香豆雌酚对膝骨关节炎患者软骨细胞OPG/RANKL及MMP-1、MMP-3表达的影响

目的:研究香豆雌酚(coumestrol)对膝骨关节炎患者软骨细胞OPG/RANKL及MMP-1、MMP-3表达的影响,初步探讨其对骨关节炎的治疗机制。方法:分离、培养因膝骨关节炎行膝关节置换患者的膝关节软骨细胞,运用Ⅱ型胶原免疫荧光染色进行软骨细胞鉴定。用不同浓度(10-9、10-8、10-7mol·L-1)香豆雌酚干预第3代细胞,并设0浓度为空白对照组,分别于培养后3、7 d用real-time PCR法测定细胞骨保护素(osteoprotegerin,OPG)、核因子κB受体活化因子配基(receptor activator of nuclear factor-κB ligand,RANKL)、基质金属蛋白酶-1(matrix metalloproteinase-1,MMP-1)和基质金属蛋白酶-3(matrix metalloproteinase-3,MMP-3)mRNA的表达。结果:干预3 d,10-8mol·L-1组OPG及RANKL mRNA表达均增加,其OPG/RANKL相对表达是空白对照组的2.59倍(P<0.05);干预7 d,10-9和10-8mol·L-1组OPG mRNA表达均增加,10-7mol·L-1组RANKL mRNA表达增加,10-8mol·L-1组OPG/RANKL相对表达是空白对照组的3.85倍(P<0.05)。干预3 d,10-8mol·L-1组MMP-1 mRNA表达减少(P<0.05);干预7 d,10-8mol·L-1组MMP-1和MMP-3 mRNA表达均减少(P<0.05)。结论:香豆雌酚能增加膝骨关节炎患者关节软骨细胞OPG表达及OPG/RANKL值,减少其MMP-1、MMP-3表达,具有一定的软骨保护作用。     

Role of receptor activator of nuclear factor-kappa B ligand (RANKL), osteoprotegerin and macrophage protein 1-alpha (MIP-1a) in monoclonal gammopathy of undetermined significance (MGUS)

The aim of this study was to evaluate the role of markers of bone remodelling, and osteoclast activation/function in patients with monoclonal gammopathy of undetermined significance (MGUS). We have measured serum levels of soluble RANKL (sRANKL), osteoprotegerin (OPG), macrophage inflammatory protein-1alpha (MIP-1alpha), markers of bone resorption [N-telopeptide of collagen type-I (NTX), and tartrate-resistant acid phosphatase isoform-5b (TRACP-5b)] and bone formation [bone-alkaline phosphatase (bALP)] in 40 MGUS patients. These parameters were compared with those of 42 newly diagnosed myeloma patients, and 45 healthy, gender- and age-matched controls. MGUS patients had elevated levels of NTX, sRANKL, and sRANKL/OPG ratio compared with controls (P < 0.0001). Furthermore, TRACP-5b, MIP-1alpha and NTX were decreased in patients with MGUS compared with myeloma patients (P < 0.001), while OPG and bALP were increased (P < 0.001). Serum levels of MIP-1alpha, as well as TRACP-5b, and sRANKL/OPG ratio were reduced, while bALP was increased in MGUS patients, even when compared with myeloma patients who had stage I/II disease. These results demonstrate that increased osteoclastogenesis leading to increased bone resorption is present in MGUS but seems to be compensated for by normal bone formation, which is absent in MM. Furthermore MIP-1alpha, bALP, and sRANKL/OPG may be useful tools for distinguishing between cases of MGUS and early myeloma.     

Osteoporosis in children and young adults

Osteoporosis is a major public health problem with serious long-term complications. In children, the definition of osteoporosis is not only based on densitometric criteria but also takes into account vertebral and long bone fragility fractures. Several factors, such as long-term high-dose steroids, chronic inflammation, malnutrition, immobility, lack of sex steroids, and medication can reduce bone density and increase the risk for fragility fractures when left untreated. Also, genetic conditions can predispose to primary bone fragility disorders, with osteogenesis imperfecta being the most common. Furthermore, since the growing skeleton is at an increased rate of bone remodeling, the ability to heal long bone fractures and reshape vertebral fractures differentiates children from adults. The scope of this chapter is to review the risk factors of osteoporosis and fragility fractures and describe the commonest causes of primary and secondary osteoporosis and their management in children and young adults.     

Investigation of osteoclastogenic signalling of the RANKL substitute LIGHT

LIGHT (TNFSF14) is a member of the TNF superfamily and is known to substitute for RANKL to induce osteoclast differentiation. LIGHT binds HVEM and LTβR, but it is not known whether these receptors play a role in osteoclast formation or whether LIGHT acts via RANKL signalling pathways. We found that both RANKL and LIGHT strongly induced phosphorylation of Akt and NFκB but not JNK in mouse osteoclast precursor cells. The addition of an Akt inhibitor showed decreased osteoclast differentiation and resorption mediated by both RANKL and LIGHT. RT-PCR and FACS analysis showed that CD14⁺ human osteoclast precursors expressed HVEM and LTβR; expression levels of HVEM increased in the course of osteoclastogenesis and a decrease in LIGHT expression was associated with an increase in HVEM suggesting that there is a feedback loop related to this receptor. Our findings show that LIGHT is not inhibited by the soluble RANKL receptor OPG and that LIGHT is a potent osteoclastogenesis factor that activates the Akt, NFκB and JNK pathways.     

Mutations in K-Ras linked to levels of osteoprotegerin and sensitivity to TRAIL-induced cell death in pancreatic ductal adenocarcinoma cells

Osteoprotegerin (OPG) is a soluble receptor expressed in the serum of patients with diabetes, arthritis and pancreatic cancer. While OPG has been considered a tumor survival factor for bone metastasizing breast and prostate cancers, the role of OPG in pancreatic cancer, which itself rarely metastasizes to bone, is not known. Pancreatic ductal adenocarcinoma (PDAC) cell lines were found to secrete OPG and the level of OPG production correlated with sensitivity to TRAIL-induced apoptosis. Silencing OPG sensitized cells to TRAIL-induced apoptosis. Interestingly, a positive correlation was noted between OPG production level and K-Ras mutation status. Earlier studies implicated K-Ras in conferring resistance to TRAIL-induced apoptosis in pancreatic cells and this study demonstrates that K-Ras mediated TRAIL resistance in pancreatic cancer cells occurs due to increased OPG production. Silencing K-Ras in pancreatic cancer cells decreased OPG levels and increased sensitivity to TRAIL-induced apoptosis. These observations indicate that OPG can play a role in both cell survival and in PDAC cell sensitivity to TRAIL-induced apoptosis, which may contribute to metastasis. Targeted inhibition of OPG binding to TRAIL may represent a therapeutic approach in the treatment of pancreatic cancer.     

Gingival Crevicular Fluid Levels of Sclerostin,Osteoprotegerin, and Receptor Activator of Nuclear Factor‐κB Ligand in Periodontitis

Background: To investigate changes in the levels and relative ratios of sclerostin, osteoprotegerin (OPG), and receptor activator of nuclear factor‐κB ligand (RANKL) in the gingival crevicular fluid (GCF) of patients with periodontitis after non‐surgical periodontal treatment. Methods: Fifty‐four individuals (27 healthy controls and 27 patients with chronic periodontitis [CP]) were enrolled in the study. Periodontitis patients received non‐surgical periodontal therapy. GCF sampling and clinical periodontal parameters were assessed before and 6 weeks after therapy. Sclerostin, OPG, and RANKL levels were measured by enzyme‐linked immunosorbent assay, and their relative ratios were calculated. Results: Total amounts and concentrations of sclerostin were significantly higher in patients with CP than in healthy individuals (P <0.025) and decreased after treatment (P <0.05). The RANKL/OPG ratio was significantly lower in healthy individuals than in patients with periodontitis before and after treatment (P <0.025), but no significant difference was observed in patients with periodontitis after treatment (P >0.05). The sclerostin/OPG and sclerostin/RANKL ratios were significantly lower in healthy individuals than in patients with periodontitis before and after treatment (P <0.025) and decreased in patients with periodontitis after treatment (P <0.05). Conclusions: The GCF sclerostin level may be more reliable than the RANKL/OPG ratio as a diagnostic and prognostic marker of periodontal disease and treatment outcome. Regulation of sclerostin levels may aid the development of new therapeutic strategies for the treatment of periodontal disease.