唑来膦酸对大鼠颌面骨和外周骨创伤后骨改建的影响

目的 研究唑来膦酸对大鼠颌面骨和外周骨创伤后骨改建的影响。方法 SD大鼠随机分为实验组和对照组,每周分别给予尾静脉注射唑来膦酸(80 μg/kg)和PBS。用药两周后,在全麻下拔除一侧上颌第一磨牙,并在同侧胫骨近心端制备骨缺损,缝合创口,继续用药至术后1、4和12周后分批处死,分离并收集骨组织样本。Micro-CT分析各组骨缺损区新骨形成情况。HE和Masson染色分析骨缺损区软组织愈合情况、新骨形成情况、有无炎症反应和死骨形成等。ELISA法检测骨改建过程中关键因子RANKL和OPG的表达。结果 Micro-CT结果显示,实验组拔牙创表面高低不平,中部浅凹处可见游离的死骨片,而对照组拔牙创新骨形成区域与周边骨质均匀连续,进行了正常的生理性骨改建。实验组胫骨缺损区愈合,骨皮质完整连续,较对照组厚且致密,骨松质内的新生骨亦明显较对照组排列紧密。术后4周和12周,实验组胫骨BV/TV值较对照组明显上升(P<0.05),实验组颌骨BV/TV值较对照组差异无统计学意义。组织学染色显示,实验组颌骨拔牙创黏膜未愈合或延迟愈合,未愈合的黏膜下方可见暴露骨坏死,骨质出现不同程度的硬化,且周围伴有大量的炎性细胞浸润,为典型的双膦酸盐相关性颌骨坏死(BRONJ)组织病理表现。对照组颌骨拔牙创上皮正常愈合,覆盖创面,拔牙窝进行了正常的生理性骨改建。实验组胫骨骨缺损已愈合,骨皮质较对照组增厚,新骨生成和骨改建速度较对照组快,松质骨内新生骨小梁数量和密度亦较对照组增高。细胞因子检测显示,实验组颌骨RANKL/OPG比值较对照组明显下降(P<0.05),实验组胫骨RANKL/OPG比值则较对照组明显上升(P<0.05)。结论 唑来膦酸抑制大鼠颌骨拔牙后骨改建,引起颌骨BRONJ样病变,却在一定程度上促进大鼠外周骨骨创后骨改建。RANKL/OPG值可能在BRONJ发生过程中起重要作用。     

骨灵片通过p38 MAPK信号通路影响成骨细胞功能的机制

目的 探讨骨灵片促进成骨细胞增殖、矿化以及基因表达的细胞信号通路.方法 用含有骨灵片的大鼠血清、p38MAPK抑制剂SB203580作用人成骨细胞MG-63,测定各组成骨细胞增殖率、矿化结节数量.采用Western-blotting及Real time RT-PCR方法观察各组成骨细胞护骨素(OPG)、护骨素配体(RANKL)以及巨噬细胞集落刺激因子(M-CSF)基因表达和磷酸化p38含量.结果 经含骨灵片药液的大鼠血清作用后,MG-63细胞的增殖和矿化结节形成数明显增加,OPG基因表达增加,RANKL基因表达减少,OPG/RANKL比值显著增加,M-CSF有下降趋势,但没有显著性差异,同时刺激p38磷酸化.结论 骨灵片可能通过p38MAPK通路促进成骨细胞增殖以及矿化结节形成,调控OPG、RANKL以及M-CSF的表达,达到治疗骨质疏松症的作用.     

甲状腺激素对血管内皮细胞护骨素表达的影响

目的:观察甲状腺激素对血管内皮细胞护骨素(OPG)表达的影响.方法:人脐静脉内皮细胞(HUVEC)分别在不同浓度甲状腺激素(0.1、1、50 nmol/L,T3)条件下培养24h.ELISA法检测细胞培养液上清OPG的含量,RT-PCR法检测内皮细胞OPG mRNA的表达水平.结果:与生理浓度组T3(1 nmol/L)比较,低浓度组T3(0.1 nmol/L)内皮细胞培养液上清OPG的含量增加(P<0.05),OPG mRNA表达水平明显增高(P<0.05);高浓度组T3(50 nmol/L)内皮细胞培养液上清OPG的含量减少(P<0.05),OPG mRNA表达水平明显降低(P<0.05).结论:HUVEC在低浓度甲状腺激素作用下OPG表达增加,提高甲状腺激素浓度可逆转该现象.     

RANK, RANKL and osteoprotegerin in bone biology and disease

Upon the discovery of RANK, RANKL and OPG in the late 1990s, their importance in the maintenance of the skeletal structure and their dramatic role in bone disease were largely unexpected. In recent years the understanding of these proteins, in particular their regulation, has greatly increased. This review aims to bring the interested reader up to date with the latest news and views on the mechanisms controlling bone resorption in normal and pathological conditions.     

Osteoprotegrin knockout mice demonstrate abnormal remodeling of the otic capsule and progressive hearing loss

OBJECTIVES: The otic capsule, when compared with other bones in the body, is unique in that it undergoes no significant remodeling of bone after development. We previously demonstrated that osteoprotegerin (OPG), which inhibits formation and function of osteoclasts, is produced at high levels in the inner ear of normal mice and secreted into the perilymph from where it diffuses into the surrounding otic capsule bone through a lacunocanalicular system. To test our hypothesis that the high level of OPG may be important in the inhibition of otic capsule remodeling, we studied the light microscopic histology of the otic capsule in OPG knockout mice for evidence of abnormal remodeling of bone. We also tested the hearing in OPG knockout mice to determine whether OPG and its influence on surrounding bone is important for auditory function. METHODS: Temporal bone histopathology and pathophysiology were compared in homozygous OPG knockout mice and C57BL/6 (B6) mice, the background strain for the knockouts. Auditory function in age-matched animals from each group was evaluated at approximately 4-week intervals from 8 to 21 weeks using frequency-specific auditory brainstem responses (ABR) and distortion product otoacoustic emissions (DPOAE). After each of the last three evaluations, the cochleae from one mouse of each group were harvested, processed, and examined by light microscopy. RESULTS: Osteoprotegerin knockout mice demonstrated abnormal remodeling of bone within the otic capsule with multiple foci showing osteoclastic bone resorption and formation of new bone. Such changes were not seen in the age-matched B6 controls. The active bone remodeling process in the knockout animals showed many similarities to otosclerosis seen in human temporal bones. Over the time period that we monitored, auditory function was significantly and progressively compromised in the knockout animals relative to B6 controls. At the earliest age of test (8 wk), the loss was apparent as a mild, high-frequency reduction in sensitivity by ABR. In contrast, DPOAE losses in the knockouts were substantial even at 8 weeks, and by 21 weeks, these losses exceeded our equipment limits. Results of ABR testing showed hearing sensitivity changes in the animals of the background strain were confined largely to the high frequencies, whereas OPG knockouts demonstrated substantial low-frequency shifts in addition to those at high frequencies. CONCLUSIONS: The histopathological and pathophysiological findings in OPG knockout mice support the hypothesis that OPG is important in the inhibition of bone remodeling within the otic capsule and the maintenance of normal auditory function. This mouse may provide a valuable animal model of human otosclerosis.     

Breast cancer-derived Dickkopf1 inhibits osteoblast differentiation and osteoprotegerin expression: implication for breast cancer osteolytic bone metastases

Most breast cancer metastases in bone form osteolytic lesions, but the mechanisms of tumor-induced bone resorption and destruction are not fully understood. Although it is well recognized that Wnt/beta-catenin signaling is important for breast cancer tumorigenesis, the role of this pathway in breast cancer bone metastasis is unclear. Dickkopf1 (Dkk1) is a secreted Wnt/beta-catenin antagonist. In the present study, we demonstrated that activation of Wnt/beta-catenin signaling enhanced Dkk1 expression in breast cancer cells and that Dkk1 overexpression is a frequent event in breast cancer. We also found that human breast cancer cell lines that preferentially form osteolytic bone metastases exhibited increased levels of Wnt/beta-catenin signaling and Dkk1 expression. Moreover, we showed that breast cancer cell-produced Dkk1 blocked Wnt3A-induced osteoblastic differentiation and osteoprotegerin (OPG) expression of osteoblast precursor C2C12 cells and that these effects could be neutralized by a specific anti-Dkk1 antibody. In addition, we found that breast cancer cell conditioned media were able to block Wnt3A-induced NF-kappaB ligand reduction in C2C12 cells. Finally, we demonstrated that conditioned media from breast cancer cells in which Dkk1 expression had been silenced via RNAi were unable to block Wnt3A-induced C2C12 osteoblastic differentiation and OPG expression. Taken together, these results suggest that breast cancer-produced Dkk1 may be an important mechanistic link between primary breast tumors and secondary osteolytic bone metastases.     

基于“肝主筋、肾主骨”理论探讨OPG/RANKL/RANK信号轴与绝经后骨质疏松症症的筋骨相关性

中医基础理论素有“肝主筋，肾主骨”之说，肝藏血主筋，筋束骨利关节，肾为先天之本，主骨而生髓，肝肾精血同源、筋骨相关，互资互生，二者共同介导机体器官、脏腑的发育、生长与衰退过程，尤其与骨细胞生长代谢机制关系密切。随着中医药分子生物学研究的深入，发现骨保护蛋白（OPG）/核因子-kB受体活化因子（RANK）/核因子-kB受体活化因子配体（RANKL）信号轴在骨代谢中发挥关键作用。本文基于“肝主筋，肾主骨”理论，从绝经后骨质疏松症症的病因病机出发，从“肝肾论治，筋骨并重”角度将OPG/RANKL/RANK信号转导系统与骨代谢内环境稳态相结合，为中医药柔肝补肾、调筋养骨机制提供分子生物学、筋骨力学依据支撑，以期进一步指导临床论治。     

大黄素治疗对去卵巢大鼠骨量流失的保护作用机制研究

目的探讨大黄素(emodin,DHS)对去卵巢大鼠骨量流失的影响,并探索可能的机制。方法通过双侧去卵巢建立骨质疏松大鼠模型;随后随机分为假手术组(Sham)、去卵巢组(OVX)以及大黄素组(DHS),每组10只;其中DHS组大鼠接受大黄素[90 mg/(kg·d)]治疗12周;待治疗结束后使用Micro-CT、HE染色切片、骨代谢指标、以及蛋白质印迹观察治疗效果以及可能的机制。结果治疗12周后,与OVX组相比,Micro-CT和HE染色切片结果显示DHS组的大鼠骨小梁数量和骨密度得到明显改善。DHS组大鼠BMD、TV/BV、Tb.N、Tb.Th和Tb.Sp较OVX组明显改善(P<0.05)。与OVX组相比,DHS组的BALP水平明显升高(P<0.05);而TRACP-5b和β-CTX水平显著降低(P<0.05)。和OVX组比较,DHS组OPG表达水平上调(P<0.05),而RANKL和β2AR表达水平下调(P<0.05)。结论大黄素可以通过降低β2AR表达和激活OPG/RANKL信号通路介导对去卵巢大鼠骨量流失的保护作用。     

Osteoporosis following heart transplantation and immunosuppressive therapy

Heart transplantation (HT) remains the ultimate final therapy for patients with end-stage heart failure, who despite optimal medical and surgical treatments exhibit severe symptoms. To prevent rejection of the transplanted organ, HT patients require life-long immunosuppressive therapy. The goal of the immunosuppression is to minimise the risk of immune-mediated graft rejection, while avoiding clinical side-effects. Current immunosuppressive agents have yielded good survival outcome, however, complications of the immunosuppressive therapy, such as impaired bone strength and increased fracture risk, are common among HT patients rendering increased morbidity and mortality rates. The main aim of the present review was to summarise current knowledge on bone strength impairment after HT and concomitant immunosuppressive therapy.