背景:糖皮质激素引起的髓内脂肪蓄积是由机体脂肪代谢异常在髓内形成的脂肪沉淀,还是由于激素直接作用于骨髓间充质细胞使细胞发生了分化,以及具体生成的过程目前还没有定论.目的:观察不同浓度地塞米松对骨髓间充质细胞成脂分化的影响.设计、时间及地点:观察性实验,于2007-01/2009-01在中国医科大学生物化学教研室完成.材料:3或4周龄SD大鼠80只,体质量(110±10)g,雌雄不拘.方法:分离培养大鼠骨髓间充质细胞,并进行传代,取第3代细胞作为实验标本.主要观察指标:①倒置显微镜下观察骨髓间充质细胞在地塞米松10-7 mol/L浓度和正常培养状态下3,7,14,21 d时的形态变化.②细胞碱性磷酸酶、Sundan Ⅲ染色后骨髓间充质细胞的形态变化.③酚试剂法测定碱性磷酸酶活性.④四甲基偶氮唑盐法检测地塞米松对细胞增殖的影响.结果:①原代细胞接种24 h后,培养瓶底即出现少许贴壁细胞,多呈纺锤形或梭形;随着培养时间延长,贴壁细胞明显增多,呈放射状排列.传代后细胞分布均匀,呈典型的成纤维细胞样.地塞米松刺激后,细胞由梭形变为多角形或不规则形,后期,随着浓度增大和作用时间延长,细胞团聚现象消失,很多贴壁的细胞悬浮、死亡.②在正常培养细胞中,没有或很少见到橘红色的脂肪颗粒着色,在含有地塞米松培养的细胞中,随着激素浓度增高和时间持续,细胞内Sundan Ⅲ着色的脂肪颗粒明显增加.⑨在21 d时,正常培养的细胞碱性磷酸酶活性值分别是10-8,10-7,10-6 mol/L地塞米松组的1.57倍,4.49倍和5.0倍.在7,14,21 d时,正常培养的细胞碱性磷酸酶分别是相应107 mol/L地塞米松组的2.93倍,3.80倍和4.39倍,差异具有显著性意义(P＜0.05).④高浓度地塞米松对细胞增殖活力的抑制效果明显.10-7 mol/L和10-6 mol/L与其他各组相比,差异具有显著性意义(P＜0.05).结论:大剂量地塞米松促进骨髓间充质细胞的脂肪生成,抑制成骨分化,并且随着浓度增高和作用时间延长,作用加强.
揹景:糖皮質激素引起的髓內脂肪蓄積是由機體脂肪代謝異常在髓內形成的脂肪沉澱,還是由于激素直接作用于骨髓間充質細胞使細胞髮生瞭分化,以及具體生成的過程目前還沒有定論.目的:觀察不同濃度地塞米鬆對骨髓間充質細胞成脂分化的影響.設計、時間及地點:觀察性實驗,于2007-01/2009-01在中國醫科大學生物化學教研室完成.材料:3或4週齡SD大鼠80隻,體質量(110±10)g,雌雄不拘.方法:分離培養大鼠骨髓間充質細胞,併進行傳代,取第3代細胞作為實驗標本.主要觀察指標:①倒置顯微鏡下觀察骨髓間充質細胞在地塞米鬆10-7 mol/L濃度和正常培養狀態下3,7,14,21 d時的形態變化.②細胞堿性燐痠酶、Sundan Ⅲ染色後骨髓間充質細胞的形態變化.③酚試劑法測定堿性燐痠酶活性.④四甲基偶氮唑鹽法檢測地塞米鬆對細胞增殖的影響.結果:①原代細胞接種24 h後,培養瓶底即齣現少許貼壁細胞,多呈紡錘形或梭形;隨著培養時間延長,貼壁細胞明顯增多,呈放射狀排列.傳代後細胞分佈均勻,呈典型的成纖維細胞樣.地塞米鬆刺激後,細胞由梭形變為多角形或不規則形,後期,隨著濃度增大和作用時間延長,細胞糰聚現象消失,很多貼壁的細胞懸浮、死亡.②在正常培養細胞中,沒有或很少見到橘紅色的脂肪顆粒著色,在含有地塞米鬆培養的細胞中,隨著激素濃度增高和時間持續,細胞內Sundan Ⅲ著色的脂肪顆粒明顯增加.⑨在21 d時,正常培養的細胞堿性燐痠酶活性值分彆是10-8,10-7,10-6 mol/L地塞米鬆組的1.57倍,4.49倍和5.0倍.在7,14,21 d時,正常培養的細胞堿性燐痠酶分彆是相應107 mol/L地塞米鬆組的2.93倍,3.80倍和4.39倍,差異具有顯著性意義(P＜0.05).④高濃度地塞米鬆對細胞增殖活力的抑製效果明顯.10-7 mol/L和10-6 mol/L與其他各組相比,差異具有顯著性意義(P＜0.05).結論:大劑量地塞米鬆促進骨髓間充質細胞的脂肪生成,抑製成骨分化,併且隨著濃度增高和作用時間延長,作用加彊.
배경:당피질격소인기적수내지방축적시유궤체지방대사이상재수내형성적지방침정,환시유우격소직접작용우골수간충질세포사세포발생료분화,이급구체생성적과정목전환몰유정론.목적:관찰불동농도지새미송대골수간충질세포성지분화적영향.설계、시간급지점:관찰성실험,우2007-01/2009-01재중국의과대학생물화학교연실완성.재료:3혹4주령SD대서80지,체질량(110±10)g,자웅불구.방법:분리배양대서골수간충질세포,병진행전대,취제3대세포작위실험표본.주요관찰지표:①도치현미경하관찰골수간충질세포재지새미송10-7 mol/L농도화정상배양상태하3,7,14,21 d시적형태변화.②세포감성린산매、Sundan Ⅲ염색후골수간충질세포적형태변화.③분시제법측정감성린산매활성.④사갑기우담서염법검측지새미송대세포증식적영향.결과:①원대세포접충24 h후,배양병저즉출현소허첩벽세포,다정방추형혹사형;수착배양시간연장,첩벽세포명현증다,정방사상배렬.전대후세포분포균균,정전형적성섬유세포양.지새미송자격후,세포유사형변위다각형혹불규칙형,후기,수착농도증대화작용시간연장,세포단취현상소실,흔다첩벽적세포현부、사망.②재정상배양세포중,몰유혹흔소견도귤홍색적지방과립착색,재함유지새미송배양적세포중,수착격소농도증고화시간지속,세포내Sundan Ⅲ착색적지방과립명현증가.⑨재21 d시,정상배양적세포감성린산매활성치분별시10-8,10-7,10-6 mol/L지새미송조적1.57배,4.49배화5.0배.재7,14,21 d시,정상배양적세포감성린산매분별시상응107 mol/L지새미송조적2.93배,3.80배화4.39배,차이구유현저성의의(P＜0.05).④고농도지새미송대세포증식활력적억제효과명현.10-7 mol/L화10-6 mol/L여기타각조상비,차이구유현저성의의(P＜0.05).결론:대제량지새미송촉진골수간충질세포적지방생성,억제성골분화,병차수착농도증고화작용시간연장,작용가강.
BACKGROUND:Lipopexia induced by glucocorticoid is fat precipitation in marrow due to abnormal lipomatabolism,or differentiation of cells resulting from hormone-affected bone marrow mesenchymal cells.The precise generating procedure remains unclear.OBJECTIVE:To Jnvastigate effects of vadous concentrations of dexamethasone on adipogenic differentiation of bone marrow mesenchymal cells.DESIGN,TIME AND SETTING:The observational study was performed at the Department of Biochemistry,China Medical University from January 2007 to January 2009.MATERIALS:A total of 80 Sprague-Dawley rats aged 3 or 4 weeks,weighing (110±10) g,of both genders,were used in this study.METHODS:Rat bone marrow mesenchymal cells were isolated and subcultured.Bone marrow masenchymal cells at the third passage were used as samples.MAIN OUTCOME MEASURES:Cellular morphologic changes after treatment of 10~(-7) mol/L dexamethasone were observed under an inverted microscope,as well as at 3,7,14 and 21 days under normal culture condition.Morphological changes of bone marrow mesenchymal cells were observed following alkaline phosphatase and Sudan Ⅲ staining.Alkaline phosphatase activity was measured using phenol reagent.Effects of dexamethasone on cell proliferation were measured using MTT assay.RESULTS:Following 24 hours of incubation,a few adherent cells were found in the bottom of the culture flask,showing spindle shape.With prolonged time,adherent cells became more,presenting radiated shape.Following passage,cells distributed uniformly,showing typical fibroblast-shape.Following dexamethasone stimulation,cells changed from spindle-shape into polygonal or irregular shape.In the late phase,with increased concentration and prolonged time,cell colonies disappeared;cells adhered,and died.In normal cultured cells,no or a few orange particles were found.In dexamethasone-cultured cells,with increased hormone concentration and prolonged time,Sundan Ⅲ stained particles increased.At 21 days,alkaline phosphatase activity under normal culture was separately 1.57-,4.49-and 5.0-fold of 10~(-8),10~(-7),10~(-6) mol/L dexamethasone group.At 7,14 and 21 days,alkaline phosphatase activity under normal culture was separately 2.93-,3.80-and 4.39-fold of 10~(-7) mol/L dexamethasone group (P ＜ 0.05).High concentration of dexamethasone had significant inhibitory effects on cell proliferation activity.Significant difference was detected as compared 10~(-7) mol/L and 10~(-6) mol/L to other groups (P ＜ 0.05).CONCLUSION:High-dose dexamethasone enhances adipogenic and suppresses osteoblastic differentiation of bone marrow mesenchymal cells.The effect will increase along with the increasing content and prolonged duration of demamethasone stimulation.
