The Klotho gene codes for a protein that is thought to influence the homeostasis of several tissues, including bone, as well as the aging process. Although the mechanism of action has not been fully elucidated, some studies in women have associated Klotho allelic variants to bone mineral density (BMD). The objective of this study was to determine the relationship of a common T/G polymorphism, resulting in a phenylalanine (F) to valine (V) substitution, with male bone mass. BMD was measured by dual-energy X-ray absorptiometry in 362 Spanish men aged 19-83 years. Klotho alleles were determined by a Taqman assay. Allele frequencies were 85% and 15% for the F and V alleles, respectively. In comparison with the most common FF genotype, young and middle-aged men (age less than 53 years) with FV/VV genotypes had higher age- and body mass index-adjusted BMD at the lumbar spine (1.059 +/- 0.017 vs. 1.016 +/- 0.011 g/cm(2), P = 0.036), the hip (1.077 +/- 0.017 vs. 1.033 +/- 0.011 g/cm(2), P = 0.028), and the calcaneus (0.599 +/- 0.125 vs. 0.547 +/- 0.108 g/cm(2), P = 0.012). Klotho alleles explained about 2-4% of BMD variance. However, Klotho genotype was not associated to BMD in older men. There were no Klotho-related differences in height, body weight, calcium intake, tobacco or alcohol consumption, or serum testosterone levels. In conclusion, these results suggest that allelic variants of Klotho constitute one of the genetic factors influencing BMD in male adults.