This study was designed to assess human bone marrow cell response and particularly cell adhesion, proliferation, and differentiation, when cultured in vitro on titanium alloy and hydroxyapatite with different values of surface roughness. A further aim was to compare the cell response on the two materials, currently used in spinal surgery. Cell adhesion was determined after 0.5, 2, 4, and 18 hours of incubation; proliferation after 8, 11, 14, and 16 days of culture; and differentiation was evaluated with the expression of alkaline phosphatase activity after 8 and 16 days of culture. This study showed that bone marrow cells grew faster on titanium alloy than on hydroxyapatite, although fewer cells attached to titanium, compared to those attached to hydroxyapatite. No statistically significant difference was observed as the expression of alkaline phosphatase activity on hydroxyapatite and titanium alloy of the same roughness. Cell adhesion, proliferation, and differentiation are dependent on surface roughness of the biomaterial, and all three increased as the roughness of titanium alloy increased. Conclusively, surface roughness of titanium and hydroxyapatite significantly influences bone marrow cell response, and therefore these biomaterials should be used with rough outer surface, if a significant cell response on them is desired. These advantages of titanium and hydroxyapatite theoretically seem to be of particular importance in the following situations: long fusions, lumbosacral fusion, revision surgery with poor bone bank, neuropathic scoliosis associated with few bone graft reserves, and adult patients with severe osteoporosis.