Our hypothesis is that rotation increases apoptosis in standard tissue culture medium at shear stresses of greater than approximately 0.3 dyn/cm2. Human MIP-101 poorly differentiated colorectal carcinoma cells were cultured for 6 d in complete medium in monolayers, on Teflon®-coated nonadherent surfaces (static three-dimensional [3D]) or in rotating 3D cultures either in microgravity in low-earth orbit (3D μg) or in unit gravity on the ground (3D 1g). Apoptosis (determined morphologically), proliferation (by MIB1 staining), and the expression of epidermal growth-factor receptor (EGF-R), TGF-α, or TGF-β were assessed by immunohistochemistry, while the expression of the differentiation marker carcinoembryonic antigen (CEA) was assessed on Western blots. Over the course of 6 d, static 3D cultures displayed the highest rates of proliferation and lowest apoptosis. This was associated with high EGF-R, TGF-α, and TGF-β expression which was greater than that of a monolayer culture. Both rotated 3D 1g and 3D μg cultures displayed lower expression of EGF-R, TGF-α, or TGF-β and proliferation than that of monolayer or static 3D cultures. However, rotated 3D μg displayed significantly less apoptosis and greater CEA expression than rotated 3D 1g cultures. When rotated cultures of MIP-101 cells were grown under static conditions for another 3 d, proliferation increased and apoptosis decreased. Thus, rotation appears to increase apoptosis and decrease proliferation, whereas static 3D cultures in either unit or microgravity have less apoptosis, and reduced rotation in microgravity increases CEA expression.
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Vol. 36 • No. 6