Effects of antisense oligonucleotides targeting VEGF on radio sensitivity of uterine cervix cancer Hela cells

Objective： To determine the impact of antisense oligonucleotides targeting vascular endothelial growth factor （VEGF） on radiosensitivity of uterine cervix cancer Hela cells. Methods： VEGF antisense oligodeoxynucleotides （ASODN） was transfected into Hela cells by liposome-mediated method. Cells transfected with the oligodeoxynuclecotide and saline were used as control groups. Cells were irradiated by 6 MV X ray at the dose of 0 Gy, 2 Gy, 4 Gy and 6 Gy respectively. The expression of VEGF mRNA was determined by RT-PCR. Apoptosis were evaluated using FCM. Cloning efficiency was determined by colony formation assay. Results： The expression of VEGF mRNAwas inhibited by ASODN （P 〈 0.01） in Hela cells. The inhibited activation which was influenced by radiation resulted in increasing apoptosis （P 〈 0.01） and inhibiting plating efficiency （P 〈 0.01）. Conclusion： The expression of VEGF induced by X irradiation in Hela cells can be blocked by VEGF ASODN. Treatment with VEGF might increase apoptosis in HeLa cells and enhance radiosensitivity.     

The Hallmarks of Cancer and the Radiation Oncologist: Updating the 5Rs of Radiobiology

A comprehensive, mechanistic understanding of radiobiological phenomena that can be integrated within the broader context of cancer biology offers the prospect of transforming clinical practice in radiation oncology. In this review, we revisit the six established biological hallmarks of cancer and examine how they have provided insights into novel therapeutic strategies. In addition, we discuss the potential of two emerging hallmarks to continue to expand our understanding beyond the narrow confines of the traditional 5Rs of radiobiology.     

咪喹莫特抑制小鼠体表黑色素瘤生长和促使肿瘤特异性放疗增敏的实验研究

目的研究咪喹莫特联合外放疗对C57BL/6小鼠黑色素瘤模型肿瘤生长的影响,并判定咪喹莫特对肿瘤特异性放疗增敏的疗效。方法建立C57BL/6小鼠(n=20)的体表黑色素瘤模型,随机分为对照组、放疗组、咪喹莫特治疗组和放疗联合咪喹莫特治疗组。测量肿瘤生长体积,观察肿瘤生长延迟效应,并采用免疫组织化学检测和生存曲线评估咪喹莫特对肿瘤放疗增敏效果。结果放疗联合咪喹莫特治疗组肿瘤生长延迟最显著,其次治疗效果依次为咪喹莫特治疗组、放疗组和对照组。4组间两两比较,差异均有显著性意义(P<0.01)。C57BL/6小鼠黑色素瘤的组织病理学检测中,发现微管相关蛋白1轻链3B(MAP1LC3B)和α-平滑肌激动蛋白(α-SMA)在放疗联合咪喹莫特治疗组呈高表达。3种不同治疗组的MAP1LC3B和α-SMA表达与对照组间比较,差异均有显著性意义(P<0.05)。C57BL/6小鼠生存表证实,放疗联合咪喹莫特治疗组生存率明显优于其他治疗组和对照组,差异有显著性意义(P<0.05)。结论咪喹莫特联合外放疗对C57BL/6小鼠黑色素瘤生长有绝对延迟效应,咪喹莫特可为新的肿瘤特异性放疗增敏诱导剂。     

靶向NBS1的RNA干扰对鼻咽癌CNE-2细胞放射增敏作用的研究

目的:研究靶向NBS1的RNA干扰对鼻咽癌CNE-2细胞的放射增敏作用。方法:靶向NBS1的小干扰RNA转染鼻咽癌CNE-2细胞48小时后,4Gy剂量X射线照射。照射后24小时,流式细胞仪分析鼻咽癌细胞凋亡率。结果:X线照射+RNAi-NBS1组的细胞凋亡率显著高于单独X线照射组和X线照射+RNAi-阴性对照组(P<0.01)。结论:靶向NBS1的RNA干扰在鼻咽癌CNE-2细胞中有显著的放射增敏作用,诱导细胞凋亡是其放射增敏的重要机制之一。     

Adenovirus-mediated expression of Tob1 sensitizes breast cancer cells to ionizing radiation

Aim： To investigate the effect of the Tob1 gene, a member of the Transducing Molecule of ErbB2/B-cell Translocation Ggene （TOB/BTG） family, by using the adenovirus-mediated expression of Tobl on radiosensitivity in a human breast cancer cell line MDA-MB-231. Methods： Cell survival was determined by clonogenic assay. Apoptosis was evaluated by DNA fragmentation gel electrophoresis and terminal deoxynucleotidyl transferase-mediated nick end labeling assay. Protein expression was analyzed by Western blot assay and DNA repair was measured by a host cell reactivation assay. Results： We demonstrated that pre-irradiation treatment with Ad5-Tob1 significantly increased radiosensitivity, accompanying the increased induction of apoptosis and the repression of DNA damage repair. Furthermore, Ad5-Tob1-mediated radiosensitivity correlates with the upregulation of the pro-apoptotic protein Bax and the downregulation of several DNA double strand break repair proteins, including DNA-dependent protein kinases, Ku70 and Ku80, and X-ray-sensitive complementation group 4. Conclusion： Tob1, as a new radiosensitizer, is a new target in the radiotherapy of breast cancer via increasing apoptosis and suppressing DNA repair.     

Radiation-induced DNA damage and repair in peripheral blood mononuclear cells from Nijmegen breakage syndrome patients and carriers assessed by the Comet assay

Nijmegen breakage syndrome (NBS) patients and carriers are predisposed to malignancy and are often treated with X-irradiation. In the present study, the single-cell gel electrophoresis (Comet) assay was used to examine radiation-induced DNA damage and repair in peripheral blood mononuclear cells from NBS patients (n=13) and carriers (n=36) of six unrelated families. Cells from apparently healthy donors (n=10) and from breast cancer patients with normal clinical radiosensitivity (n=10) served as controls. Cells were irradiated with 5 Gy of X-rays and assayed for initial DNA damage and for residual DNA damage after 40 min of repair; the kinetics of DNA repair also was estimated. In addition, the nuclear area of unirradiated cells was extracted from the Comet data. The initial radiation-induced DNA fragmentation indicated that cells from members of two out of six NBS families were significantly more sensitive to X-irradiation than cells from the controls. Cells from four NBS families had longer DNA repair half-time values, while cells from five NBS families had significantly increased residual DNA damage following repair. The mean nuclear area of unirradiated cells processed in the Comet assay was 1.3-fold higher in cells from all NBS families than in the controls (P<0.05). Notably, the Comet assay parameters (initial and residual DNA damage and the repair kinetics) of irradiated NBS cells predicted the carrier status of the majority (86%) of blindly tested individuals. The prediction of NBS status was higher if the nuclear area of unirradiated cells was used as the endpoint. The results of this study suggest that the impaired radiation response of NBS cells should be taken into account if radiotherapy of NBS patients and carriers is required.     

MnSOD表达特性与食管癌细胞放射敏感性的关系

目的:研究锰超氧化物歧化酶(manganese superoxide dismutase,MnSOD)在一氧化氮(mtric oxide,NO)作用下对食管癌TE-1细胞放射敏感性的影响。方法:采用慢病毒转染法将MnSOD重组质粒转染食管癌TE-1细胞,分别建立中、高水平过量表达MnSOD的稳定转染细胞(TE-1Mm、TE-1Mh)及空载体细胞(TE-1Mn)。进一步采用RT-PCR、Western blot检测转染后TE-1细胞、空载体TE-1Mn细胞、稳定转染细胞TE-1Mm及TE-1Mh中MnSOD mRNA和蛋白的表达;四甲基偶氮唑蓝(MTT)比色实验评价NO供体硝普钠(sodium nitroprusside,SNP)、放射线及二者联合对以上4组细胞的抑制;流式细胞术(FCM)及Western blot检测各组细胞凋亡、活性氧(reactive oxygen species,ROS)荧光强度及蛋白表达的变化。结果:成功建立了稳定中、高水平过量表达MnSOD的TE-1细胞株TE-1Mm和TE-1Mh;RT-PCR及Western blot证实上述2种细胞中含有不同表达水平的MnSOD。MTT法及FCM显示,TE-1Mm细胞的存活率下降,细胞凋亡率上调,而TE-1Mh细胞的存活率上调,细胞凋亡率下降,相应组间相比差异具有统计学意义(P0.05)。0.5～2mmol/L SNP及2、4、6Gy放射线处理48 h,TE-1Mm细胞生长抑制率明显增加,细胞生长明显减慢,放射增敏比增加,细胞凋亡率上调;TE-1Mh细胞抑制率反而降低,细胞生长加速,放射增敏比降低,细胞凋亡率下降,相应组间相比差异均具有统计学意义(P0.05)。Western blot结果及FCM显示:0.5～2 mmol/L SNP及2、4、6 Gy放射线处理48 h,与TE-1Mm细胞相比,TE-1Mh细胞MnSOD蛋白表达量及细胞内ROS相对降低,相应组间相比差异均具有统计学意义(P0.05)。结论:中等水平过量表达MnSOD增加食管癌细胞放射敏感性,抑制增殖,诱导凋亡,而高水平过量表达MnSOD则与之相反,为今后通过MnSOD提高放射敏感性来抑制食管癌或通过其抗拒放射来保护正常细胞提供了可能性。