考福新或咪唑二酮增强内皮素—1升高大鼠主动脉腺苷环—磷酸

硝苯啶（１００ｎｍｏｌ．Ｌ＾－１）或无细胞外钙抑制内皮素－１（ＥＴ－１）收缩大鼠胸主动脉８０％以上。ＥＴ－１（２０ｎｍｏｌ．Ｌ＾－１；１５．２５ｍｉｎ）增加血管环腺苷酸（ｃＡＭＰ）含量，该作用被咪唑二酮（１００μｍｏｌ．Ｌ＾－１）增强。ＥＴ－１还能增加福斯科林的增加ｃＡＭＰ作用。本文证明ＥＴ－１收缩大鼠胸主动脉于少涉及两种信息转导机制，即开放硝苯啶敏感性钙通道和增加ｃＡＭＰ。     

抗抑郁剂吗氯贝胺的行为效应

加强５－羟色氨酸及β－苯乙胺致小鼠刻板行为实验表明，ｉｇ吗氯贝胺（Ｍｏｃ）对单胺氧化酶（ＭＡＯ）－Ａ的选择性抑制作用（ＥＤ５０＝０．３３ｍｇｋｇ－１，２ｈ）较ＭＡＯ－Ｂ（ＥＤ５０＝１２．７ｍｇｋｇ－１，２ｈ）强约３９倍．２４ｈ内ｉｐＭｏｃ３００ｍｇｋｇ－１有一定镇静作用．对运动系统无明显影响，不引起记忆及定向障碍．Ｍｏｃ６００ｍｇｋｇ－１不能对抗氧化震颤素引起的小鼠震颤和分泌增加．Ｍｏｃ的ＬＤ５０为小鼠：１．３－１．５ｇｋｇ－１ｉｇ；１９８－１９９ｍｇｋｇ－１ｉｖ；大鼠：５４１－５６２ｍｇｋｇ－１ｉｐ．Ｍｏｃ剂量依赖地对抗利血平引起的小鼠和大鼠睑下垂，活动减少和体温降低；使慢性应激刺激引起的小鼠活动性增高及穿箱逃避电击反应失败率增高得到恢复，提示其抗抑郁作用．     

N-甲基-N′-硝基-N-亚硝基胍致大鼠气管上皮细胞体外转化及细胞遗传学改变

利用原代大鼠气管上皮（ＲＴＥ）细胞研究Ｎ－甲基－Ｎ′－硝基－Ｎ－亚硝基胍（ＭＮＮＧ）的致转化作用，并对转化细胞进行细胞遗传学研究．结果表明：增殖旺盛细胞（ＥＧＶ）集落是ＲＴＥ细胞体外转化最早期的特征，ＥＧＶ转化频率与ＭＮＮＧ剂量呈明显的剂量反应关系．从０．３ｍｇＬ－１ＭＮＮＧ剂量组分离到２个ＥＧＶ集落，经消化，传代形成了永生化的细胞系（ＲＴＥＳ１，ＲＴＥＳ２），ＲＴＥＳ１为多倍体核型，２号和７号染色体数目增加至４个且呈高频发生（１００％）；ＲＴＥＳ２呈二倍体核型．电镜结果证实转化细胞来自大鼠气管上皮组织．以上结果提示，原代ＲＴＥ细胞体外培养模型是研究致癌物定量及致癌机理的理想模型系统．     

Effects of platelet activating factor and ginkgolid B on the cAMP levels in washed rabbit platelets

研究了血小板激活因子（ＰＡＦ）和ＰＡＦ拮抗剂银杏内酯Ｂ对洗涤兔血小板中ｃＡＭＰ含量的作用．结果表明ＰＡＦ（０．１－１．０μｍｏｌ·Ｌ－１）对血小板的基础ｃＡＭＰ水平无影响，但对前列腺素Ｅ１（ＰＧＥ１）２μｍｏｌ·Ｌ－１及４，５－二氢－６－［４－（１Ｈ－咪唑－１－）苯基］－５－甲基－３－（２Ｈ）－哒嗪酮（ＣＩ－９３０）２０μｍｏｌ·Ｌ－１引起的ｃＡＭＰ升高有显著的抑制作用．银杏内酯Ｂ能完全拮抗ＰＡＦ抑制ＰＧＥ１和ＣＩ－９３０升高ｃＡＭＰ的作用，ＩＣ５０分别为４．７和１２．５μｍｏｌ·Ｌ－１．合用磷酸肌酸／磷酸肌酸激酶和阿司匹林对ＰＡＦ和银杏内酯Ｂ的作用均无影响．提示ＰＡＦ对磷酸二酯酶的激活作用及腺苷酸环化酶的抑制作用是ＰＡＦ的直接作用，与其同ＰＡＦ受体结合有关．     

8－氯腺苷对小鼠肝癌H_（22）细胞的抑瘤作用研究

８－氯腺苷对小鼠肝癌Ｈ２２实体瘤的抑瘤率最高可达７１．７±１３．３％（ｉＰ）和６６．１±４．４６％（ｉｖ）（１００ｍｇ·ｋｇ－１ｄ－１×７ｄ）。腹水型Ｈ２２细胞经８－氯腺苷作用２４ｈ后，ｃＡＭＰ含量增加，ＤＡＧ含量下降；ＰＫＡＩ型调节亚基在药物作用后明显下降；凝胶阻滞分析表明，反式转录因子ＣＲＥＢ结合活性增加；原位杂交结果显示Ｐ５３抗癌基因表达增强。以上结果提示，８－氯腺苷抑瘤作用机制与第二信使系统和基因表达调控过程有关。     

一种新二萜类化合物的体外抗肿瘤研究

目的研究一种新二萜类化合物凤厥内酯Ａ（Ｆ－Ａ）的抗肿瘤活性。方法采用体外培养的人癌细胞株——早幼粒细胞白血病ＨＬ－６０和红白细胞白血病Ｋ５６２作对象，观察Ｆ－Ａ对其抑瘤活性、诱导分化、细胞周期的影响。结果Ｆ－Ａ对ＨＬ－６０和Ｋ５６２细胞２４ｈ的半数抑制浓度ＩＣ５０分别为７．６ｍｇｇ－１和９．１ｍｇｇ－１，在４ｍｇＬ－１时即明显抑制ＨＬ－６０细胞的对数生长；对３Ｈ－ＴｄＲ参入作用在培养４８ｈ后有一定抑制作用；透视电镜显示肿瘤细胞有损伤表现；细胞周期分析显示Ｇ２＋Ｍ期细胞增多；ＮＢＴ还原反应试验阴性。结论Ｆ－Ａ明显抑制ＨＬ－６０和Ｋ５６２细胞生长，抗肿瘤活性强，对细胞周期的影响提示Ｍ期阻滞，研究初步提示Ｆ－Ａ不具细胞诱导分化作用。     

8－氯腺苷对小鼠肝癌H_（22）细胞的抑瘤作用研究

８－氯腺苷对小鼠肝癌Ｈ２２实体瘤的抑瘤率最高可达７１．７±１３．３％（ｉＰ）和６６．１±４．４６％（ｉｖ）（１００ｍｇ·ｋｇ－１ｄ－１×７ｄ）。腹水型Ｈ２２细胞经８－氯腺苷作用２４ｈ后，ｃＡＭＰ含量增加，ＤＡＧ含量下降；ＰＫＡＩ型调节亚基在药物作用后明显下降；凝胶阻滞分析表明，反式转录因子ＣＲＥＢ结合活性增加；原位杂交结果显示Ｐ５３抗癌基因表达增强。以上结果提示，８－氯腺苷抑瘤作用机制与第二信使系统和基因表达调控过程有关。     

茶多酚对细胞免疫功能的影响及抑瘤作用

ＥＡＣ或Ｓ１８０鼠分别ｉｇ４０、８０和１２０ｍｇ／ｋｇ·ｄ－１的ＴＰ对ＥＡＣ的抑制率分别为６５．３％、５１．０％和４８．７％；对Ｓ１８０的抑制率为６６．３％、６８．５％和５９．８％。ｉｇＴＰ的ＥＡＣ鼠的胸腺指数比对照组分别提高５０．７％、４０．６％和１９．８％；脾指数提高３４．４％、３１．１％和２０．０％；Ｓ１８０鼠的胸腺指数也分别提高了６６．２％、４７．２％和４８．１％；脾指数略有增高。离体细胞培养测定ＴＰ对致敏Ｔ淋巴细胞的粘瘤和钻瘤活性结果表明：各ＴＰ组ＥＡＣ鼠的胸腺或脾淋巴细胞对自体瘤细胞的ＡＴＩ和ＥＴＩ均明显高于对照组（Ｐ＜０．０１）。正常小鼠ｉｇ　６０ｍｇ／ｋｇ·ｄ－１的ＴＰ，连续给药１２ｄ后、腹腔Ｍ吞噬ＣＲＢＣ的百分率和吞噬指数比对照组分别增加６９．６％和６８．７％。     

17β-雌二醇对人成骨细胞株HOS TE85细胞增殖及胞内cAMP,cGMP,iNOS影响

目的观察１７β－雌二醇（Ｅ２）对ＨＯＳＴＥ８５细胞增殖及胞内ｃＡＭＰ，ｃＧＭＰ，ｉＮＯＳ影响。方法［３Ｈ］－ＴｄＲ参入，放射免疫及Ｌ－３Ｈ－精氨酸转化法。结果Ｅ２１．０ｍｍｏｌ·Ｌ－１处理４８ｈ，细胞计数为（９４．３×１０７±７．５×１０７）个·Ｌ－１，对照组为（６５．０×１０７±５．９×１０７）个·Ｌ－１，［３Ｈ］－ＴｄＲ参入量增加４５．０％；细胞ｉＮＯＳ活性为（７３．０±６．６）ｐｍｏｌ·Ｌ－１·ｇ－１·ｍｉｎ－１（比对照组增加４６１．５％），ｃＧＭＰ为（０．６９±０．０３）ｐｍｏｌ·Ｌ－１·１０－８ｃｅｌ（比对照组增加３０５．９％）；抑制ｈＰＴＨ诱导胞内ｃＡＭＰ升高。结论Ｅ２能刺激成骨细胞增殖。Ｅ２可能通过影响胞内ｃＡＭＰ，ｃＧＭＰ，ｉＮＯＳ含量和活性而发挥作用。     

Effects of tanshinone Ⅱ-A sullfonate on adhesion molecule expression of endothelial cells and platelets in vitro

探讨丹参酮ⅡＡ磺酸钠（Ｔａｎ）对培养人脐静脉内皮细胞（ＨＵＶＥＣ）和人血小板表达粘附分子的影响．方法：用流动血细胞计数仪测定肿瘤坏死因子（ＴＮＦα）诱导人脐静脉内皮细胞ＩＣＡＭ１和凝血酶诱导人血小板Ｐ选择素的表达．结果：ＨＵＶＥＣ经ＴＮＦα处理后，明显增加细胞表面ＩＣＡＭ１的表达，增加ＨＬ６０细胞粘附到内皮细胞表面达加入细胞总数的３０％±６％（对照组为４６％±０７％）．在ＴＮＦα处理前，用Ｔａｎ（２５－２００μｍｏｌ·Ｌ－１）与ＨＵＶＥＣ共孵育，则Ｔａｎ剂量依赖性地抑制ＴＮＦα的作用．Ｔａｎ（２５－２００μｍｏｌ·Ｌ－１）与人血小板孵育后，可剂量依赖性地抑制凝血酶诱导人血小板表面Ｐｓｅｌｅｃｔｉｎ的表达．结论：Ｔａｎ可抑制内皮细胞和血小板表达粘附分子．     

赖氨酰氧化酶样-4与丝氨酸蛋白酶基因在人宫颈癌细胞中的表达

目的探讨赖氨酰氧化酶样-4(LOXL4)和丝氨酸蛋白酶(Matriptase)在三株人宫颈癌细胞中的表达情况。方法采用qRT-PCR及Western blot方法检测三株体外培养的宫颈癌细胞株Hela、ME180、HCC94中赖氨酰氧化酶样-4和Matriptase mRNA的表达及LOXL4蛋白的翻译水平。结果 LOXL4 mRNA在三株人宫颈癌细胞中存在不同程度的表达,蛋白翻译水平与mRNA基因表达一致,以Hela表达为最高,ME180次之,HCC94表达最低;而ME180和HCC94中MatriptasemRNA相对表达丰度分别是Hela的4.09倍和12.40倍。结论本研究成功建立了LOXL4及Matriptase在体外培养宫颈癌细胞中表达的检测平台,LOXL4、Matriptase的相对表达水平可能与宫颈癌细胞的侵袭和迁移能力有关。     

赖氨酰氧化酶样-4与尿激酶型纤溶酶原激活物及其受体在人宫颈癌细胞中的表达及意义

目的探讨赖氨酰氧化酶样基因-4（LOXL4）、尿激酶型纤溶酶原激活物（uPA）及其受体（uPAR）在人宫颈癌细胞株中的表达。方法采用半定量RT-PCR方法检测人宫颈癌细胞株Hela、ME180、HCC94中LOXL4、uPA和uPAR mRNA的表达,同时运用Western blot技术检测各细胞中LOXL4蛋白的翻译情况。结果 LOXL4mRNA在三株人宫颈癌细胞株中表达差异有统计学意义（P〈0.01）,其中在Hela细胞株中表达浓度最高,ME180细胞株表达浓度次之,而HCC94细胞株表达浓度最低;而uPA、uPAR mRNA表达浓度水平在HCC94细胞中最高,明显高于Hela、ME180细胞的表达浓度水平（P〈0.05）。结论 LOXL4、uPA、uPAR基因表达可能与宫颈癌细胞的侵袭和迁移有关。     

全反式维A酸对单纯疱疹病毒胸苷激酶基因结合更昔洛韦旁观者效应增强作用及其机制

目的　通过研究全反式维A酸 (ATRA )对单纯疱疹病毒胸苷激酶基因结合更昔洛韦 (HSV tk/GCV)旁观者效应的增强作用及与Cx4 3表达的关系 ,为提高TK自杀基因的疗效及探索增强作用机制提供实验基础。方法　应用Polybrene转染、荧光定量RT PCR等技术 ,观察ATRA对宫颈癌细胞ME180 ,ME180 /TK转化细胞旁观者效应及诱导Cx4 3mRNA表达的作用。结果　ME180 ,ME180 /TK不同比例的混合细胞随ATRA浓度的增加而存活率降低。ME180 +ME180 /TK混合细胞的存活率随ME180 /TK细胞比例的增加而降低。在 2mg·L- 1更昔洛韦 (GCV)作用下 ,以 10 - 8mol·L- 1ATRA与不加ATRA (0mol·L- 1)的作用相比较 ,在ME180与ME180 /TK不同比例混合的各组中 ,细胞的存活率明显降低 (P <0 .0 5 )。并观察到10 - 8及 10 - 10 mol·L- 1ATRA对GCV旁观者效应的增强作用。RT PCR结果表明 ,经ATRA处理的ME180细胞 ,其Cx4 3mRNA的相对拷贝数比值增高约 1.84倍及 2 .6 5倍。结论　宫颈癌ME180细胞中 ,ATRA在10 - 8～ 10 - 10 mol·L- 1范围内 ,具有明显增强HSV tk/GCV旁观者效应的作用。其主要机制是通过ATRA在转录水平诱导Cx4 3mRNA表达上调 ,导致旁观者效应的增强。     

金粉蕨素体外诱导人宫颈癌HeLa 细胞凋亡的机制

目的：探讨金粉蕨素（ONY）体外诱导人宫颈癌HeLa细胞凋亡的作用及其分子机制。方法体外培养人宫颈癌HeLa、CaSki、SiHa、ME180细胞,采用MTT法检测ONY对人宫颈癌HeLa、CaSki、SiHa、ME180细胞生长抑制率的影响;Hoechst33258染色检测ONY对人宫颈癌HeLa细胞凋亡的形态学变化;流式细胞术（FCM）检测ONY对HeLa细胞凋亡率的影响;DNA琼脂糖电泳检测HeLa细胞凋亡的DNA条带;Western blot分析凋亡相关蛋白表达变化。结果 ONY对人宫颈癌HeLa、CaSki、SiHa、ME180细胞生长有较强的抑制作用,呈剂量和时间依赖性,以HeLa细胞对ONY最为敏感,作用24 h的IC50值为10.48μg·mL-1。经ONY作用于HeLa细胞24 h后,细胞出现典型的凋亡形态学改变,表现出典型的凋亡特征的亚二倍体峰,且呈剂量依赖性,同时出现典型的凋亡DNA条带;同时,cytochrome c、caspase-9和caspase-3蛋白表达增加,bcl-2蛋白表达下调,而bax蛋白表达不变, Bcl-2/Bax比值下调（P〈0.05）。结论 ONY可能通过调控线粒体途径相关蛋白抑制宫颈癌HeLa细胞增殖并诱导其凋亡。     

Growth inhibition of lung cancer cells by adenosine 5′‐triphosphate

Preliminary clinical data suggest that adenosine 5′‐triphosphate (ATP) may inhibit lung tumor growth. Because studies of ATP on lung cancer cells are lacking, the aim of the present study was to explore effects of extracellular ATP on the growth and morphology of human lung tumor cells. Five human lung tumor cell lines derived from tumors with different cellular characteristics, i.e., a small cell carcinoma (GLC4), a large cell carcinoma (H460), a squamous cell carcinoma (H520), a mesothelioma (MERO82), and a papillary adenocarcinoma (H441), were exposed to 0, 0.5, 1, 2, and 3 mM ATP. Total cell numbers and dead or damaged cells were measured on days 1, 2, and 3. ATP induced a significant, dose‐dependent growth inhibition in GLC4, H460, H520, and MERO82 cells. In contrast, H441 cells showed already maximal inhibition at 0.5 mM. Compared to untreated control cell lines, a significant growth inhibition (mean ± SEM) of 65 ± 5% (GLC4), 59 ± 5% (H460), 45 ± 5% (H520), 38±2% (MERO82), and 55 ± 8% (H441) was shown after 3 days incubation with 3 mM ATP. ATP also induced changes in morphology and attachment to the substratum. Although not demonstrated by the Trypan Blue exclusion test, on photographs it seems that ATP induces death of GLC4 and H460 cells at higher concentrations. In conclusion, in four out of five explored lung tumor cell lines, ATP induces a dose‐dependent growth inhibition. Lung adenocarcinoma cells show already maximal inhibition at the lowest tested ATP dose. There is a relationship between growth inhibition and morphology changes. Drug Dev. Res. 60:196–203, 2003. © 2003 Wiley‐Liss, Inc.     

基质金属蛋白酶-9基质金属蛋白酶-2在宫颈癌细胞株中的表达及意义

目的探讨基质金属蛋白酶(MMP)-9及MMP-2在人子宫颈癌细胞株Siha、CaSki中的表达及意义。方法选择两种宫颈癌细胞(未转移的宫颈鳞癌细胞株Siha、宫颈癌转移细胞株CaSki细胞),采用细胞免疫化学SABC法和酶联免疫吸附试验(ELISA)法对两种宫颈癌细胞及细胞上清液中MMP-9、MMP-2表达情况进行检测和分析。结果 MMP-9、MMP-2在两种宫颈癌细胞中都高表达,且两者在肠转移的CaSki细胞中及细胞上清液中表达明显高于未转移的宫颈癌细胞Siha细胞及细胞上清液,差异有统计学意义(P<0.01)。结论 MMP-9、MMP-2在宫颈癌细胞株CaSki中的高表达与其细胞的转移能力密切相关。     

低氧诱导因子-1α和Survivin在宫颈鳞癌组织中的表达及意义

目的探讨低氧诱导因子-1α（hypoxia-inducible factor-1α,HIF-1α）和Survivin在宫颈鳞癌中的表达及其临床意义。方法采用免疫组化SP法检测50例宫颈鳞癌及10例正常宫颈组织中HIF-1α和Survivin蛋白的表达情况,并结合临床病理特征进行分析。结果（1）HIF-1α和Survivin在宫颈鳞癌中的表达阳性率分别为64%、70%,明显高于正常对照组（二者在正常宫颈组织中均无表达）,P〈0.05,差异有显著性。（2）HIF-1α与临床分期显著相关（χ^24.67,P〈0.05）,与分化程度显著相关（χ^29.04,P〈0.05）;Survivin与临床分期显著相关（χ^25.36,P〈0.05）,与分化程度有明显相关性（χ^26.32,P〈0.05）。（3）HIF-1α和Survivin在宫颈鳞癌中阳性表达呈正相关关系。结论HIF-1α与Survivin均在宫颈鳞癌的发生、发展中起重要作用,且两者在宫颈鳞癌的演进中可能起着协同作用,故联合检测可作为判断宫颈鳞癌恶性程度及评估预后的重要指标。     

A cationic cytofectin with long spacer mediates favourable transfection in transformed human epithelial cells

The synthesis and transfection potential of a novel cationic cholesterol cytofectin with a dimethylamino head group and a long 12 atom, 15A spacer incorporating relatively polar amido and dicarbonyl hydrazine linkages are reported. Thus N,N-dimethylaminopropylamidosuccinylcholesterylformylhydrazide (MS09) in equimolar admixture with dioleoylphosphatidylethanolamine (DOPE) forms stable unilamellar liposomes (80-150 nm) which cluster into very effective transfecting, serum nuclease-resistant, lipoplexes with DNA (180-200 nm) at a liposome+/DNA- molar charge ratio of 2.8:1 (12:1, w/w). Gel retardation and ethidium displacement assays confirmed that DNA was fully liposome-associated and maximally compacted at this ratio. Transfection levels in three human transformed epithelial cell lines, as established by luciferase transgene activity, was found to be optimal at this charge ratio and in the following order: cervical carcinoma (HeLa)>oesophageal carcinoma (SNO)>hepatoblastoma (HepG2). Activity in the murine fibroblast line NIH-3T3 was comparable to that in HepG2 cells. MS09 lipoplexes achieved approximately three-times and two-times greater activity than Lipofectin complexes in HeLa and SNO cells, respectively, whilst comparable levels were recorded in HepG2 and NIH-3T3 cells. MS09 lipoplexes were well tolerated by HepG2, HeLa and SNO cells with cell numbers found to be 80, 85 and 75% of untreated cultures, respectively, at the optimal transfection concentration. These lipoplexes also exhibited high activity in the presence of 10% foetal bovine serum (FBS) in HeLa (17% inhibition) and HepG2 (33% inhibition) cells.     

Bcl-2 overexpression protects human keratinocyte cells from Ukrain-induced apoptosis but not from G2/M arrest

Exposure of ME180 and A431 carcinoma cells to Ukrain (NSC-631570), a semisynthetic compound consisting of alkaloids isolated from Chelidonium majus L. (Papaveracea), results in cell cycle arrest at the G2/M phase. Ukrain selectively inhibits growth of ME180 and A431 cells at a concentration range from 3.5 microM to 7.0 microM and induces apoptosis. In contrast, normal human keratinocytes showed no difference in the kinetics of progression through the cell cycle in response to this compound. We found that at a concentration of 7.0 microM of this drug Bcl-2 protein overexpression protected HaCaT cell line keratinocytes against apoptosis induced by Ukrain but did not prevent G2/M arrest. Following exposure of normal keratinocytes to Ukrain, we detected an increase in Bcl-2 protein levels and a significant change in protein modification as suggested by observation of its different isoform with shifted electrophoretic mobility. Bcl-2 protein expression and its isoform distribution did not change substantially in ME180 and A431 carcinoma cells. We also suggest that drug-induced mitotic arrest and apoptosis represent dual Ukrain action on cell cycle progression machinery and Bcl-2-involved program cell death in the cell.     

Lopinavir shows greater specificity than zinc finger ejecting compounds as a potential treatment for human papillomavirus-related lesions

Non-surgical, antiviral treatment options are desirable for HPV-related lesions within the genitourinary and upper digestive tract. We compared the toxicity of three zinc finger-ejecting (ZFE) compounds (4,4-dithiodimorpholine, azodicarbonamide, and diamide) to the HIV protease inhibitor lopinavir using HPV-positive SiHa, CaSki, HeLa, ME180, and HPV-negative C33A cervical carcinoma cell lines as well as primary human foreskin keratinocytes (PHFKs). Colorimetric growth assays revealed selective toxicity when treated with lopinavir. All carcinoma cell lines, except CaSki, were sensitive to 20 μM lopinavir whereas primary PHFKs were highly resistant. In contrast, 4,4-dithiodimorpholine was uniformly toxic to all cells tested while azodicarbonamide and diamide showed no effect at all. It is concluded that lopinavir may be an attractive candidate to treat pre-cancerous and cancerous HPV-positive lesions.