[摘 要] 目的：探讨丹皮酚（Pae）通过调控果蝇zeste基因增强子同源物2/神经外营养蛋白4抗体/细胞周期蛋白依赖性激酶抑制因子2A（EZH2/NXPH4/CDKN2A）轴对肺癌A549细胞增殖、迁移和侵袭能力的影响。方法：以梯度浓度（0、6.25、12.5、25、50、100、200 µg/mL）的Pae处理肺癌A549细胞，采用CCK-8法确定干预浓度；将A549细胞分为对照组（未经任何处理的A549细胞）、Pae组（12.5 µg/mL Pae）、Pae + siEZH2组（转染siEZH2 + 12.5 µg/mL Pae）、Pae+siNC组（转染siNC + 12.5 µg/mL Pae）、Pae + vector NC组（转染vectorNC + 12.5 µg/mL Pae）、Pae + vectorEZH2组（转染vector EZH2+12.5 µg/mL Pae），予以相应的处理后，采用克隆形成实验检测细胞克隆数，流式细胞术检测细胞周期分布，Transwell实验检测细胞侵袭能力的变化，划痕实验检测细胞迁移能力的变化，流式细胞仪检测细胞凋亡情况，WB法检测EZH2、NXPH4、CDKN2A、Bcl-2和caspase-3蛋白表达量。在A549细胞中单独转染siEZH2或siNC，采用流式细胞仪测量细胞凋亡，WB法检测Bcl-2和caspase-3蛋白表达。建立A549细胞裸鼠移植瘤模型，评估Pae的体内抗肿瘤作用及其对相关蛋白表达的影响。结果：选择12.5 µg/mL为后续实验干预浓度。与对照组相比，Pae组细胞克隆数、S期细胞比例、迁移、侵袭能力以及EZH2、NXPH4和Bcl-2蛋白表达量显著降低，G0/G1期细胞比例、细胞凋亡率以及CDKN2A和caspase-3蛋白表达量明显升高（均P < 0.05）；与Pae + siNC组相比，Pae + siEZH2组细胞克隆数、S期细胞比例、迁移侵袭能力以及EZH2、NXPH4和Bcl-2蛋白表达量下降幅度更大，G0/G1期细胞比例、细胞凋亡率以及CDKN2A和caspase-3蛋白表达量升高幅度更大（P < 0.05）；与Pae+vectorNC组相比，Pae + vectorEZH2组细胞克隆数、S期细胞比例、迁移侵袭能力以及EZH2、NXPH4和Bcl-2蛋白表达量显著升高，G0/G1期细胞比例、细胞凋亡率以及CDKN2A和caspase-3蛋白表达量明显下降（P < 0.05）。敲低EZH2后A549细胞凋亡率和caspase-3表达明显上升，Bcl-2表达明显下降（P < 0.05）。体内实验表明Pae可显著降低肿瘤体积和质量且抑制EZH2/NXPH4/CDKN2A通路的活性。结论：Pae通过抑制EZH2/NXPH4/CDKN2A通路抑制A549细胞增殖、迁移和侵袭，诱导细胞凋亡。

Objective : To study the differential expression and diagnostic value of miR-26a-5p and miR-151a-3p in plasma exosomes of tuberculosis. Methods : Differentially expressed miRNAs ( miR-26a-5p and miR-151a-3p) in tuberculosis exosomes were identified through bioinformatics website．Furthermore，70 tuberculosis patients (tuber- culosis group) and 58 healthy subjects ( healthy control group) were selected for clinical validation : Plasma exo- somes，extracted by differential centrifugation，were identified by Western blot，transmission electron microscopy， nanoparticle tracking analysis．The expression levels of miR-26a-5p and miR-151a-3p in plasma exosome were de- tected by RT-qPCR in the tuberculosis group and the control group．The diagnostic value of miR-26a-5p and miR- 151a-3p in tuberculosis was evaluated by ROC curve. Results : The characteristics of exosomes were identified by transmission electron microscopy，nanoparticle tracking analysis and Western blot，which showed that plasma exo- somes were extracted successfully．In clinical validation，the internal and external parameters were calibrated sim- ultaneously，the expression level of miR-26a-5p in plasma exosome of patients with tuberculosis was lower than that in control group (P＜0. 000 1) ，and the expression level of miR-151a-3p in plasma exosome of patients with tuber- culosis was higher than that in control group (P＜0. 000 1) ，which were consistent with the bioinformatics results. As a diagnostic marker，taking internal reference as the benchmark，the AUC of plasma exosome miR-26a-5p and miR-151a-3p for tuberculosis differentiation was 0. 872 and 0. 709，the AUC of the combined miR-26a-5p and miR- 151a-3p was 0. 915 (P ＜0. 000 1 ) ． Taking external reference as the benchmark ，the AUC of plasma exosomes miR-26a-5p and miR-151a-3p were 0. 829 and 0. 854 respectively ，the AUC of the combined miR-26a-5p and miR-151a-3p was 0. 911 (P＜0. 000 1) ． Conclusion The expression level of plasma exosome-derived miR-26a- 5p decreases in tuberculosis patients and the expression levelof miR-151a-3p increases，and the clinical diagnostic efficiency of bothis high，which may be a potential biomarker for the diagnosis of tuberculosis.

[Abstract] Objective: To investigate the expression of miR144-3p in bladder cancer tissues and cells and its effect on the proliferation and invasion of T24 cells. Methods: A total of 36 cases of bladder cancer tissue specimens and 10 cases of normal bladder epithelial tissue specimens were collected from Tangdu Hospital of Air Force Medical University during February 2018 and December 2018. In addition, bladder cancer T24 cell line and normal urothelial cell line SV-HUC-1 were also collected for this study. The levels of miR144-3p in bladder cancer tissues and cells were detected by qPCR methods. The miR-144-3p mimics and miR-NC were transfected into T24 cells by LipofectamineTM 2000, respectively. The proliferation, cell cycle distribution and invasion abilities were detected by MTT, Flow cytometry and Transwell chamber methods, respectively. TargetScan software was used to predict the binding site between miR-144-3p and E2F3 (E2F transcription factor 3); Dual luciferase reporter gene assay was used to verify the relationship between miR-144-3p and E2F3; and WB was used to detect the expression levels of miR-144-3p and E2F3 in cells. Results: The expression of miR-144-3p was downregulated in bladder cancer tissues and cells (all P<0.01). In addition, the expression level of miR-144-3p in muscular invasive bladder cancer tissues was significantly lower than that in non-muscular invasive bladder cancer tissues (P<0.05). Dual luciferase reporter gene assay confirmed that there was a targeted relationship between miR-144-3p and E2F3. Overexpression of miR-144-3p inhibited the proliferation and invasion of T24 cells (all P<0.01) and downregulated the expression of E2F3 (P<0.01); upregulation of E2F3 could reverse the inhibitory effect of miR-144-3p overexpression on proliferation and invasion of T24 cells. Conclusion: miR-144-3p has low expression level in bladder cancer tissues. It inhibits proliferation and invasion of bladder cancer cells by downregulating E2F3.

[Abstract] Objective: To study the effect and possible mechanism of TGF-β2 on the invasion of glioma stem cells (GSCs). Methods: Tumor tissues of 8 patients with glioblastoma multiforme, who underwent resection at Department of Neurosurgery of the FirstAffiliated Hospital of China Medical University duringApril 2016 toApril 2017, were collected. The primary culture of glioma cells were conducted with trypsin digestion. Partial primary glioma cells were seeded into serum-free DMEM/F12 culture medium containing EGF, bFGF and B27 to obtain suspension of tumor spheres. Immunoflurenscent staining and differentiation assay were used to detect whether the tumor spheres were GSCs. TGF-β2 secretion ability of GSCs was determined by ELISAassay.After transfection of TGF-β2 siRNA, the invasion ability of glioma stem cells was determined by Transwell assay. Western blotting was used to examine the effect of TGF-β2 on expression of matrix metalloproteinases (MMP) in glioma stem cells. Results: The suspended tumor spheres were proved to be GSCs by immunofluorescent staining and differentiation assay; the tumor spheres expressed the marker of GSCs（CD133）and had the ability to multi-differentiate (glia and neuronal cells). Compared with the primary glioma cells, Glioma stem cells exerted significantly improved TGF-β2 secretion ability ([74.13±3.63] vs [46.13±2.61] pg/ml, P<0.05); and TGF-β2 silencing significantly reduced the invasion ability of glioma stem cells ([105.71±8.69] vs [63.67±5.93], P<0.05) and inhibited MMP-2 and MMP-9 expressions. Conclusion: TGF-β2 can promote the invasiveness of glioma stem cells through MMP-2 and MMP-9 pathway.