Effect Of Four Different Osteoporosis Drugs On The Expression Of Matrix Gla Protein In Primary Osteoblasts Of SD Rat

Objective:To observe the expression of matrix gla protein(MGP) mRNA in cultured primary osteoblasts of Sprague-Dawley(SD) rat in vitro after treatment with osteoporosis drugs(vitamin K2,PTH,1,25(OH)2D3 and alendronate), to investigate the potential role of MGP in the pathogenesis of osteoporosis.Methods:1.Primary cultured osteoblasts(OBs) were derived from sequential trypsin/ collagenase-digested calvaria isolated from newborn SD rat (postnastal day 1-3).2.OBs of the second generation were identified by Van Gieson collagen staining, alkaline phosphorase(ALP) staining and calcified nodules staining.3.OBs of the fourth generation were selected to interfere with vitamin K2,PTH (1-34),1, 25 (OH) 2D3 and alendronate, then cultured for 24 h in mediums which contained various concentrations of vitamin K2(10-7,10-6,10-5mol/L, respectively), PTH(1-34) (10-9,10-8,10-7mol/L, respectively),1,25 (OH) 2D3(10-10,10-9,10-8mol/L respectively), alendronate (10-6,10-5,10-4mol/L, respectively). After cultured 24h, total RNA was extracted and examined by real-time quantitative RT-PCR.Results:l.The primary cultured cells had typical morphological characters of osteoblasts.2.Van Gieson collagen staining, ALP staining and calcified nodules staining were all positive.3. Vitamin K2,PTH(1-34),1,25(OH)2 D3 and alendronate could modulate the expression of MGP mRNA in calvarial osteoblasts, and in a dose-dependent manner.①.MGP mRNA was induced 24h after vitamin K2 treatment, with a 2.56-fold induction seen after administration with vitamin K2 at the concentration of 10-5mol/L (2.12-fold induction after 10-6mol/L vitamin K2 addition, 1.57-fold induction after 10-7mol/L vitamin K2 addition).②.MGP mRNA was remarkely induced 24h after PTH (1-34) treatment, with a 6.78-fold induction seen after administration with PTH (1-34) at the concentration of 10-7mol/L (5.31-fold induction after 10-8mol/L PTH (1-34)addition,2.23-fold induction after 10-9mol/L PTH (1-34) addition).③.MGP mRNA was induced 24h after 1,25 (OH) 2D3 treatment, with a 8.93-fold induction seen after administration with 1,25 (OH)2D3 at the concentration of 10-8mol/L (6.95-fold induction after 10-9mol/L 1,25 (OH) 2 D3 addition,3.47-fold induction after 10-10mol/L 1,25 (OH) 2D3 addition)。④.MGP mRNA was induced 24h after alendronate treatment, with a 3.47-fold induction seen after administration with alendronate at the concentration of 10-4mol/L (2.49-fold induction after 10-5mol/L alendronate addition,1.98-fold induction after 10-6mol/L alendronate addition)Conclusion:MGP mRNA was detected in OBs from primary culture of newborn SD rat calvaria by real-time quantitative RT-PCR. Vitamin K2,PTH,1,25 (OH) 2D3 and alendronate can upregulate MGP mRNA expression in calvarial osteoblasts, and in a dose-dependent manner. MGP may be a potent target of osteoporosis drugs, which could be involved in the pathogenesis of osteoporosis.