balance. INTRODUCTION: Although it has been suggested that the matrix metalloproteinases (MMPs) may play a role in initiating the bone resorption process in vitro, there is no evidence that they play any role in in vivo bone maintenance. MATERIALS AND METHODS: We used an artificial promoter specifically driving cells of the osteoblastic lineage to overexpress the tissue inhibitor of MMPs (TIMP-1) cDNA in mice. Densitometric analysis, using DXA and pQCT, and static and dynamic histomorphometry were used to evaluate the bone phenotype both in male and female transgenic mice. We evaluated osteoblastic differentiation using a primary osteoblast culture and osteoclast activity using an ex vivo organ culture. RESULTS AND CONCLUSION: We showed that at 1 and 2.5 months of age, only the female mice exhibited a bone phenotype. These mice displayed specific increases in the BMD and bone volume of trabecular bone. This increase was accompanied by decreased trabecular separation, suggesting a decrease in bone resorption. Using an ex vivo resorption assay, we demonstrated that parathyroid hormone (PTH)-stimulated bone resorption was reduced in these mice. Evaluation of the bone histomorphometric dynamic parameters showed that the mineralizing surfaces and bone formation rate were both reduced. There was no change in the mineralization lag time or number of osteocyte lacunae. Using primary osteoblast culture and molecular analysis, we showed that the differentiation and function of osteoblasts from transgenic mice were normal, but that the ex vivo formation of mineralized nodules was delayed. This model is the first to show that in vivo MMPs play a role in bone remodeling and bone balance. Moreover, our data suggest that MMP activity could be involved in the hormonal regulation of bone resorption by osteoblasts.

A combination of cis-acting elements is required to activate the pro-alpha 1(I) collagen promoter in tendon fibroblasts of transgenic mice.

The genes encoding the two type I collagen chains are selectively activated in few cell types, including fibroblasts and osteoblasts. By generating transgenic mice, we have previously shown that the activity of the mouse pro-alpha1(I) promoter was controlled by separate cell-specific cis-acting elements. In particular, a sequence located between -3.2 and -2.3 kb was needed to induce expression of the reporter gene at high levels in tendon fibroblasts. In the present work, by using the same transgenic approach, we have identified two short elements in this sequence, named tendon-specific element (TSE) 1 and TSE2, that were necessary to direct reporter gene expression selectively in tendon fibroblasts. Gel shift assays showed that TSE1 and TSE2 bound proteins specifically present in nuclear extracts from tendon fibroblasts and that the sequence of TSE2 binding a tendon-specific protein corresponded to an E-box. Analysis of transgenic mice further indicated that TSE1 and TSE2 needed to cooperate not only with each other but also with other cis-acting elements of the proximal promoter to activate reporter gene expression in tendon fibroblasts. Similarly, it pointed out that the so-called osteoblast-specific element had to interact with downstream sequences to drive reporter gene expression in osteoblasts of transgenic mice. Thus, expression of the mouse pro-alpha1(I) collagen gene in tendon fibroblasts appears to be the result of a unique combination of different cis-acting elements, including TSE1 and TSE2.