Hsa-miR-4282对肝癌细胞系SMMC-7721生长的影响

目的  探讨Hsa-miR-4282在肝癌细胞系SMMC-7721中的表达及其对细胞生长的影响。方法 采用实时荧光定量PCR法检测Hsa-miR-4282在人正常肝上皮细胞系HL-7702和人肝癌细胞系MHCC97-H、SMMC-7721，以及 20例肝癌组织及其相应癌旁组织中的表达。采用瞬时转染法将Hsa-miR-4282 mimics（上调组）和Hsa-miR-4282 inhibitor（下调组）分别转染肝癌SMMC-7721细胞，上调组和下调组分别设置相应阴性对照组。转染后采用MTT法检测细胞增殖能力，平板克隆形成实验检测细胞克隆形成能力，流式细胞仪检测细胞凋亡能力。结果 Hsa-miR-4282在肝癌组织、肝癌细胞MHCC97-H及肝癌细胞SMMC-7721中的表达均低于癌旁组织及正常肝细胞HL-7702（P＜0.05）。MTT实验结果显示，Hsa-miR-4282上调后肝癌SMMC-7721细胞的OD值低于其阴性对照组，而下调后OD值高于其阴性对照组 （P＜0.05）。平板克隆形成实验显示，下调组的细胞克隆数高于其阴性对照组［（240±7） 个 vs （191±10） 个，P=0.005）］，而上调组细胞克隆数低于基阴性对照组［（146±10） 个 vs （193±12） 个，P=0.013）］。流式细胞仪检测结果显示，Hsa-miR-4282上调组细胞凋亡率较其阴性对照组升高［（23.89±1.89）% vs（16.6±1.14）%，P=0.009）］，下调组细胞凋亡率较期阴性对照组降低［（14.98±0.46）% vs （17.79±0.73）%，P=0.010］。结论 Hsa-miR-4282上调可抑制肝癌SMMC-7721细胞增殖，促进细胞凋亡，可能与肝癌的发病机制有关。     

“T” is for “transgender”: an analysis of children’s picture books featuring transgender protagonists and narrators

ABSTRACT

This project examined 55 picture books featuring transgender, gender expansive or genderqueer protagonists or narrators published between 2008 and 2018. The purpose of the study was to determine how this genre of children’s literature supports and challenges four gender assumptions: the gender binary, gender essentialism, sex/gender congruency and gender stability. Additionally, this critical analysis explored misgendering within this genre and themes of social rejection and acceptance. Protagonists and narrators were permitted a degree of gender nonconformity, however, the majority of picture books missed opportunities for a more complete exploration of gender possibilities.     

Recent advances in preclinical in vitro approaches towards quantitative prediction of hepatic clearance and drug-drug interactions involving organic anion transporting polypeptide (OATP) 1B transporters

Hepatic uptake mediated by organic anion transporting polypeptide (OATP) 1B1 and 1B3 can serve as a major elimination pathway for various anionic drugs and as a site of drug-drug interactions (DDIs). This article provides an overview of the in vitro approaches used to predict human hepatic clearance (CL_h) and the risk of DDIs involving OATP1Bs. On the basis of the so-called extended clearance concept, in vitro–in vivo extrapolation methods using human hepatocytes as in vitro systems have been used to predict the CL_h involving OATP1B-mediated hepatic uptake. CL_h can be quantitatively predicted using human donor lots possessing adequate OATP1B activities. The contribution of OATP1Bs to hepatic uptake can be estimated by the relative activity factor, the relative expression factor, or selective inhibitor approaches, which offer generally consistent outcomes. In OATP1B1 inhibition assays, substantial substrate dependency was observed. The time-dependent inhibition of OATP1B1 was also noted and may be a mechanism underlying the in vitro–in vivo differences in the inhibition constant of cyclosporine A. Although it is still challenging to quantitatively predict CL_h and DDIs involving OATP1Bs from only preclinical data, understanding the utility and limitation of the current in vitro methods will pave the way for better prediction.     

miR-26b通过调节Notch1信号通路参与原发性肝细胞肝癌侵袭的机制研究

目的:探讨miR-26b参与原发性肝细胞肝癌（HCC）侵袭的机制。方法:在细胞培养液中培养人肝细胞系HL-7702和HCC细胞各系Hepb-3、HuH-7、MHCC97-L、MHCC97-H。实时荧光定量PCR法（qRT-PCR）检测miR-26b的表达水平；用miR-26b mimics、miR-26b inhibitors和Notch1-siRNA分别转染HCC细胞；MTT实验检测转染后HCC细胞的活力；采用Western blot检测Notch1受体蛋白表达水平的变化；Transwell小室测定不同处理后的HCC细胞的侵袭能力。结果:人正常肝细胞系HL-7702和HCC细胞系Hepb-3、HuH-7、MHCC97-L、MHCC97-H中的miR-26b相对表达含量随其侵袭和迁移能力的升高而依次下降；抑制miR-26b的表达，Notch1受体蛋白表达明显增高，而此时HCC细胞的侵袭性显著增强；相反，上调miR-26b的表达，Notch1受体蛋白表达明显降低，而HCC细胞侵袭性显著下降；miR-26b可能通过调控Notch1信号通路调节HCC细胞侵袭性。结论:miR-26b通过负调控Notch1信号通路抑制HCC细胞侵袭能力，为HCC侵袭的机制奠定了理论基础，miR-26b可能成为HCC治疗的新靶点。     

miR-30c Impedes Glioblastoma Cell Proliferation and Migration by Targeting SOX9

miR-30c has been acknowledged as a tumor suppressor in various human cancers, such as ovarian cancer, gastric cancer, and prostate cancer. However, the role of miR-30c in glioblastoma (GBM) needs to be investigated. In our study, we found that the expression of miR-30c was significantly downregulated in GBM tissues and cell lines. We found that overexpression of miR-30c inhibited cellular proliferation of GBM cells in vitro and in vivo. More GBM cells were arrested in the G₀ phase after miR-30c overexpression. Moreover, we showed that miR-30c overexpression suppressed the migration and invasion of GBM cells. Mechanistically, we found that SOX9 was a direct target of miR-30c in GBM cells. Overexpression of miR-30c inhibited the mRNA and protein levels of SOX9 in GBM cells. Moreover, there was a negative correlation between the expression of miR-30c and SOX9 in GBM tissues. Finally, we showed that restoration of SOX9 in GBM cells reversed the proliferation, migration, and invasion of GBM cells transfected with miR-30c mimic. Collectively, our results demonstrated that miR-30c suppressed the proliferation, migration, and invasion of GBM cells via targeting SOX9.     

凉血通瘀方干预高血压大鼠急性脑出血模型对脑组织中差异miRNA表达的影响＊

目的 研究凉血通瘀方对高血压大鼠急性脑出血模型脑组织miRNA表达的影响，对差异表达的miRNA靶基因进行分析，探索凉血通瘀方可能的药效机制。方法 将自发性高血压大鼠随机分成对照组（B）和实验组（C）。适应性饲养一周后，C组灌胃凉血通瘀方，B组灌胃等体积生理盐水，连续5天，每天1次。构建脑出血模型后收集脑组织，借助全转录组测序技术获得miRNA表达量，与miRBase数据库比对获取已知miRNA，使用miRDeep2预测新miRNA。差异分析软件为DESeq2，筛选阈值为|log₂FC| ≥1 并且P <0.05。对显著差异表达的miRNA进行靶基因预测，对靶基因进行GO功能、KEGG通路富集和PPI网络分析。结果 实验组和对照组对比，共发现21个显著差异表达的miRNA，上调有9个，下调有12个，共预测得到1243个有统计学意义的靶基因。GO富集分析发现，生物过程中突触囊泡分泌的调节、神经递质分泌的调节和神经递质运输的调节占前三位，神经元投射终点、全膜、质膜区域和细胞投射则是主要的细胞成分。分子功能分别为小GTPase绑定、底物特异性跨膜转运蛋白活性和离子跨膜转运体活性。通路分析结果显示，靶基因在癌证通路、pI3K-Akt信号通路、人类乳头瘤病毒感染、神经活性配体-受体相互作用和MAPK通路等分布广泛。采用STRING网站和Cytoscape软件，根据MCC算法筛选出ADRA2C、CASR、CCL28、CCR1、DRD2、GNAT3、GRM2、DYNC1LI1、GABBR1、GNAI1等核心靶基因。结论 凉血通瘀方对脑出血急性期鼠脑组织内miRNA的表达有重要影响；显著差异表达miRNAs可能通过靶向核心基因调控凉血通瘀方干预急性脑出血的病理过程及预后。