Modifying RANKL/OPG mRNA expression in differentiating and growing human primary osteoblasts

Description

The OPG / RANKL system in primary cultures of human osteoblasts has been studied by di ff erent authors. However, very few studies have been performed on gene expression of RANKL and OPG at di ff erent stages of maturation on human osteoblast cultures.The e ff ect of 17-estradiol and 1,25dihydroxyvitamin D3 on the OPG / RANKL system is not known during the di ff erent states of cellular maturation. In this work we quantifi ed OPG and RANKL protein levels (ELISA) and the mRNA of OPG, RANKL, collagen type I, alkaline phosphatase, and osteocalcin (semi-quantita-tive RT-PCR) in human osteoblasts. We analyzed these in basal conditions and after incubation with 17- -estradiol and 1,25dihydroxyvitamin D3 in the first and second phases. We found that OPG secretion and expression levels increased throughout cellular growth.RANKL proteins were detected only in the first stage, and the expression increased throughout the first phase. Thus, the RANKL / OPG ratio was higher in immature osteoblasts than in mature osteoblasts. The evolution of RANKL gene expression was related to collagen I and alkaline phosphatase, while OPG was related to osteocalcin. We observed no modi fi cations after estradiol and 1,25dihydroxyvitamin D3 treatment. Our results suggest that the OB is a positive stimulator at precocious stages of diff erentiation on osteoclastogenic modulates.

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