去卵巢大鼠骨髓微环境中RUNX2\PPARγ基因及OPG\RANKL基因蛋白表达变化的实验研究

Changes in RUNX2\PPARγ and OPG\RANKL expression in the bone marrow microenvironment from ovariectomized rats

[目的]探讨在去卵巢大鼠骨质疏松模型骨髓微环境中成骨细胞、脂肪细胞和破骨细胞分化的相互关系及其机制。[方法]将3个月龄雌性SD大鼠16只随机分为2组:假手术组(SHAM)和去卵巢组(OVX),每组8只。采用双侧卵巢切除术复制骨质疏松大鼠模型。术后14周,应用双能X线吸收仪法(DXA)测第4腰椎和股骨骨密度(BMD)。qRT-PCR法测量骨髓细胞RUNX2、PPARγ、OPG和RANKL mRNA表达量。石蜡组织切片HE染色测量第3腰椎和胫骨近端骨组织脂肪细胞数目。免疫组织化学染色测定胫骨近端骨组织OPG/RANKL蛋白表达量。[结果]与SHAM比较,OVX组腰椎和股骨BMD下降(P<0.05)。OVX组股骨骨髓细胞成骨分化转录因子RUNX2mRNA水平比SHAM组增高(P<0.05),成脂分化转录因子PPARγmRNA表达水平比SHAM组增高(P<0.05)。OVX组胫骨和第3腰椎脂肪细胞数目比SHMA组增多(P<0.05)。与SHAM组比较,OVX组股骨骨髓细胞RANKLmRNA和胫骨RANKL蛋白表达量增加(P<0.05)且OPG/RANKL的比率都降低(P<0.05),而两者OPG的mR-NA和蛋白表达量无统计学差异(P>0.05)。[结论]去卵巢大鼠骨量丢失可能是骨髓微环境中成骨细胞分化、脂肪细胞分化和破骨细胞分化紊乱导致。

[ Objective] To explore the interrelationship and mechanism among osteoblastogenesis, adipogenesis and oste- oclastogenesis in the bone marrow microenvironment from ovariectomized rat model of osteoporosis. [ Method ] Sixteen healthy female SD rats ,3 months old,were randomly divided into two groups（ n = 8 each） ： sham group and OVX group. The osteoporotic rat model was established by bilateral ovariectomy . Fourteen weeks after ovariectomized, the bone mineral density （BMD） of fe- mur and the fourth lumbar was measured by DXA. QRT - PCR were used for mRNA levels of RUNX2, PPAR~, OPG and RANKL in bone marrow cells. The number of adipoeytes in the paraffin sections of bone tissues of proximal tibia and the third lumbar were measured with HE staining. The protein expression for OPG and RANKL in bone tissues of proximal tibia were measured by immunohistochemical staining. [ Result ] Compared with the sham group, BMD in the lumbar vertebra and femur from ovariectomized rats significantly decreased （P 〈 0.05 ）. The mRNA level of RUNX2 in bone marrow from the OVX rat fe- mur increased statistically compared with the sham group （ P 〈 0.05 ）. Compared with the sham guoup, The mRNA level of PPAR~ also increased statistically in the OVX （ P 〈 0.05 ）. The number of adipocytes in the tibia and the third lumbar vertebra were greater in the OVX group than in the sham group （P 〈 0.05 ）. Compared with the sham guoup, the mRNA level of RANKL in bone marrow from the femur and the protein expression for RANKL in the tibia were higher in the OVX group （P 〈 0.05 ） , and the ratio of OPG/RANKL were lower in both mRNA and protein （ P 〈 0.05 ）. But no significant differences was found for the mRNA and the protein of OPG between the two groups （ P 〉 0.05 ）. [ Condusion I The ovarieetomized rats exhibited loss of bone mass. This may be caused by the imbalances among osteoblastogenesis, adipogenesis and osteoclastogenesis in the bone marrow mlcroenvlronment.