Antimutagenicity and Antioxidative DNA Damage Properties of Oligomeric Proanthocyanidins from Thai Grape Seeds in TK6 Cells

Oligomeric proanthocyanidins (OPCs) are found mostly in red grape seeds. Many publications have reportedthat OPCs possess an excellent anti-oxidant effects. Since it could against cellular damage from reactive oxygenspecies (ROS) led to reduce the risk of chronic disease and cancers. We carried out this study on the ThaiOPCs to evaluate the mutagenicity/ anti-mutagenicity and anti-oxidative DNA damage effects in TK6 cells bymicronucleus (MN) and comet assays. In the MN assay, OPCs-treatment of TK6 cells at concentrations rangingfrom 10-200 μg/ml (4 and 24 h) did not cause micronucleus induction over the negative control group but revealeda significant reduction the micronucleus frequencies against the known mutagen (mitomycin C). In the cometassay, OPCs-treated TK6 cells at concentrations of 100, 250, 500, and 1,000 μg/ml could inhibit DNA damageinduced by H2O2 as indicated by 18.7, 36.4, 30.6, and 60.1%, respectively. Our results suggest that OPCs possessthe anti-mutagenic and anti-oxidative DNA damage effects in TK6 cells under the conditions of this assay.     

The increment of micronucleus frequency in cervical carcinoma during irradiation in vivo and its prognostic value for tumour radiocurability.

A potential usefulness of micronucleus assay for prediction of tumour radiosensitivity has been tested in 64 patients with advanced stage (II B-IV B) cervical carcinoma treated by radiotherapy. The study of cellular radiosensitivity in vitro was conducted in parallel with the study of cellular damage after tumour irradiation in vivo. Radiosensitivity of in vitro cultured primary cells isolated from tumour biopsies taken before radiotherapy was evaluated using cytokinesis-block micronucleus assay. Frequency of micronuclei per binucleated cell (MN/BNC) at 2 Gy was used as a measure of radiosensitivity. Radiation sensitivity in vivo was expressed as per cent increment of micronucleus frequency in cells isolated from biopsy taken after 20 Gy (external irradiation, 10 x 2 Gy) over the pre-treatment spontaneous micronucleus level and was called MN20. Very low correlation (r = 0.324) was observed between micronucleus frequency in vitro and in vivo. Although micronucleus frequency at 2 Gy differed widely between tumours evaluated (mean MN/BNC was 0.224; range 0.08-0.416), no significant correlation was observed between this parameter and clinical outcome. The average increment of micronucleus frequency after 20 Gy amounted to 193% of spontaneous level (range 60-610%) and was independent of spontaneous micronucleation before radiotherapy. In contrast to in vitro results, these from in vivo assay seem to have a predictive value for radiotherapy of cervix cancer. The micronucleus increment in vivo that reached at least 117.5% of pretreatment value (first quartile for MN20 data set) correlated significantly with better tumour local control (P < 0.008) and overall survival (P < 0.045). Our results suggest that evaluation of increment of micronucleus frequency during radiotherapy (after fixed tested dose of 20 Gy) offers a potentially valuable approach to predicting individual radioresponsiveness and may be helpful for individualization of treatment strategy in advanced stage cervical cancer.     

Anticancer Potential of Cratoxylum formosum Subsp. Pruniflorum (Kurz.) Gogel Extracts Against Cervical Cancer Cell Lines

Background: Most northeast Thai vegetables may play roles in human health by acting as antioxidant andanticancer agents. Recent study showed that Cratoxylum formosum subsp. pruniflorum (Kurz.) Gogel. (Teawdang)could inhibit growth of liver cancer cell lines. Cervical cancer, which has human papilloma virus as its maincause, is found at high incidence in Thailand. Due to increasing drug resistance, searches for potential anticancercompounds from natural source are required. Therefore, our purpose was to evaluate the cytotoxicity of Teawdangextracts in cervical cancer cell lines. Materials and Methods: Teawdang edible parts, purchased from KhonKaen market during July-October 2013 was extracted with organic solvent. Phenolic profiles of crude hexane(CHE), ethyl acetate (CEE), methanol (CME) and water (CWE) extracts were performed by high performanceliquid chromatographic (HPLC) techniques. Their cytotoxic effects on cervical cancer cells were investigatedwith HPV-non infected (C-33A) and HPV-infected (HeLa and SiHa) cell lines. Results: HPLC profiles showedthat all crude extracts contained caffeine, ferulic acid and resveratrol. CME and CEE had high contents of gallicacid and quercetin. Catechin was found only in CWE. Cytotoxicity test showed that CEE had the lowest IC50on HeLa (143.18±13.35 μg/mL) and SiHa cells (106.45±15.73 μg/mL). C-33A cells were inhibited by CWE (IC50= 130.95±3.83 μg/mL). Conclusions: There were several phenolic compounds in Teawdang extracts which mayhave cytotoxic effects on cervical cancer cell lines. Investigation of these bioactive compounds as new sources ofanticancer agents is recommended.     

基于MATLAB和HTML混合编程的多幅图像显示与校验

CB法微核图像自动分析系统的研究过程中,至少要有一千幅甚至几千幅双核细胞图像需要人工检视,并对每个双核细胞内所含的微核个数进行核对,以便进行系统性能测试和结果对比分析。本文采用MATLAB和HTML语言混合编程的方法 ,首先将自动识别出的双核细胞图像及其内部微核个数等信息自动写进HTML的表单中,并将多幅双核细胞图像和分析结果以网页的形式显示,然后再利用由VBScript编写的嵌入网页中的脚本程序实现每幅细胞图像中微核识别结果的校验。这为采用MATLAB语言显示多幅图像提供了一种可行的方法 ,对于采用GUI设计其它图像分析软件也是有益的。     

沙利度胺对人肝癌细胞株SMMC-7721体外生长的抑制作用

目的 研究沙利度胺对人肝癌细胞株SMMC-7721体外生长的抑制作用及其可能的机制.方法 将不同浓度的沙利度胺作用于人肝癌细胞株SMMC-7721,采用四甲摹偶氮唑蓝(MTT)法检测沙利度胺对SMMC-7721细胞的增殖抑制作用.将SMMC-7721细胞培养至对数生长期,采用DNA琼脂糖凝胶电泳、荧光显微镜观察、流式细胞仪检测等方法 观察沙利度胺处理后SMMC-7721细胞的凋亡梯度、形态学变化和凋亡率,并对凋亡调控蛋白caspase-3的表达进行测定.采用酶联免疫吸附(ELISA)法测定不同浓度的沙利度胺处理后SMMC-7721细胞表达血管内皮生长因子(VEGF)的变化.结果 沙利度胺的浓度从3.125μg/ml增至200μg/ml时,其对SMMC-7721细胞的增殖抑制率从11.7%增至34.2%;当沙利度胺的浓度>25 μg/ml时,其对SMMC-7721细胞的增殖抑制作用明显强于空白对照组(P<0.05).200 μg/ml的沙利度胺处理SMMC-7721细胞24 h后,行琼脂糖凝胶电泳,可见到DNA梯形条带;48 h后梯形条带更明显,并且在荧光显微镜下可见SMMC-7721细胞出现核固缩和核裂解现象.200μg/ml的沙利度胺处理SMMC-7721细胞12、24、48和72 h时,碘化丙啶(PI)法检测SMMC-7721细胞的凋亡率分别为3.1%±0.5%、8.4%±1.3%、19.4%±3.5%和25.8%±2.1%,24 h起的凋亡率均明显高于空白对照组SMMC-7721细胞48 h的自然凋亡率(1.6%±0.6%,均P<0.05).50、100和200μg/ml的沙利度胺处理SMMC-7721细胞48 h时,Annexin V-FITC/PI双标法检测SMMC-7721细胞的凋亡率分别为8.7%±1.2%、16.8%±2.5%和25.4%±4.5%,均明显高于空白对照组SMMC-7721细胞48 h的自然凋亡率(2.1%±0.5%,均P<0.05).随着沙利度胺浓度的增加,表达caspase-3蛋白的SMMC-7721细胞数量不断增加,而SMMC-7721细胞中VEGF的含量却逐渐下降.结论 沙利度胺可能通过诱导肝癌细胞的凋亡、抑制肿瘤血管的生成而发挥双重抗肿瘤生长的作用.     

Cytotoxic Activity of Propolis Extracts from the Stingless Bee Trigona Sirindhornae Against Primary and Metastatic Head and Neck Cancer Cell Lines

Background: Propolis, a resinous substance produced by the honeybee, has a wide spectrum of potent biologicalactivities. However, anti-cancer activity of propolis obtained from Trigona sirindhornae, a new species of stinglessbee, has not yet been reported. This study concerned cytotoxicity of propolis extracts from T. sirindhornae against twohead and neck squamous cell carcinoma (HNSCC) cell lines. Materials and Methods: A dichloromethane extractof propolis (DMEP) was prepared generating 3 fractions: DMEP-A, DMEP-B, and DMEP-C. Genetically-matchedHNSCC cell lines derived from primary (HN30) and metastatic sites (HN31) in the same patient were used to studycytotoxic effects of the DMEPs by MTT assays. The active compounds in the DMEPs were analyzed by reversephasehigh performance liquid chromatography. Results: DMEP-A exhibited cytotoxic activity on HN30 cells withsignificantly decreased viability at 200 μg/ml compared with the control (p<0.05). However, no significant cytotoxiceffect was evident in HN31 cells. DMEP-B and DMEP-C significantly decreased the viability of both cell lines from100–200 μg/ml and 50–200 μg/ml, respectively (p<0.05). Interestingly, HN31 cells were more toxically sensitivecompared with the HN30 cells when treated with DMEP-B and DMEP-C. IC50 values for DMEP-B with HN30 andHN31 cells were more than 200 μg/ml and 199.8±1.05 μg/ml, respectively. The IC50 of DMEP-C to HN30 and HN31cells was found to be 114.3±1.29 and 76.33±1.24 μg/ml, respectively. Notably, apigenin, pinocembrin, p-coumaric acid,and caffeic acid were not detected in our propolis extracts. Conclusion: T. sirindhornae produced propolis displayscytotoxic effects against HNSCC cells s. Moreover, DMEP-B and DMEP-C differentially inhibited the proliferationof a metastatic HNSCC cell line.     

三氯生的体外遗传毒性评价

目的：应用体外碱性彗星试验、体外胞质分裂阻滞微核细胞组学试验和细菌回复突变试验（Ames试验）评价三氯生（TCS）的体外遗传毒性。方法：采用TK6人淋巴母细胞进行体外彗星试验和体外微核试验,共设定5个剂量（3.5、8.8、17.5、26.3和35 μmol/L）组。同时,应用鼠伤寒沙门氏菌TA98、TA100和DNA修复酶缺陷型YG7108（Ogt^-/Ada^-）菌株对TCS进行Ames试验,共设定8个剂量（0.000 5、0.001 67、0.005、0.016 7、0.05、0.167、0.5和1.67 μg/皿）组。结果：与对照组比较,彗星试验结果显示,TCS在各个剂量下均能引起TK6细胞的尾长、尾部DNA强度和尾矩的增加,差异均具有统计学意义（P均 < 0.01）,且尾部DNA强度呈现剂量效应（r=0.943,P=0.017）;微核试验表明,TCS未引起TK6细胞微核率升高（P > 0.05）;Ames试验结果表明TCS未引起TA98、TA100和YG7108菌株的回复突变菌落数增加（P > 0.05）。结论：TCS主要引起细胞DNA损伤,但未对TK6细胞造成染色体损伤,对Ames试验菌株不具有致突变作用。     

体内碱性彗星试验与微核试验联合测定甲磺酸乙酯遗传毒性方法的建立

目的：建立彗星试验与微核试验结合的检测方法,应用其评价甲磺酸乙酯（EMS）对大鼠的遗传毒性。方法：试验设置阴性对照组（0.9%的生理盐水）及EMS低（50 mg/kg）、中（100 mg/kg）、高（200 mg/kg）剂量组,共4组,每组5只雄性SD大鼠,分别在0、24、48和69 h经口灌胃1次性给予受试物,在末次给药后3 h进行外周血采样,外周血经固定、洗脱、荧光染色后进行微核试验采用流式细胞术测定网织红细胞微核率;然后处死动物进行肝脏、肾脏和胃组织的取样,样品经单细胞悬液制备、制片、裂解、解旋电泳及染色后,应用Comet Assay IV软件进行彗星试验数据的收集。结果：彗星试验中,50、100和200 mg/kg EMS染毒组肝脏、肾脏和胃的尾DNA百分含量与阴性对照组相比均显著增加,且呈剂量反应关系（r=0.93,P<0.05）;微核试验中,EMS高剂量组骨髓毒性太大未收集到足够的细胞用于检测,仅EMS中剂量组外周血网织红细胞微核率较对照组显著增加（P<0.01）。结论：初步建立了大鼠多靶器官彗星试验方法;在同一试验中彗星试验和微核试验联合可节省动物数量,提高试验效率。     

沙利度胺对人宫颈癌Hela细胞周期的影响及其作用机制

目的:探讨沙利度胺对人宫颈癌Hela细胞周期的影响及其作用机制。方法:沙利度胺（50 μg/ml、100 μg/ml和200 μg/ml）处理人宫颈癌Hela细胞24 h后,运用CCK-8法检测细胞活力;采用流式细胞术检测细胞周期分布;采用蛋白免疫印迹法检测Cyclin E、p53、p-p53和GAPDH蛋白表达水平。结果:沙利度胺(50 μg/ml、100 μg/ml和200 μg/ml)作用Hela细胞24 h后,细胞活力显著降低,且呈浓度依赖性(P<0.05)。50 μg/ml、100 μg/ml和200 μg/ml沙利度胺作用24 h后,Hela细胞G0/G1期细胞数明显增加,S期细胞数显著减少(P<0.05)。此外,沙利度胺作用Hela细胞24 h可明显下调Cyclin E蛋白的表达(P<0.05)。沙利度胺处理Hela细胞24 h后,p-p53/p53比值明显增大(P<0.05)。结论:沙利度胺可能通过促进p53蛋白的活化从而抑制人宫颈癌细胞周期进程。     

Free Radical Scavenging Properties of Annona squamosa

Annona squamosa has extensively been used in the traditional and folkloric medicine and found to possess many biological activities. Different solvents, petroleum ether, chloroform, ethyl acetate and methanol extracts of Annona squamosa seeds (ASPE, ASCH, ASEA, ASME) have been used to prepare plant extracts. The present investigations dealt with the free radical scavenging activity of four extracts using various techniques such as total reducing power estimation, total phenolic count, 1,1-diphenyl-2-picryl hydrazyl (DPPH) radical scavenging effect, evaluation of ABTS cation decolorisation capacity, FRAP assay, hdroxyl radical scavenging assay, super oxide assay and Nitric oxide radical scavenging assay of the extracts. The results showed that the four extracts of Annona squamosa showed significant reducing power in four extracts. The total phenolic contents in petroleum ether, chloroform, ethyl acetate, methanol extracts and positive control were 0.64±0.17, 0.54±0.27, 0.49±0.24, 0.57±0.22 and 0.66±0.33. The antioxidant capacity by ABTS assay of ASPE, ASCH, ASEA, ASME and positive control, trolox showed 77.75±0.5,73.25±1.7,78.5± 1.2 , 80 ± 0.8 μg/ml and 94.2 ± 0.9 respectively. The (50 % scavenging activity) SA50 of ASPE and ASCH, ASEA and ASME was found to be 34.4 μg/ml, 43.8 μg/ml 34.7 μg/m and 28.8 μg/ml respectively by DPPH assay. The percentage of hydroxyl radical scavenging increased with the increasing concentration of the extracts. ASPE, ASCH, ASEA and ASME showed superoxide radical scavenging activity, as indicated by their values 66 ± 0.5, 68 ± 1 ,63 ± 1 and 70 ± 0.5 μg/ml respectively compared to gallic acid which was 97 ± 0.5 μg/ml. The values for scavenging of nitric oxide for ASPE, ASCH, ASEA and ASME were 91.0 ± 1.0, 66.75 ± 0.5, 71.75 ± 1.1 and 75.75 ± 1.15 μg/ml while value for standard ascorbic acid was 91.0 ± 1.0 μg/ml. The results revealed strong antioxidants in four extracts may lead to the development of potent antioxidant agents from Annona squamosa seeds.     

β-榄香烯联合X线照射对肺腺癌A549细胞DNA损伤及修复影响

目的 探讨β-榄香烯联合放射对肺腺癌A549细胞DNA损伤及修复影响.方法 10、20 μg/mlβ-榄香烯作用于肺腺癌A549细胞24 h后进行X线照射.通过克隆形成实验观察β-榄香烯对A549细胞放射敏感性影响,采用彗星分析法探讨损伤修复的机制.结果 克隆形成实验结果显示β-榄香烯能增加A549细胞放射敏感性,10 μg/ml和20 μg/mlβ-榄香烯联合照射组的放射增敏比分别为1.55和1.64(D0值比)及1.43和1.75(Dq值比).20 μg/mlβ-榄香烯联合X线照射组与单独照射组和单独药物组0、2、6、24 h的彗星尾力矩相比有所增加,分别为7.16±2.61与0.95±0.65和1.81±1.23(F=231.24,P<0.01)、3.65±2.06与0.11±0.07和1.58±1.40(F=90.22,P<0.01)、2.09±0.83与0.1±0.05和0.45±0.25(F=238.44,P<0.01)、1.45±1.37与0.11±0.08和0.60±0.40(F=38.94,P<0.01),表明β-榄香烯与放射线联合对A549细胞DNA损伤增加和修复的抑制作用.结论 β-榄香烯对A549细胞放射敏感性的影响可能与其对DNA放射损伤及修复作用有关.

Abstract：

Objective To study if β-elemene can increase radiation-induced deoxyribonucleic acid (DNA) damage and decrease the damage repair.Methods Exponentially growing human lung adenocarcinoma cells (A549) were exposed to 10 or 20 μg/ml β-elemene for 24 h before irradiation.The effect of β-elemene on the in vitro radiosensitivity of A549 cells was evaluated using clonogenic assay.DNA damage and repair were evaluated using comet assay.Results Exposure to β-elemene before irradiation increased the radiosensitivity of A549 cells.The SERD0 for 10 μg/ml and 20 μg/ml β-elemene was 1.55 and 1.64, respectively.The SERDq for 10 μg/ml and 20 μg/ml β-elemene was 1.43 and 1.75, respectively.Combined treatment, comparing to irradiation or β-elemene treatment alone, induced higher levels of DNA damage and slower rate of damage repair.A549 cells exposed to 20 μg/ml β-elemene followed by irradiation showed a higher levels of tail moment (TM) than those exposed to irradiation or β-elemene alone at 0 h,2 h,6 h and 24 h after irradiation.The TM of the three groups at 0 h,2 h,6 h and 24 h after irradiation was 7.16±2.61,0.95±0.65 and 1.81±1.23(F=231.24,P<0.01), 3.65±2.06,0.11±0.07 and 1.58±1.40(F=90.22,P<0.01), 2.09±0.83,0.1±0.05 and 0.45±0.25(F=238.44,P<0.01), 1.45±1.37,0.11±0.08 and 0.60±0.40(F=38.94,P<0.01), respectively. Conclusions β-elemene can enhance the radiosensitivity of A549 cells through the enhancement of DNA damage and the inhibition of DNA damage repair.     

Radiosensitization Effects of a Zataria multiflora Extract on Human Glioblastoma Cells

Background: Although radiotherapy is one of the most effective strategies in the treatment of cancers, it isassociated with short and long term side effects on normal tissues. Zataria multiflora Boiss (Laminacea) (ZM)has several biological properties such as antioxidant and anti-inflammation activities.Here we investigated cellkilling effects of a hydroalcoholic Zataria multiflora extract on cell death induced by ionizing radiation in a humanglioblastoma cell line (A172) and human non-malignant fibroblasts (HFFF2) in vitro. Materials and Methods: A172and HFFF2 cells were treated with a hydroalcoholic extract of dried aerial parts of Zataria multiflora at differentconcentrations (25, 50, 100, 150 and 200 μg/ml) and then exposed to ionizing radiation (IR). Cell proliferationand DNA fragmentation were evaluated. Thymol content in the extract was analyzed and quantified by HPLCmethods. Results: A172 cell proliferation was significantly inhibited by ZM. The percentage cell survival was91.8±8.57 for cells treated with 200 μg/ml of ZM extract alone while it was 76.0±4.27 and 66.2±8.42 for cellstreated with ZM and exposed to IR at doses of 3Gy and 6Gy, respectively. Radiation-induced apoptosis in A172cells was significantly increased following treatment with ZM at doses of 200 μg/ml. ZM extract did not exhibitany enhanced cell killing effects and apoptosis caused by IR on HFFF2 cells. Conclusions: These data showselective radiosensitization effects of ZM in A172 cells apparently due to increased radiation-induced apoptosis.     

大黄素和大黄酸的体外遗传毒性评价

目的：评价大黄素和大黄酸的体外遗传毒性。方法：使用不同浓度的大黄素和大黄酸（均分别为20、40、80、120μg/ml）处理人的类淋巴母细胞WTK1后,进行彗星实验、体外微核试验和TK基因突变试验。并设溶剂对照组和甲基甲烷磺酸（mthylmethane sulfonate,MMS）阳性对照组。结果：大黄素80和120μg/ml剂量组TK基因突变频率、细胞拖尾率及平均尾长均增高（P〈0.05）;大黄酸120μg/ml剂量组TK位点总突变频率增高（P〈0.05）。结论：在本实验条件下,大黄素和大黄酸均表现出弱致突变作用。     

CHO细胞体外微核高内涵筛选方法的建立及应用

目的：探讨微核试验高内涵筛选方法在遗传毒性评价中的应用价值,建立简便、经济、快速、高效的微核试验高通量筛选方法。应用微核高内涵筛选方法对已知的遗传毒性阳性物质进行验证分析。方法：选择8种典型的断裂剂或非整倍体诱导剂进行试验。应用Cellomics ArrayScan高内涵筛选系统检测药物诱发的CHO细胞微核发生率。结果：苯并芘在25.15和50.3μg/ml可诱发微核显著增加（P〈0.05）;依托泊苷在0.16～20μg/ml浓度范围均可诱发微核发生率显著增加（P〈0.01）,在1.25～20μg/ml浓度,细胞存活率低于50%;2-氨基芴在0.1～1.26μg/ml的浓度范围内微核发生率和阴性对照组相比差异均无统计学意义（P〉0.05）;2-氨基蒽在6.44、12.88、25.75和51.5μg/ml,秋水仙素在0.08～20.4μg/ml,甲基磺酸乙酯在0.754、1.509和3.018 mg/ml,氯化铬在2.344μg/ml,甲基磺酸甲酯在0.101 mg/ml浓度时,均可诱发CHO细胞微核发生率显著增加（P〈0.05或P〈0.01）。结论：微核高内涵筛选方法可以满足快速、高效、经济、可靠检测微核的要求。     

多细胞系胞质分裂阻滞微核细胞组学试验法的建立与应用

目的: 采用不同组织来源的细胞系建立微核细胞组学,比较其遗传毒性生物标志物的敏感性,并研究雷公藤甲素(TPL)和甘草酸二铵(DG)对大鼠成纤维肺细胞(CHL)和犬肾小管上皮细胞系(MDCK)细胞微核率的影响。方法:首先建立方法,采用不同浓度丝裂霉素C(MMC)与CHL、L02、MDCK、Bhas42细胞作用6 h后给予细胞松弛素B(CytoB)继续培养约1.5个细胞倍增时间。收获细胞、制片、染色及镜检。计算每个剂量胞质分裂增殖指数(CBPI)、复制指数(RI)、坏死(Nec)及凋亡(Apop)细胞的千分率、双核细胞中微核(MN)、核芽(NBUD)和核质桥(NPB)出现的千分率。然后进行方法的验证,采用CHL细胞经不同浓度(1、10和100 μg/mL)DG预处理24 h后观察DG对上述细胞毒性和微核细胞组相关指标的影响。为进一步了解微核细胞组学对遗传毒性靶器官评估的效果,MDCK细胞经DG(10 μg/mL)预处理24 h后采用不同浓度(1和5 μg/mL)TPL染毒1 h,观察其对遗传毒性的影响。结果:MMC诱发的不同细胞系细胞毒性(CBPI、RI、Apop与Nec)均随MMC浓度升高呈升高趋势。各组细胞的MN、NBUD和NPB均出现不同程度的剂量相关性升高,但不同细胞系对MMC诱发的生物标志物峰值出现的敏感度不同。在验证实验中,与对照组相比,DG(10和100 μg/mL)对MMC(0.2 μg/mL)诱发的MN、NBUD和NPB升高有显著的抑制作用(P均<0.05)。TPL可诱发MDCK细胞NBUD和MN显著上升(P<0.05),且DG预处理可有效拮抗该效应(P<0.05)。结论:多细胞系胞质分裂阻滞微核细胞组学实验法可同时对多项遗传毒性指标进行评价。DG对CHL及MDCK细胞产生的染色体断裂有保护作用,在临床配伍中可发挥抗细胞DNA损伤的功效。     

Anticancer Effects of Curcuma C20-Dialdehyde against Colon and Cervical Cancer Cell Lines

Background: Recent attention on chemotherapeutic intervention against cancer has been focused ondiscovering and developing phytochemicals as anticancer agents with improved efficacy, low drug resistanceand toxicity, low cost and limited adverse side effects. In this study, we investigated the effects of CurcumaC20-dialdehyde on growth, apoptosis and cell cycle arrest in colon and cervical cancer cell lines. Materials andMethods: Antiproliferative, apoptosis induction, and cell cycle arrest activities of Curcuma C20-dialdehydewere determined by WST cell proliferation assay, flow cytometric Alexa fluor 488-annexin V/propidium iodide(PI) staining and PI staining, respectively. Results: Curcuma C20 dialdehyde suppressed the proliferation ofHCT116, HT29 and HeLa cells, with IC50 values of 65.4±1.74 μg/ml, 58.4±5.20 μg/ml and 72.0±0.03 μg/ml,respectively, with 72 h exposure. Flow cytometric analysis revealed that percentages of early apoptotic cellsincreased in a dose-dependent manner upon exposure to Curcuma C20-dialdehyde. Furthermore, exposure tolower concentrations of this compound significantly induced cell cycle arrest at G1 phase for both HCT116 andHT29 cells, while higher concentrations increased sub-G1 populations. However, the concentrations used in thisstudy could not induce cell cycle arrest but rather induced apoptotic cell death in HeLa cells. Conclusions: Ourfindings suggest that the phytochemical Curcuma C20-dialdehyde may be a potential antineoplastic agent forcolon and cervical cancer chemotherapy and/or chemoprevention. Further studies are needed to characterizethe drug target or mode of action of the Curcuma C20-dialdehyde as an anticancer agent.     

芒柄花黄素对白血病HL-60细胞增殖、凋亡和miR-21/PTEN/AKT信号通路的影响

目的:探讨芒柄花黄素对白血病HL-60细胞增殖、凋亡的影响及其机制。方法:以不同浓度（0、12.5、25、50、100和200 μg/ml）芒柄花黄素处理HL-60细胞24、48和72 h后，采用MTT法检测细胞存活率，并计算出IC50以筛选出合适的作用浓度。采用流式细胞仪检测芒柄花黄素对HL-60细胞周期和凋亡的影响，实时荧光定量PCR检测芒柄花黄素对HL-60细胞中miR-21表达的影响，免疫印迹法检测芒柄花黄素对HL-60细胞中PTEN、AKT和p-AKT蛋白表达的影响。结果:与0 μg/ml处理组相比，25、50、100和200 μg/ml芒柄花黄素处理24 h后HL-60细胞的存活率明显降低（P<0.05）；同时，12.5、25、50、100和200 μg/ml芒柄花黄素分别处理48 h和72 h后HL-60细胞的存活率明显低于0 μg/ml处理组（P<0.05）；芒柄花黄素作用HL-60细胞24、48和72 h后的IC50值分别为146.50 μg/ml、120.10 μg/ml和89.00 μg/ml。50、100和150 μg/ml芒柄花黄素处理后HL-60细胞在G0/G1期的百分比、细胞凋亡率和PTEN蛋白的表达水平均逐渐升高，而细胞在S期和G2/M期的百分比、miR-21的表达水平以及p-AKT蛋白的表达水平均逐渐降低，且与0 μg/ml相比，差异均有统计学意义（P<0.05），但HL-60细胞中AKT蛋白的表达水平与0 μg/ml组比较差异无统计学意义（P>0.05）。结论:芒柄花黄素可抑制白血病HL-60细胞增殖并诱导细胞凋亡，其作用机制可能与调控miR-21/PTEN/AKT信号通路有关。     

Impact of mitochondrial DNA on hypoxic radiation sensitivity in human fibroblast cells and osteosarcoma cell lines

The purpose of this study was to evaluate the impact of mitochondrial DNA (mtDNA) on radiation sensitivity under hypoxic conditions. The cell lines used were rho+ and rho0, which carry wild-type mtDNA and no mtDNA, respectively. The rho0 cells do not utilize oxygen because they lack the capacity to carry out oxidative phosphorylation. To confirm the role played by mtDNA in different cell lines, two types of cell line were used: human fibroblast and osteosarcoma cells. Radiosensitivity was evaluated by the colony formation assay, micronucleus (MN) formation assay and comet assay. Hypoxia lowered radiosensitivity in all three experiments for all four cell lines. Between rho+ and rho0 cells, no difference was found in the results from the colony formation assay and comet assay. However, higher MN formation was found in rho+ cells than in rho0 cells, not only under room air conditions in both the fibroblast and osteosarcoma cell lines, but also under hypoxic conditions. Therefore, although hypoxia lowers the radiosensitivity in general, the impact of mtDNA persists under hypoxic conditions.     

Genotoxic and Anti-Genotoxic Effects of Vanillic Acid Against Mitomycin C-Induced Genomic Damage in Human Lymphocytes In Vitro

Vanillic acid, a vegetable phenolic compound, is a strong antioxidant. The aim of the present study was todetermine its effects on mitomycin C-induced DNA damage in human blood lymphocyte cultures in vitro, bothalone and in combination with mitomycin C (MMC). The cytokinesis block micronucleus test and alkalinecomet assay were used to determine genotoxic damage and anti-genotoxic effects of vanillic acid at the DNAand chromosome levels. MMC induced genotoxicity at a dose of 0.25 μg/ml. Vanillic acid (1 μg/ml) significantlyreduced both the rates of DNA damaged cells and the frequency of micronucleated cells. A high dose of vanillicacid (2 μg/ml) itself had genotoxic effects on DNA. In addition, both test systems showed similar results whentested with the negative control, consisting of dimethyl sulfoxide (DMSO) in combination with vanillic acid (1μg/ml)+MMC. In conclusion, vanillic acid could prevent oxidative damage to DNA and chromosomes when usedat an appropriately low dose.     

去甲斑蝥素对人类胆管癌细胞系QBC939增殖和凋亡作用的实验研究

 【摘要】　目的　研究去甲斑蝥素（NCTD）对人类胆管癌细胞系QBC939的生长抑制作用,初步探讨其作用机制。方法　将NCTD作用于经体外培养的QBC939细胞系,分别采用四甲基偶氮唑蓝比色法（MTT法）,流式细胞术及免疫组织化学SP法进行检测。结果　0.125、0.75、2.5、10、120 μg／ml NCTD作用48 h,对QBC939细胞系均有抑制作用（P＜0.05）,且具有浓度依赖性,绘制浓度效应曲线,得出半数抑制浓度（IC50）为（3.66±1.14）μg／ml;在分别作用12、24、36、48、72 h后,生存率有随时间增加而下降的趋势（P＜0.05）。流式细胞术结果表明,随着NCTD浓度的增加,凋亡率逐渐增高,各浓度组分别为（8.6±0.4）%、（17.6±0.3）%、（22.9±0.4）%、（25.5±0.9）%、（31.1±1.5）%（P＜0.001）;质量浓度为2.5 μg／ml的NCTD作用QBC939细胞48 h后,出现G2／M期阻滞现象,NCTD与对照组分别为（14.1±1.0）%和（5.7±0.3）%（P＜0.05）。不同浓度的NCTD作用于QBC939细胞后,与对照组相比Caspase-3蛋白表达均增加。结论　NCTD具有抑制人类胆管癌细胞系QBC939增殖的作用,这种作用可能与诱导细胞凋亡,干扰细胞生长周期有关。