High expression of XPA confers poor prognosis for nasopharyngeal carcinoma patients treated with platinum-based chemoradiotherapy

In this study, we tried to explore if xeroderma pigmentosum complementation group-A (XPA) expression is likely a prognostic prediction factor for locally advanced nasopharyngeal carcinoma (NPC) patients treated with platinum-based chemoradiotherapy, which was considered to bring chemotherapy-related severe toxicity compared with radiotherapy alone. Firstly, MTT assay revealed that downregulating XPA expression in NPC HONE1 and CNE1 cells decreased IC₅₀ of cisplatin and sensitized cells to cisplatin. XPA expression was detected by immunohistochemistry in cancer tissues from locally advanced NPC patients treated with platinum-based chemoradiotherapy. The relationships between XPA expression and clinicopathologic features, overall survival and progression-free survival of patients were evaluated. The results showed that XPA expression was not associated with clinicopathologic parameters, but was likely an independent prognostic factor for patient survival. High XPA level predicts a poor prognosis, and the prediction values were higher in subgroups of younger, higher EBV antibody titer, or treated with concurrent chemoradiotherapy. Combining XPA levels and T/N classifications, we successfully classified these patients into low, medium and high risk groups for platinum-based chemoradiotherapy. These findings suggest that XPA levels may be a potential predictor of prognosis in locally advanced NPC patients treated with platinum-based chemoradiotherapy, and helpful for selecting patients likely to need and benefit from this treatment in future.     

EGFR mono-antibody salvage therapy for locally advanced and distant metastatic penile cancer: Clinical outcomes and genetic analysis

Purpose

There are limited therapeutic options for patients with advanced penile squamous cell carcinoma (PSCC) after chemotherapy failure. Thus, we evaluated the feasibility of salvage treatment using the epidermal growth factor receptor (EGFR) mono-antibody nimotuzumab in chemotherapy-failed PSCC patients and explored potential response or resistance biomarkers.

垂直向控制在矫正成人骨性Ⅱ类高角错畸形应用中的研究进展

颌骨与牙包含三个维度：矢状向、横向和垂直向,以往临床上错畸形的矫治常聚焦于前突或后缩问题,而易忽略垂直向问题。垂直向问题往往对矢状向产生影响,如垂直向发育过度患者多呈现高角骨面型,易引起下颌骨顺时针旋转,形成骨性Ⅱ类错畸形。早期由于相应的矫正手段有限,正颌手术一直是骨性Ⅱ类高角面型患者的首选治疗方式。随着种植支抗的出现及应用,使用微钛板和（或）微螺钉种植体可以提高单纯正畸治疗的垂直向控制效果,减小了部分轻、中度垂直向发育过度患者的治疗风险和创伤。本文旨在回顾近年关于正畸代偿及正颌-正畸联合治疗矫正成人骨性Ⅱ类高角病例的研究,从不同方式的控制效果、稳定性分析以及临界病例的治疗方式等方面,对应用垂直向控制纠正骨性Ⅱ类高角错畸形的研究进展进行综述,为临床诊疗和相关研究提供一定的思路和依据。     

重组人内皮抑素腺病毒抑制肝癌裸鼠移植瘤生长

目的 观察重组人内皮抑素腺病毒(Ad/hEndo)对人肝癌裸鼠移植瘤生长的影响。方法 人脐静脉内皮细胞ECV-304经Ad/hEndo感染后,western印迹检测人内皮抑素的表达。人肝癌BEL-7402细胞移植到裸鼠背脊部后,检测Ad/hEndo对肝癌移植瘤生长的抑制作用。逆转录聚合酶链反应(RT-PCR)检测肿瘤组织中内皮抑素mRNA的表达。分析人内皮抑素在裸鼠体内的表达分布。结果 Western印迹检测到人内皮抑素基因在ECV-304细胞内高效表达。Ad/hEndo明显抑制人肝癌BEL-7402裸鼠移植瘤生长(F=4.061,P<0.05)。Ad/hEndo组血管密度计数为6.88±1.08,DMEM组为13.60±1.71(t=9.216,P<0.01)。瘤内注射Ad/hEndo后3d,RT-PCR在肿瘤组织检测到内皮抑素mRNA的表达,7d后表达不明显。人内皮抑素蛋白主要分布在肿瘤组织。结论 腺病毒介导的人内皮抑素基因在体内、体外获得高效表达,并明显抑制肝癌裸鼠移植瘤的生长与血管生成。     

Evaluation of long-term toxicity of Ad/hIFN-, an Adenoviral vector encoding the human interferon-gamma gene, in nonhuman primates

Interferon-gamma(IFN-gamma) plays an important role in the immunomodulation and growth inhibition of many tumor cells, but its clinical application is limited by its systemic toxicity. Ad/hIFN-gamma, a nonreplicating adenoviral vector encoding human IFN-gamma, has been reported to inhibit tumor growth in vitro and in a xenograft model. In this study, the long-term toxicity of Ad/hIFN-gamma was assessed in cynomolgus macaques (Macaca fascicularis). Thirty animals were enrolled into 5 groups, and administered intramuscularly, respectively, Ad/hIFN-gamma (3.3 x 10(10), 3.3 x 10(11), or 3.3 x 10(12) VP/kg), Ad/LacZ (vector control, 3.3 x 10(11) VP/kg), or excipient 3 times per week for 8 weeks, followed by a 4-week recovery period. At 12 weeks all experimental animals appeared generally healthy, and there were no statistically significant differences in body weight, urinalysis, hemogram, blood biochemistry, and electrocardiogram results between the treatment and control groups. No significant toxic effects were noted on macroscopic and microscopic examinations of organs and tissues. Preliminary investigation of the immunotoxicity of Ad/IFN-gamma indicated that anti-adenoviral and anti-hIFN-gamma antibodies were generated. These data demonstrate that long-term, high-dose intramuscular administration of Ad/IFN-gamma was not notably toxic and might be safe for clinical therapeutic use.     

一种用流式细胞术检测基因转移效率的新方法

背景与目的：基因转移效率一直以来都是用表达报告基因的细胞（显色或发荧光）占细胞总数的百分比（即转染率）来表达,此百分比是由研究者借助显微镜（或荧光显微镜）计数得出来的。该方法（本文称为人工计数法）存在客观性、准确性不够和工作量大、且不能同时检测基因表达效率等问题。本研究拟寻找一种客观、准确、简便地检测基因转移和表达效率的新方法。方法：以绿色荧光蛋白（GFP）基因为目的基因、重组腺病毒（AdCMV/GFP)为基因载体,用人工计数法测定其对肝癌细胞株HepG2的转染率;用流式细胞术（flow cytometry,FCM)测定其对肝癌细胞株HepG2、Bel7402、Hep3B、SMMC7721和鼻咽癌细胞株CNE－2的转染效率和GFP（绿色荧光蛋白）基因的表达效率（以荧光指数表示）。结果：以FCM测得AdCMV/GFP对HepG2的转染率与载体量呈对数相关（与人工法相似,但前者略高）,荧光指数与载体量呈线性相关（γ＝0．9984）,其它细胞株的结果与HepG2相似。结论：采用FCM法测定基因转移效率克服了主观因素的影响,测定结果客观准确,可以用于测定基因载体对真核细胞的转染效率,或研究转录调控元件的功能。     

人神经营养素-3基因重组腺病毒载体的构建、表达和生物活性检测

目的构建具有生物活性的人神经营养素-3（NT-3）基因重组腺病毒表达载体.方法从人脑组织mRNA中扩增NT-3基因全长cDNA,定向克隆于穿梭质粒pShuttle中,获得一个带有CMV启动子的质粒.再与腺病毒骨架DNA（Adeno-X viral DNA）体外连接,形成重组腺病毒质粒（pAd-NT-3）.用pAd-NT-3转染人胚肾293细胞后包装成有感染能力的重组腺病毒颗粒（Adeno-NT-3）. 结果NT-3基因RT-PCR扩增产物约801 bp.Adeno-NT-3 PCR鉴定为正确重组子.RT-PCR、免疫细胞化学、ELISA及Western blot检测显示,Adeno-NT-3能在其转染的293细胞表达和分泌NT-3.这种NT-3能使体外培养的神经干细胞更多地分化为神经元样细胞. 结论应用体外连接法成功构建了人NT-3基因重组腺病毒表达载体,其表达产物具有促进神经干细胞分化为神经元样细胞的活性作用.     

维生素E对L-NAME致大鼠高血压的影响

目的:观察维生素E对L-NAME所致大鼠高血压模型的降压作用。方法:用L-NAME复制稳定SD大鼠高血压模型。建模成功后,将大鼠随机分为空白对照组、低剂量组(维生素E 400mg.kg-1.d-1)和高剂量组(维生素E 1 200mg.kg-1.d-1)。大鼠每日口服维生素E,每2周以尾动脉测压法测量血压,统计数据采用重复测量的方差分析。结果:L-NAME复制高血压模型稳定可行,服用维生素E后,三组大鼠血压都有下降。下降程度:对照组<低剂量组<高剂量组(P<0.001)。结论:维生素E可降低L-NAME致大鼠高血压模型的血压。     

Adenovirus-mediated delivery of interferon-γ gene inhibits the growth of nasopharyngeal carcinoma

Background

Epidermal growth factor receptor (EGFR) is suggested to predict the radiosensitivity and/or prognosis of human esophageal squamous cell carcinoma (ESCC). The objective of this study was to investigate the efficacy of Nimotuzumab (an anti-EGFR monoclonal antibody) on ESCC radiotherapy (RT) and underlying mechanisms.

Methods

Nimotuzumab was administrated to 2 ESCC cell lines KYSE30 and TE-1 treated with RT. Cell growth, colony formation and apoptosis were used to measure anti-proliferation effects. The method of RNA interference was used to investigate the role of insulin-like growth factor binding protein-3 (IGFBP-3) in ESCC cells radiosensitivity treated with Nimotuzumab. In vivo effect of Nimotuzumab on ESCC radiotherapy was done using a mouse xenograft model.

Results

Nimotuzumab enhanced radiation response of KYSE30 cells (with high EGFR expression) in vitro, as evidenced by increased radiation-inhibited cell growth and colony formation and radiation-mediated apoptosis. Mechanism study revealed that Nimotuzumab inhibited phosphorylated EGFR (p-EGFR) induced by EGF in KYSE30 cells. In addition, knockdown of IGFBP-3 by short hairpin RNA significantly reduced KYSE30 cells radiosensitivity (P<0.05), and even after the administration of Nimotuzumab, the RT response of IGFBP-3 silenced KYSE30 cells was not enhanced (P>0.05). In KYSE30 cell xenografts, Nimotuzumab combined with radiation led to significant tumor growth delay, compared with that of radiation alone (P=0.029), and also with IGFBP-3 up-regulation in tumor tissue.

Conclusions

Nimotuzumab could enhance the RT effect of ESCC cells with a functional active EGFR pathway. In particular, the increased ESCC radiosensitivity by Nimotuzumab might be dependent on the up-regulation of IGFBP-3 through EGFR-dependent pathway.

     

MSC-CNTF靶向性治疗变态反应性脑脊髓炎动物对病情和炎症的影响

目的探讨重组腺病毒转导的骨髓间充质细胞（MSC）导向的睫状神经营养因子（CNTF）基因靶向性治疗C57小鼠实验性变态反应性脑脊髓炎（EAE）,对动物病情和和炎症的影响。方法用成功构建的Ad-CNTF-IRES-GFP载体,体外观察Ad-CNTF-IRES-GFP转染MSC（MSC-CNTF）对MSC表达CNTF的影响,再用MOG35-55建立C57BL/6小鼠的EAE模型,将MSC-CNTF移植治疗EAE小鼠,观察EAE动物病情评分;ELISA法观察外周血的促炎症因子TNF-α、IFN-γ,IL-12p35和抗炎症因子IL-10的水平,HE染色观察脑和脊髓炎症细胞浸润,用免疫荧光和免疫组化观察病变部位CD3（＋）T细胞、炎性细胞因子TNF-α、IFN-γ的表达水平。结果（1）MSC-CNTF在MOI等于100的情况下,分泌的CNTF较DIL染色的MSC（MSC-DIL）、或Ad-eGFP转染的MSC（MSC-GFP）增高20倍（P〈0.01）。（2）MSC-CNTF治疗的EAE小鼠病情显著减轻。不仅平均发病时间缩短,发病率下降,病情减轻,而且病情严重程度也明显减轻。（2）MSC-CNTF治疗明显减低EAE小鼠外周血TNF-α和IFN-γ水平,明显升高外周血IL-10水平。（3）MSC-CNTF治疗EAE小鼠可明显降低病变部位的炎症细胞数量和炎性细胞因子TNF-α水平,而对IFN-γ表达几乎影响不大。结论MSC-CNTF移植治疗的EAE动物病情明显减轻,其机制是：减低外周血TNF-α和IFN-γ水平,降低病变部位CD3（＋）细胞数量和炎性细胞因子TNF-α水平。