基于MAPK和β-catenin/BMP信号通路探讨天然药物对感染性骨疾病骨生长重建的调控机制研究

Our previous studies have found that LPS and IL-1 induced the inhibition of osteoblast differentiation and the apoptosis in MC3T3-E1 osteoblasts; at the same time, LPS also promoted preosteoclast differentiation into mature osteoclasts, and enhance bone resorption of the differentiated osteoclast. We also found that quercetin, a flavonoid, significantly triggered the apoptosis and inhibited osteoclastic formation and bone resorption in LPS-induced osteoclasts through the activation of JNK pathway; meanwhile quercetin reversed the inhibition of osteogenesis gene expression induced by LPS/IL-1 through inhibition of JNK and enhancement of Wnt/beta-catenin signaling pathway in osteoblasts. But with the increase of dose, the risk of inhibiting osteoblast proliferation aroused. Icariin, a herbal drug, also reversed the apoptosis and the inhibition of osteogenesis related genes of osteoblasts induced by LPS in our studies. .In future project when exposed to LPS/IL-1 for an indicated periods, either osteoclasts or osteoblasts were treated with natural plant medicines including icariin, astragaloside IV, oleanic acid or diosgenin, and cell functions were observed. The expression of osteoclastogenesis or osteogenesis related genes, the phosphorylation and signaling proteins of Wnt/β-catenin、Smad/BMPs、MAPK and NF-κB were measured using real-time QPCR and western blot analysis. Co-culture with osteoclasts and osteoblasts, cells were treated with LPS/IL-1 and natural plant medicines. Effects of natural plant medicines on cells interactions between osteoclasts and osteoblasts were assessed. Administration of natural medicines treated into rats subjected to osteomyelitis; we evaluated the formation of osteoclasts or osteoblasts of staining skull, the inflammatory factors and signaling proteins in rats. Our project will explore potential natural medicine system to improve inflammatory bone regeneration and bone remodeling, and provide new way for the treatment of inflammatory bone disease.

前期研究发现LPS和IL-1抑制成骨细胞分化和促成骨细胞凋亡；同时LPS促进破骨前体细胞分化，并增强已分化的破骨细胞骨吸收。发现槲皮素通过激活JNK通路抑制LPS诱导源性破骨细胞的活性并致破骨细胞凋亡；槲皮素又通过抑制JNK、激活Wnt/β-catenin通路逆转LPS/IL-1对成骨细胞相关基因表达的抑制。随着剂量增加，槲皮素有抑制成骨细胞增殖的风险。还发现淫羊藿甙逆转LPS促成骨细胞凋亡及对成骨相关基因的抑制。项目以天然植物性药物处理LPS/IL-1介导破骨/成骨细胞，观察破骨/成骨细胞的活性、相关因子表达及Wnt/β-catenin、Smad/BMPs、MAPK及NF-κB活性水平。对LPS/IL-1 介导细胞共培养和大鼠骨髓炎模型，采用天然植物性药物处理，观察破骨/成骨细胞形成、炎症因子及相关信号蛋白表达。探讨改善炎症性骨生长重建的天然药物系统，必将为感染性骨疾病的治疗提供新思路。

骨髓炎易造成炎症性骨溶解、骨不愈合。前期发现槲皮素通过激活JNK通路抑制LPS诱导源性破骨细胞的活性并致破骨细胞凋亡；槲皮素又通过抑制JNK、激活Wnt/β-catenin通路逆转LPS/IL-1对成骨细胞相关基因表达的抑制。随着剂量增加，槲皮素有抑制成骨细胞增殖的风险。项目在LPS/ IL-1介导破骨/成骨细胞分化过程中，并采用槲皮素、黄芪甲甙等治疗，探讨LPS/IL-1导致骨形成失平衡确切机制及天然药物改善炎症介质引起骨丢失的作用和它们对Wnt/β-catenin、Smad/BMPs、MAPK及NF-κB的活性水平影响。主要的研究发现：①黄芪甲甙 (Astragaloside IV) 在体外具有恢复LPS抑制下成骨细胞分化和成骨活性的作用机制；同时黄芪甲甙具有抑制LPS对破骨细胞分化和骨吸收的作用机制。②槲皮素(Quercetin)、黄芪甲甙在体外具有抑制IL-1 α促破骨细胞分化和骨吸收的作用。③淫羊藿甙(Icariin)治疗具有逆转LPS促成骨细胞凋亡及对成骨相关基因的抑制的作用机制。④槲皮素和黄芪甲甙治疗LPS炎症模型小鼠：发现体内小剂量天然药物不能减轻炎症性骨溶解。相关高剂量天然药物治疗LPS/IL-1炎症小鼠的研究继续开展中。研究为槲皮素和黄芪甲甙治疗炎症性骨疾病能够改善骨重建和代谢提供依据。