摘要
背景:绿原酸是干李子中含量最丰富的多酚类物质,作为一种食源性药物有望用于骨质疏松治疗之中。目的:探究绿原酸对成骨前体细胞成骨分化的影响及其机制,并在骨质疏松小鼠模型中进行相关验证。方法:(1)体外实验:以含不同质量浓度(0,0.1,1,10,100 mg/L)绿原酸的培养基培养MC3T3-E1细胞,采用CCK-8法检测细胞增殖。采用含0,10,20 mg/L绿原酸的成骨诱导培养基培养MC3T3-E1细胞,进行碱性磷酸酶染色、茜素红染色、成骨基因表达与BMP-2/RUNX2/SMAD4信号通路蛋白检测。(2)体内实验:将32只雌性C57BL/6小鼠随机分成4组,假手术组、去卵巢组及低、高剂量绿原酸组,每组8只,低、高剂量绿原酸组去卵巢后腹腔注射25,50 mg/(kg·d)的绿原酸,连续给药8周。给药8周后,进行骨组织Micro-CT扫描与组织形态学观察。结果与结论:(1)体外实验:当绿原酸质量浓度≤10 mg/L时,对MC3T3-E1细胞的增殖无明显影响。碱性磷酸酶染色与茜素红染色显示,随着绿原酸质量浓度的增加,MC3T3-E1细胞的碱性磷酸酶活性、钙沉积和钙结节生成能力提高,成骨相关基因碱性磷酸酶、骨桥蛋白、骨钙素、骨形态发生蛋白2、RUNX2、SMAD4表达升高,骨形态发生蛋白2、RUNX2、SMAD4蛋白表达升高。(2)体内实验:Micro-CT扫描显示,去卵巢组小鼠骨密度、骨体积分数、骨小梁厚度及数量均低于假手术组(P <0.05),骨小梁分离度高于假手术组(P <0.05);经过不同质量浓度绿原酸干预后,去卵巢小鼠的上述指标均有所改善,其中高剂量绿原酸组改善效果有显著性意义(P <0.05)。苏木精-伊红染色显示,去卵巢组小鼠骨量少于假手术组,经过绿原酸干预后骨量增加。(3)绿原酸可能通过BMP-2/RUNX2/SMAD4信号通路促进成骨分化,预防去卵巢小鼠的骨丢失。
BACKGROUND:Chlorogenic acid is the most abundant polyphenol in dried plums,which,as a food-derived drug,is expected to be used in the treatment of osteoporosis.OBJECTIVE:To investigate the effect of chlorogenic acid on osteogenic differentiation of osteogenic precursor cells and its mechanism and to verify its effect on bone loss in a mouse model of osteopo rosis.METHODS:(1)In vtro test:MC3 T3-E1 cells we re cultured in vitro with different concentrations of chlorogenic acid(0,0.1,1,10,100 mg/L) and cell proliferation was detected using cell counting kit-8.MC3 T3-E1 cells were cultured in osteogenic induction media containing 0,10,20 mg/L chlorogenic acid,followed by alkaline phosphatase staining,alizarin red staining,and detection of osteogenic genes and BMP-2/RUNX2/SMAD4 signaling pathway-related proteins.(2) In vivo test:Thirty-two female C57 BL/6 mice were randomly divided into four groups,sham ope ration group,ovariectomized group,and low-and high-dose chlorogenic acid groups,with 8 mice in each group.An ovariectomized mouse model was established,and chlorogenic acid was then administe red at high(50 mg/kg/d) and low(25 mg/kg/d) concentrations in the latter two groups for 8 continuous weeks,respectively.Mic ro-CT scan and histomorphology analysis were performed on the femur and tibia of the mice 8 weeks later.RESULTS AND CONCLUSION:(1) In vitro test:Chlorogenic acid at a concentration of ≤ 10 mg/L had no obvious effects on the proliferation of MC3 T3-E1 cells.With the increased concentration of chlorogenic acid,the alkaline phosphatase activity,calcium deposition and calcium nodule formation ability of MC3 T3-E1 cells increased;the mRNA expression levels of osteogenesis-related genes,alkaline phosphatase,osteopontin,osteocalcin,bone morphogenetic protein 2,RUNX2 and SMAD4 increased;and the protein expression levels of bone morphogenetic protein 2,RUNX2,and SMAD4 increased.(2) In vivo test:Micro-CT scan results showed that compared with the sham operation group,the bone mineral density,bone volume fraction,
作者
朱灿
何家恒
陈迟迟
刘波
罗宗平
孙杰
施勤
Zhu Can;He Jiaheng;Chen Chichi;Liu Bo;Luo Zongping;Sun Jie;Shi Qin(Department of Orthopedics,the First Affiliated Hospital of Soochow University,Institute of Orthopedics of Soochow University,Soochow University Medical School,Suzhou 215123,Jiangsu Province,China)
出处
《中国组织工程研究》
CAS
北大核心
2023年第14期2170-2175,共6页
Chinese Journal of Tissue Engineering Research
基金
国家自然科学基金面上项目(81972059),项目负责人:施勤。
关键词
绿原酸
成骨基因
成骨分化
细胞增殖
成骨诱导
成骨矿化
信号通路
骨丢失
chlorogenic acid
osteogenic gene
osteogenic differentiation
cell proliferation
osteogenic induction
osteogenic mineralization
signaling pathway
bone loss
