Proliferation and differentiation sequence of osteoblast histogenesis under physiological conditions in rat periodontal ligament.

To define the mechanism of osteoblast histogenesis, nuclear morphometry was utilized as a marker for precursor cell differentiation. One hour after 3H-thymidine injection, groups of 7-week-old rats were killed at hourly intervals over one complete 24-hr photoperiod (LD 12:12). S-phase and mitosis were assessed in autoradiographs of 3-micron sections of molar periodontal ligament (PDL) adjacent to a physiological bone-forming surface. Labeled nuclei were divided into four categories according to morphometry of nuclear size: A (40-79 micron3), B (80-119 micron3), C (120-169 micron3), and D (greater than or equal to 170 micron3) cells. C and D cells synthesize DNA during the light and divide in the following dark phase; the rhythm for A cells is the opposite. B cells demonstrated no preference and were subsequently determined to be nonosteogenic. Compared to A cells the S-phase photoperiod of C and D cells (combined) is approximately a one-to-one reciprocal relationship, suggesting two proliferating progenitors in series. Based on arrest points in the histogenesis sequence, five compartments are defined: 1) A cells, less differentiated, self-perpetuating precursors; 2) A' cells, committed osteoprogenitors; 3) C cells, G1 stage preosteoblasts; 4) D cells, G2 stage preosteoblasts; and 5) Ob cells, morphologically distinct osteoblasts. Minimal elapsed time for the A----A'----C----D----Ob sequence is about 60 hr (five alternating dark/light cycles). A stress/strain-mediated increase in nuclear volume (A'----C) is an important, rate-limiting step in osteoblast differentiation.