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CHARACTERISTICS OF TUMOR-DERIVED CELLS IN 3-D ANALOGUE MICROGRAVITY ENVIRONMENT.  V. Chopra¹, T. Reed¹, T. V. Dinh¹, T. G. Wood², N. R. Pellis³, and E. V. Hannigan¹. Department of Ob-Gyn¹, and Molecular Science Center², UTMB, Galveston, TX and Cellular Biotechnology, NASA, JSC³, Houston, TX.

   Co-cultures of three-dimensional (3-D) constructs of one cell type with dispersed cells of a second cell type in low-shear rotating suspension cultures in analogue microgravity environment have been used to investigate invasive properties of normal and malignant cells. Immunohistochemical staining procedures of cocultured harvests of tumor-derived cells from gynecological cancer patients along with fibroblasts and endothelial cells demonstrate various markers of interest.

   Human umbilical vein-derived endothelial cells (HUVEC) were used to study the mitogenic response of the conditioned medium collected from 3-D monocultures and cocultures using proliferation and migration assays. The production of interleukin-2, -6, and -8, vascular endothelial cell growth factor (VEGF), and basic fibroblast growth factor were studied by using ELISA and RT-PCR. The conditioned medium collected from 3-D cocultures showed increased expression of 1) the message level of VEGF and its receptor flt-1 and KDR and 2) intracellular and vascular cell adhesion molecules, when measured by using Live cell ELISA assays and immunofluorescence staining as compared with 3-D monocultures. There was an increase in production of 1) enzymatic activity that could generate bioactive angiostatin from purified human plasminogen, and 2) fibrin, mucin, and elastic fiber by cell aggregates of 3-D cocultures of patient-derived cells as compared with 3-D monocultures

   We have shown that the epithelial and endothelial cells undergo a switch in characteristics when grown in an in vitro 3-D environment, that mimics the in vivo host environment as compared with conventional two-dimensional cultures. This coculture provides new insights into the invasive process and its effects on both invading and invaded cells and can be used to study the effectiveness of various antiangiogenic agents.

This project was funded by NASA, contract # NCC8-170.

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