Icariin inhibits RANKL-induced osteoclastogenesis via modulation of the NF-κB and MAPK signaling pathways

The receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL)-RANK regulatory axis is a major regulator of osteoclast differentiation and activation. Icariin, a flavonol glycoside isolated from the Epimedium herb, has been reported to prevents bone loss in ovariectomized mice and inhibits wear particle-induced osteolysis. However, the molecular mechanism through which icariin inhibits RANKL-induced osteoclastogenesis has not been fully understood. Therefore, we aimed to investigate the effects of icariin on RANKL-induced osteoclastogenesis and to elucidate the mechanism underlying this effect. Our results showed that RANKL-induced osteoclastogenesis was inhibited by icariin in bone marrow macrophages (BMMs) and RAW264.7?cells, and that this effect was due to suppression of NF-κB and mitogen-activated protein kinase (MAPK) activation. In addition, icariin inhibited F-actin ring formation and attenuated the bone resorption ability of mature osteoclasts. Collectively, our results indicate that icariin may be a promising potential candidate for the treatment of osteolytic diseases such as osteoporosis. Moreover, our findings lay the foundation for understanding and intervening in osteoclast-related diseases at the molecular level.     

淫羊藿苷缓解腹部皮瓣缺血再灌注损伤模型大鼠的作用及机制研究

摘要 目的：研究淫羊藿苷缓解腹部皮瓣缺血再灌注损伤（IRI）模型大鼠的作用及机制。方法：取30只SD级大鼠作为研究对象,将其按照随机数字表法分作假手术组、模型组以及淫羊藿苷组,每组各10只。其中模型组和淫羊藿苷组大鼠均制作大鼠腹部皮瓣IRI模型,假手术组以及模型组大鼠予以生理盐水腹腔注射,淫羊藿苷组大鼠则予以淫羊藿苷腹腔注射。对比各组大鼠皮瓣存活面积及存活率、血清炎症因子以及氧化应激指标水平、皮瓣组织中p38丝裂原活化蛋白激酶（p38 MAPK）信号通路相关蛋白表达情况。结果：模型组、淫羊藿苷组大鼠的皮瓣存活面积及存活率均低于假手术组,但淫羊藿苷组大鼠的皮瓣存活面积及存活率均高于模型组（P＜0.05）。模型组、淫羊藿苷组大鼠血清白细胞介素-10（IL-10）均低于假手术组,但淫羊藿苷组高于模型组;模型组、淫羊藿苷组大鼠血清肿瘤坏死因子-?琢（TNF-?琢）均高于假手术组,但淫羊藿苷组低于模型组（P＜0.05）。模型组、淫羊藿苷组大鼠血清超氧化物歧化酶（SOD）、谷胱甘肽（GSH）水平均低于假手术组,但淫羊藿苷组大鼠血清SOD、GSH水平均高于模型组;模型组、淫羊藿苷组大鼠血清丙二醛（MDA）水平均高于假手术组,但淫羊藿苷组大鼠血清MDA水平低于模型组（P＜0.05）。模型组、淫羊藿苷组大鼠皮瓣组织p38 MAPK、丝裂原活化蛋白激酶磷酸酶-2（MKP-2）相对表达量均高于假手术组,但淫羊藿苷组皮瓣组织p38 MAPK相对表达量低于模型组,而MKP-2相对表达量高于模型组（P＜0.05）。结论：淫羊藿苷可通过调控p38 MAPK信号通路缓解炎症反应及氧化应激,发挥减轻腹部皮瓣IRI的作用。     

Icariin stimulates angiogenesis by activating the MEK/ERK- and PI3K/Akt/eNOS-dependent signal pathways in human endothelial cells

We investigated the molecular effect and signal pathway of icariin, a major flavonoid of Epimedium koreanum Nakai, on angiogenesis. Icariin stimulated in vitro endothelial cell proliferation, migration, and tubulogenesis, which are typical phenomena of angiogenesis, as well as increased in vivo angiogenesis. Icariin activated the angiogenic signal modulators, ERK, phosphatidylinositol 3-kinase (PI3K), Akt, and endothelial nitric oxide synthase (eNOS), and increased NO production, without affecting VEGF expression, indicating that icariin may directly stimulate angiogenesis. Icariin-induced ERK activation and angiogenic events were significantly inhibited by the MEK inhibitor PD98059, without affecting Akt and eNOS phosphorylation. The PI3K inhibitor Wortmannin suppressed icariin-mediated angiogenesis and Akt and eNOS activation without affecting ERK phosphorylation. Moreover, the NOS inhibitor NMA partially reduced the angiogenic activity of icariin. These results suggest that icariin stimulated angiogenesis by activating the MEK/ERK- and PI3K/Akt/eNOS-dependent signal pathways and may be a useful drug for angiogenic therapy.     

综述:中药治疗阿尔茨海默病的作用特点

阿尔茨海默病(AD)是一种多因素相关的复杂性疾病,目前临床治疗效果不佳．仅针对单靶点或单致病途径的药物不易取得好的疗效．另一个重要原因是干预时机太晚,当诊断出痴呆时患者脑内已有大量神经元死亡．因此,应当针对多靶点、多途径治疗,同时将治疗时机提前到痴呆发生前,才有可能在AD的药物干预领域实现新的突破．本文综述了作者近十多年来在中药治疗AD方面的研究工作,包括中药新复方参乌胶囊、中药提取物何首乌二苯乙烯苷、山茱萸环烯醚萜苷、淫羊藿黄酮和淫羊藿苷对多种拟AD动物模型和细胞模型的影响及其作用机制．这些中药的特点是作用在AD复杂发病机制的多靶点和多途径,尤其是具有神经保护和神经营养/再生作用,且对线粒体和突触具有明显的保护作用,可望用于AD的早期干预或轻度认知障碍期(MCI)的治疗,从而阻止或延缓痴呆的发生与进程．     

HPLC法测定前列舒乐颗粒中淫羊藿苷的含量

目的:建立前列舒乐颗粒中淫羊藿苷的HPLC测定方法。方法:以C18(4.6 mm×15 mm,5μm)为色谱柱,以乙腈-水(30∶70)为流动相,流速1 mL/min,测定波长270 nm。结果:线性范围在0.40～4.00μg(r=0.9999),平均回收率为99.79%,RSD=2.5%(n=6)。结论:本方法简便可行,重现性好,结果可靠,可以用来测定淫羊藿苷的含量,控制前列舒乐颗粒的质量。     

Variation of Epimedins A – C and Icariin in Ten Representative Populations of Epimedium brevicornu Maxim., and Implications for Utilization

The concentration variations of main flavonoids, epimedins A–C and icariin, among ten representative populations of Epimedium brevicornu Maxim . were assessed by HPLC. The populations were collected during the flowering stage and included 419 individual samples. Remarkable variations within and among populations were detected. SXXA Population (see Fig. 1) was an outlier due to its significant low concentrations (<1.00–4.46 mg/g). But even without SXXA, significant concentration differences among populations were still observed in epimedin A (2.31–8.42 mg/g), epimedin B (6.67–55.7 mg/g), epimedin C (5.39–23.0 mg/g), icariin (8.50–39.9 mg/g), and their total (29.1–123 mg/g). All populations except SXXA showed much higher concentrations than the recommended standards (i.e. 5 mg/g for icariin and 13 mg/g for the total). A high‐concentration‐population structure, estimated both by principal component analysis (PCA) and unweighted pair group method with averaging (UPGMA) cluster analysis, based on Euclidean distances, was observed. Both methods allowed separation of the populations in four groups defined by the concentrations of four main flavonoids. The populations (SXLC and SXQS) located in north of Yellow River were clustered together and characterized by highest concentrations of epimedin B, icariin, and their total. Considering of the high concentrations of main flavonoids and abundant resources, E. brevicornu could be exploited as a good medical resource for Herba Epimedii and would offer a tremendous potential for commercial development, but SXXA population should be paid special attention, and further study is needed.     

Icaritin and its glycosides enhance osteoblastic, but suppress osteoclastic, differentiation and activity in vitro

Icariin, a principal flavonoid glycoside in Herba Epimedii, is hypothesized to possess beneficial effects on bone mass. Icariin is metabolized to icariside II and then to icaritin in vivo. In the present study, we investigated the in vitro effects of icariin, icariside II and icaritin on both osteoblasts and osteoclasts. After treatment with these compounds at concentrations 10(-5)-10(-8) mol/l, osteoblasts were examined for proliferation, alkaline phosphatase activity, osteocalcin secretion and matrix mineralization, as well as expression levels of bone-related proteins. The formation of osteoclasts was assessed by counting the number of multinucleated TRAP-positive cells. The activity of isolated rat osteoclasts was evaluated by measuring pit area, actin rings and superoxide generation. Icariside II and icaritin increased the mRNA expression of ALP, OC, COL-1 and OPG, but suppressed that of RANKL. In addition, these compounds reduced the number of multinucleated TRAP-positive cells and the osteoclastic resorption area. Also decreases were observed in superoxide generation and actin ring formation that are required for osteoclast survival and bone resorption activity. These findings suggest that icaritin, which was more potent than icariin and icariside II, enhanced the differentiation and proliferation of osteoblasts, and facilitated matrix calcification; meanwhile it inhibited osteoclastic differentiation in both osteoblast-preosteoclast coculture and osteoclast progenitor cell culture, and reduced the motility and bone resorption activity of isolated osteoclasts.     

Interaction of the Bioactive Flavonol,Icariin, with the Essential Human Solute Carrier Transporters

Solute carrier transporters (SLCs), in particular the organic anion transporting polypeptides (OATPs) and organic anion/cation transporters (OATs/OCTs), are responsible for the cellular entry of many clinically important drugs in body. They largely influence drug safety and efficacy. Icariin is a flavonol widely present in many herbal preparations, which is used to improve sexual function and prevent osteogenesis. However, precautions are necessary in therapies containing icariin due to its involvement in drug–drug/herb interactions, possibly mediated through competing drug uptake via membrane‐transporter proteins. This study is the first to comprehensively evaluate the interactions between icariin and a range of essential SLCs. Our data demonstrated that icariin can significantly inhibit OATP1B3‐ and OATP2B1‐mediated cellular uptake of specific substrates (IC₅₀ of 3.0 ± 1.3 and 6.4 ± 1.9 μM, respectively). Our study revealed that icariin can potentially compete with coadministrated drugs for particular SLCs, which may impact the therapeutic outcome of regimens.