反义端粒酶催化亚单位抑制喉癌细胞生长的离体实验研究

目的：探讨人端粒酶催化亚单位（hTERT)反义寡核苷酸对人喉癌细胞株Hep-2的生长抑制作用。方法；用脂质体包裹hTERT反义寡核苷酸转染体外培养的Hep-2细胞，分别于转染后24、48、72h用MTT法检测细胞增殖活性，转染的72h行HE染色及端粒酶活性测定。结果：转染后细胞增殖活性降低；细胞生长密度下降；端粒酶活性受到抑制。结论：hTERT反义寡核苷酸短期内可以降低端粒酶活性、抑制喉癌细胞的生长。     

近5年国家自然科学基金资助耳鼻咽喉科学研究结题分析及成果简介

This paper reviewed and analyzed the 72 fulfilled projects in the discipline of Otorhinolar-yngology supported in division Ⅱ of basic research of clinical medicine by the National Natural Science Foun-dation of China (NSFC) from year 2002 to 2006, and gave a brief introduction to the part of research a-chievements. The science foundation promotes Otorhinolaryngology to a quick development, and produces some outstanding research achievements with international recognition. Thus, it plays an important role in el-evating the level of discipline and bringing up specialists. According to the retrospection and analysis of the fulfilled projects and their research achievements, we hope it can objective and overall reflect research pro-gress in the field of Otorhinolaryngology, and offer as a reference to the relevant researchers and scientific re-search managers.     

靶向PD-1及PD-L1免疫治疗所致肿瘤爆发性进展的研究进展

靶向程序性死亡受体1/程序性死亡受体配体1（programmed cell death1/programmed cell death ligand 1，PD-1/PD-L1）免疫治疗对实体瘤和血液肿瘤疗效明显，但是部分患者反而出现疾病超进展（hyperprogressive disease，HPD）。因此，建立统一的HPD诊断标准、筛选强有效的HPD预测因素、根据患者自身情况开展个体化治疗已然成为免疫治疗亟待解决的重要问题。本文对HPD定义、潜在机制、鉴别诊断、预测因素及临床管理等方面进行综述，旨在为更好地开展肿瘤免疫治疗提供有益借鉴。     

GATA3+Treg细胞在变应性鼻炎中的表达及意义

目的 研究GATA3⁺Treg细胞在变应性鼻炎(AR)患者和小鼠中的表达和意义。方法 收集6例对照组患者(Control组)和6例变应性鼻炎患者(AR组)的鼻黏膜,检测其鼻黏膜炎症反应。同期收集12例Control组和12例AR组患者外周血单个核细胞(PBMC),检测其Treg细胞和GATA3⁺Treg细胞比例。将C57BL/6小鼠分为正常对照组(Control组)和变应性鼻炎组(AR组)。比较两组小鼠AR症状学评分、外周血OVA-sIgE水平、鼻黏膜炎症反应。收集小鼠脾脏,检测两组小鼠体内Treg细胞和GATA3⁺Treg细胞比例以及Th2细胞因子表达。结果 与Control组相比,AR组患者鼻黏膜中嗜酸性粒细胞浸润和杯状细胞增生明显增多(P<0.01),PBMC中Treg细胞和GATA3⁺Treg细胞表达下降(P<0.05)。动物实验发现：与Control组相比,AR组小鼠过敏症状评分、外周血OVA-sIgE水平、鼻黏膜嗜酸性粒细胞浸润和杯状细胞增生均明显增加(P<0.01)。...     

小干扰RNA对喉鳞状细胞癌裸鼠移植瘤增殖细胞核抗原及p53蛋白表达的影响

目的：应用小干扰RNA（siRNA）技术抑制裸鼠喉癌皮下移植瘤人端粒酶逆转录酶（hTERT）基因表达,探讨移植瘤中增殖细胞核抗原（PCNA）及p53蛋白表达的变化。方法：根据hTERT cDNA序列构建表达短发夹RNA（shRNA）的、靶向hTERT mRNA的真核表达质粒pshRNA,其载体质粒含荧光素报告基因。建立人喉鳞状细胞癌Hep-2细胞株裸鼠皮下接种模型,将pshRNA转染入荷瘤裸鼠瘤体内,观察肿瘤生长情况。以激光共聚焦显微镜观察质粒在瘤体内的表达;以免疫组织化学SP法检测PCNA及p53蛋白在肿瘤内的表达。结果：所有裸鼠均接种成功,5d后可见皮下肿瘤形成,14d左右肿瘤直径达5～7mm。pshRNA及空质粒载体转染入瘤体后,共聚焦显微镜下见癌组织中有绿色荧光表达。病理学检查发现：pshRNA组肿瘤生长受到抑制,细胞分裂少见,可见大量癌细胞坏死。与注射生理盐水的对照组比较,转染质粒完毕后7d抑瘤率为76．50％,P〈0．01。pshRNA治疗后瘤体内PCNA蛋白表达显著下调（P〈0．05）,p53蛋白表达显著上调（P〈0．05）。结论：靶向hTERT mRNA的shRNA可显著抑制人喉癌裸鼠移植瘤的生长,其机制可能是诱导PCNA下调,促进p53蛋白表达所致。     

短发夹RNA对人骨髓间充质干细胞生长增殖的影响

目的:观察靶向人端粒酶逆转录酶(hTERT)的短发夹RNA(shRNA)对人骨髓间充质干细胞(hMSCs)生长增殖的影响,探讨靶向hTERT的RNA干扰技术临床应用的安全性。方法:根据RNA干扰原理,利用构建的表达shRNA的靶向hTERT mRNA的真核表达质粒(shRNA1),非特异性的shRNA真核表达质粒(shRNA2),转染试剂(德国metafectene)和正常培养液处理细胞。24h后在共聚焦显微镜下检测荧光表达情况;24h、48h、72h用MTT法检测细胞增殖活性;48h后进行HE染色、RT-PCR及端粒酶活性检测。结果:①共聚焦显微镜下见shRNA1及shRNA2组有大量的细胞表达荧光。②相应时间点各组细胞生长增殖无明显差异(P>0.05);HE染色发现,同等培养条件下,各组细胞生长密度及形态无明显变化。③RT-PCR显示各组细胞均无hTERT mRNA的表达;端粒酶活性为阴性。结论:靶向hTERT mRNA的shRNA对正常hMSCs的生长增殖无明显影响,该方法对不表达hTERT的正常体细胞可能是安全的。     

近5年国家自然科学基金资助耳鼻咽喉科学研究结题分析及成果简介

This paper reviewed and analyzed the 72 fulfilled projects in the discipline of Otorhinolar-yngology supported in division Ⅱ of basic research of clinical medicine by the National Natural Science Foun-dation of China (NSFC) from year 2002 to 2006, and gave a brief introduction to the part of research a-chievements. The science foundation promotes Otorhinolaryngology to a quick development, and produces some outstanding research achievements with international recognition. Thus, it plays an important role in el-evating the level of discipline and bringing up specialists. According to the retrospection and analysis of the fulfilled projects and their research achievements, we hope it can objective and overall reflect research pro-gress in the field of Otorhinolaryngology, and offer as a reference to the relevant researchers and scientific re-search managers.     

Treg细胞亚型在首诊癌症患者外周血中的表达及意义

目的 探讨癌症患者外周血中调节性T细胞(regulatory T cells, Tregs)亚型即1型辅助性T细胞(type 1 helper T cell, Th1)样Treg细胞(T-bet⁺Treg)、Th2样Treg细胞(GATA3⁺Treg)、Th9样Treg细胞(Pu-1⁺Treg)、Th17样Treg细胞(RORγ⁺Treg)及滤泡型辅助性T细胞(follicular helper T cell, Tfh)样Treg细胞(CXCR5⁺Treg)的表达情况及其对应的CD4⁺T细胞子集：Th1、2型辅助性T细胞(type 2 helper T cell, Th2)、9型辅助性T细胞(type 9 helper T cell, Th9)、17型辅助性T细胞(type 17 helper T cell, Th17)、Tfh的表达。方法 将23例癌症患者设为研究组，23例门诊健康体检者设为正常对照组，采用流式细胞术检测患者外周血中T-bet^+...     

近5年国家自然科学基金资助耳鼻咽喉科学研究结题分析及成果简介

This paper reviewed and analyzed the 72 fulfilled projects in the discipline of Otorhinolar-yngology supported in division Ⅱ of basic research of clinical medicine by the National Natural Science Foun-dation of China (NSFC) from year 2002 to 2006, and gave a brief introduction to the part of research a-chievements. The science foundation promotes Otorhinolaryngology to a quick development, and produces some outstanding research achievements with international recognition. Thus, it plays an important role in el-evating the level of discipline and bringing up specialists. According to the retrospection and analysis of the fulfilled projects and their research achievements, we hope it can objective and overall reflect research pro-gress in the field of Otorhinolaryngology, and offer as a reference to the relevant researchers and scientific re-search managers.