Single Nucleotide Polymorphisms in the Human Gene for Osteoprotegerin are Not Related to Bone Mineral Density or Fracture in Elderly Women

Osteoprotegerin (OPG), a secreted member of the tumor necrosis factor receptor family, is a potent inhibitor of osteoclast activation and differentiation. In animal models OPG prevents bone loss, and in humans bone resorption can be reduced by injections of OPG. OPG may also play a role in cardiovascular disease since mice lacking the OPG gene display arterial calcification. In a screening effort of the OPG gene, we recently discovered a single nucleotide polymorphism in the promoter region of OPG (T950C), and reported an association with vascular morphology and function in 59 healthy individuals. Due to the pronounced effect of OPG on bone turnover, the present study was conducted to investigate whether OPG polymorphisms are also associated with bone mineral density or with fracture. The relationship between single nucleotide polymorphisms in the promoter region of OPG (T950C) and the first intron (C1217T), and bone mineral density, measured by DXA in the hip or spine or ultrasound of the heel, was investigated in the Malmö OPRA-study of 1044 women, all 75 years old. The possible relation to fracture incidence was also analyzed. Among the 858 and 864 individuals respectively, genotyped, no significant associations between the investigated single nucleotide polymorphisms and bone mineral density measurements (T950C P = 0.50–0.64, C1217T P = 0.51–1.00), quantitative ultrasound measurements of the calcaneus, or fractures (T950C P = 0.61–0.66, C1217T P = 0.14–0.33) were found. Thus, our results show that polymorphisms in the OPG gene, one of which has previously been found to be associated with cardiovascular morphology and function, are not associated with bone mineral density in elderly Swedish women.     

rhIGF-1对成骨细胞增殖、分化及骨保护素、骨保护素配体基因mRNA表达的影响

目的 观察不同剂量rhIGF-1对成骨细胞增殖，分化及骨保护素，骨保护素配体mRNA基因表达的影响，为明确骨保护素，骨保护素配体在骨质疏松症发病的作用机理及rhIGF-1在临床中应用提供实验依据。方法 取2代培养的大鼠成骨细胞，在rhIGF-1为0ng/ml,10ng/ml,20ng/ml,50ng/ml的浓度中培养。观察细胞的生长及钙结节的形成，MTT法，碱性磷酸酶（ALP），骨钙素（OCN）测定细胞增殖和分化，RT－PCR测定rhIGF-1对成骨细胞骨保护素，骨保护素配体基因mRNA表达的影响。结果 成骨细胞在7d可铺满瓶壁，30d可形成钙结节，rhIGF-1可促进成骨细胞的增殖，ALP和OGN的分泌，促进骨保护素，骨保护素配体基因的表达，以促进骨保护素表达明显，在rhIGF-1为10ng/ml时作用明显。结论 rhIGF-1可促进大鼠成骨细胞的增殖，分化，及骨保护素，骨保护素配体基因mRNA的表达，骨保护素mRNA表达显。rhIGF-1可能通过影响骨保护素，骨保护素配体而调节成骨细胞破骨细胞的平衡，使骨重建，从而防治骨质疏松症。     

Regulatory effects of osteoprotegerin on cellular and humoral immune responses

The interaction between receptor activator of nuclear factor-kappaB ligand (RANKL) and RANK has been reported to regulate immunity in addition to bone metabolism. The aim of this study was to determine if osteoprotegerin (OPG), an inhibitor of the RANKL-RANK interaction and possibly a new drug against osteoporosis, would adversely affect immunity. OPG was used to treat mice developing different models of cellular and humoral immune responses and also in vitro in T and B cell assays. In mice, OPG does not affect cell-mediated reactions such as contact hypersensitivity to the hapten oxazolone and liver damage, granuloma formation, and infectious load induced by mycobacterial infection. However, OPG increases humoral reactions such as the production of IgM, IgG, and IgE against the T cell dependent antigen keyhole limpet hemocyanin and the production of IgM against the T cell independent antigen Pneumovax. In vitro, OPG modestly co-stimulates T cells but does not affect the proliferation of B cells. OPG has modest immunoregulatory effects that seem to be confined to the humoral response to specific antigens.     

Raji-MG63细胞共培养体系中心肌营养蛋白样细胞因子1(CLCF1)对RANKL/OPG比值及成骨细胞分化的影响

目的探讨Raji-MG63细胞共培养体系中,心肌营养蛋白样细胞因子1(cardiotrophin-like cytokine factor 1,CLCF1)基因对骨保护素(osteoprotegerin,OPG)、核因子κB受体活化因子配体(receptor activator of NF-κB ligand,RANKL)和成骨细胞分化的影响。方法通过CRISPR/Cas9基因编辑技术构建CLCF1基因敲除Raji细胞系,实验分2组:对照组(Control组)和CLCF1敲除组(CLCF1-KO组)。应用Transwell技术建立Raji-MG63细胞共培养体系,通过蛋白质印迹法(western blot,WB)检测CLCF1、OPG、RANKL及JAK2/STAT3蛋白的变化。运用碱性磷酸酶(ALP)活性检测和茜素红染色观察成骨细胞分化能力。结果WB结果显示,与对照组比较,CLCF1敲除组OPG(P<0.01)蛋白明显下调,RANKL/OPG比值升高(P<0.001),磷酸化JAK2(P<0.001)和STAT3(P<0.01)蛋白明显下调,差异具有统计学意义;ALP活性检测和茜素红染色结果显示,Raji-MG63细胞共培养体系中,CLCF1敲除组MG63细胞ALP活性和矿化形成能力降低,与对照组比较差异具有统计学意义(P<0.001)。结论CLCF1基因能通过调控RANKL/OPG比值和JAK2/STAT3通路影响成骨细胞分化。     

虾青素对去卵巢糖尿病大鼠骨质流失的防治作用

目的探讨虾青素能否防治去卵巢糖尿病大鼠的骨质流失以及可能的机制。方法 3月龄雌性SD大鼠分为3组(每组6只):对照组CON(假手术),模型组OVX/T1DM(去卵巢糖尿病大鼠),药物组OVX/T1DM-ASX(去卵巢糖尿病大鼠,给予虾青素100 mg/(kg·d)。结果连续治疗60 d后,与OVX/T1DM组相比,OVX/T1DM-ASX组骨密度(BMD)明显升高(P0.01),血清I型胶原蛋白(CTX-1)、骨钙素、I型前胶原蛋白n端前肽(PINP)、抗酒石酸磷酸酶5b(TRACP 5b)水平均显著升高(P0.01)。虾青素治疗能抑制去卵巢糖尿病大鼠骨组织形态学的改变,减少骨髓脂肪细胞增加,提高OPG/RANKL的比值。结论虾青素对绝经后糖尿病骨质流失有保护作用,这种作用与调控OPG/RANKL轴有关。     

从中医“虚”论治原发性骨质疏松症与OPG/RANK/RANKL信号通路的相关性

原发性骨质疏松症(primary osteoporosis,POP)是现代临床中常见的骨科疾病,其归属于中医学之"骨痿""骨痹""骨枯"等范畴。随着中医药事业的发展,中医学在POP的治疗中应用愈加广泛,且疗效显著,已受到学界的广泛认可。但遗憾的是,其缺乏现代研究理论依据的支撑。随着分子生物学的发展,OPG/RANK/RANKL信号通路的发现对中医学治疗此病具有重要意义。故笔者将从中医"虚"理论出发,并以中医学脏腑辨证理论为基础,借助现代分子生物学的研究成果OPG/RANK/RANKL信号通路,揭示中医治疗POP的OPG/RANK/RANKL信号通路表达机制,以期为中医学治疗POP提供科学理论依据。