长链非编码RNA-8439促进肝癌细胞多能因子nanog的表达

目的 探讨长链非编码RNA（lncRNA）-8439与多能因子nanog的关系,及lncRNA对肝癌细胞生长的影响。方法 在肝癌原发灶与转移灶组织中,通过lncRNA测序与生物信息学分析方法筛选与多能因子nanog、oct4、sox2相关的差异表达lncRNA。采用实时荧光定量PCR（qPCR）法检测差异表达的lncRNA在人肝癌细胞系Hep3B、Huh7细胞及对应的肿瘤悬浮球中的表达量。通过生物信息学分析明确lncRNA-8439与nanog结合的可能性,采用荧光原位杂交方法观察lncRNA-8439在Hep3B和Huh7细胞中的定位。敲低lncRNA-8439后,采用qPCR和蛋白质印迹法检测2种肝癌细胞中nanog的表达,观察悬浮球生长情况。过表达lncRNA-8439后,采用qPCR和蛋白质印迹法检测2种肝癌细胞中nanog的表达。结果 筛选到9种与多能因子相关的差异表达lncRNA,但仅lncRNA-8439在2种肝癌细胞悬浮球中呈一致的上调表达,与对应的贴壁细胞相比差异均有统计学意义（P均<0.01）。LncRNA-8439的3''端有nanog结合位点。LncRNA-8439在细胞核中表达,细胞质中并无分布。敲低lncRNA-8439后,2种肝癌细胞中nanog表达量在RNA与蛋白水平均降低（P均<0.01）,肝癌肿瘤悬浮球数量减少。过表达lncRNA-8439后,2种肝癌细胞中nanog表达量在RNA与蛋白水平均增加（P均<0.01）。结论 LncRNA-8439通过促进多能因子nanog的表达影响肿瘤细胞的自我更新能力,可能是导致肝癌侵袭和转移的原因。     

lncRNA-135调控miR-592维持小鼠胚胎干细胞多能性

目的筛选并确定调控miR-592作用的关键lncRNA,探讨其对miR-592沉默靶基因cep135的调控作用。方法利用lncRNA基因芯片筛选小鼠胚胎干细胞(mouse embryonic stem cells, mESCs)及KO miR-592 mESCs中差异的lncRNA从而找出关键调控分子lncRNA-135。利用转染试剂干扰lncRNA-135的表达,使用Western印迹法、双荧光素酶报告系统及免疫细胞化学染色等技术检测lncRNA-135对miR-592及其靶基因cep135的表达影响,进而阐明lncRNA-135对mESCs多能性的调控作用。结果lncRNA基因芯片分离确定了527条差异表达的lncRNA,经过生物信息学分析及qRT-PCR验证,确定了评分最高的lncRNA-135为关键作用分子。双荧光素酶报告系统证实了lncRNA-135能够结合miR-592以干扰其沉默靶基因cep135。而Western印迹法和免疫细胞化学染色结果则证实lncRNA-135通过作用于miR-592,阻止其对cep135的抑制作用,进而促进mESCs的多能性。结论lncRNA-135可以通过充当miR-592的“分子海绵”从而调节与mESCs多能性维持有关的多个基因的表达,这将可能帮助建立调控性miRNA网络与mESCs多能性之间的联系。     

Treatment duration-dependent efficacy of Japanese cedar pollen sublingual immunotherapy: Evaluation of a phase II/III trial over three pollen dispersal seasons

BackgroundWe conducted a randomized, placebo-controlled, double-blind clinical trial to investigate the optimal dose and long-term efficacy and safety of Japanese cedar (JC) pollen tablets for SLIT (JapicCTI-142579). Here, we report details of the effects of the JC pollen SLIT tablet on rhinitis and conjunctivitis symptoms over three pollen dispersal seasons.MethodsA total of 1042 JC pollinosis patients (aged 5–64 years) were randomized to receive tablets containing placebo (P), 2000, 5000, or 10,000 Japanese allergy units (JAU) of JC pollen for 15 months to identify an optimal dose. Patients receiving P (n = 240) and the optimal dose (5000 JAU; A, n = 236) were then randomized to receive P or A for an additional 18 months (AA, AP, PA, and PP groups, allocation ratio 2:1:1:2). Nasal and ocular symptoms, rescue medication use, and quality of life (QOL) were assessed on quantitative scales.ResultsIn the second and third seasons, the AA, AP, and PA groups exhibited significantly better improvements in nasal, ocular, and medication scores compared with the PP group in the order AA > AP > PA > PP during the second season and AA > PA > AP > PP during the third season. Rescue medication use and QOL scores were also significantly better in the AA, AP, and PA groups compared with the PP group.ConclusionsThe JC pollen SLIT tablet relieved nasal and ocular symptoms and medication use and improved QOL in a treatment duration-dependent manner. Continuous dosing regimens appear to enhance the efficacy of the drug.