抗内甲基转移酶单克隆抗体的制备及临床应用研究

目的：建立抗人甲基转移酶单克隆抗体细胞株并建立相应的蛋白测定方法,观察脑瘤组织中O^6-甲基鸟嘌呤－DNA甲基转移酶（MGMT）表达水平与亚硝脲药物疗效的关系,方法：采用细胞融合技术建立抗人甲基转移酶单克隆抗体细胞胞株,采用免疫组织化染色方法,检测60例脑瘤组织中MGMT表达水平,结果：得到了7株分泌抗人甲基转移酶单克隆抗体的杂交瘤细胞株,并建立了测定MGMT免疫组化方法,观察的60例脑瘤组织中有27例MGMT阴性,经亚硝脲药物治疗,其中24例好转,3例复发,另3例MGMT可疑阳性的患者,经亚硝脲药物治疗,其中2例好转,1例复发,25例MGMT阳性患者,经亚硝脲药物治疗,5例好转,12例复发,8例死亡,5例MGMT为强阳性患者,使用亚硝脲药物治疗无效,全部死亡,结论：建立了7株稳定分泌抗人甲基转移酶单克隆抗体的杂交瘤细胞株及MGMT蛋白测定方法,为临床开展亚硝脲预见性化疗提供了必要手段,初步结果显示,脑瘤组织中MGTM表达水平与亚硝脲药物疗效及预后有关,是肿瘤细胞对亚硝脲药物产生耐药性的基础。     

培养和冷冻对人胎胰岛素功能的影响

胰腺组织进行培养以及应用冷冻技术选择性保存组织活力,能达到纯化胰岛和改变胰岛免疫原性的目的。低温保存可以贮存大量胰岛,为临床移植创造条件。本文报道培养和冷冻条件对人胎胰碎片活性的影响。     

破解癌细胞化疗耐药之谜

由中国人民解放军军事医学科学院章扬培教授等人共同完成的“06-甲基鸟嘌呤-DNA-甲基转移酶与肿瘤预见性化疗新策略的研究”，荣获2004年国家科技进步奖二等奖。     

MGMT与肿瘤的预见性化疗

O^6-甲基鸟嘌呤DNA甲基转移酶（O^6-methylguanine DNA-transferase，MGMT），是细胞修复DNA损伤的一种酶，这种酶又被称为O^6-烷基鸟嘌呤DNA烷基转移酶（AGAT）。顾名思义。此酶可以修复由各种烷化剂（alkylating agents）造成的细胞DNA烷基化损伤，特别是DNA分子中鸟嘌呤第6位氧原子上的甲基乃至烷基化损伤。根据MGMT基因及其表达水平的不同，将细胞特别是肿瘤细胞分成Mer^＋和Mer^-两种类型：Mer^＋对烷化剂耐药，Mer^-对烷化剂敏感，即细胞中的MGMT水平决定了细胞对烷化剂的耐药程度；接着科学家集中精力研究如何克服Mer^＋细胞耐药性的问题，发现O^6-BG可以有效克服Mer^＋细胞对烷化剂耐药性。这些研究为开展甲基转移酶MGMT与肿瘤预见性、个体化治疗奠定了基础。     

EXPRESSION OF HUMAN α-GALACTOSIDASE AND α1,2-FUCOSYL-TRANSFERASE GENES MODIFIES THE CELL SURFACE GALα1,3GAL ANTIGEN AND CONFERS RESISTANCE TO HUMAN SERUM-MEDIATED CYTOLYSIS

Objective To explore the strategies which reduce the amount of xenoantigen Galα1, 3 Gal. Methods Human α-galactosidase gene and α 1,2-fucosyltransferase gene were transferred into cultured porcine vascular endothelial cells PEDSV.15 and human α-galactosidase transgenic mice were produced. The Galα 1,3Gal on the cell surface and susceptibility of cells to human antibody-mediated lysis were analyzed. Results Human α-galactosidase gene alone reduced 78% of Galα1,3Gal on PEDSV.15 cell surface while human α-galactosidase combined with α 1,2-fucosyltransferase genes removed Galα 1,3Gal completely. Decrease of Galα1,3Gal could reduce susceptibility of cells to human antibody-mediated lysis, especially during co-expression of α-galactosidase gene and α1,2-fucosyltransferase gene. RT-PCR indicated positive human α-galactosidase gene expression in all organs of positive human α-galactosidase transgenic F1 mice including heart, liver, kidney, lung, and spleen, the amount of Galα1,3Gal antigens on which was reduced largely. 58% of spleen cells from F1 mice were destroyed by complement-mediated lysis compared with 24% of those from normal mice. Conclusions Human α-galactosidase gene and α1,2-fucosyltransferase gene effectively reduce the expression of Galα1,3Gal antigens on endothelial cell surface and confers resistance to human serummediated cytolysis. The expression of human α-galactosidase in mice can also eliminate the Galα1,3Gal antigens in most tissues and decrease the susceptibility of spleen cells to human serum-mediated cytolysis.     

肝病患者血清碱性谷胱甘肽S－转移酶的变化

肝病患者血清碱性谷胱甘肽Ｓ－转移酶的变化范开春，黄英才，任会明，谭子兴，李春海谷胱甘肽Ｓ－转移酶（ＧＳＴｓ）是一组多基因家族蛋白，胞液中ＧＳＴｓ同功酶可分为碱性（即ａ类）、中性和酸性３大类。ＧＳＴｓ占肝细胞胞浆蛋白的３．７％，ａ类又占肝ＧＳＴｓ的８４...     

EXPRESSION OF HUMAN α-GALACTOSIDASE AND α1,2-FUCOSYLTRANSFERASE GENES MODIFIES THE CELL SURFACE GALα1,3GAL ANTIGEN AND CONFERS RESISTANCE TO HUMAN SERUM-MEDIATED CYTOLYSIS

Objective To explore the strategies which reduce the amount of xenoantigen Galα1, 3 Gal. Methods Human α-galactosidase gene and α1,2-fucosyhransferase gene were transferred into cultured porcine vascular endothelial cells PEDSV. 15 and human α-galactosidase transgenic mice were produced. The Galα1,3Gal on the cell surface and susceptibility of cells to human antibody-mediated lysis were analyzed. Results Human α-galactosidase gene alone reduced 78% of Galα1,3Gal on PEDSV. 15 cell surface while human α-galactosidase combined with α1,2-fucosyltransferase genes removed Galα1,3Gal completely. Decrease of Galα1,3Gal could reduce susceptibility of cells to human antibody-mediated lysis, especially during co-expression of α-galactosidase gene and α1,2-fucosyltransferase gene, RT-PCR indicated positive human α-galactosidase gene expression in all organs of positive human α-galacto-sidase transgenic F1 mice including heart, liver, kidney, lung, and spleen, the amount of Galα1,3Gal antigens on which was reduced largely. 58% of spleen cells from F1 mice were destroyed by complement-mediated lysis compared with 24% of those from normal mice. Conclusions Human α-galactosidase gene and α1,2-fucosyltransferase gene effectively reduce the expression of Galα1,3Gal antigens on endothelial cell surface and confers resistance to human serummediated cytolysis. The expression of human α-galactosidase in mice can also eliminate the Galα1,3Gal antigens in most tissues and decrease the susceptibility of spleen cells to human serum-mediated cytolysis.     

人肝碱性谷胱甘肽S-转移酶与肝细胞损伤性疾病关系的初步研究

应用血清ＧＳＴ－α酶联检测试剂盒测定了甲型肝炎、急性乙型肝炎、慢性活动性肝炎（慢活肝）、慢性迁延性肝炎（慢迁肝）和肝硬化病人血清ＧＳＴ－α水平，结果均比正常人组和非肝胆疾病组显著升高。除慢迁肝外，其它各型肝炎组ＧＳＴ－α水平与ＡＬＴ显著正相关（Ｐ＜０．０１）。联合测定ＧＳＴ－α和ＡＬＴ可提高阳性检出率。在慢性迁延性肝炎组，ＧＳＴ－α检测敏感度显著高于ＡＬＴ（Ｐ＜０．０１）。在观察的７９例肝癌病人中，ＧＳＴ－α阳性率为８１％，而ＡＬＴ阳性率为６１％，经统计学分析，血清ＧＳＴ－α检测敏感度显著高于ＡＬＴ（Ｐ＜０．００１）。在３０例乙肝表面抗原（ＨＢｓＡｇ）阳性无症状携带者中，ＧＳＴ－α阳性率为７０％，而ＡＬＴ仅为３７％。结果表明：①血清ＧＳＴ－α与ＡＬＴ联合测定可提高肝细胞损伤性疾病的阳性检出率。②在肝癌发生时，血清ＧＳＴ－α水平升高，一方面由于肝细胞受损，释放ＧＳＴ－α增多，更主要的原因是肝细胞表达ＧＳＴ－α增强所致，因此，ＧＳＴ－α可作为肝癌肿瘤酶学标志物。③在轻微肝细胞损害时，ＧＳＴ－α检测敏感度高于ＡＬＴ，血清ＧＳＴ－α可作为肝细胞损伤的敏感、早期和器官特异性较好的血清酶学标志。     

α半乳糖苷酶的c DNA克隆及序列测定

咖啡豆中存在的α半乳糖苷酶可作用于人Ｂ型血红细胞表面糖链末端的α半乳糖苷键，造成糖链末端的α半乳糖水解，使Ｂ型血表面的糖链结构与Ｏ型血相同，从而将Ｂ型血转变为Ｏ型血，在野战输血和自然灾害急救中具有重要的意义。为实现以基因工程方法生产α半乳糖苷酶的目的，我们克隆了海南兴隆咖啡豆的α半乳糖苷酶全长ｃＤＮＡ，并与已报道的桑托斯咖啡豆的α半乳糖苷酶ｃＤＮＡ序列进行了比较。根据已报道的桑托斯咖啡豆α半乳糖苷酶ｃＤＮＡ序列设计了５′、３′端上、下游引物和与全长ｃＤＮＡ中间部位完全匹配的引物，分别与上、下游引…     

人肝碱性谷胱甘肽S—转移酶与肝细胞损伤性疾病关…

应用血清ＧＳＴ－α酶联检测试剂盒测定了甲型肝炎、急性乙型肝炎、慢性活动性肝炎（慢活肝）、慢性迁延性肝炎（慢迁肝）和肝硬化病人血清ＧＳＴ－α水平，结果均比正常人组和非肝胆疾病组显著升高。除慢迁肝外，其它各型肝炎组ＧＳＴ－α水平一ＡＬＴ显著正相关。联合测定ＧＳＴ－α和ＡＬＴ可提高阳性检出率。在慢性延性肝炎组，ＧＳＴ－α检测敏感度显著高于ＡＬＴ。在观察的７９例肝癌病人中，ＧＳＴ－α阳性在８１％，而ＡＬＴ