Treatment of rat proximal and distal colonic cells with sodium orthovanadate enhances their adhesion and survival in primary culture.

We have studied the effect of sodium orthovanadate, an inhibitor of protein tyrosine phosphatases, on primary cultures of colonocytes and stromal cells. Everted proximal and distal colonic tissue of adult rats were disintegrated by a collagenase/dispase solution for 60 min at 37 degrees C to prepare viable gland fragments and isolated cells. Cell preparations were inoculated onto plastic substratum or cytodex-3 microcarriers in a defined maintenance medium or in 1% fetal calf serum media. Incorporation of sodium orthovanadate (> or = 50 microM) in these media constantly enhanced the survival (cell enumeration and trypan blue exclusion P < 0.05) and the adhesion (up to four-fold by crystal violet staining, P < 0.01) of colonocytes (characterized by cytokeratin-18, transforming growth factor-alpha or alkaline phosphatase expression) and stromal cells. Removal of sodium orthovanadate from culture media restored cellular death processes. Incorporation of 10 mM n-butyric acid did not promote cell adhesion and survival except for distal cells exposed to 2 mM sodium orthovanadate. Besides studies in the regulation of anoikis in primary culture, the model will help to assay the influences of dietary and growth factors on the biology of non-cancerous colonic cells.