To determine the osteoblastic dysfunction that may be involved in the pathophysiology of osteoporosis in men we have compared histomorphometric indices of bone formation with in vitro characteristics of osteoblastic cells isolated from the trabecular bone surface in 23 untreated men with eugonadal osteoporosis. In most patients (n = 14), trabecular bone loss resulted from decreased bone formation evidenced by a lower than normal osteoblast surface, double tetracycline labeled surface, bone formation rate, and mean wall thickness. In these patients, DNA synthesis by cultured osteoblastic cells was altered. The peak of [3H]thymidine incorporation into DNA, the maximal DNA synthesis, and the area under the curve of cell proliferation were lower than the values in normal bone cells from age-matched controls. Parameters of bone cell growth were decreased in correlation with the extent of actively bone forming surfaces. By contrast, in patients (n = 9) with normal histomorphometric indices of bone formation, bone cell proliferation in vitro was not different from normal. Parameters of osteoblastic differentiation in vitro such as osteocalcin production and alkaline phosphatase activity were normal in the two groups of patients. This study shows that the trabecular bone loss resulting from defective bone formation in eugonadal osteoporotic men is associated with a lower than normal proliferative capacity of osteoblastic cells lining the trabecular bone surface.