Several mesenchymally derived cells, including osteoblasts, secrete HGF. 1,25(OH)₂-vitamin D₃ (1,25D) inhibits proliferation and induces differentiation of MG-63 osteoblastic cells. Here we show that MG-63 cells secrete copious amounts of HGF and that 1,25D inhibits HGF production. MG-63 cells also express HGF receptor (c-Met) mRNA, suggesting an autocrine action of HGF. Indeed, while exogenous HGF failed to stimulate cellular proliferation, neutralizing endogenous HGF with a neutralizing antibody inhibited MG-63 cell proliferation; moreover, inhibiting HGF synthesis with 1,25D followed by addition of HGF rescued hormone-induced inhibition of proliferation. Non-neutralized cells displayed constitutive phosphorylation of c-Met and the MAP kinases MEK1 and Erk1/2, which were inhibited by anti-HGF antibody. Constitutive phosphorylation of Erk1/2 was also abolished by 1,25D. Adding HGF to MG-63 cells treated with neutralizing HGF antibody induced rapid phosphorylation of c-Met, MEK1 and Erk1/2. Thus endogenous HGF induces a constitutively active, autocrine mitogenic loop in MG-63 cells. The known anti-proliferative effect of 1,25D on MG-63 cells can be accounted for by the concomitant 1,25D-induced inhibition of HGF production.