中华口腔医学杂志
中華口腔醫學雜誌
중화구강의학잡지
Chinese Journal of Stomatology
2014年
10期
619-624
,共6页
王燕萍%宋卫健%景双林%于金华
王燕萍%宋衛健%景雙林%于金華
왕연평%송위건%경쌍림%우금화
雌激素类%间充质干细胞%细胞增殖%细胞分化
雌激素類%間充質榦細胞%細胞增殖%細胞分化
자격소류%간충질간세포%세포증식%세포분화
Estrogens%Mesenchymal stem cells%Cell proliferation%Cell differentiation
目的 探讨雌激素缺乏对大鼠颌骨骨髓间充质干细胞(mandibular bone marrow stem cells,mBMSC)增殖及成骨向分化能力的影响,以期为组织工程颌骨再生及种子细胞选择提供实验室依据.方法 将10只8周龄成年雌性SD大鼠用抽签法随机分为假手术组及卵巢去势组(去势组),每组5只.全麻下将去势组大鼠双侧卵巢摘除,假手术组行相同切口,保留双侧卵巢.1个月后处死大鼠,分离下颌骨,全骨髓培养法分离培养两组mBMSC.采用甲基噻唑基四唑(methyl thiazolyl tetrazolium,MTT)法、碱性磷酸酶(alkaline phosphotase,ALP)活性检测、茜素红染色、实时反转录PCR和蛋白质印迹法检测雌激素缺乏对mBMSC增殖及成骨向分化能力的影响.结果 MTT生长曲线显示去势组mBMSC的增殖能力显著低于假手术组.两组细胞培养3d后去势组ALP活性[(0.710±0.011) Sigma单位/蛋白]显著低于假手术组[(1.512±0.021) Sigma单位/蛋白],差异有统计学意义(P＜0.01).14d时茜素红染色结果显示去势组mBMSC形成的矿化结节较假手术组少,钙离子浓度定量检测结果显示去势组mBMSC钙结节中的钙离子含量[(0.433±0.045) μg/g]显著低于假手术组[(1.453±0.131) μg/g](P＜0.01).实时反转录PCR及蛋白质印迹法检测结果显示,去势组mBMSC在培养7 d后,各项骨向分化相关指标的表达均显著低于假手术组(P＜0.05).结论 雌激素缺乏显著降低大鼠mBMSC体外增殖及成骨向分化能力.
目的 探討雌激素缺乏對大鼠頜骨骨髓間充質榦細胞(mandibular bone marrow stem cells,mBMSC)增殖及成骨嚮分化能力的影響,以期為組織工程頜骨再生及種子細胞選擇提供實驗室依據.方法 將10隻8週齡成年雌性SD大鼠用抽籤法隨機分為假手術組及卵巢去勢組(去勢組),每組5隻.全痳下將去勢組大鼠雙側卵巢摘除,假手術組行相同切口,保留雙側卵巢.1箇月後處死大鼠,分離下頜骨,全骨髓培養法分離培養兩組mBMSC.採用甲基噻唑基四唑(methyl thiazolyl tetrazolium,MTT)法、堿性燐痠酶(alkaline phosphotase,ALP)活性檢測、茜素紅染色、實時反轉錄PCR和蛋白質印跡法檢測雌激素缺乏對mBMSC增殖及成骨嚮分化能力的影響.結果 MTT生長麯線顯示去勢組mBMSC的增殖能力顯著低于假手術組.兩組細胞培養3d後去勢組ALP活性[(0.710±0.011) Sigma單位/蛋白]顯著低于假手術組[(1.512±0.021) Sigma單位/蛋白],差異有統計學意義(P＜0.01).14d時茜素紅染色結果顯示去勢組mBMSC形成的礦化結節較假手術組少,鈣離子濃度定量檢測結果顯示去勢組mBMSC鈣結節中的鈣離子含量[(0.433±0.045) μg/g]顯著低于假手術組[(1.453±0.131) μg/g](P＜0.01).實時反轉錄PCR及蛋白質印跡法檢測結果顯示,去勢組mBMSC在培養7 d後,各項骨嚮分化相關指標的錶達均顯著低于假手術組(P＜0.05).結論 雌激素缺乏顯著降低大鼠mBMSC體外增殖及成骨嚮分化能力.
목적 탐토자격소결핍대대서합골골수간충질간세포(mandibular bone marrow stem cells,mBMSC)증식급성골향분화능력적영향,이기위조직공정합골재생급충자세포선택제공실험실의거.방법 장10지8주령성년자성SD대서용추첨법수궤분위가수술조급란소거세조(거세조),매조5지.전마하장거세조대서쌍측란소적제,가수술조행상동절구,보류쌍측란소.1개월후처사대서,분리하합골,전골수배양법분리배양량조mBMSC.채용갑기새서기사서(methyl thiazolyl tetrazolium,MTT)법、감성린산매(alkaline phosphotase,ALP)활성검측、천소홍염색、실시반전록PCR화단백질인적법검측자격소결핍대mBMSC증식급성골향분화능력적영향.결과 MTT생장곡선현시거세조mBMSC적증식능력현저저우가수술조.량조세포배양3d후거세조ALP활성[(0.710±0.011) Sigma단위/단백]현저저우가수술조[(1.512±0.021) Sigma단위/단백],차이유통계학의의(P＜0.01).14d시천소홍염색결과현시거세조mBMSC형성적광화결절교가수술조소,개리자농도정량검측결과현시거세조mBMSC개결절중적개리자함량[(0.433±0.045) μg/g]현저저우가수술조[(1.453±0.131) μg/g](P＜0.01).실시반전록PCR급단백질인적법검측결과현시,거세조mBMSC재배양7 d후,각항골향분화상관지표적표체균현저저우가수술조(P＜0.05).결론 자격소결핍현저강저대서mBMSC체외증식급성골향분화능력.
Objective To determine the effects of estrogen deficiency on the proliferation and osteogenic differentiation of mandibular bone marrow stromal cells(mBMSC).Methods Ten 8-week-old female SD rats were randomly divided into two groups,ovariectomized group(OVX,n=5) and sham-operation group(n=5).All rats were anesthetized and both ovaries of OVX-rats were gently removed.Sham-operation rats were treated with the same incisions to expose the ovaries but without any hurt to them.One month after the operation,the mandibular bones were gently separated and mBMSC were isolated.Methyl thiazolyl tetrazolium(MTT) assay,alkaline phosphotase(ALP) activity,alizarin red staining,real-time reverse transcription(RT)-PCR and Western blotting were respectively used to examine the proliferative activity and osteogenic potential of mBMSC.Results MTT results showed that OVX-mBMSC exhibited the decreased proliferative activity as compared with Sham-mBMSC.ALP activity of OVX-mBMSC[(0.710±0.011) Sigma unit/protein] was lower than that of Sham-mBMSC[(1.512±0.021) Sigma unit/protein](P＜0.01).Alizarin red staining showed that OVX-mBMSC formed less calcified nodules than Sham-mBMSC.Ca2+ concentration analysis showed Ca2+ of OVX-mBMSC[(0.433±0.045) μg/g] was less than Sham-mBMSC[(1.453±0.131) μg/g](P＜0.01).Real-time RT-PCR and Western blotting results showed that the expression of osteogenic markers (Alp,Runx2/RUNX2,Osx/OSX,Ocn/OCN) in OVX-mBMSC was significantly inhibited as compared with Sham-mBMSC(P＜0.05).Conclusions Estrogen deficiency significantly inhibits the proliferation and osteogenic capacity of mBMSC.
