维生素E拮抗氯化汞致小鼠微核作用的研究

本文采用微核试验方法对维生素Ｅ（ＶｉｔＥ）拮抗氯化汞（ＨｇＣｌ２）致小鼠胸骨骨髓多染红细胞微核作用进行了研究，其主要结果有：（１）给小鼠ＶｉｔＥ５ｍｇ／ｋｇ体重及以上剂量时，可显著地降低ＨｇＣｌ２１．０ｍｇ／ｋｇ体重的致小鼠微核作用（Ｐ〈０．００１）；（２）在ＨｇＣｌ２（１．０ｍｇ／ｋｇ）给小鼠染毒前４ｈ和染毒后２ｈ内补充ＶｉｔＥ２０ｍｇ／ｋｇ体重时，均显示明显地拮抗微核作用（Ｐ〈０．００１～Ｐ〉     

环氧乙烷的遗传毒性研究IV.小鼠骨髓细胞染色体畸变亚急性试验

以４组小鼠各１０只为一组，分别给予ＥｔＯ吸入剂量为５４，１８０，５４０ｍｇ／ｍ＾３２ｈｒ／ｄ连续５天。观察了ＥｔＯ对小鼠骨髓细胞染色体畸变的畸变率，发现总畸变率分别为０．５，１．２，３．２，７．０％，高浓度（５４０ｍｇ／ｍ＾３）中等浓度（１８０ｍｇ／ｍ＾３）二组与对照组比较有统计显著性差异；而低浓度（５４ｍｇ／ｍ＾３）组与对照组比较无显著差异。结果表明ＥｔＯ使小鼠骨髓细胞分裂中的染色体畸变，主要为     

聚乙二醇修饰重组人白细胞介素－2的体内抗小鼠宫颈癌作用

聚乙二醇（ＰＥＧ－８０００）化学修饰重组人白细胞介素－２（ＰＥＧ－ｒＩＬ－２）显著地延长了ｒＩＬ－２的循环半衰期。本研究比较了不同剂量方案的ＰＥＧ－ｒＩＬ－２和ｒＩＬ－２的体内抗小鼠宫颈癌作用，采用局部用药抗移植的小鼠腹水瘤型和皮下实体瘤型宫颈癌Ｕ１４。结果表明：腹腔内注射一定剂量的ＰＥＧ－ｒＩＬ－２（４５００ＩＵ，ＱＤ×５）能较相同剂量的ｒＩＬ－２显著延长荷腹水瘤小鼠的存活期（２３．１±３．６天比１６．５±２．０天，Ｐ＜０．０１），但ＰＥＧ－ｒＩＬ－２剂量过低则无明显疗效。在小鼠皮下移植肿瘤的局部并于肿瘤移植后的第４天开始注射给药，不同剂量的ＰＥＧ－ｒＩＬ－２（１５００～１３５００ＩＵ，ＱＤ×５）对皮下移植瘤的生长抑制作用显著强于相同剂量的ｒＩＬ－２（Ｐ＜０．０１），且抗肿瘤作用呈剂量依赖效应。本研究提示，ＰＥＧ－ｒＩＬ－２的抗小鼠宫颈癌作用优于ｒＩＬ－２，对人宫颈癌的局部免疫治疗具有潜在的临床价值。     

聚乙二醇修饰重组人白细胞介素—2的体内抗小鼠宫颈癌作用

聚乙二醇（ＰＥＧ－６０００）化学修饰竽组人白细胞介素－２（ＰＥＧ－ｒＩＬ－２）显地延长了ｒＩＬ－２的循不半衰期。本研究比较了不同剂量方案析ＰＥＧ－ｒＩＬ－２和ｒＩＬ－２的体内抗小鼠宫颈癌作用。采用局部用药抗移植的小鼠腹水瘤型和皮下实体瘤型宫颈癌Ｕ１４。结果表明：腹腔内注射一定剂量的ＰＥＧ－ｒＩＬ－２＊（４５００ＩＵ，ＱＤ×５）能较相同剂的ｒＩＬ－２显延长荷腹水瘤小鼠的存活期（２３．１±３．６天     

ENU诱发xy1E,C57BL/6J转基因小鼠体内基因突变及微核效应的实验研究

目的：用诱变剂乙基亚硝基脲（ Ｅ Ｎ Ｕ） 验证基于ｐ Ｕ Ｃ１１８ Ｎ Ｘ 质粒载体的转基因小鼠用于研究体内基因突变的可行性。方法：用酶切、环化方式从经和未经 Ｅ Ｎ Ｕ 诱变处理的ｘｙ１ Ｅ， Ｃ５７ Ｂ Ｌ／６ Ｊ转基因小鼠脾脏组织 Ｄ Ｎ Ａ 中分离ｐ Ｕ Ｃ１１８ Ｎ Ｘ 质粒载体，使其转化 Ｄ Ｈ１０ Ｂ 宿主菌并铺 Ａｍｐｒ 平板培养后喷洒邻苯二酚溶液，通过菌落黄白颜色不同筛选突变体，并对突变靶基因ｘｙ１ Ｅ 测序分析以确定突变类型和位点。同时取该转基因小鼠外周血及骨髓细胞，观察 Ｅ Ｎ Ｕ 对转基因小鼠微核形成率的影响。结果：（１） 转基因小鼠脾脏组织ｘｙ１ Ｅ 基因自发突变率小于４ ．７９ ×１０ － ５ ，经 Ｅ Ｎ Ｕ（５０ｍ ｇ／ｋｇ ×５） 处理后，脾组织诱发基因突变率为１９ ．８３ ×１０ － ５ ，两者相差显著；（２） Ｅ Ｎ Ｕ 诱发小鼠脾组织基因突变的类型包括颠换（５０ ％ ） 、单／ 双碱基插入（３７ ．５ ％ ）和转换（１２ ．５ ％ ） ；（３） 经 Ｅ Ｎ Ｕ（５０ ｍｇ／ｋｇ ×５） 处理后，转基因小鼠外周血和骨髓细胞的微核率均明显增加，分别为７ ．６ ‰和８ ．８ ‰，与溶剂对照组相比有显著性差异（ Ｐ＜ ０ ．０１） ，表明 Ｅ Ｎ Ｕ 可明显诱发     

大刺猴头（88）多糖抗突变,抗氧化及对免疫功能影响的研究

本实验观察了大刺猴头多糖（Ｈｅｒｉｃｌｉｕｍｐｏｌｙｓａｃｃｈａｒｉｄｅ（ＨＥＰＳ）对大肠杆菌ＳＯＳ应答的抑制作用；径口给大鼠ＨＥＰＳ（３０ｄ）测定大鼠血液和肝组织中超氧化物歧化酶（ｓｕｐｅｒｏｘｘｉｄｅｄｉｓｕｍｕｔａｓｅＳＯＤ），脂质过氧化物丙三醛（Ｍａｌｏｎｙｌｄｉａｌｄｅｇｙｄｅ（ＭＤＡ）含量的变化；经口给小鼠（ＨＥＰＳ）（３０ｄ），其对小鼠腹腔吞噬率、吞噬指数，脏器指数的影响。结果表明：ＨＥＰＳ可以抑制由（ＭｉｔｏｍｙｃｉｎｃＭＭＣ）诱导的ＳＯＳ应答，其抑制率大肠杆菌ＰＱ３５为６４％，ＰＱ３７为５６％；经口给大鼠ＨＥＰＳ其血液及肝匀浆中ＳＯＤ含量均高于对照组。ＭＤＡ的含量均低于对照组，各自２组比较分别有非常显著性差异及显著性差异（Ｐ＜０．０１，ｐ＜０．０５）。ＨＥＰＳ组小鼠的腹腔吞噬细胞吞噬率、吞噬指数，胸腺指数，腺指数与对照组比较也有非常显著性差异（ｐ＜０．０１）。结果提示：大刺猴头（８８）（ＨＥＰＳ）在本实验体系表现有抗突变，抗氧化及增强免疫功能的作用。     

维生素E对色拉油烟冷凝物致小鼠毒性作用的影响

本文采用微核试验和精子畸形分析方法，观察维生素Ｅ对色拉油烟冷凝物致小鼠微核率和精子畸形率作用的影响，并作Ｐｏｉｓｓｏｎ 分布资料的Ｕ 检验。结果显示色拉油烟冷凝物５ｍｌ／ｋｇ 和１０ｍｌ／ｋｇ 体重给小鼠灌胃染毒后，其微核率和精子畸形率均明显地高于２－５ｍｌ／ｋｇ 组和单纯色拉油对照组（Ｐ＜ ０．０１） ，而２ ．５ｍｌ／ｋｇ 组未显示致小鼠微核率和精子畸形率升高作用（Ｐ＞ ０．０５） ；给小鼠补充维生素Ｅ２０ｍｇ／ｋｇ 体重后，可显著地降低色拉油烟冷凝物致微核率和精子畸形率升高的作用（Ｐ＜ ０．０１）。故认为色拉油烟冷凝物对小鼠具有明显地毒性作用，而补充维生素Ｅ则可降低其作用。     

可溶性白介素－2受体水平和红细胞C_3b受体活性与恶性肿瘤病情相关性的研究

为探讨血清可溶性白细胞介素—２受体（ＳＩＬ—２Ｒ）水平和红细胞Ｃ３ｂ受体活性与恶性肿瘤病情严重程度的关系，本文检测了１０８例恶性肿瘤病人血清ＳＩＬ—２Ｒ水平，红细胞Ｃ３ｂ受体花环率（Ｅ—Ｃ３ｂＲＲ）和肿瘤红细胞花环率（ＴＥＲＲ）的变化。结果显示，恶性肿瘤病人ＳＩＬ—２Ｒ明显增高，（Ｅ—Ｃ３ｂＲＲ）和ＴＥＲＲ明显降低，与对照组比较差异有显著性（Ｐ＜０．００１）；有淋巴结转移和病死者ＳＩＬ—２Ｒ增高和Ｅ—Ｃ３ｂＲＲ，ＴＥＲＲ降低较无转移和存活者更明显（Ｐ＜０．００１）；手术前ＳＩＬ—２Ｒ增高、Ｅ—Ｃ３ｂＲＲ和ＴＥＲＲ降低较手术后更明显（Ｐ＜０．００１，０．０１）。恶性肿瘤病人ＳＩＬ—２Ｒ水平与Ｅ—Ｃ３ｂＲＲ和ＴＥＲＲ呈负相关（Ｐ＜０．００１）。提示血清ＳＩＬ—２Ｒ水平增高和红细胞ｃ３ｂ受活性降低恶性肿瘤病情严重程度，淋巴结转移和预后有密切关系。     

β2—微球蛋白,唾液酸及癌胚抗原联合检测消化道癌

对７１例消化道癌（食管 癌、胃癌、大肠癌）患者和２４例消化道良性疾病患者进行血β２－微球蛋白（β２－ＭＧ）、唾液酸（ＳＡ）和癌胚抗原（ＣＥＡ）检测。结果发现，癌症病人血β２－ＭＧ、ＳＡ和ＣＥＡ含量较良性疾病明显增高（Ｐ〈０．０５）；其β２－ＭＧ阳性率为６２．０％，ＳＡ为２９．６％，ＣＥＡ２８．２％；三项标记物联合检测阳性率可提高到８０．３％。实验证实，２－ＭＧ、ＳＡ、ＣＥＡ的检测对消化道癌的诊断有     

肝癌,肝硬变患者的戊型肝炎病毒感染

目的研究肝癌和肝硬变患者的肝炎病毒感染情况。方法用酶联免疫吸附测定（ＥＬＩＳＡ）法测定患者血清中的乙型肝炎病毒表面抗原（ＨＢｓＡｇ）、丙型肝炎病毒抗体（抗ＨＣＶ）和戊型肝炎病毒抗体（抗ＨＥＶ）。结果肝癌患者中抗ＨＥＶ阳性率为５８．９％（６３／１０７），ＨＢｓＡｇ阳性率为６９．２％（７４／１０７），抗ＨＣＶ阳性率为１０．３％（１１／１０７），肝硬变患者的阳性率依次为６３．０％（１７／２７）、７４．１％（２０／２７）、７．４％（２／２７）。只有抗ＨＥＶ阳性而ＨＢｓＡｇ和抗ＨＣＶ阴性的肝癌患者有１３例（１２．２％）。仅ＨＢｓＡｇ阳性而抗ＨＥＶ和抗ＨＣＶ阴性的有２４例（２２．４％），仅抗ＨＣＶ阳性而抗ＨＥＶ和ＨＢｓＡｇ阴性的有３例（２．８％）。全部阴性的有１０例（９．４％）。肝硬变患者中仅抗ＨＥＶ阳性而抗ＨＣＶ和ＨＢｓＡｇ阴性的有５例（１８．５％），仅ＨＢｓＡｇ阳性而抗ＨＥＶ和抗ＨＣＶ阴性的有９例（３３．３％），全部阴性的有１例（３．７％）。结论除ＨＢＶ和ＨＣＶ外，ＨＥＶ感染似乎在肝癌变及肝硬变中也起着一定的作用     

E838联合γ射线对淋巴瘤荷瘤小鼠的协同抑瘤作用

 目的 本研究主要观察E838对小鼠移植性淋巴瘤的体内抑瘤效果及相关的生物学指标，探讨合用137Cs γ射线是否具有抑瘤增效作用。方法 取2～3mm3淋巴瘤瘤块接种于IRM 2小鼠腋部皮下，24h后将荷瘤小鼠随机分为对照组、单放组、E838低、中、高药物组及药物合用照射组、环磷酰胺组。药物组与药物合用照射组对应性腹腔注射相同剂量E838，每日1次，连续7天，环磷酰胺隔日1次×4。合用照射组于给药的第4天进行全身1Gy照射，每日1次，连续5天。观察各组小鼠骨髓有核细胞数和肿瘤抑制率。结果 E838 3个剂量对小鼠移植性淋巴细胞瘤的抑瘤率分别为(44.14±15.96)%、(70.74±11.17)%和(50.00±18.09)%，与对照组比较差异有统计学意义(P<0.001)，骨髓有核细胞数与对照组相比则明显提高。E838合用137Csγ射线能提高抑瘤效果，抑瘤率分别为(65.43±2.13)%、(77.13±6.38)%和(67.55±11.17)%，(P<0.001)，对肿瘤的杀伤作用高于单放组和单药治疗组。结论 E838对小鼠肿瘤细胞具有良好的抑制作用，E838合用γ射线具有协同抑瘤作用，在适当剂量范围内可以促进荷瘤小鼠放疗后骨髓损伤修复。     

E838对CTX化疗损伤小鼠的保护作用

 目的 观察E838对环磷酰胺所致小鼠损伤的保护作用。 方法 对小鼠采用连续5天腹腔注射E838药物,从第3天同时给予环磷酰胺（CTX）,阳性对照为茜草双酯,观察外周血白细胞、骨髓有核细胞数、内源性脾结节形成（CFU-S）、脾脏指数及胸腺指数的变化。 结果 E838可明显减轻化疗所致小鼠免疫功能的抑制,并可使化疗后小鼠的体重恢复增长, 白细胞、骨髓有核细胞和CFU-S明显高于对照组,差异有统计学意义（P<0.05）。中、高剂量组白细胞与茜草双酯组比较,差异有统计学意义（P<0.05）。 结论 提示E838具有对抗化疗损伤的作用。     

E838对辐射所致小鼠淋巴细胞DNA双链断裂的防护作用

目的：探讨E838对辐射导致小鼠淋巴细胞DNA双链断裂的防护作用。方法：小鼠照射前给予E838,观察其存活率和动物平均存活时间;用中性单细胞凝胶电泳技术,检测1 Gyγ射线照射后小鼠淋巴细胞DNA双链断裂,并与给予同类药物小鼠进行比较,用CASP软件分析彗星图像,SPSS12．0进行数据的统计学分析。结果：E838预防用药能够明显提高损伤动物的30天存活率,E838低、中、高剂量组存活率分别比对照组提高55%、85%、75%,平均生存天数与对照组比较,差异有统计学意义（P<0.001）,保护指数分别为1.99、2.56、2.46。E838与炔雌醇比较,中、高剂量组的动物平均生存天数明显延长,差异有统计学意义（P<0.001）。E838能明显减轻淋巴细胞DNA双链断裂损伤,E838低、中、高组,TDNA百分比（11.68、6.24、9.05）、TL（18.53、10.09、16.02）、TM（2.13、0.64、1.21） 和OTM（2.52、0.99、1.59 ）残余损伤逐渐减少,与对照组比较,差异有统计学意义(P<0.001)。E838与炔雌醇、尼尔雌醇比较差异有统计学意义(P<0.05)。结论：E838对辐射损伤具有明显的保护作用。     

E838联合环磷酰胺抗白血病L1210细胞的作用

目的 观察E838及其与化疗药物环磷酰胺(CTX)联合应用对白血病L1210细胞荷瘤小鼠的肿瘤抑制及生命延长作用。方法 以L1210荷瘤IRM-2小鼠为模型,实验分对照组、CTX组、E838组、E838+CTX组,分别检测肿瘤抑制率、骨髓有核细胞数、胸腺与脾指数、生命延长率等指标,比较各组间的差别。结果 E838治疗组瘤重明显小于对照组,联合用药组对荷瘤小鼠白血病L1210有较强抑制作用。生命延长也有提高作用。结论 E838对白血病L1210有抑制作用,与化疗药物CTX合用时,能提高疗效及机体免疫功能。     

Roles of tyrosine residues 845, 892 and 922 in constitutive activation of murine FLT3 kinase domain mutant

FLT3 tyrosine kinase domain (TKD) mutations are detected in approximately 7% of acute myeloid leukemia patients, and suggested to correlate with poor prognosis and confer resistance to FLT3 inhibitors. To explore activation mechanism of FLT3 TKD mutation, we analysed critical tyrosine residues for the constitutive activation and downstream signaling of the mutant by generating a series of single Tyr --> Phe substitution mutant of all 22 cytoplasmic tyrosine residues of murine FLT3 TKD-mutant (mFLT3Asp838Val). Tyr845Phe, Tyr892Phe and Tyr922Phe substitutions suppressed the phosphorylation of mFLT3Asp838Val itself, the activation of Erk1/2, STAT3 and STAT5, and the factor-independent cell proliferation and survival. In contrast, these three Tyr --> Phe mutations partially suppressed but maintained the ligand-dependent activation and anti-apoptotic activity of wild-type FLT3, suggesting that these tyrosine residues were more critical for the constitutive activation and signaling of mFLT3Asp838Val. These three Tyr --> Phe mutations also inhibited the constitutive activation of other FLT3 mutants bearing internal tandem duplication, Asp838Tyr or Ile839del. The suppression of mFLT3Asp838Val activation and signaling by these substitutions was partially recovered by shifting the culture temperature from 37 to 33 degrees C, or by the introduction of Cdc37 and Hsp90. Taken together, Tyr845, Tyr892 and Tyr922 are the critical residues in mFLT3Asp838Val activation, possibly through stabilizing the active conformation of mFLT3Asp838Val.     

Effect of promoter and intron 2 polymorphisms on murine lung K-ras gene expression

Differences in tumor formation among inbred mouse strains with high (A/J) and low (C3H) susceptibility for lung cancer have been linked to a repetitive element within the second intron of the K-ras gene. The purpose of this investigation was to determine whether differences within the K-ras gene promoter region or the intron 2 polymorphism affect K-ras gene expression in lung tumors and target alveolar type II cells isolated from A/J and C3H mice. Ribonuclease protection assays were performed using RNA isolated from 4-(methylnitrosamino)-1-(3- pyridyl)-1-butanone (NNK)-induced lung tumors from each mouse strain and alveolar type II cells isolated from A/J and C3H mice. An 838 bp fragment of the murine K-ras gene promoter region was amplified by PCR, cloned and sequenced from both mouse strains. Promoter regions from both mouse strains were inserted into a luciferase reporter gene vector, with and without the second intron polymorphism, and transfected into sensitive, intermediate and resistant lung tumor cell lines. No significant differences in K-ras gene promoter activity was found between the two strains using these specific reporter gene constructs. Consistent with these results, levels of K-ras expression did not differ between alveolar type II cells, whole lung or tumors induced by NNK in A/J or C3H mice. Moreover, in lung tumor cell lines derived from mice with differing susceptibility for lung cancer, K-ras expression did not correlate with the growth rate of tumors induced in nude mice from these cell lines. These results indicate that factors involved in modulating the rapid clonal expansion of the mutated K-ras allele from A/J mice are not directly linked to expression of this gene. Other genetic changes or losses in conjunction with hypothesized modifier loci, such as the Par1 locus, must play a significant role in establishing the phenotypic strain differences for lung tumor formation.      

Vaccination of full-length HPV16 E6 or E7 protein inhibits the growth of HPV16 associated tumors

Cervical cancer is the second most common cancer in women worldwide. Human papillomavirus (HPV) is the primary etiologic agent of cervical cancer. Two HPV16 proteins, E6 and E7, are consistently expressed in tumor cells. Most therapeutic vaccines target one or both of these proteins. Taking the advantages of safety and no human leukocyte antigen restriction, protein vaccine has become the most popular form of HPV therapeutic vaccines. Here we demonstrate that immunization with full-length HPV16 E6 or E7 protein elicited specific immunological effect and inhibition of TC-1 cell growth using TC-1 mouse model. HPV16 E6 and E7 genes were cloned into pET-28a(+) and introduced into E. coli Rosetta. Expression of the genes was induced by IPTG. Proteins were purified by Ni-NTA agarose and they were detected by SDS-PAGE and Western blotting. C57BL/6 mice were vaccinated with 1.5 nmol HPV16 E6 or E7 protein. Then they were implanted with 1x10(5) TC-1 cells. No tumor was detected in any mouse vaccinated with E7 protein. Forty days later, the tumor-free mice and control mice were challenged with 2x10(5) TC-1 cells. All control mice developed tumors 6 days later, but E7 immunized mice were tumor free until 90 days. Tumor growth was slow in the E6 immunized mice, but 83% of the mice developed tumors and the survival percentage was not significantly different from the control. An adoptive immune model was used to demonstrate the therapeutic effect. Results showed that the development of TC-1 cells was obviously reduced by transfusion of T-cells but not serum from mice immunized with E7 protein. T-cells from E7 immunized mice also induced the lysis of TC-1 cells in the cytotoxic T lymphocyte assay. These findings show that immunization with HPV16 E6 or E7 protein was able to elicit specific protective immunity against TC-1 tumor growth.     

Cabbage and vitamin E: their effect on colon tumor formation in mice

The effects of cabbage and vitamin E on colon carcinogenesis were investigated in Swiss mice treated with 1,2-dimethylhydrazine. Throughout the experiment the mice were fed a laboratory chow diet (46 mg vitamin E per kg) or chow containing 13 g cabbage per 100 g or 180 mg vitamin E per kg. Starting after 31 days of diet treatment the mice received 7 weekly s.c. injections of DMH. They were sacrificed 17 weeks after the first dose of DMH. While diet did not significantly alter colon tumor response, some trends were observed. Female mice given cabbage had a higher incidence (percent of mice with a tumor) and multiplicity (tumors per tumor bearing mouse) of colon tumors. Males were little affected by cabbage apart from a lower incidence of adenocarcinomas. Compared with mice fed the control diet those given vitamin E had a higher colon tumor incidence. This effect, which was stronger in females, was due to an increased incidence of adenomas. Vitamin E had little apparent affect on tumor multiplicity apart from a reduction in adenocarcinomas in females and adenomas in males. The data do not support the view that cabbage and vitamin E are protective against colon cancer.