Clinorotation inhibits chondrogenesis in micromass cultures of embryonic mouse limb cells

Studies of the response of mammalian chondrocytes to gravitational changes in vivo, in organ culture, and in cell culture show that chondrogenesis is reduced in microgravity or by unloading, and increased by low levels of excess g. To investigate the cellular responses to microgravity using a ground based model, micromass cultures were exposed to simulated weightlessness on two clinostats. For rotation on the large clinostat, cultures were set up in Rose chambers, and cells were videotaped and photographed at several time periods after rotation began. For the smaller clinostat, cultures were set up in T-flasks, and two axes of rotation for clinostated cultures were used. Stationary controls [+1 g, -1 g (upside-down), and sideways] as well as rotation controls were employed. Rotation rate was 30 rpm for both clinostatted cultures and rotation controls. Chondrocyte differentiation was assessed by cartilage specific alcian blue staining. Significantly fewer alcian blue stained nodules were present in clinostatted cultures than in stationary controls or rotation controls. Nodules that did not stain with alcian blue, probably due to unsulfated matrix were present in all cultures. The number of nodules in sideways controls was greater than in any other culture (108% of +1 g controls), probably due to ongoing stimulus of the cell via cytoskeletal components. The results show that chondrocytes in culture respond to changes in the gravity vector in a predictable manner, and that carefully controlled clinostat studies can be useful adjuncts to and predictors for spaceflight experiments.