Expression of mRNAs for type-I collagen, bone sialoprotein, osteocalcin, and osteopontin at different stages of osteoblastic differentiation and their regulation by 1,25 dihydroxyvitamin D-3

We have used in situ hybridization to evaluate the effects of 1,25 dihydrox yvitamin D-3 (1,25 (OH)(2) D-3) on the expression of mRNA for bone-matrix p roteins and to determine whether mature osteoblasts respond differently to 1,25 (OH)(2) D-3 than younger, newly differentiated osteoblasts. Rat calvar ia cells were cultured for 7, 12, 15, and 19 days to obtain a range of nodu les from very young to very mature. At each time point, some cultures were treated with 10 nM 1,25 (OH)(2) D-3 for 24 h prior to fixation. In control cultures, type-I collagen mRNA was detectable in osteoblastic cells in very young nodules and increased with increasing maturity of the nodules and th e osteoblasts lining them. The bone sialoprotein mRNA signal was weak in yo ung osteoblasts, increased in older osteoblasts, and decreased in mature os teoblasts. Weak osteocalcin and osteopontin signals were seen only in osteo blasts of intermediate and mature nodules. 1,25 (OH)(2) D-3 treatment marke dly upregulated osteocalcin and osteopontin mRNAs and downregulated mRNA le vels of bone sialoprotein and, to a lesser extent, type-I collagen in both young and mature osteoblasts. However, a marked diversity of signal levels for bone sialoprotein, osteocalcin, and osteopontin existed between neighbo ring mature osteoblasts, particularly after 1,25 (OH)(2) D-3 treatment, whi ch may therefore selectively affect mature osteoblasts, depending on their differentiation status or functional stage of activity.