Virus Production in Cultures of Simian Virus 40-Transformed Human Amnion Cells

Summary

Cultures of epithelial human amnion cells, infected and transformed by simian virus 40 (SV40), produced virus at all stages of cultivation before and during the period of “crisis,” as reported for human fibroblasts. The virus production pattern was tri phasic, including an initial period of high virus production (phase 1), a period of low yields (phase 2), and a subsequent increase in yields before “crisis” (phase 3). Variation of the multiplicity of the initial virus exposure and culture age, and of the transfer dilutions, did not change this pattern. However, the time of phase 3 was related to approaching “crisis” and therefore depended on the multiplicity of exposure and transfer dilution factors. Shortly after infection the number of virus-producing cells (infectious centers or V-antigen-containing cells) was high, as reported for fibroblasts, but a subsequent period with nearly all cells producing virus was not observed for the amnion cultures. During the next period (phase 2 for amnion cells) very few cells (amnion: 0.01–0.3%) were virus producers in both systems; the output per amnion cell, however, was relatively higher (up to 2500 ID 50/cell). Before “crisis,” virus-producing cells increased significantly in the amnion cultures, in contrast to a decrease reported for fibroblasts. Inclusion-body-containing amnion cells were observed both initially and during serial cultivation, even after many culture passages; their frequency was correlated with the virus titers in the culture supernatant. Virus production and cell recovery at culture transfer were inversely related, and, as previously reported for chromosome changes, the morphology and growth pattern of the transformed cells differed in the three virus phases. The triphasic virus production pattern in the present system is interpreted as the results of a series of shifts in the balance between the amount of virus available and the degree of susceptibility of the transformed cells to reinfection. “Crisis” is correlated with a cellular change toward a decreased resistance to SV40. Since premature “crisis” could not be induced by superinfection with SV40 during the period of low virus production, it is apparent that “crisis” requires a certain number of cell divisions.