Perturbation of gastric mucosa in mice expressing the temperature-sensitive mutant of SV40 large T antigen. Potential for establishment of an immortalised parietal cell line

Abstract

Gastric parietal cells have a unique secretory membrane system that undergoes a profound transformation when the parietal cell is stimulated to secrete acid. Understanding this process has been hindered by the lack of an immortalised parietal cell line. Here we have explored a strategy for the development of a parietal cell line by the generation of transgenic mice bearing the temperature-sensitive mutant of the SV40 large T antigen (SV40 tsA58) under the control of the regulatory sequences of the gastric H+/K+ ATPase beta-subunit (H/Kbeta-tsA58). Three H/ Kbeta-tsA58 transgenic mouse lines were established, namely 218, 224 and 228, all of which expressed the tsA58 T antigen in the gastric mucosa. Unexpectedly, the gastric mucosae of all lines were hypertrophic indicating that the temperature-sensitive large T antigen was partially active at 37 degrees C. Immunofluorescence together with light and electron microscopic studies revealed that mature parietal and zymogenic cells were absent in H/Kbeta-tsA58 transgenic lines 218 and 224, and small undifferentiated cells were the dominant cell type in the gastric units. On the other hand, a few mature parietal cells were detected in line 228 together with an increased proportion of undifferentiated cells and, normally rare, pre-parietal cells. As line 228 represented a rich source of pre-parietal cells, gastric cells from line 228 were isolated and cultured at 33 degrees C, the permissive temperature for tsA58. Gastric epithelial cells, expressing the T antigen, were maintained in culture for over 6 weeks. Upon a temperature shift to 39 C the cultured gastric cells developed characteristics of differentiated parietal cells, including the presence of a nascent canaliculus and dramatically increased production of the gastric H+/K+ ATPase beta-subunit. Therefore, this system shows the potential to generate an immature parietal cell line that can be induced to differentiate in vitro.