The rumen epithelium increases in size and surface area in response to increased feed intake, presumably due to increased epithelial cell proliferation. Isolated ruminal cells were cultured in vitro to determine the effect of various growth factors on ruminal cell proliferation. The growth factors evaluated included insulin, insulin-like growth factor-I and epidermal growth factor. VFA propionate and butyrate were also evaluated. Ruminal epithelial cells were isolated from five lambs via serial tryptic digestion and prepared for sterile culture by serial washes in sterile buffered salts solution. Cell proliferation was assessed by [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetra-zoliumbromide] (MTT) assay. Cell growth was stimulated to 75, 96 and 97% of the growth of cells cultured in DMEM supplemented with 5% FCS (for Ins, IGF-I and EGE respectively). Propionate and butyrate addition resulted in an overall decline in cell growth to below controls (61 and 49%, respectively). When butyrate (1 mmol) was present in the media Ins, IGF-I and EGF overcame the butyrate inhibition, but did not attain the growth observed in uninhibited cultures (67, 57 and 60%, respectively). Although butyrate has been implicated as a possible mediator of ruminal cell growth in vivo, growth factors such as insulin, IGF-I and EGF are more effective inducers of growth in vitro. (C) 1999 Elsevier Science B.V. All rights reserved.