槟榔碱可抑制体外 RANKL 诱导的破骨细胞分化，并减轻体内 LPS 诱导的骨丢失。
Arecoline suppresses RANKL-induced osteoclast differentiation in vitro and attenuates LPS-induced bone loss in vivo.
机构信息
Division of Allergy-Immunology-Rheumatology, Taipei Veterans General Hospital, Taipei, Taiwan; Biobank Management Center of the Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan; Department of Biomedical Engineering, National Defense Medical Center, Taipei, Taiwan.
Graduate Institutes of Life Sciences, National Defense Medical Center, Taipei, Taiwan.
出版信息
Phytomedicine. 2020 Apr;69:153195. doi: 10.1016/j.phymed.2020.153195. Epub 2020 Feb 22.
BACKGROUND
Areca nut has anti-inflammatory, antiparasitic, antihypertensive, and antidepressant properties. The pathological hallmarks of inflammatory joint diseases are an increased number of osteoclasts and impaired differentiation of osteoblasts, which may disrupt the bone remodeling balance and eventually lead to bone loss.
PURPOSE
The present study assessed the effects of arecoline, the main alkaloid found in areca nut, on osteoclast and osteoblast differentiation and function.
METHOD
M-CSF/RANKL-stimulated murine bone marrow-derived macrophages (BMMs) were incubated with several concentrations of arecoline, and TRAP staining and pit formation were assessed to monitor osteoclast formation. Quantitative real-time RT-PCR and western blot analyses were used to analyze the expression of osteoclast-associated genes and signaling pathways. The effects of arecoline on bone were investigated in an in vivo mouse model of lipopolysaccharide (LPS)-induced trabecular bone loss after oral administration of arecoline. Alizarin red S staining and assays to measure ALP activity and the transcription level of osteoblast-related genes were used to evaluate the effects of arecoline on osteoblast differentiation and bone mineralization.
RESULTS
In a dose-dependent manner, arecoline at concentrations of 50-100 μM reduced both the development of TRAP-positive multinucleated osteoclasts and the formation of resorption pits in M-CSF/RANKL-stimulated BMMs. In M-CSF/RANKL-stimulated BMMs, arecoline also suppressed the expression and translocation of c-Fos and NFATcl, and osteoclast differentiated-related genes via interference with the AKT, MAPK, and NF-kB activation pathways. Femur bone loss and microcomputed tomography parameters were recovered by oral administration of arecoline in the mouse LPS-induced bone loss model. Lastly, arecoline increased ALP activity, bone mineralization, and the expression of osteoblast differentiation-related genes, such as ALP and Runx2, in MC3T3-E1 cells.
CONCLUSION
Our data suggest that arecoline may attenuate or prevent bone loss by suppressing osteoclastogenesis and promoting osteoblastogenesis. These findings provide evidence supporting arecoline's use as a potential therapeutic agent in bone-loss disorders and diseases.
背景
槟榔具有抗炎、抗寄生虫、降血压和抗抑郁作用。炎症性关节疾病的病理特征是破骨细胞数量增加和成骨细胞分化受损，这可能破坏骨重塑平衡，最终导致骨质流失。
目的
本研究评估了槟榔中主要生物碱槟榔碱对破骨细胞和成骨细胞分化和功能的影响。
方法
用几种浓度的槟榔碱孵育 M-CSF/RANKL 刺激的鼠骨髓来源巨噬细胞（BMM），通过 TRAP 染色和陷窝形成来监测破骨细胞形成。采用定量实时 RT-PCR 和 Western blot 分析检测破骨细胞相关基因和信号通路的表达。通过口服槟榔碱在脂多糖（LPS）诱导的小梁骨丢失的小鼠模型中研究槟榔碱对骨的影响。用茜素红 S 染色和测定碱性磷酸酶（ALP）活性和骨形成相关基因转录水平的实验来评估槟榔碱对成骨细胞分化和骨矿化的影响。
结果
槟榔碱呈剂量依赖性地降低了 50-100 μM 浓度的 M-CSF/RANKL 刺激的 BMM 中 TRAP 阳性多核破骨细胞的发育和吸收陷窝的形成。在 M-CSF/RANKL 刺激的 BMM 中，槟榔碱还通过干扰 AKT、MAPK 和 NF-kB 激活通路，抑制 c-Fos 和 NFATcl 的表达和易位以及破骨细胞分化相关基因的表达。在 LPS 诱导的小鼠骨丢失模型中，口服槟榔碱可恢复股骨骨丢失和微计算机断层扫描参数。最后，槟榔碱增加了 MC3T3-E1 细胞中 ALP 活性、骨矿化和骨分化相关基因（如 ALP 和 Runx2）的表达。
结论
我们的数据表明，槟榔碱通过抑制破骨细胞生成和促进成骨细胞生成来减轻或预防骨质流失。这些发现为槟榔碱作为骨丢失疾病潜在治疗剂的应用提供了证据。
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