西瑞香素通过NF-κB途径抑制RANKL诱导的破骨细胞分化

摘要：目的:研究西瑞香素作为一种具有抗炎抗氧化作用的中药成分对破骨细胞分化的影响及机制。方法:从BALB/c小鼠骨髓中分离获得骨髓单核细胞,使用GST-rRANKL诱导形成破骨细胞并分为3组:对照组、RANKL诱导组和RANKL+西瑞香素组,采用TRAcP染色评估西瑞香素对破骨细胞成熟与融合的影响,用Western Blot和qRT-PCR检测破骨细胞相关表型蛋白和核因子κB（NF-κB）信号通路蛋白,构建荧光素酶报告基因检测转录因子NF-κB活性。结果:西瑞香素能够显著抑制RANKL诱导的破骨细胞形成,抑制RANKL刺激后的Acp5、V-ATPase-d2和CTSK的转录水平,并抑制与破骨细胞分化密切相关的两个转录因子NFATc-1和c-Fos的表达量。西瑞香素能促进IκB-α的表达,从而抑制NF-κB活性。结论:西瑞香素通过促进IκB-α的表达,抑制NF-κB活性,使破骨细胞分化相关的关键转录因子NFATc-1和c-Fos的表达量下调,从而通过抑制NF-κB途径发挥抑制RANKL诱导的破骨细胞分化的作用。     

丹参酮ⅡA调节骨关节炎小鼠骨代谢的作用机制

目的 探讨丹参酮ⅡA(TanⅡA)通过介导Yes激酶相关蛋白（Yes-associated protein,YAP）、核因子-κB受体活化因子配基（receptor activator of nuclear factor-κB ligand,RANKL）/核因子κB受体活化因子（eceptor activator of nuclear factor-κB,RANK）/骨保护蛋白（osteoprotegerin,OPG）调节骨关节炎小鼠骨代谢的作用机制。方法 建立骨关节炎小鼠模型,将60只小鼠随机分成假手术组、模型组、TanⅡA低剂量组和TanⅡA高剂量组,每组15只,造模成功后灌胃给药,连续4周。HE和番红O固绿染色观察软骨组织病理损伤并进行Mankin评分。酶联免疫吸附试验（enzyme-linked immunosorbent assay, ELISA）检测血清骨碱性磷酸酶（bone alkaline phosphatase,BALP）、骨钙素（osteocalcin,OC）、Ⅰ型胶原交联羧基末端肽（C-telopeptide of typeⅠcollagen,CTX）、白细胞介素...     

资木瓜总苷对RA模型小鼠的骨保护作用及机制

目的 研究资木瓜总苷对类风湿关节炎（RA）模型小鼠的骨保护作用及可能机制，为进一步将其开发为抗RA药物提供参考。方法 将70只雄性DBA/1小鼠按随机数字表法分为正常组、模型组、资木瓜总苷低剂量组（60 mg/kg）、资木瓜总苷高剂量组（240 mg/kg）和雷公藤多苷片组（阳性对照，30 mg/kg），每组14只。除正常组外，其余各组小鼠均采用葡萄糖-6-磷酸异构酶混合多肽诱导制备RA小鼠模型。观察并记录各组小鼠的体质量、后足趾厚度、关节炎评分；采用苏木精-伊红染色法、抗酒石酸酸性磷酸酶染色法及番红固绿染色法观察小鼠踝关节组织的滑膜炎症及骨、软骨破坏情况；采用酶联免疫吸附测定法检测小鼠血清中白细胞介素6(IL-6)和踝关节组织中肿瘤坏死因子α(TNF-α)、IL-4、IL-10的含量；采用Western blot法检测小鼠踝关节组织中核因子κB受体活化因子配体（RANKL）、核因子κB受体活化因子（RANK）、骨保护素（OPG）、肿瘤坏死因子受体相关蛋白6(TRAF6)、活化T细胞核因子1(NFATC1)的蛋白表达水平。结果 在给药结束时，与正常组比较，模型组小鼠的体质量显著降低（P...     

亚砷酸对大鼠类风湿关节炎模型作用机制的探讨

目的：研究亚砷酸在类风湿关节炎（RA）大鼠中的治疗作用.方法：40只大鼠随机分成4组：正常对照组（C组）、模型组（M组）、低剂量亚砷酸组（LSA 1.5 mg·kg^-1·d^-1）、高剂量亚砷酸组（HSA 3.0 mg·kg^-1·d^-1）.原位杂交检测各组核因子-κB（NF-κB）、细胞核因子-κB受体活化因子配体（RANKL）mRNA的表达.结果：与C组比较,M组NF-κB、RANKL mRNA大量表达（P＜0.01）;与M组比较,LSA组及HSA组NF-κB、RANKL mRNA表达降低（P＜0.01）,且HSA组更明显.结论：RANKL、NF-κB在类风湿关节炎发生发展中具有重要作用,亚砷酸对类风湿关节炎有治疗作用.     

强直性脊柱炎骨破坏中RANKL-RANK-OPG系统及中药干预研究进展

摘 要强直性脊柱炎（AS）是风湿性疾病中的一种慢性炎症性疾病,主要累及脊柱和骶髂关节。破骨细胞在AS骨破坏中发挥重要作用,细胞核因子κB受体活化因子配体/细胞核因子κB受体活化因子/骨保护素（RANKL-RANK-OPG）系统通过影响破骨细胞的生成和激活进而在骨病变中发挥重要作用。本文就近年来该系统对强直性脊柱炎骨质破坏的影响以及中药的干预作用进行综述,为抗AS新药发现和机制研究提供依据。     

穿心莲内酯促进骨折骨愈合作用机制的研究进展

骨折后的愈合状态直接影响患者的生活质量,如何提高骨折患者骨愈合效果是亟待解决的问题。穿心莲内酯是穿心莲中主要活性成分,可能通过促进成骨细胞增殖,抑制核因子κB（NF-κB）信号通路激活,激活Wnt/β-连环蛋白（Wnt/β-catenin）信号通路,调节护骨素/细胞核因子κB受体活化因子配基（OPG/RANKL）信号通路,调节成骨基因表达产物,改善软骨细胞功能,抑制雌激素相关受体α（ERRα）信号通路等多途径促使骨愈合。总结了穿心莲内酯促进骨折骨愈合作用及其作用机制,希望为穿心莲内酯的临床使用提供依据。     

RANK/RANKL/OPG系统及其相关疾病和抗RANKL疗法的研究进展

骨骼是一个动态组织,在生物的整个生命周期中不断地进行构建、吸收、重建,核因子(nuclear factor,NF)κB受体活化因子(receptor activator of NF-κB,RANK)/NF-κB受体活化因子配体(receptor activator of NF-κB ligand,RANKL)/护骨素(osteoprotegerin,OPG)系统是调控这一过程的关键系统.RANK/RANKL/OPG属于肿瘤坏死因子及其受体超家族,三者通过调控破骨细胞的分化和活化来影响骨吸收和重建的过程.另外,免疫细胞也参与和调节RANK/RANKL的表达和分泌,而免疫细胞本身亦受到该系统的影响,因此,RANK/RANKL/OPG系统成为骨和免疫之间的重要关联.RANK/RANKL/OPG系统的失衡与多种骨代谢性疾病及免疫系统疾病引发的继发性骨病密切相关,该系统的发现为研究相关疾病的病理机制和治疗方法提供了基础.由此建立的抗RANKL疗法已经开始应用于临床试验和研究.此文对RANK/RANKL/OPG系统、系统相关疾病以及抗RANKL治疗的进展做一综述.     

淫羊藿苷联合GM6001治疗酒精性股骨头坏死大鼠的作用机制及对OPG/RANK/RANKL通路的影响

目的 研究淫羊藿苷联合GM6001治疗酒精性股骨头坏死大鼠的作用机制,同时探究其对骨保护素（OPG）/核因子-κB受体活化因子（RANK）/核因子-κB受体活化因子配体（RANKL）通路的调节作用。方法 大鼠随机分为对照组、模型组、淫羊藿苷组、GM6001组及联合组（淫羊藿苷＋GM6001）,每组12只。采用ig给予大鼠白酒建立酒精性股骨头坏死大鼠模型,淫羊藿苷组按照60 mg/kg ig给予大鼠淫羊藿苷,GM6001组按照100 mg/kg尾iv GM6001,联合组同时给予淫羊藿苷（60 mg/kg）及GM6001（100 mg/kg）,1次/d,连续8周,对照组及模型组给予生理盐水。旷场实验测定大鼠活动次数、活动总距离,抓力测定仪测定大鼠最大抓力;显微CT仪测定股骨头骨体积分数、骨小梁间距、骨小梁数目、骨小梁厚度;酶联免疫吸附法（ELISA）测定血清磷、钙水平,血清及股骨头骨形成蛋白-2（BMP-2）、转化生长因子-β（TGF-β）、碱性成纤维细胞生长因子（bFGF）水平。苏木精–伊红（HE）染色法测定股骨头组织病理学变化;实时定量聚合酶链式反应（PCR）测定股骨头OPG、RANK、RANKL mRNA水平;免疫印迹法测定股骨头OPG、RANK及RANKL蛋白水平。结果 与模型组比较,淫羊藿苷组、GM6001组、联合组活动次数、活动总距离、最大抓力、骨体积分数、骨小梁数目、骨小梁厚度,血清磷、钙、BMP-2、TGF-β、bFGF水平,股骨头BMP-2、TGF-β、bFGF水平,股骨头OPG mRNA及蛋白水平显著升高（P＜0.05）;骨小梁间距、股骨头RANK和RANKL的mRNA及蛋白水平显著降低（P＜0.05）,且联合组降低更为显著（P＜0.05）。结论 淫羊藿苷联合GM6001能够显著修复大鼠酒精性股骨头坏死,缓解股骨头组织病理学改变,其机制可能与调节OPG/RANK/RANKL有关。     

霉酚酸抑制内皮细胞核因子—κB的活性

目的：研究霉酚酸对内皮细胞核因子－κB（NF－κB）活力及其抑制因子IκBa的影响。方法：利用凝胶迁移率实验（EMSA）检测不同浓度的霉酚酸对一般培养状态和佛波脂（PMA）激活的内皮细胞的NF－κB活力影响。并用Western blot方法检测霉酚酸对NF－κB抑制因子IκBa的作用。结果：PMA显著激活内皮细胞中的NF－κB活性。降低IκBα的蛋白水平,霉酚酸抑制PMA刺激的内皮细胞NF－κB活性升高,并抑制内皮细胞胞浆IκBα蛋白的降解,结论：霉酚酸可影响内皮细胞NF－κB的活性,这可能是MPA影响内皮细胞功能的作用机制之一。     

鲫鱼卵唾液酸糖蛋白抑制小鼠破骨细胞的分化机制研究

目的：探讨鲫鱼卵唾液酸糖蛋白(Carassius auratus sialyglycoproteins,Ca-SGP)对核因子κB受体活化因子配体(receptor activator of NF-κB ligand,RANKL)诱导形成的破骨细胞的抑制作用及机制。方法：初断乳ICR雄性小鼠,无菌取股骨,分离骨髓造血细胞,通过诱导剂巨噬细胞集落刺激因子(macrophage colony stimulating factor,M-CSF)和RANKL诱导破骨细胞的分化与成熟。MTT法检测Ca-SGP对破骨细胞及其前体细胞活力的影响;通过抗酒石酸酸性磷酸酶(tartrate-resistant acid phosphatase,TRAP)染色、TRAP检测试剂盒和骨吸收陷窝实验测定Ca-SGP在破骨细胞分化与成熟过程中的作用;实时荧光定量PCR法(qRT-PCR)测定破骨细胞形成过程中核因子κB(Nuclear factor,NF-κB)和丝裂原活化蛋白激酶(mitogen-activated protein kinases,MAPK)信号通路关键因子的表达。结论：Ca-SGP可有效抑制由RANKL诱导的破骨细胞分化及成熟,降低细胞TRAP活性及其表达,抑制骨吸收陷窝形成;分子机制方面,Ca-SGP可明显降低信号传导关键蛋白TRAP6的活性,进而抑制NF-κB和MAPKs信号通路关键因子mRNA的表达,达到抑制破骨细胞分化成熟的效果。结果：Ca-SGP可通过NF-κB和MAPKs信号通路抑制由RANKL诱导的破骨细胞形成和分化成熟。     

Environmental level of cadmium exposure stimulates osteoclasts formation in male rats

Low level of cadmium (Cd) exposure may enhance osteoclasts formation in vitro. The aim of the study was to observe the effects of Cd on osteoclasts formation in vivo. Sprague-Dawley male rats were divided into 4 groups which were given Cd via drinking water at concentrations of 0, 2, 10 and 50 mg/L for 12 weeks. At the 12th week, urine samples were collected from all of the rats. All rats were then sacrificed and the blood was collected for biomarkers assay. Bone tissues were dissected for mineral density determinations, histological investigation, tartrate resistant acid phosphatase staining and immunohistochemical staining. The bone mineral density and bone microstructure index of rats treated with 50 mg Cd/L were obviously lower than in control rats. Histochemical investigation showed that Cd could induce osteoclasts formation in a dose-dependent manner. Tartrate resistant acid phosphatase 5b levels in rats treated with Cd were higher than the control. Immunohistochemical investigation showed that Cd could enhance receptor-activated nuclear factor kappa B ligand expression (RANKL) and inhibit osteoprotegerin (OPG) expression. Our study evidences in vivo that excessive bone resorption mediated via osteoclasts is an important way for Cd toxic effects on bone and OPG/RANKL may play an important role.     

Inhibition of osteoclast differentiation and bone resorption by tanshinone IIA isolated from Salvia miltiorrhiza Bunge

Osteoclasts, multinuclear cells specialized for bone resorption, differentiate from the monocyte/macrophage lineage of hematopoietic cells. Intervention in osteoclast differentiation is considered an effective therapeutic approach to the treatment of bone diseases involving osteoclasts. In this study, we found that tanshinone IIA, originating from Salvia miltiorrhiza Bunge, inhibited the differentiation of osteoclasts. Addition of tanshinone IIA to the osteoclast precursor culture caused a significant decrease in the level of calcitonin receptor, c-Src, and integrin beta3 mRNA, which are normally upregulated during the osteoclast differentiation dependent on RANKL (receptor activator of nuclear factor kappa B ligand). RANKL activated the ERK, Akt, and NF-kappaB signal transduction pathways in osteoclast precursor cells, and tanshinone IIA suppressed this activation. Tanshinone IIA also inhibited the bone resorptive activity of differentiated osteoclasts, which was accompanied with the disruption of the actin ring. Thus, tanshinone IIA has the potential to ameliorate bone-resorption diseases in vivo by reducing both the number and activity of osteoclasts.     

Dosing time‐dependent effect of raloxifene on plasma plasminogen activator inhibitor‐1 concentrations in post‐menopausal women with osteoporosis

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地塞米松对树突状细胞Toll样受体4信号通路的影响

目的：观察地塞米松对树突状细胞（den-dritic cells,DC）表面分子Toll样受体4（toll-like re-ceptor 4,TLR4）及转录因子NF-κB表达的影响,以探讨地塞米松抑制DC分化成熟和抗原提呈功能的作用机制。方法：常规培养小鼠DC,在培养体系中加入地塞米松作用后,通过流式细胞术分析DC TLR4表达情况;凝胶电泳迁移实验（EMSA）分析DC NF-κB活化情况。结果：地塞米松以剂量依赖方式抑制NF-κB表达,但对TLR4表达无抑制作用。结论：地塞米松影响DC分化成熟和抗原提呈功能的机制可能与其对NF-κB活化的抑制作用有关。     

Salvia miltiorrhiza compounds protect the liver from acute injury by regulation of p38 and NFκB signaling in Kupffer cells

Context: Salvia miltiorrhiza Bunge is a traditional Asian medicine used to treat cerebral and cardiac ischemia. However, the effects of the active compounds of S. miltiorrhiza on liver damage are unclear.

Objective: In this study, we tested the effects on acute liver injury of crude S. miltiorrhiza extracts from roots as well as neotanshinone B, dehydromiltirone, tanshinol A, tanshinone I, dihydrotanshinono I, neotanshinone A, cryptanshinono, tanshinone II A, and salvianolie acid B from purified S. miltiorrhiza extracts.

Materials and methods: Various compounds or ethanol extract of S. miltiorrhiza (50, 100, and 200?mg/kg, p.o.) were administered to rats for five consecutive days. After acute carbon tetrachloride (CCl₄)-induced liver injury by treatment of rats with a single dose of CCl₄ (0.75?mL/kg, p.o), rat liver function was tested by measuring serum biochemical parameters. Serum cytokine concentrations were assessed by enzyme-linked immunosorbent assay (ELISA). Expression of p38 and NFκB was evaluated by western blot.

Results: All S. miltiorrhiza components showed their effects on liver function from the dose from 50 to 200?mg/kg. At the dose of 200?mg/kg, they reduced serum levels of alkaline phosphatase (ALP) by 34–77%, alanine aminotransferase (ALT) by 30–57%, aspartate aminotransferase (AST) by 43–72%, creatine total bilirubin (BIL-T) by 33–81%, albumin (ALB) by 37–67%, indicating that S. miltiorrhiza extracts protected liver from CCl₄-induced damage. Moreover, S. miltiorrhiza extracts at 200?mg/kg reduced the increase in the proinflammatory cytokines tumor necrosis factor-α (TNF-α) by 25–82%, interleukin-1 (IL-1) by 42–74% and interleukin-6 (IL-6) by 67–83%, indicating an effect on alleviating liver inflammation. Furthermore, in vitro, S. miltiorrhiza extracts inhibited p38 and NFκB signaling in Kupffer cells. This effect could be a main mechanism by which S. miltiorrhiza protects against acute liver toxicity.

Discussion and conclusion: Active compounds of S. miltiorrhiza protected the liver from CCl₄-induced injury. Protection might have been due to inhibition of p38 and NFκB signaling in Kupffer cells, which subsequently reduced inflammation in the liver.     

胎盘组织核因子-κB与妊娠并发症关系研究进展

核因子-κB(NF-κB)是一种细胞内重要的转录因子,调控包括炎症、凋亡在内多种基因的表达,在胎盘中分布广泛。本文对胎盘组织中NF-κB与先兆子痫、HELLP综合征和胎膜早破等妊娠并发症的关系研究进行综述,概括了胎盘NF-κB在这些妊娠并发症发生过程中的作用以及依赖NF-κB途径治疗相关疾病的研究进展,为胎盘相关疾病的深入研究奠定基础,为妊娠并发症疾病的诊治提供新思路。     

Antibodies to receptor activator of nuclear factor-κ B ligand (RANKL)

Patent WO02095012A1 claims for the development of antibodies against receptor activator of nuclear factor-κ B ligand (RANKL) and immunologically functional fragments that neutralize RANKL. These molecules can be used to detect RANKL in biological samples, allowing the identification of pathological conditions in which RANKL alteration contributes to their pathophysiology. The use of anti-RANKL antibodies is of high importance in the treatment of bone disorders characterized by the stimulation of osteoclast function and subsequent bone loss. Postmenopausal and steroid-induced osteoporosis, myeloma bone disease, cancer bone metastases with lytic lesions and bone loss due to rheumatoid disorders (including rheumatoid arthritis) are candidates for anti-RANKL therapy, as RANKL is implicated in their pathogenesis.     

外源性骨生长因子对骨质疏松症的治疗效果

目的观察外源性骨生长因子(骨肽片)对骨质疏松症的治疗效果,为临床治疗提供可行性途径和方法。方法选择2003-02～2005-05求治的50例骨质疏松性患者,口服骨肽片0.3g,3次/d,连续服用1年,治疗前后分别测量跟骨骨密度及骨代谢生化指标自身对照。结果经随访,50例患者中,临床症状缓解显效32例,占64.0%;有效18例,占36%。总有效率100%。超声骨密度三项指标宽波段超声衰减(BUA)、声速(SOS)以及由两者演算而来的硬度指数(Stiffness)水平显著提高(P<0.01)。骨代谢生化指标血清骨碱性磷酸酶(BALP)、Ⅰ型前胶原羧基端肽(PICP)、骨钙素(BGP)水平显著增高(P<0.05或<0.01)。结论补充外源性骨生长因子可促进成骨代谢、改善骨结构、提高骨密度,是治疗骨质疏松症的有效途径和方法,并对骨质疏松性骨折的预防有积极意义。