人胰岛素样生长因子Ⅱ反义RNA对肝癌细胞恶性表型的抑制作用

研究人胰岛素样生长因子Ⅱ（ＩＧＦ－Ⅱ）反义ＲＮＡ对肝癌细胞株ＳＭＭＣ－７７２１的抑制效应。方法人ＩＧＦ－ⅡｃＤＮＡ０．１ｋｂ反向插人真核细胞表达载体ｐｃＮＡ３,获得ＩＧＦ－Ⅱ反义ＲＮＡ表达载体ｐＩＧＦ－ⅡＡＳ,将其导人人肝癌细胞株ＳＭＭＣ-７７２１,观察软琼脂培养集落形成的能力。流式细胞仪（ＦＣＭ）测定ｐＩＧＦ－ⅡＡｓ表达ＩＧＦ－Ⅱ反义ＲＮＡ对ＳＭＭＣ-７７２１细胞生长的影响。结果转导人ｐＩＧＦ－ⅡＡｓ的ＳＭＭＣ－７７２１细胞,不能在软琼脂上形成集落,ＦＣＭＭ实导人ＰＩＧＦ－ⅡＡｓ细胞Ｓ期增多,而对照组ＳＭＭＣ－７７２１细胞和带ＤＣＤＮＡ３空载体的ＳＭＭＣ－７７２１细胞则无明显变化．结论ｐＩＧＦ-ⅡＡＳ＆义ＲＮＡ体外可抑制肝癌细胞株ＳＭＭＣ－７７２１的致瘤性。     

Fas基因转导胃癌细胞的表达

目的将Ｆａｓ基因导入胃癌细胞，建立Ｆａｓ基因表达株，并比较转导前后ｍＲＮＡ与蛋白的表达水平．方法采用分子克隆技术将Ｆａｓ基因插入真核表达载体ｐＢＫＣＭＶ的多克隆克隆位点之间，以脂质体介导法将目的基因导入受体细胞ＳＧＣ７９０１，用Ｇ４１８筛选克隆细胞；以Ｓｏｕｔｈｅｒｎｂｌｏｔ，Ｎｏｒｔｈｅｒｎｂｌｏｔ，Ｗｅｓｔｅｒｎｂｌｏｔ检测Ｆａｓ基因的表达．结果成功地构建了真核表达载体ｐＢＫＦａｓｃＤＮＡ．转导细胞后，从１×１０５细胞中筛选出１００个抗性克隆以上，转导率大于０１％，随机挑选２个克隆扩增培养，获得了１株稳定的抗性细胞，从而有效地建立了Ｆａｓ基因表达株（ＳＧＣ７９０１Ｆａｓｃｅｌｓ）．杂交结果表明，转导株与非转导株均有ｃＤＮＡ的表达，但转导株在ｍＲＮＡ及蛋白水平的表达均明显高于非转导株．结论Ｆａｓ基因在胃癌细胞中处于低表达状态；通过真核表达载体的介导，Ｆａｓ基因导入胃癌细胞后，能有效地表达ＦａｓｍＲＮＡ及其蛋白．     

日本血吸虫26kDa谷胱甘肽S—转移酶DNA疫苗的研究及其保 …

本文对大陆株日本血吸虫２６ｋＤａ谷胱甘肽Ｓ－转移酶（Ｓｊ２６ＧＳＴ）的ＤＮＡ疫苗进行研究，并观察了其免疫保护效果。分别构建含Ｓｊ２６ＧＳＴ基因的重组质粒ｐＣＤ－Ｓｊ２６和ｐＢＫ－Ｓｊ２６，采用脂质体介导的基因转移将ｐＣＤ－Ｓｊ２６和ｐＢＫ－Ｓｊ２６分别导入ＮＩＨ３Ｔ３成纤维细胞，用免疫荧光法（ＩＦＡ）证实Ｓｊ２６ＧＳＴ在真核细胞中的表达。分别将ｐＣＤ－Ｓｊ２６和ｐＢＫ－Ｓｊ２６及ｐＢＫ－ＣＭＶ肌注     

转化生长因子β1正,反义基因对人肝癌细胞SMMC—7721增殖的影响

目的探讨人转化生长因子β１（ＴＧＦβ１）正、反义基因对人肝癌细胞 ＳＭＭＣ－７７２１增殖的影响。方法将ＴＧＦβ１正、反义基因分别导入ＳＭＭＣ－７７２１细胞,观察各转染细胞生长情况、凋亡特征及Ｆａｓ、ｐ５３的表达。结果与空质粒转染细胞相比,正义ＴＧＦβ１基因转染细胞生长速度减慢,出现明显的细胞凋亡形态学改变,３－羟基末端标记法（ＴＵＮＥＬ）为阳性,而反义基因转染细胞未发生上述改变,两者都可检测到ｐ５３信号,但未检测到Ｆａｓ表达。结论ＴＧＦβ１可通过诱导ＳＭＭＣ－７７２１凋亡而对其生长起负调控作用,其机制可能与Ｆａｓ／ＦａｓＬ、ｐ５３无关。     

人类HLA—B7可诱导启动子调控IRES连接的人TNFα和IFN？…

目的：为增强肿瘤细胞表面ＭＨＣＩ类分子表达，并调动免疫系统更有效地抑制和杀伤肿瘤细胞，构建在ＨＬＡ－Ｂ７启动子调控下，ＩＲＥＳ连接的人ＴＮＦα和ＩＦＮβ基因的重组逆转录病毒载体，方法与结果：将ＴＮＦαｃＤＮＡ克隆入ｐＢｌｕｅｓｃｒｉｐｔｓｋ（＋）中，改换酶切位点后定向克隆入含ＨＬＡ－Ｂ７启动子的质粒中，构建成ｐＧＬ３Ｂ７ＴＮＦ。ＩＦＮβｃＤＮＡ经定向插入ｐＧＥＭ－７Ｚｆ（＋）改换酶切点切下，定向克     

HBV反义全基因真核细胞表达载体的构建及在肝癌细胞内的表达

１．材料与方法：乳哺动物细胞表达载体ｐＢＫ－ＣＭＶ,长约４．５ｋｂ,带有Ｔ_７公用序列。ｐＣＰ１０是ｐＢＲ３２２ＥｃｏＲＩ位点中插入双拷贝头尾相接的ＨＢＶａｙｗ亚型全基因ＨＢＶＤＮＡ重组质粒。ＰＣＲ引物ＸＩ１：１４２８－１４４８ｎｔ５'ＣＧＴ　ＣＣＣＧＴＣ　ＧＧＣＧＣＴＧＡＡＴＣＣ３',ＸＩ_２：１６７２－１６５３ｎｔ５'ＡＧＴＣＣＡＡＧＡＧＴＣＣＴＣＴＴＡＡＧ３'。人肝癌细胞系ＳＭＭＣ－７７２１,出上海细胞所提供。将ＥｃｏＲＩ酶切后回收的全基因ＨＢＶ片段。在Ｔ_４ＤＮＡ连接酶作用下,与ＥｃｏＲＩ酶切…     

病毒胸苷激酶基因治疗人胃癌的实验研究

为观察单纯疱疹病毒脱氧胸苷激酶（ＨＳＶ－ＴＫ）／丙氧鸟苷（ＧＣＶ）基因治疗系统对胃癌细胞的杀伤作用，将ＨＳＶ－ＴＫｃＤＮＡ定向克隆入逆转录病毒载体ｐＤＯＲ－ｎｅｏ中，用ＬＩＰＯＦＥＣＴＡＭＩＮＥ法将该ＨＳＶ－ＴＫ基因转染胃癌细胞系ＳＧＣ－７９０１，测定阳性转染细胞（ＳＧＣ－７９０１／ＴＫ）在体内外对ＧＣＶ的敏感性。结果示ＧＣＶ在体外对ＳＧＣ－７９０１／ＴＫ细胞有明显的杀伤作用，其作用呈现剂量和时间依赖性特点，且同时表现出对周围亲代ＳＧＣ－７９０１细胞的杀伤效应（旁观者效应）。体内实验表明ＧＣＶ可抑制ＨＳＶ－ＴＫ阳性胃癌细胞的生长，使已形成的肿瘤逐渐消失。提示逆转录病毒载体介导的ＨＳＶ－ＴＫ基因转移胃癌细胞，配合以ＧＣＶ治疗，是胃癌基因治疗的又一途径。     

Bax基因对人胃癌耐药细胞多药耐药性的逆转作用

目的 研究凋亡相关基因Ｂax对胃癌多药耐药细胞的多药耐药性的逆转作用。方法 构建含有人Ｂax cＤＮＡ全长的真核表达载体pＢＫ－Ｂax,经脂质体导入缺乏Ｂax蛋白表达的人 胃癌多药耐药细胞系ＳＧＣ７９０１/ＶＣＲ中。Ｗestern印迹观察Ｂax基因在转导细胞中的表达,ＭＴＴ法检测Ｂax转导细胞与空载体转导细胞对化疗药物的敏感性。结果 成功构建了Ｂax的真核表达载体,Ｂav转导细胞能稳定表达Ｂax蛋白,与空载体转导细胞相比。Ｂ     

含日本血吸虫脂肪酸结合蛋白（SjFABPc）基因DNA重组质粒 …

目的 观察裸ＤＮＡ重组质粒ｐＣＤ－ＳｊＦＡＢＰｃ在体外转染ＨｅｐＧ２细胞以及质粒ＤＮＡ免疫鼠肌组织中的表达，为进一步应用该质粒进行ＤＮＡ免疫实验奠定基础。方法 用脂质体介导ＤＮＡ转染法将ｐＣＤ－ＳｊＦＡＢＰｃ转染贴壁细胞ＨｅｐＧ２，Ｇ４１８加压筛选获得阳性克隆细胞并传代培养，ＳＤＳ－ＰＡＧＥ及Ｗｅｓｔｅｒｎ－ｂｌｏｔ鉴定目的基因在ＨｅｐＧ２细胞中的表达，将ｐＣＤ－ＳｊＦＡＢＰｃ质粒肌注免疫ＢＡＬＢ     

人尿激酶基因治疗的研究

人肝癌ＳＭＭＣ－７７２１细胞的高密度脂蛋白－载脂蛋白的受体介导的逆向胞饮代谢途径魏强，吴满平，梅美珍，庄庆祺（上海医科大学药学院生物化学教研室，上海２０００３２）人肝癌ＳＭＭＣ－７７２１细胞的细胞膜上具有高密度脂蛋白受体。为了阐明ＨＤＬ蛋白部分的代谢...     

HUMAN INSULIN1

Two methods of synthesis of human insulin have been developed to the stage of commercial production. One entails synthesis of human insulin by bacteria (“biosynthetic” human insulin) and one entails the conversion of pork insulin into human insulin by an amino acid substitution (“semisynthetic” human insulin). Both forms of human insulin have been shown to be safe, and to have similar efficacy and pharmacokinetics to purified pork insulin. Human insulin given by subcutaneous injection has been shown to elicit antibody formation in man, although the extent of this may be slightly less than in the case of purified pork insulin, and is certainly less than in the case of beef insulin and beef-pork combinations. The clinical significance of these differences in immunogenicity is doubtful. However there are rare defined situations, such as allergy to pork insulin and antibody-mediated insulin resistance where human insulin has been shown to have advantages over purified pork insulin. Bacterial synthesis of insulin provides an assured supply for the world's future needs, and it is conceivable that further technological refinement may make bacterial synthesis cheaper than extraction and purification of animal insulins. However, current evidence provides no basis for recommending human rather than purified animal insulin for the routine management of insulin dependent diabetes.     

DIFFERENTIAL EFFECTS OF OVINE AND HUMAN CORTICOTROPHIN-RELEASING FACTOR IN HUMAN SUBJECTS

Ten healthy subjects received 200 micrograms of human CRF (hCRF) and 200 micrograms of ovine CRF (oCRF) as an intravenous bolus injection on two different occasions. After hCRF plasma ACTH levels rose significantly (P less than 0.0005, by Friedman's nonparametric analysis of variance) from a basal value of 35 +/- 3 pg/ml (mean +/- SEM) to a peak value of 80 +/- 7 pg/ml 30 min after hCRF administration. This ACTH response was followed by a rise in plasma cortisol levels (P less than 0.0005, by Friedman's test) from a baseline value of 0.32 +/- 0.03 mumol/l to a peak value of 0.56 +/- 0.02 mumol/l 60 min after hCRF. Ovine CRF elicited similar rises in the plasma ACTH and cortisol levels. However, as derived from the faster rate of decline of ACTH and cortisol after hCRF than after oCRF, human CRF had a significantly shorter duration of action than ovine CRF in humans. Human CRF not only stimulated ACTH release by the human pituitary gland but also prolactin release. After hCRF administration prolactin levels rose significantly (P less than 0.005, by Friedman's test) from a basal value of 179 +/- 18 mU/l to a peak value of 288 +/- 34 mU/l at 10 min.     

POST-MAMMOPLASTY HUMAN ADJUVANT DISEASE

Morphoea and keratoconjunctivitis sicca developed in a womanwith seronegative rheumatoid arthritis six years after augmentationmammoplasty. Previous reports of post-mammoplasty connectivetissue disease have followed the use of silicone gel, whereasthe more ‘biologically inert’ saline-filled silasticimplants were used in this case. KEY WORDS: Mammoplasty, Silicone, Adjuvant disease, Morphoea