Retroviral transduction of alveolar bone cells with a temperature-sensitive SV40 large T antigen

We have transduced adult human alveolar bone (AB) cells with a gene constru ct encoding a temperature-sensitive mutation of the SV40 large T antigen (t sT). Such cells divided rapidly, for more than 50 passages thus far, at a p ermissive low temperature (34.5 degreesC), comparable to the non-transduced parental cells at 37 degreesC. However, the tsT-transduced AB cells failed to grow at a non-permissive high temperature (39 degreesC) at which the T antigen is inactivated. Nevertheless, the cells formed mineralised nodules in vitro at both the low and high temperatures. Flow cytometry analysis sho wed that the transduced cells cultured at 34.5 degreesC, like the parental cells at 37 degreesC, were smaller and less granular than the transduced ce lls incubated at 39 degreesC. Moreover, the transduced cells grown at 34.5 degreesC were also found to express bone sialoprotein. osteopontin and type I collagen at levels similar to those of the parental cells at 37 degreesC , although osteonectin and fibronectin were down-regulated. When the transd uced cells were incubated at 39 degreesC, the expression of all antigens wa s up-regulated, particularly osteonectin. Thus, we have obtained long-term cultures of tsT-transduced AB cells whose growth is temperature-dependent a nd which express certain features characteristic of bone-derived cells.