中国免疫学杂志
中國免疫學雜誌
중국면역학잡지
CHINESE JOURNAL OF IMMUNOLOGY
2015年
9期
1173-1177
,共5页
陶弋婧%朱顺飞%陈超%赵娟娟%郭萌萌%张忆雄%秦娜林%徐林
陶弋婧%硃順飛%陳超%趙娟娟%郭萌萌%張憶雄%秦娜林%徐林
도익청%주순비%진초%조연연%곽맹맹%장억웅%진나림%서림
微小RNA-7%基因敲减%胸腺%CD4+SP细胞
微小RNA-7%基因敲減%胸腺%CD4+SP細胞
미소RNA-7%기인고감%흉선%CD4+SP세포
miRNA-7%Knock down%Thymus%CD4+SP cell
目的:研究microRNA-7( miRNA-7,miR-7)基因敲减对小鼠CD4+SP细胞胸腺发育的影响。方法:检测miR-7基因敲减(miR-7 knock down,miR-7KD)和野生型(Wild-type,WT)小鼠胸腺重量及细胞总数变化;HE染色观察miR-7KD小鼠胸腺的形态学结构改变;FACS检测胸腺CD4+单阳性(Single positive,SP)细胞的比例并计算细胞总数,同时检测CD4+SP细胞CD44及CD62L表达变化;核抗原Ki-67染色法检测CD4+SP细胞的增殖情况;FACS检测CD4+SP细胞的凋亡变化;Western blot 技术检测胸腺中总ERK1/2和磷酸化ERK1/2表达水平变化。结果:与WT小鼠相比,miR-7KD小鼠胸腺体积、重量以及细胞总数均显著减少,且形态学结构发生改变( P<0.05);FACS结果显示,miR-7KD小鼠胸腺CD4+SP细胞比例明显降低,且细胞总数减少( P<0.05)。此外,CD4+SP细胞的CD44表达水平显著增加,而CD62 L表达水平明显减少( P<0.05);同时,CD4+SP细胞增殖及凋亡比例均显著增加(P<0.01);最后,miR-7KD小鼠胸腺中ERK1/2以及磷酸化ERK1/2的表达均明显下调( P<0.05)。结论:miR-7基因敲减后可显著影响CD4+SP细胞的胸腺发育,这可能与细胞活化水平及ERK1/2信号通路改变有关。
目的:研究microRNA-7( miRNA-7,miR-7)基因敲減對小鼠CD4+SP細胞胸腺髮育的影響。方法:檢測miR-7基因敲減(miR-7 knock down,miR-7KD)和野生型(Wild-type,WT)小鼠胸腺重量及細胞總數變化;HE染色觀察miR-7KD小鼠胸腺的形態學結構改變;FACS檢測胸腺CD4+單暘性(Single positive,SP)細胞的比例併計算細胞總數,同時檢測CD4+SP細胞CD44及CD62L錶達變化;覈抗原Ki-67染色法檢測CD4+SP細胞的增殖情況;FACS檢測CD4+SP細胞的凋亡變化;Western blot 技術檢測胸腺中總ERK1/2和燐痠化ERK1/2錶達水平變化。結果:與WT小鼠相比,miR-7KD小鼠胸腺體積、重量以及細胞總數均顯著減少,且形態學結構髮生改變( P<0.05);FACS結果顯示,miR-7KD小鼠胸腺CD4+SP細胞比例明顯降低,且細胞總數減少( P<0.05)。此外,CD4+SP細胞的CD44錶達水平顯著增加,而CD62 L錶達水平明顯減少( P<0.05);同時,CD4+SP細胞增殖及凋亡比例均顯著增加(P<0.01);最後,miR-7KD小鼠胸腺中ERK1/2以及燐痠化ERK1/2的錶達均明顯下調( P<0.05)。結論:miR-7基因敲減後可顯著影響CD4+SP細胞的胸腺髮育,這可能與細胞活化水平及ERK1/2信號通路改變有關。
목적:연구microRNA-7( miRNA-7,miR-7)기인고감대소서CD4+SP세포흉선발육적영향。방법:검측miR-7기인고감(miR-7 knock down,miR-7KD)화야생형(Wild-type,WT)소서흉선중량급세포총수변화;HE염색관찰miR-7KD소서흉선적형태학결구개변;FACS검측흉선CD4+단양성(Single positive,SP)세포적비례병계산세포총수,동시검측CD4+SP세포CD44급CD62L표체변화;핵항원Ki-67염색법검측CD4+SP세포적증식정황;FACS검측CD4+SP세포적조망변화;Western blot 기술검측흉선중총ERK1/2화린산화ERK1/2표체수평변화。결과:여WT소서상비,miR-7KD소서흉선체적、중량이급세포총수균현저감소,차형태학결구발생개변( P<0.05);FACS결과현시,miR-7KD소서흉선CD4+SP세포비례명현강저,차세포총수감소( P<0.05)。차외,CD4+SP세포적CD44표체수평현저증가,이CD62 L표체수평명현감소( P<0.05);동시,CD4+SP세포증식급조망비례균현저증가(P<0.01);최후,miR-7KD소서흉선중ERK1/2이급린산화ERK1/2적표체균명현하조( P<0.05)。결론:miR-7기인고감후가현저영향CD4+SP세포적흉선발육,저가능여세포활화수평급ERK1/2신호통로개변유관。
Objective:To investigate the influence of miR-7 knock down on the development of CD 4+SP cells in the thymus in mice,and preliminary explore its possible mechanism.Methods:The changes of volume ,weight and total cell counts of thymus in miR-7 knock down (miR-7KD) mice were observed compared with Wild-type(WT)mice;the pathological changes of thymus were observed by HE staining.FACS analysis was performed on the proportion ,as well as the expression level of CD44 and CD62L,of thymus CD4+single positive (SP) cells.Meanwhile,the proliferation percentage of CD4+SP cells was measured by Ki-67 staining.The apoptosis percentage of CD4+SP cells was analyzed by FACS.The changes on the transduction of ERK 1/2 pathways were determined by Western blot.Results:Compared with WT mice ,the size,weight and total cell number of thymus were marked reduced in miR-7KD mice( P<0.05 );moreover ,pathological change also was presented.The proportion and total cell number of thymus CD 4+SP cells were marked decreased ( P<0.05 ).Furthermore ,the expression level of CD 44 and proliferation percentage ,as well as apoptosis percentage ,of CD4+SP cells were obviously increased (P<0.05),however,the expression level of CD62L of CD4+SP cells were decreased (P<0.05). Finally,the level of total ERK1/2 and phosphor-ERK1/2 was decreased obviously ( P<0.05 ).Conclusion: miR-7 knock down can affect the development of CD 4+SP cells in the thymus , which might be closely related to the cell activation state and altered the transduction of ERK1/2 pathways.