Growth of mammalian cells at high oxygen concentrations.

Oxygen, although essential to the growth of mammalian cells in vitro and in vivo, has been widely reported to be toxic at concentrations at or above the oxygen concentration in culture medium equilibrated with air (approximately 200 microM). We were therefore surprised to note that a diploid human B-cell line (TK6) was able to proliferate normally while exposed to 380 microM-oxygen. This observation was extended to Vero (African Green monkey kidney) cells, and Sp2/0-derived murine transfectomas producing antibody. Using an experimental system with a high capacity for oxygen transfer, we determined the growth rates of the three cell lines at controlled oxygen concentrations ranging from 80 microM to 910 microM. Each of these cell types was able to grow normally at oxygen concentrations up to 360-380 microM. At oxygen concentrations above 380 microM, a significant increase in the apparent doubling times of the cells was observed. No adverse effect of oxygen on TK6 cell survival was seen for concentrations ranging from 60 microM to 410 microM. We conclude that exponential growth at nearly maximum growth rates was observed for Sp2/0-derived cells, TK6, and Vero cells at constant oxygen concentrations up to 400 microM (twice air saturation). These findings have substantial implications for process control in the production of cells and cell-derived materials. The intrinsic oxygen transfer rate of any cell culture vessel can be increased severalfold by raising the oxygen gas concentration.(ABSTRACT TRUNCATED AT 250 WORDS)

Mass cultivation of bovine ocular pigment epithelial cells in microcarrier suspension culture.

Recent developments in tissue culture technology permit the mass cultivation of anchorage-dependent cells in a compact and easily maintained system by utilizing the large surface area of suspended microspheres for cellular attachment and proliferation. The present study describes the successful cultivation of pure bovine iris pigment epithelium and mixed bovine retinal pigment epithelium-choroid by microcarrier suspension culture. Ocular pigment epithelial cells were collected from enucleated calf eyes and cultured in conventional monolayer fashion on flat plates. Subsequently, cells were harvested by trypsinization and were inoculated into microcarrier culture. Confluence was obtained in 8 to 12 days and final cell densities were 1 x 10(6) cells/ml for iris pigment epithelium and 8 x 10(5) cells/ml for combined retinal pigment epithelium--choroid; the microcarrier density was 5 gm/L. Samples from ongoing microcarrier cultures were processed for light and electron microscopic examination. Culture of bovine ocular pigment epithelial cells by microcarrier suspension technique provides a simplified method of cultivation of large numbers of cells, with additional advantages in ongoing culture maintenance and accessibility.