靶向Bcl-xL的miRNAs在机采血小板储存过程中的表达及对血小板凋亡的影响

Detection of miRNAs which targeted gene Bcl- xL in storaged human apheresis platelets and the effect of miRNAs on platelet apoptosis

目的观察靶向Bcl-x L的miRNAs在机采血小板储存过程中的表达量变化,结合生物信息学分析探讨miRNA对血小板凋亡的可能影响。方法利用实时荧光定量PCR(qRT-PCR)对储存第1 d、第3 d和第5 d的机采血小板中可能靶向Bcl-x L的miRNAs及Bcl-x L mRNA的表达量进行检测。评估高表达的miRNA与Bcl-x L的种子序列匹配度(seed match)、热力学稳定性、保守型及靶向性预测分值,并对miRNA表达与Bcl-x L mRNA变化之间的相关性进行分析。结果在血小板储存过程中,靶向Bcl-x L的hsa-miR-133a,hsa-miR-326,hsa-miR-491-5p,hsa-miR-98表达呈上升趋势(P<0.05),其中hsa-miR-326表达量显著增高(P<0.001);hsa-let-7家族(除hsa-let-7c),hsa-miR-133b,hsa-miR-342-3p,hsa-miR-342-5p表达水平无明显改变(P>0.05);hsa-let-7c,hsa-miR-140-5p,hsa-miR-184未检测到;而Bcl-x L mRNA在储存过程中表达呈下降趋势。hsa-miR-133a的表达变化与Bcl-x L mRNA呈显著负相关,生物信息学分析显示hsa-miR-133a和hsa-miR-326更有可能靶向调控Bcl-x L。结论Bcl-x L是血小板存活的关键调控蛋白,hsa-miR-133a,hsa-miR-326有可能调控Bcl-x L mRNA的表达,从而影响血小板的凋亡。

Objective To observe the expression changes of microRNAs（ miRNAs） which targeted Bcl- xL in storing apheresis platelets,and to explore the potential effect of miRNAs on platelet apoptosis with bioinformatics. Methods Quantitative real- time PCR（ qRT- PCR） was used to detect the expression levels of miRNAs and mRNA Bcl- xL in apheresis platelets at different preservation time（ 1 d,3 d,5 d）. To evaluate the seed match between high expressive miRNA and Bc1- xL,thermodynamic stability,conservative or targeting direction prediction scores,and the correlation between expression of miRNA and Bcl- xL were analyzed between mRNA changes. Results Our observation suggested that apheresis platelets contained highly abundant miRNAs. In which,the expression of hsa- miR- 133 a,hsa- miR- 326,hsa- miR- 491- 5p,hsa- miR- 98 were of increasing trend during storage（ P〈0. 05）. Noteworthily,the expression of hsa- miR- 326 distinctly demonstrated a marked increase in all platelets（ P 〈0. 001）. The hsa- let- 7 family（ except hsa- let- 7c）,hsa- miR- 133 b,hsa- miR- 342- 3p and hsa- miR- 342- 5p had no significant changes at different preservation time（ P〉 0. 05）; hsa- let- 7c,hsa- miR-140- 5p and hsa- miR- 184 were not detected. However,the expression of Bcl- xL was down- regulated during storage.Further,changes of expression of hsa- miR- 133 a and Bcl- xL mRNA showed a significant negative correlation,bioinformatics analysis showed that hsa- miR- 133 a and hsa- miR- 326 were more likely to regulate the expression of Bcl- xL. Conclusion Since Bcl- xL is the key regulator protein of platelet survival. hsa- miR- 133 a and hsa- miR- 326 possibly regulate the expression of Bcl- xL mRNA,so as to affect the platelet apoptosis.