DEMONSTRATION OF CELLULAR AGING AND SENESCENCE IN SERIALLY PASSAGED LONG-TERM CULTURES OF HUMAN TRABECULAR OSTEOBLASTS

The proliferative capacity and cellular and biochemical characteristic s of human trabecular bone osteoblasts were analysed throughout their replicative lifespan in vitro. Like several other cell types, human os teoblasts demonstrated a typical Hayflick phenomenon of cellular aging comprising a period of rapid proliferation until cumulative populatio n doubling level (CPDL) 22 to 24, followed by a phase of slow growth a nd the final cessation of cell division at CPDL 32 to 34. Comparing yo ung cells (less than 20% lifespan completed) and old cells (more than 90% lifespan completed) revealed a progressive increase in population doubling (PD) time, a decrease in attachment frequency, a decrease in the number of S-phase positive cells, a decrease in the rates of DNA, RNA and protein synthesis, an increase in the protein content per cell and an increased proportion of senescence-specific beta-galactosidase positive cells. While osteoblastic production of collagen type I decr eased progressively during aging, alkaline phosphatase activity droppe d rapidly after the first few passages and then remained constant duri ng the rest of the proliferative lifespan. Significant morphological c hanges from thin and spindle-shaped early passage young cells to large , flattened and irregularly shaped late passage old cells full of intr acellular debris were observed. In comparison, osteoblasts established from an osteoporotic bone sample showed a maximum CPDL of less than 5 , had a longer PD time and exhibited abnormal senescent morphology. Th us, we have demonstrated for the first time that human osteoblasts, li ke several other diploid cell types, have a limited proliferative capa city in vitro and undergo aging and senescence as measured by various cellular and biochemical markers. In addition, preliminary studies sho w that cells from osteoporotic bone have a severely reduced proliferat ive capacity. This model of bone cell aging facilitates study of the m olecular mechanisms of osteoblast senescence as well as factors relate d to osteoblast dysfunction in patients with osteoporosis.