NRP -1 b1/b2单克隆抗体对裸鼠胃癌移植瘤生长的影响

目的：探讨神经纤毛蛋白1（neuropilin -1,NRP -1）在胃癌细胞 BGC823中的表达情况,并观察自主研发的抗 NRP -1 b1/ b2单克隆抗体（NRP -1 mAb）对裸鼠胃癌移植瘤生长的影响。方法：基于实验室前期已成功制备出纯度和浓度均较高的 NRP -1 mAb。利用流式细胞术检测该抗体亲和性、细胞免疫荧光定位检测 NRP -1的分布及 NRP -1 mAb 特异性。建立胃癌移植瘤裸鼠模型,向移植瘤中注射不同浓度的 NRP -1 mAb,研究抗体对胃癌裸鼠皮下移植瘤生长的影响。2周后根据瘤体大小得出抑瘤率。最后免疫组化分析癌组织内血管内皮生长因子（VEGF）的表达情况。结果：胃癌细胞 BGC -823上存在 NRP -1,且主要分布在细胞膜上,能与自制的 NRP -1 mAb 有效结合。通过动物实验,NRP -1 mAb 能有效抑制移植瘤的生长,高浓度抗体较低浓度对移植瘤抑制作用更明显,NRP -1 mAb 治疗组中 VEGF 表达较空白组低（P <0.01）。结论：胃癌细胞 BGC -823细胞膜上表达 NRP -1,NRP -1 mAb 能特异性结合胃癌细胞上 NRP -1蛋白,NRP -1 mAb 能抑制胃癌裸鼠移植瘤的生长,且与浓度呈正相关,可能与下调 VEGF 表达有关。     

Suramin联合DDP对人鼻咽癌裸鼠移植瘤生长的抑制作用

目的探讨suramin对人鼻咽低分化鳞癌裸鼠移植瘤生长的影响。方法建立人鼻咽低分化鳞癌细胞株（CNE-2）裸鼠移植瘸模型。将24只荷瘤裸鼠随即机分为4组；生理盐水对照组、suramin组、DDP组和suramin＋DDP组。每组6只。各组裸鼠分别经腹腔注射生理盐水、suramin（0．1mg／g·d^-1）、DDP（0．01mg／g·d^-1）、suramin和DDP（suramin0．1mg／g·d^-1，DDP0．01nag／g·d^-1），每天1次，连续10d后改为每周给药2次。共4w。将移植瘤完整切除。并称其重量。在光学显微镜和电子显微镜下观察移植瘤细胞形态变化，采用免疫组化法检测瘤组织中血管内皮生长因子（vascular endotheliai growth factor,VEGF）表达情况，TUNEL法检测移植瘤细胞凋亡。结果腹腔分别注射suramin、DDP和suramin＋DDP后，人鼻咽癌BALB／C裸鼠移植瘤生长被抑制，各组抑瘤率分别为28．42％、57．52％和71．35％，与对照组相比有明显差异，suramin联合DDP其抑瘤率明显增加。对照组VEGF表达为（75．17±9．0）％。suramin组、DDP组和suramin＋DDP组的VEGF表达明显减少，分别为（43．83±7．5）％、（67．33±8．3）％和（35．50±6．8）％，与对照组相比。suramin组和suramin＋DDP组的VEGF表达有明显差异（P〈0．01）；suramin组和suramin＋DDP组的细胞凋亡率分别为（23．83±7．5）％和（35．50±6．8）％。与对照组（6．17±4．0）％相比有明显差异（P〈0．01）。结论suramin可抑制人鼻咽低分化鳞癌BALB／C裸鼠移植瘤的生长，suramin对人鼻咽低分化鳞癌BALB／C裸鼠移植瘤的生长抑制作用可能与其诱导癌细胞凋亡增加、抑制VEGF表达有关。suramin与传统化疗药物联用可起协同作用。     

裸鼠肺癌移植瘤中NRP-1的表达及其意义

目的:探讨非小细胞肺癌(non-smallcelllungcancer,NSCLC)中神经纤毛蛋白-1(neuropilin-1,NRP-1)的表达与肿瘤发展及血管新生的关系。方法:用RT-PCR方法体外检测人肺腺癌细胞株A549中血管内皮生长因子(vascularendo-thelialgrowthfactor-165,VEGF165)mRNA的表达。此细胞株皮下接种裸鼠,建立两组荷瘤时间不同的肺癌移植瘤模型,两组移植瘤块均经常规HE染色鉴定。用免疫组化S-P法检测两组移植瘤块中NRP-1的表达,并用全自动图像分析系统分析其平均吸光度,同时对两组移植瘤内微血管密度(microvesseldensity,MVD)值进行测定。结果:A549细胞株表达VEGF165mRNA,经常规HE染色鉴定裸鼠肺癌皮下移植瘤模型建立成功。NRP-1在瘤内血管内皮细胞及部分A549细胞的胞浆中有表达。两组移植瘤中NRP-1的平均吸光度分别为0·11±0·02,0·18±0·02(P<0·05),MVD值分别为13±1·58,24±2·92(vessels/mm2)(P<0·05),且NRP-l的平均吸光度与MVD值呈线性正相关(Pearson相关系数r=0·92,P=0·0002)。结论:两组荷瘤时间不同的裸鼠肺癌皮下移植瘤中NRP-1的表达及MVD值随时间进展增加,肿瘤中NRP-1的表达与血管新生有关。     

裸鼠肺癌移植瘤中NRP-1的表达及其意义

Objective:To establish angiogenesis model of xenografts of lung cancer cell in nude mouse and investigate the expression of the neuropilin-1 (NRP-1) protein in tumors and its role in progression and angiogenesis of lung cancer.Methods:Human lung adenocarcinoma cells A549 were analyzed for the expression of vascular endothelial growth factor165(VEGF165)mRNA using RT-PCR in vitro.TWo groups of nude mice were subcutaneously inoculated with A549 at different tumor-loading time.Two groups of xenografts were jdentified by hematoxylin and eosin (HE) staining.their microvessel density (MVD) were analyzed meanwhile.Two groups were analyzed for the expression of NRP-1 protein and their mean absorbency by using immunohistochemistry and automatic image analysis system respectively.Results:A549 expressed VEGF165 mRNA,and xenografts of A549 in nude mice were successfully established and confirmed by HE staining.The atypia of cancer cells and angiogenesis were occurred in two groups.Two groups of MVD were 13.06±1158.23.61±3.11(vessels/mm2)(P＜0.01).NRP-1 protein was expressed in cytoplasm of vascular endothelium cells and partial tumor cells.Two groups of mean absorbency of NRP-1 were 0.1095±O.0228,0.1784±0.0151 (P＜0.01).Conclusion:The angioqenesis models of xenografts in nude mice with lung cancer cell A549 expressing VEGF165 mRNA at different tumor-loading times were established successfully.The expression of NRP-1 protein and MVD were increased with the tumor progression.Our results demonstrate that NRP-1 protein in lung cancer is related to angiogenesis.     

姜黄素下调HIF-1α/MMP-9通路抑制人胃癌裸鼠移植瘤生长

目的 研究姜黄素对人胃癌细胞株BGC-823裸鼠皮下移植瘤的生长抑制作用及其机制.方法 以0、1、5、10、20、50mg/L的姜黄素处理BGC-823细胞24或48 h,甲基噻唑盐(MTT)法及Transwell 分析细胞增殖和侵袭能力;荷瘤裸鼠检测姜黄素对肿瘤生长影响,同时Western blot检测瘤体内HIF-1α、MMP-9蛋白的表达;荧光素酶报告基因系统检测姜黄素对HIF-1α及HIF-1α对MMP-9启动子区域调控.结果 姜黄素处理人胃癌细胞BGC-823后,在体内体外的生长均受到抑制且作用呈量效依赖关系;细胞侵袭能力明显降低;HIF-1α、MMP-9蛋白的表达降低.姜黄素通过转录水平抑制HIF-1α及MMP-9的启动子活性.结论 姜黄素在体内外均抑制胃癌的生长,它可以通过降低HIF-1α活性,下调MMP-9水平,从而抑制肿瘤侵袭,发挥抗肿瘤作用.     

裸鼠的人原发性胃癌移植瘤生物学及分子遗传学性质研究

A human primary gastric cancer tissue (adenocarcinoma II-III) was transplanted into nude mice (SWISS/DF. nu/nu). It has been transferred for 8 generations at 56 sites in 28 nude mice with transplantable rate of 100%. The transplanted tumor is designated as transplantable human primary gastric cancer-1 in nude mice (THPGC-1). The growth of THPGC-1 is rather rapid and the size of transplanted tumor reaches 1 cm2, 4-5 weeks after transfer. The morphology and histochemistry of the original tumor were retained well in the initial and serial transplanted tumors. THPGC-1 could secret carcinoembryonic antigen (CEA). After intravenous or intraperitoneal injection of 131I-antiCEA monoclonal antibody into the THPGC-1 bearing nude mice, the radiolabeled antibody was concentrated and localized in the tumor as shown by gamma-camera analysis. Similar pattern of lactate dehydrogenase isoenzyme was observed both in primary gastric cancer tissue and THPGC-1 tissue. Chromosomal examination revealed that THPGC-1 was human aneuploid ones. Southern blot analysis showed that the pattern of repetitive DNA bands and the structures of 28s, rDNA, c-H-ras and c-myc genes in THPGC-1 were identical to the original primary gastric cancer DNA. The results suggest that THPGC-1 be a reliable model for the research of the molecular biology of cancer cells and experimental gastric cancer diagnosis and treatment.     

人胃癌转移型裸鼠移植瘤cDNA文库构建及分析

从人转移型胃癌裸鼠移植瘤中抽提出总RNA,分离出多聚腺苷酸RNA,合成双链cDNA,接上ECoRI接头,以pT7T3 18U质粒为载体,以大肠杆菌NM522为受体菌,构建了人转移型胃癌裸鼠移植瘤cDNA文库。该文库含1.5×10~6转化子,插入片段cDNA长度为0.4～6.0Kb,重组效率为80％,并用四种癌基因探针筛选了阳性克隆。     

贝伐单抗与重组人血管内皮抑素对人乳腺癌MCF-7细胞裸鼠移植瘤生长的影响

[目的]探讨贝伐单抗、重组人血管内皮抑素对人乳腺癌MCF-7细胞裸鼠移植瘤生长的影响。[方法]建立人乳腺癌裸鼠移植瘤模型,随机分为对照组、低剂量贝伐单抗组、高剂量贝伐单抗组、低剂量莺组人血管内皮抑素组、高剂量重组人血管内皮抑素组、低剂量联合组以及高剂量联合组,用药3周。检测裸鼠体重、移植瘤体积、移植瘤重量,计算抑瘤率。[结果]与对照组相比,低剂量贝伐单抗组、高剂量贝伐单抗组、低剂量联合组、高剂量联合组裸鼠移植瘤生长曲线较平缓,移植瘤重量明显下降（P〈0．01）,抑瘤率分别为67．69％、68．88％、78.32％和79．26％。低剂量联合组与低剂量贝伐单抗组移植瘤重量存在统计学差异（P〈0．05）。低剂量重组人血管内皮抑素组、高剂量重组人血管内皮抑素组移植瘤生长与对照组无统计学差异（P〉0．05）。[结论]贝伐单抗能抑制人乳腺癌MCF-7细胞裸鼠移植瘤生长,低剂量贝伐单抗联合重组人血管内皮抑素能进一步提高抗肿瘤作用。     

姜黄素下调HIF-1α/MMP-9通路抑制人胃癌裸鼠移植瘤生长北大核心CSCD

目的研究姜黄素对人胃癌细胞株BGC-823裸鼠皮下移植瘤的生长抑制作用及其机制。方法以0、1、5、10、20、50mg/L的姜黄素处理BGC-823细胞24或48h,甲基噻唑盐(MTT)法及Transwell分析细胞增殖和侵袭能力;荷瘤裸鼠检测姜黄素对肿瘤生长影响,同时Western blot检测瘤体内HIF-1α、MMP-9蛋白的表达;荧光素酶报告基因系统检测姜黄素对HIF-1α及HIF-1α对MMP-9启动子区域调控。结果姜黄素处理人胃癌细胞BGC-823后,在体内体外的生长均受到抑制且作用呈量效依赖关系;细胞侵袭能力明显降低;HIF-1α、MMP-9蛋白的表达降低。姜黄素通过转录水平抑制HIF-1α及MMP-9的启动子活性。结论姜黄素在体内外均抑制胃癌的生长,它可以通过降低HIF-1α活性,下调MMP-9水平,从而抑制肿瘤侵袭,发挥抗肿瘤作用。     

常规化疗联合节拍化疗对乳腺癌裸鼠移植瘤的实验研究

背景与目的:乳腺癌治疗应从全局出发,将针对肿瘤细胞本身和肿瘤微环境相结合,而研究发现节拍化疗在抗血管生成等改善肿瘤微环境方面有明显优势.本文旨在观察环磷酰胺(cyclophosphamide,CTX)常规化疗联合节拍化疗对乳腺癌裸鼠移植瘤的抑瘤效应,探讨其对血管新生和细胞增殖、凋亡的影响.方法:建立乳腺癌裸鼠原位移植瘤模型,随机分成4组:节拍化疗(LDM)组、常规化疗(MTD)组、联合(LDM+MTD)组和0.9％NaCl溶液对照组,治疗期间观察裸鼠一般状况、隔日称重并测量皮下移植瘤体积,每周尾静脉采血白细胞计数(white blood cell counts,WBC).实验结束后处死小鼠取瘤称重,计算各组抑瘤率.免疫组织化学法检测移植瘤组织中微血管密度(microvessel density,MVD),以及血管内皮细胞生长因子(vascularendothelial growth factor,VEGF)、凝血酶敏感蛋白1(thrombospondin-1,TSP-1)、增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)表达,TUNEL检测肿瘤细胞凋亡.结果:CTX 3种化疗方案均可不同程度抑制移植瘤生长,其中LDM组和MTD组移植瘤生长曲线类似,LDM+MTD组移植瘤生长明显缓于其他各组;LDM组、MTD组和LDM+MTD组的抑瘤率分别为32.95％、41.57％和69.15％; LDM组和LDM+MTD组MVD、VEGF相对低表达、TSP-1相对高表达,而组间比较差异无统计学意义(P＞0.05),但与其余两组比较差异有统计学意义(P＜0.05).MTD组和LDM+MTD组PCNA相对低表达,与其余两组比较差异有统计学意义(P＜0.05),LDM+MTD组凋亡指数(apoptosis index,AI)明显高于其他各组,差异有统计学意义(P＜0.05).结论:CTX常规化疗联合节拍化疗兼有抗血管生成、抑制细胞增殖、促进肿瘤细胞凋亡作用,抑瘤作用较单一传统化疗和节拍化疗更为明显,不良反应不明显.     

西妥昔单抗单用及联合应用ECF化疗方案对胃癌裸鼠移植瘤生长的影响

目的:探讨表皮生长因子受体特异性单抗西妥昔单抗(C225)单药及联合应用ECF(表柔比星、顺铂和5-FU)化疗方案对胃癌裸鼠移植瘤生长的影响.方法:将胃癌SGC-7901细胞制成细胞悬液后接种于裸鼠皮下,待成瘤后将裸鼠随机分为生理盐水组、ECF化疗组、单纯西妥昔单抗组和西妥昔单抗联合ECF化疗组.定期测量每只裸鼠肿瘤的长、短径计算肿瘤体积.用药结束后,处死裸鼠, 完整剥取肿瘤组织,称重,计算抑瘤率.用免疫组化方法检测肿瘤组织中表皮生长因子受体和增殖细胞核抗原的表达.结果:与生理盐水组相比,ECF化疗组抑瘤率为22.5%,西妥昔单抗组抑瘤率为20%,西妥昔单抗联合ECF化疗组抑瘤率为25%,各组肿瘤体积变化及治疗后平均肿瘤重量差异均无统计学意义.免疫组化显示,西妥昔单抗可以抑制胃癌细胞膜表皮生长因子受体的活化,西妥昔单抗联合ECF化疗可以抑制胃癌细胞膜增殖细胞核抗原的增殖活性.结论:西妥昔单抗单药及联合应用ECF化疗方案对胃癌裸鼠移植瘤生长无明显影响,在免疫组化方面发现一些有意义的影响.     

单克隆抗体CH12抑制头颈部鳞状细胞癌裸鼠种植瘤的生长

目的:观察嵌合型单克隆抗体CH12对头颈部鳞状细胞癌(head and neck squamous cell carcinomas,HNSCC)裸鼠种植瘤生长的抑制作用,为进一步研究CH12单抗在肿瘤治疗中的作用提供参考数据。方法:Western blotting检测5种HNSCC细胞系A253、CAL27、Detroit 562、FaDu和RPMI 2650中表皮生长因子受体(epidermal growth factor receptor,EGFR)的表达,流式细胞术检测CH12单抗同这5种细胞系的结合能力。皮下接种CAL27和A253细胞,建立HNSCC裸鼠种植瘤模型。模型鼠腹腔注射CH12单抗,以PBS作为阴性对照,观察肿瘤生长情况,绘制肿瘤生长曲线。结果:EGFR在CAL27、A253、FaDu及Detroit 562细胞中均有不同程度的表达,其中CAL27细胞中EGFR的表达水平最高,A253细胞次之。CH12单抗与5种HNSCC细胞系的结合能力由高到低依次为CAL27、FaDu、A253、Detroit 562和RPMI 265细胞。CH12单抗对CAL27和A253细胞裸鼠种植瘤的生长均有显著抑制作用,抑瘤率分别为56.8%(P=0.022)和59.7%(P=0.015)。结论:单克隆抗体CH12对EGFR高表达的HNSCC细胞种植瘤的生长具有明显的抑制作用。     

Low-dose docetaxel combined with (-)-epigallocatechin-3-gallate inhibits angiogenesis and tumor growth in nude mice with gastric cancer xenografts

Low-dose metronomic (LDM) chemotherapy represents a new strategy to treat solid tumors by stronger antiangiogenic activity and less side-effects, especially in combination with other antiangiogenic agents. The aim of the study is to investigate the antiangiogenic effect of docetaxel alone and combined with (-)-epigallocatechin-3-gallate (EGCG) in preclinical settings of gastric cancer. BGC-823 human gastric cancer xenograft model was used, and tumor growth, side-effects of mice were closely monitored. Expression of vascular endothelial growth factor and CD31 were observed by immunohistochemistry, and microvessel density of the tumor tissues was assessed by CD31 immunohistochemical analysis. Our results indicated that LDM docetaxel inhibited angiogenesis and growth of gastric cancer with less toxicity, and the effects were further enhanced by the concurrent administration of EGCG. Our study, for the first time, rationally demonstrated that LDM docetaxel treatment used alone or combined with EGCG is effective and safe in preclinical settings of gastric cancer. Our data suggest that LDM docetaxel used alone or combined with EGCG may be an innovative and promising therapeutic strategy in the experimental treatment of human gastric cancer.     

CXCR4单克隆抗体抑制乳腺癌MCF-7细胞裸鼠移植瘤的实验研究

目的：研究CXC趋化因子受体4（CXC chemokine receptor 4,CXCR4）单克隆抗体（CXCR4 mAb）对人乳腺癌MCF-7细胞裸鼠皮下移植瘤生长的影响,并初步探讨CXCR4 mAb抗肿瘤的作用机制。方法：采用8周龄Balb/c雌性裸鼠,建立乳腺癌MCF-7细胞裸鼠皮下移植瘤模型。运用CXCR4 mAb进行干预,从整体水平观察CXCR4 mAb对肿瘤生长的影响,采用免疫组织化学法检测肿瘤组织中增殖细胞核抗原（PCNA）、半胱氨酸天冬氨酸酶3（Caspase-3）和血管内皮细胞生长因子（VEGF）的表达情况。结果：CXCR4 mAb可明显抑制移植瘤的生长,瘤体抑制率达到71.4%;CXCR4 mAb治疗后的肿瘤组织中PCNA和VEGF表达明显下降,而Caspase-3表达上升。结论：CXCR4 mAb可能是通过抑制肿瘤细胞增殖、促进肿瘤细胞凋亡及抑制肿瘤血管形成而发挥抗肿瘤生长的作用。     

Synergistic antitumor activity of combination chemotherapy with mitomycin C and cisplatin against human gastric cancer xenografts in nude mice

A new combined cancer chemotherapy regimen of mitomycin C (MMC) and cisplatin (DDP) showed synergistic antitumor activity against human gastric cancer xenografts St-40 and SC-1-NU in BALB/c nu/nu mice. The drugs were administered intraperitoneally at doses of 2 or 4 mg/kg for MMC and 3 or 6 mg/kg for DDP, respectively. To clarify the schedule-dependent antitumor activity of MMC and DDP against St-40 and SC-1-NU, different sequential therapies were conducted. Simultaneous administration of these agents showed the highest antitumor activity against SC1-NU among the three regimens used, whereas the sequence of MMC followed by DDP showed higher antitumor activity than the reverse sequence against St-40. The intratumoral concentration of platinum was significantly increased in St-40 treated with the sequence MMC to DDP, in comparison with the sequence DDP to MMC. The maximum tolerated dose (MTD) of this combination was 4 mg MMC plus 6 mg DDP per kg in all the combinations, and these MTDs were 2/3 of the corresponding values for their single use. Since this combination increased the antitumor activity of each single agent without any increase in their toxicity, it would appear to be useful clinically. © 1994 Wiley-Liss, Inc.     

CXCR4单克隆抗体抑制乳腺癌MCF-7细胞裸鼠移植瘤的实验研究

目的 研究CXC趋化因子受体4(CXC chemokine receptor 4,CXCR4)单克隆抗体(CXCR4 mAb)对人乳腺癌MCF-7细胞裸鼠皮下移植瘤生长的影响,并初步探讨CXCR4 mAb抗肿瘤的作用机制.方法 采用8周龄Balb/c雌性裸鼠,建立乳腺癌MCF-7细胞裸鼠皮下移植瘤模型.运用CXCR4 mAb进行干预,从整体水平观察CXCR4 mAb对肿瘤生长的影响,采用免疫组织化学法检测肿瘤组织中增殖细胞核抗原(PCNA)、半胱氨酸天冬氨酸酶3(Caspase-3)和血管内皮细胞生长因子(VEGF)的表达情况.结果 CXCR4 mAb可明显抑制移植瘤的生长,瘤体抑制率达到71.4%;CXCR4 mAb治疗后的肿瘤组织中PCNA和VEGF表达明显下降,而Caspase-3表达上升.结论 CXCR4 mAb可能是通过抑制肿瘤细胞增殖、促进肿瘤细胞凋亡及抑制肿瘤血管形成而发挥抗肿瘤生长的作用.     

Survivin基因沉默抑制结肠癌生长的动物实验研究

目的 探究survivin基因沉默对人结肠癌Lovo细胞裸鼠移植瘤生长的抑制作用.方法 构建靶向survivin的shRNA载体SUR和阴性对照质粒Neg,并将其转染人结肠癌Lovo细胞,分别种植到裸鼠皮下建立人结肠癌裸鼠移植瘤模型.随后观察各组裸鼠移植瘤生长情况,免疫组化方法检测移植瘤survivin蛋白的表达,TUNEL法检测肿瘤细胞的凋亡情况.结果 移植转染细胞8周,与空白对照组比较,SUR组移植瘤的体积和质量均有显著缩小(P<0.05),体积和质量抑瘤率分别为48.9%和51.2%.与空白对照组比较,SUR组移植瘤survivin表达显著下调,表达指数为31.9%;SUR组肿瘤细胞凋亡显著增加,凋亡指数18.47%(P<0.05).阴性对照Neg组的上述指标与空白对照组比较,差异均无统计学意义.结论 Survivin基因沉默能够抑制人结肠癌Lovo细胞裸鼠移植瘤的生长.     

Suramin inhibits growth of human osteosarcoma xenografts in nude mice

The effect of suramin on tumor growth and morphology in two different human osteosarcoma xenografts (L-I OSM and L-II OSM) grown in BALB/cA-nu/nu mice was studied. Suramin (total dose, 720 mg/kg) given by i.p. injection (60 mg/kg/dose) for up to 9 weeks significantly inhibited osteosarcoma cell growth in both tumors, suramin-treated tumors showing only one-third or less of the volume of nontreated controls. Cell cycle distribution of tumor cells measured by DNA flow cytometry demonstrated that suramin treated caused accumulation of cells in the S and G2 phases of the cell cycle, in both L-I OSM and L-II OSM. In the aneuploid L-II OSM tumor suramin preferentially inhibited the growth of aneuploid cells, leading to a decrease in the ratio of aneuploid to diploid cells. Both osteosarcomas retained their histological appearance and the liver, spleen, heart, and kidneys of the treated animals were unaffected by suramin. These results are compatible with the view that suramin inhibits the growth of human osteosarcomas by cytostatic effects.