Decreased bone resorption, osteoclast differentiation, and expression of vacuolar H+-ATPase in antisense DNA-treated mouse metacarpal and calvaria cultures ex vivo.
ABSTRACT Expression and function of vacuolar H(+)-ATPase, a key enzyme in bone resorption, were monitored in antisense DNA-treated bone organ cultures ex vivo. A novel fluoroimmunoassay was used to quantitate mRNA levels after treatment with various antisense, sense, or random DNA oligonucleotides. Conventional slot blots and in vitro translation experiments were used to monitor the efficiency of the antisense molecules. In cell cultures, the used antisense molecules were transported into osteoclasts and a population of mononuclear cells. A significant decrease in bone resorption and in the expression of the 16 kDa, 31 kDa, 42 kDa, 60 kDa, 70 kDa, and 116 kDa subunits of V-ATPase was seen after antisense treatment. Also, osteoclast differentiation was decreased in antisense-treated mouse metacarpal cultures. These data show that the proper function of V-ATPase in osteoclasts requires expression of the 16 kDa, 31 kDa, 42 kDa, 60 kDa, 70 kDa, and 116 kDa subunits of V-ATPase. Antisense DNA molecules can be used to inhibit osteoclast differentiation and function in tissue cultures, in which the physical and chemical cellular environment resembles that in vivo. However, more studies are needed to learn if antisense DNA molecules can be used for inhibiting bone resorption also in vivo.
Article: Prevention of wear particle-induced osteolysis by a novel V-ATPase inhibitor saliphenylhalamide through inhibition of osteoclast bone resorption.[show abstract] [hide abstract]
ABSTRACT: Wear particle-induced peri-implant loosening (Aseptic prosthetic loosening) is one of the most common causes of total joint arthroplasty. It is well established that extensive bone destruction (osteolysis) by osteoclasts is responsible for wear particle-induced peri-implant loosening. Thus, inhibition of osteoclastic bone resorption should prevent wear particle induced osteolysis and may serve as a potential therapeutic avenue for prosthetic loosening. Here, we demonstrate for the first time that saliphenylhalamide, a new V-ATPase inhibitor attenuates wear particle-induced osteolysis in a mouse calvarial model. In vitro biochemical and morphological assays revealed that the inhibition of osteolysis is partially attributed to a disruption in osteoclast acidification and polarization, both a prerequisite for osteoclast bone resorption. Interestingly, the V-ATPase inhibitor also impaired osteoclast differentiation via the inhibition of RANKL-induced NF-κB and ERK signaling pathways. In conclusion, we showed that saliphenylhalamide affected multiple physiological processes including osteoclast differentiation, acidification and polarization, leading to inhibition of osteoclast bone resorption in vitro and wear particle-induced osteolysis in vivo. The results of the study provide proof that the new generation V-ATPase inhibitors, such as saliphenylhalamide, are potential anti-resorptive agents for treatment of peri-implant osteolysis.PLoS ONE 01/2012; 7(4):e34132. · 4.09 Impact Factor