Spaceflight leads to osteopenia in both humans and animals, principally as a result of decreased bone formation, which might be the consequence of impaired osteoblast differentiation. The effect of microgravity on osteoblast differentiation in vitro was investigated using the human osteosarcoma cell line MG-63. Genes related to matrix formation and maturation were quantified both at the protein and mRNA level in untreated and hormone-treated (dihydroxyvitamin D 3 [1,25(OH) 2 D 3 ], 10 ؊7 M; transforming growth factor ␤ 2 (TGF-␤ 2 ), 10 ng/ml) cells cultured for 9 days under microgravity conditions aboard the Foton 10 satellite and compared with ground and inflight unit-gravity cultures. The expression of alkaline phosphatase (ALP) activity following treatment at microgravity increased only by a factor of 1.8 compared with the 3.8-fold increase at unit-gravity ( p < 0.01), whereas no alteration was detected in the production of collagen type I between unit-and microgravity. In addition, gene expression for collagen I␣1, ALP, and osteocalcin following treatment at microgravity was reduced to 51, 62, and 19%, respectively, of unit-gravity levels ( p < 0.02). The lack of correlation between collagen type I gene and protein expression induced by microgravity is most likely related to the different kinetics of gene and protein expression observed at unit-gravity: following treatment with 1,25(OH) 2 D 3 and TGF-␤ 2 , collagen I␣1 mRNA increased gradually during 72 h, but collagen type I production was already maximal after treatment for 48 h. In conclusion, microgravity decreases the activity of osteoblasts in vitro; in particular the differentiation of osteoblasts in response to systemic hormones and growth factors is reduced by microgravity. (J Bone Miner Res 1997;12:786-794)

Gelatinase B is a regulated matrix metalloproteinase with an important role in the remodelling of extracellular matrices and of basement membranes. To study the structure and function of gelatinase B in the mouse, the cDNA was cloned from a macrophage cell line (WEHI-3). Using this cDNA, a cosmid clone with the mouse gene was isolated. The complete gene (8 kbp) was sequenced and compared with the human gene structure. There was 78% similarity at the cDNA level and the exodintron structure of the murine gene was similar to the human counterpart. At the 5' untranslated side, 1200 bp of the promoter/enhancer region were sequenced and found to contain several transacting-factor-binding sites. The mRNA transcription-initiation site was determined by non-isotopic primer-extension analysis. Polymerase-chain-reaction amplification of cDNAs yielded indirect evidence for a reverse-transcription stop in WEHI-3 cell mRNA. The DNA-derived mouse-protein structure exhibited 82% similarity with the human one. This similarity was functionally reflected by cross-reactivity of the mouse protein with an antiserum against human gelatinase B. The production of murine gelatinase B was studied at the protein level by zymography and at the mRNA level by Northern blot analysis. In WEHI-3 cells the gelatinase B protein is induced by bacterial lipopolysaccharide, phorbol ester, double-stranded RNA and the cytokine interleukin-1 . Regulation of activity and structural heterogeneity of gelatinase B in WEHI-3 cells were shown to occur at the gene regulatory level, by expression of the matrix metalloproteinase inhibitor TIMP-1, and by glycosylation of the secreted protein.Matrix metalloproteinases (MMP) belong to a class of proteolytic enzymes which act together in degrading most components of extracellular matrices and of basement membranes. Physiological processes in which MMP play a role include embryonic growth and development, migration of blood leukocytes into tissues and tissue remodelling (Matrisian, 1992). Elevated levels of certain MMP are believed to be associated with various pathological states such as tumorcell invasion and metastasis and inflammatory processes such as rheumatoid arthritis and multiple sclerosis (Opdenakker and Van Damme, 1992a,b Note. The novel nucleotide sequence data published here have been deposited with the EMBL GenBank and DDBT sequence data banks and are available under the accession numbers X72794 and X72795, for the gene and cDNA, respectively.

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