Rumen fluid inhibits proliferation and stimulates expression of cyclin-dependent kinase inhibitors 1A and 2A in bovine rumen epithelial cells

It has been known for decades that microbial fermentation within the rumen is critical to postnatal rumen epithelial growth and maturation in ruminants, but the underlying mechanism is largely unknown. In this study, we determined the effect of rumen fluid, which should contain all products from rumen fermentation, on growth of rumen epithelial cells in vitro. Addition of 10% rumen fluid from cows to the culture medium inhibited (P < 0.05), whereas addition of 6.5 mM acetate, 2.5 mM propionate, or 1 mM butyrate had no effect (P > 0.1) on, the proliferation of rumen epithelial cells isolated from newborn calves. Flow cytometric assays showed that 10% rumen fluid inhibited (P < 0.05) the transition of rumen epithelial cells from the G1 phase to the S phase during the cell cycle. Real-time RT-PCR analyses of mRNA for key cell cycle regulators indicated that 10% rumen fluid did not change (P > 0.1) the expression of cyclin D1, D2, D3, E1, or E2 mRNA or that of cyclin-dependent kinase inhibitor 1B or 2B mRNA, but increased (P < 0.05) the expression of cyclin-dependent kinase inhibitors 1A and 2A mRNA in rumen epithelial cells. These mRNA data support the possibility that rumen fluid inhibits proliferation of rumen epithelial cells in vitro by increasing the expression of cyclin-dependent kinase inhibitors 1A and 2A. The result that rumen fluid inhibits proliferation of bovine rumen epithelial cells in culture indicates that rumen fermentation does not stimulate the postnatal rumen epithelial growth in cattle by directly stimulating proliferation of rumen epithelial cells.