The Effects Of Calcium, Phosphorous And OPG On The Formation And Activation Of Osteoclasts

Osteocyte's biological function is very important to metabolic balance and health of bone. Many diseases caused low bone mass through increasing the number of osteoclasts and (or) reinforcing the activation of osteoclasts, therefore, understanding the regulative mechanisms involved in osteoclasts formation and activation has the value both in theory and in practice. Calcium and phosphorus are necessary to body. The amount of them is also the most in body. Intaking enough calcium and phosphorus is vital to maintain bone's health, In our work, we study effects of calcium and phosphorus on the formation and activation of osteoclasts, which is induced by a co-culture system with rat spleen cells and osteoblasts, as thus, we offer theory basis to rational utilization of calcium and phosphorus. Moreover, we also prove that they are two mechanisms involved in the osteoclasts formation and activation through the way of cell culture in vitro, so we should take two mechanisms into account in treating osteoporosis and bone malnutrition.1. The effects of calcium and phosphorus on osteoclasts formatio and activationOsteoclasts are obtained by a co-culture system with rat spleen and osteoblast. In the conversion process, different strengths of calcium, phosphorus and different concentration ratios of calcium to phosphorus were added to the co-culture system. The formation and activation of osteoclasts is detected by ways of morphologic observation, cytochemical staining for tartrate-resistant acid phosphatase (TRAP) that is a marker of osteoclasts and detection of lacunar resorption through scanning electron micrograph. The results show that high concentration of calcium and phosphorus inhibit the formation and activation of osteoclasts (P<0.05) . Moreover, we also find that experimental group, in which the ratio of calcium to phosphorus is 2:1, has the most powerful inhibitory action to osteoclasts formation and activation.2. The effects of osteoprotegerin(OPG) on osteoclasts formatio and activationOsteoclasts are obtained by a co-culture system with rat spleen and osteoblast. In the conversion process, different strengths of OPG were added to the co-culture system. The formation and activation of osteoclasts is detected by ways of morphologic observation, cytochemical staining for tartrate-resistant acid phosphatase (TRAP) that is a marker of osteoclasts and detection of lacunar resorption through scanning electron micrograph. The results show that the number of TRAP(+) osteoclasts is decrease when concentration of OPG is increase, the number and area of lacunar resorption is as also. Discrepancy between experimental group and control group has statistical significance(P<0.01). In experimental group (50ng/ml, 75ng/ml), we can not detect lacunar resorption.3. Studies on two regulative mechanisms involved in osteoclast formation and activationGM-CSF and TNF α ( ± IL-1 a ) were added to spleen cells obtained from C57 female rat, meanwhile, OPG and sRANKL were also added to these cultures to distinguish the pathway of osteoclastogenesis. The formation and activation of osteoclasts is detected by ways of morphologic observation, cytochemical staining for tartrate-resistant acid phosphatase (TRAP) that is a marker of osteoclasts and detection of lacunar resorption through scanning electron micrograph. The results indicate that in the presence of GM-CSF, TNF α is sufficient for inducing osteoclast differentiation from spleen cells and that TNF α acts synergistically with IL-1 α to stimulate lacunar resorption. This process is distinct from the RANKL/RANK/OPG signalling pathway.