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INTEGRIN-MEDIATED BONE CELL RESPONSE TO HYPERGRAVITY  W. Vercoutere¹, C. Roden¹, M. Parra¹, C. Damsky², E. Holton¹, N, Searby¹, R. Globus^1,2, E. Almeida^{1,2 1}NASA Ames Research Center, and ² University of California, San Francisco.

   When vertebrates are placed in altered gravity environments they show   physiological responses, including loss or gain of bone and muscle tissue.  The underlying cellular mechanisms, however, are still poorly understood.   Existing studies of cellular mechanical stimulation by other forces such as flow and substrate deformation suggest that integrin adhesion to extra-cellular matrix (ECM) may play a key role in mechano-transduction.  Bone and muscle cells may also respond to gravity via ECM/integrin-initiated signaling pathways.  Using cell culture centrifugation as a model for hypergravity we found that primary rat osteoblast and fibroblast adhesion to fibronectin at 50-g promotes increased integrin focal adhesion number, focal adhesion kinase phosphorylation, cell survival and proliferation.  To determine if the proliferative response of osteoblasts to hypergravity is mediated by particular integrins, we examined osteoblast growth in various ECMs.  The NASA-developed cell culture centrifuge consists of a modi-fied 1-ft-diameter vented Eppendorf 5804 centrifuge inside an environ-mental cell culture chamber with heating, refrigeration, humidity and CO₂ controls.  We hypothesized that specific ECMs but not uncoated tissue-culture plastic would stimulate gravity-induced hyperproliferation.  If true, hyper-proliferation response is likely to be mediated by specific integrins and their signaling complexes.  Conversely, if cells subjected to hyper-gravity all exhibit the same increase in proliferation on various ECMs and plastic, then the process is unlikely to be mediated by specific integrins.

   In our current experiments, proliferation of primary osteoblasts grown on fibronectin, laminin, collagen Type I, and collagen Type IV was enhanced by continuous 24 hour 50-g hypergravity stimulus as compared with the 1-g control.  No such increase was observed for cells grown on uncoated surfaces.  Furthermore the proliferative response was greatest for cells on collagen Type I (1.4 fold increase), suggesting that α₁β₁, α₂β₁, α₃β₁ and α_νβ₁ integrins may be significantly involved.  These results suggest that specific ECM-integrin signaling in hypergravity conditions upregulate cell survival and proliferation pathways.

(Supported by NASA: 00-OBPR-01-066).

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