~(125)I放射性粒子对荷人结肠癌裸鼠治疗效应的实验研究

目的观察125I放射性粒子组织间植入对裸鼠皮下人结肠癌移植瘤的局部抑制作用。方法雌性Balb/c裸鼠20只,以人Colo205型结肠癌细胞建立裸鼠皮下人结肠癌实体肿瘤模型,待肿瘤体积长至理想大小,随机分成2组,对照组不予任何处理,治疗组给予移植瘤以125I放射性粒子组织间植入,每周测量2次肿瘤体积,治疗27d后,处死裸鼠,观察移植瘤的相对肿瘤体积(RTV)、相对肿瘤增殖率(T/C)、ABC免疫组化染色观察肿瘤增殖率[Brdu标记指数(LI)]。结果治疗27d后,裸鼠均存活。治疗组(125I粒子组)移植瘤的RTV为(251.59±72.24),明显小于对照组的RTV(1087.94±510.72),相对肿瘤增殖率为23.13%,明显减少,治疗组的LI为(15.00±5.00),明显小于对照组的LI(38.75±6.29)。结论①人Colo205型结肠癌模型是放射性粒子植入治疗大肠癌基础研究的良好模型之一;②125I放射性粒子对人结肠癌细胞增殖有明显的局部抑制作用;③125I放射性粒子在肿瘤体内累积一定剂量才会逐渐显现治疗效应。     

人外周血γδT 细胞免疫治疗裸鼠皮下人肝癌细胞移植瘤的体内实验研究

目的:研究不同剂量的人外周血γδT 细胞对裸鼠人肝癌细胞(SMMC-7721)移植瘤模型的免疫治疗作用。方法: 用人肝癌细胞株SMMC-7721 接种BALB/ c 裸鼠皮下,建立肝癌裸鼠模型。于从健康人外周血中提取单核细胞,体外特异性扩增γδT 细胞。将已建立的肝癌裸鼠模型随机分为5 组,阳性对照组为5-氟尿嘧啶(5-Fu),阴性对照组为生理盐水,治疗组用不同剂量的γδT 细胞 分别注入裸鼠尾静脉,阳性对照组用5-Fu 裸鼠腹腔注射,阴性对照组用生理盐水裸鼠尾静脉注射。观察不同剂量的γδT 细胞对肿瘤的抑制效果,包括治疗前后的体重、食物摄取量及生长状况等,并与阳性对照组和阴性对照组比较肿瘤体积(TV)、相对肿瘤体积(RTV)和相对肿瘤增殖率[T/ C(%)]变化。结果:不同剂量的γδT 细胞对裸鼠移植瘤的生长有不同程度的抑制,RTV 与生理盐水阴性对照组比较差异有统计学意义(P<0.05);与5-Fu 阳性对照组比较,TV 增长明显低于5-Fu 阳性对照组,RTV 各剂量组抑制程度相似,均略高于5-Fu 阳性对照组,T/ C(%)各剂量组比5-Fu 对照组的相对肿瘤增殖率稍低,但无显著性差异。结论:人外周血γδT 细胞对肝癌裸鼠移植瘤具有显著的抑瘤作用,为建立肝癌免疫治疗新方法提供实验依据。     

不同输注途径对CIK细胞体内分布和抑瘤作用的影响

目的 研究不同输注途径(皮下、腹腔、瘤周注射)对CIK细胞在活体内的分布规律和抑瘤活性的影响.方法 以同位素32P-adATP标记CIK细胞,而后经尾静脉途径和腹腔途径注射入裸鼠体内,用放射性定量检测方法分析CIK细胞在各器官的动态分布.裸鼠皮下注射结肠癌细胞建立肿瘤活体模型,分别按瘤周、腹腔和尾静脉注射途径接受CIK细胞治疗,观察肿瘤的体积和重量变化.结果 CIK细胞输入裸鼠体内后,迅速分布至肝、脾、肾、肺、胃、肠等器官,其中肝、脾、肾的分布浓度最高,CIK细胞在各脏器的分布规律与不同的输注途径存在一定的联系.裸鼠皮下接种结肠癌细胞系HCT-8后,瘤周和尾静脉注射CIK细胞可以明显抑制皮下移植肿瘤的生长,而腹腔注射CIK细胞对于皮下肿瘤的抑制没有统计学意义.结论 CIK细胞体内分布的广泛性表明它可以用于身体各部位恶性肿瘤的治疗;针对不同解剖部位的肿瘤可以选择不同的CIK细胞输注途径以最大限度地提高疗效.     

人外周血γδT细胞免疫治疗裸鼠皮下人肝癌细胞移植瘤的体内实验研究北大核心CSCD

目的:研究不同剂量的人外周血γδT细胞对裸鼠人肝癌细胞(SMMC-7721)移植瘤模型的免疫治疗作用。方法:(1)用人肝癌细胞株SMMC-7721接种BALB/c裸鼠皮下,建立肝癌裸鼠模型。(2)从健康人外周血中提取单核细胞,体外特异性扩增γδT细胞。(3)将已建立的肝癌裸鼠模型随机分为5组,阳性对照组为5-氟尿嘧啶(5-Fu),阴性对照组为生理盐水,治疗组用不同剂量的γδT细胞(1×105、5×10~5及25×10~5)分别注入裸鼠尾静脉,阳性对照组用5-Fu裸鼠腹腔注射,阴性对照组用生理盐水裸鼠尾静脉注射。观察不同剂量的γδT细胞对肿瘤的抑制效果,包括治疗前后的体重、食物摄取量及生长状况等,并与阳性对照组和阴性对照组比较肿瘤体积(TV)、相对肿瘤体积(RTV)和相对肿瘤增殖率[T/C(%)]变化。结果:不同剂量的γδT细胞对裸鼠移植瘤的生长有不同程度的抑制,RTV与生理盐水阴性对照组比较差异有统计学意义(P<0.05);与5-Fu阳性对照组比较,TV增长明显低于5-Fu阳性对照组,RTV各剂量组抑制程度相似,均略高于5-Fu阳性对照组,T/C(%)各剂量组比5-Fu对照组的相对肿瘤增殖率稍低,但无显著性差异。结论:人外周血γδT细胞对肝癌裸鼠移植瘤具有显著的抑瘤作用,为建立肝癌免疫治疗新方法提供实验依据。     

人食管癌细胞系的建立及其转移相关基因PCNA的表达

目的:建立人食管癌细胞系,并对其肿瘤标志物(caycinoembryanic antigen,CEA)和转移相关基因增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)的表达进行研究。方法:将人食管癌手术标本移植至裸鼠皮下,成瘤后取组织块体外原代培养,光镜和电镜观察细胞形态,测定细胞周期和染色体众数。取部分裸鼠皮下移植瘤接种至裸鼠食管,观察裸鼠体内肿瘤转移特征,采用免疫组化法测定肝转移灶PCNA基因的表达。结果:裸鼠皮下移植瘤组织16d,可形成明显的瘤结节,体外培养获得大量的肿瘤细胞,命名为YXA-1,其形态和超微结构符合鳞状细胞癌的形态学特征,染色体保持人类肿瘤染色体的特点,众数为70～76,细胞中肿瘤标志物CEA。食管癌标本原位移植裸鼠21d时,发生了广泛的肝内侵袭和转移,转移灶中PCNA表达为阳性。结论:人食管癌细胞系YXA-1维持了原发肿瘤的特征,而裸鼠其原位移植模型体现了肿瘤的转移特性,可用于肿瘤的转移研究。     

RNA干扰Slug基因对HCT116细胞裸鼠移植瘤生长及转移的影响

目的：探讨通过靶向RNA干扰（RNA interference,RNAi）抑制Slug基因的表达,观测对荷瘤裸鼠结直肠癌生长及转移的影响。方法：利用结肠癌细胞株HCT116对24只5周龄裸鼠皮下种植,建立结肠癌裸鼠皮下移植瘤模型,设立空白对照组、阴性对照组及实验组三组,每组8只。分别注射生理盐水、阴性对照质粒及慢病毒载体,观察肿瘤生长情况,绘制肿瘤生长曲线,观察各组间肿瘤生长及淋巴结转移的变化,应用免疫组化、qRT-PCR和Western blot检测Slug基因和蛋白表达情况。结果：Slug基因shRNA实验组与空白对照组和阴性对照组相比,瘤体生长减缓,移植瘤质量明显减小（3.08±0.31 vs 7.37±1.18,7.46±1.16,P＜0.01）,Slug蛋白表达明显降低（P＜0.05）;实验组淋巴结阳性率为36.3％（4/11）,与阴性对照组77.8％（14/18）及空白对照组68.4％（13/19）相比较（P＜0.01）。结论：靶向Slug的RNA干扰可以显著抑制结肠癌裸鼠模型的生长、淋巴结转移以及癌组织中Slug基因蛋白的表达,可能成为结肠癌基因治疗的潜在分子靶点。     

自体肿瘤特异性CTLs对裸鼠人乳腺癌移植瘤的抑瘤作用

目的:研究乳腺癌患者腋下淋巴结细胞诱导产生的肿瘤特异性CTLs在裸鼠体内对患者自身乳腺癌生长的抑制作用.方法:在裸鼠胸垫处种植人乳腺癌组织, 建立裸鼠乳腺癌移植瘤模型.以细胞因子联合诱导乳腺癌患者腋窝淋巴结单个核细胞成为DC和肿瘤特异性CTL(sCTL)、非特异性CTL(nCTL)和异体CTL(vCTL);将不同CTLs注射到荷瘤裸鼠的肿瘤周围, 观察其对肿瘤生长的抑制作用.结果:人乳腺癌移植瘤在裸鼠体内的成活率为100%.组织病理学证实移植瘤为乳腺浸润性导管癌.与对照组相比, 荷瘤裸鼠移植瘤的体积在sCTL、nCTL和vCTL治疗组均明显减小(P＜0.05), sCTL对自体移植瘤的生长抑制作用最强[平均瘤体积(82.70±2.09) mm3], 抑瘤率50.5%, 明显高于nCTL组[(96.15±5.35) mm3, 26.9%]和vCTL组[(96.93±4.51) mm3, 25.7%], (P＜0.01);不同CTL治疗组瘤组织内可见大量淋巴细胞和树突状细胞(DC)浸润.结论:乳腺癌裸鼠移植瘤模型成功率高, 病理学证实与人乳腺癌特征相符, 且有DC和大量特异性CTL浸润.自体特异性CTL对裸鼠体内自体乳腺癌移植瘤有较强的抑制作用.     

下调CDH17基因对那可丁耐药性人胃癌细胞裸鼠移植瘤模型的影响

研究下调CDH17基因对那可丁耐药的人胃癌MGC-803细胞裸鼠移植瘤模型的影响,为临床上寻求治疗胃癌新方案提供理论依据。以人胃癌MGC-803细胞作为研究材料,建立该细胞的裸鼠移植瘤模型。同时,用HE染色法观察瘤体组织病理学形态。结果显示,RT-PCR和Western blotting实验中siCDH17慢病毒对胃癌MGC-803细胞中CDH17的转录水平和蛋白质表达水平均具有干扰效果(P<0.05)。随后将siCDH17慢病毒干扰的人胃癌MGC-803细胞以及对照细胞进行裸鼠皮下移植瘤实验。移植后第6天各组均见皮下肿瘤结块,采用那可丁药物处理并实时检测肿瘤生长情况。结果发现,siCDH17组肿瘤体积和质量明显小于对照组和siNC组(P<0.05)。且HE染色也发现与对照组比较,siCDH17组的瘤体肿瘤细胞生长受到抑制。因此,下调CDH17基因可以增加胃癌MGC-803细胞对那可丁的敏感性,抑制胃癌MGC-803细胞的生长,最终阻滞裸鼠移植瘤的生长。     

人肺腺癌裸鼠皮下移植瘤淋巴转移模型的构建与筛选

目的构建人肺腺癌的裸鼠皮下移植瘤模型,探讨不同注射部位对肿瘤生长和淋巴转移的影响。方法 20只BALB/c裸鼠随机分为4组,每组5只。分别于左侧背部近腋窝、右侧背部近腋窝、左侧后肢和左后爪垫皮下注射A549细胞株,建立人肺腺癌移植瘤模型,考察各组裸鼠肿瘤生长和淋巴结转移情况。结果各组裸鼠成瘤明显,模型构建成功。背部近腋窝种植2组裸鼠肿瘤比后肢和爪垫种植的肿瘤出现时间早,生长速度快(P<0.05)。左侧背部近腋窝、右侧背部近腋窝、左侧后肢和左后爪垫皮下移植瘤的淋巴结转移率分别为41.7%、42.9%、23.1%和21.4%。其中左侧背部近腋窝注射最为方便。结论裸鼠左侧背部近腋窝皮下注射A549细胞操作方便,肿瘤易生长,淋巴转移率高,是构建人肺腺癌皮下移植瘤淋巴转移模型的优选方案。     

生长分化因子11对鼻咽癌转移抑制及其放射处理后免疫功能的保护作用北大核心CSCD

目的:探究生长分化因子11(GDF11)对鼻咽癌转移的影响,及其对裸鼠鼻咽癌移植瘤模型放射处理后免疫功能的作用。方法:使用不同浓度的GDF11(10、50、100μg/L)处理鼻咽癌5-8F细胞,以正常培养的5-8F细胞作为对照组,CCK-8法检测细胞增殖活性,流式细胞术检测细胞周期分布,Transwell实验检测细胞迁移与侵袭,细胞划痕实验检测划痕愈合情况。裸鼠左后肢皮下接种5-8F细胞悬液构建鼻咽癌移植瘤模型,将建模成功的15只裸鼠随机分为3组,每组5只,并进行相应处理:6 Gy组裸鼠给予6 Gy局部照射,1次/6 d,共3次,同时腹腔注射生理盐水;6 Gy^(+)GDF11组裸鼠给予6 Gy局部照射,1次/6 d,共3次,同时腹腔注射0.5 mg/kg GDF11;对照组裸鼠不给予照射,腹腔注射生理盐水。每隔2 d测量裸鼠移植瘤体积,18 d末次照射给药结束后,麻醉裸鼠,腹主动脉取血,处死裸鼠取其肿瘤、淋巴结、胸腺、脾脏、肺、肝、肾组织,电子天平称量肿瘤、胸腺、脾脏组织重量,HE染色观察肺、肝、肾组织病理学变化,全自动动物血液分析仪测定血常规指标红细胞(RBC)、白细胞(WBC)、血小板(PLT),计算胸腺指数和脾脏指数,流式细胞术检测T淋巴细胞亚群CD4^(+)、CD8^(+)表达比例。结果:与对照组5-8F细胞比较,经过10、50、100μg/L GDF11处理48 h、72 h、96 h后细胞增殖活性下降,细胞发生G0/G1期阻滞,细胞迁移数目和侵袭数目减少,细胞划痕愈合率降低,差异均具有统计学意义(P<0.01);与对照组裸鼠比较,在处理6 d、9 d、12 d、15 d、18 d后6 Gy组和6 Gy^(+)GDF11组的裸鼠肿瘤体积变小,18 d测量发现裸鼠肿瘤块和质量减小,均未发生淋巴结转移,RBC、PLT和WBC降低,T淋巴细胞亚群CD4^(+)、CD8^(+)比例降低,差异均具有统计学意义(P<0.01);相较于6 Gy组裸鼠,6 Gy^(+)GDF11组的裸鼠肿瘤体积变小,18 d的裸鼠肿瘤块和质量减小,RBC、PLT和WBC水平升高,T淋巴细胞亚群CD4^(+)、CD8^(+)比例升高,差异均具有统计学意义(P<0.01)。结论:GDF11能够抑制鼻咽癌细胞的生长与转移,抑制裸鼠鼻咽癌移植瘤模型的肿瘤生长,并在放射处理后发挥裸鼠免疫功能的保护作用。     

上调miR-187*表达对人结肠癌细胞株增殖活性的影响

 目的：研究微小RNA-187*(miR-187*)在人结肠癌细胞株及正常结肠组织中的表达,同时分析miRNA-187*上调对人结肠癌细胞增殖和细胞周期的影响。方法：选取3例结直肠癌组织及配对正常黏膜组织进行miRNA芯片检测,筛选出大肠癌组织中异常表达的miRNA;提取8株结肠癌细胞株和10例正常结肠组织中总RNA,采用Taqman 实时定量PCR方法检测miR-187*的表达;预测miR-187*的靶基因,采用实时定量PCR方法检测靶基因的表达;采用脂质体介导的转染方法将miR-187*模拟物转染人结肠癌细胞株HCT116;检测转染后miR-187*和靶基因的表达;通过MTS试剂盒检测细胞增殖能力,流式细胞术检测miR-187*对细胞周期的影响。结果：miRNA芯片检测结直肠癌组织中miR-187*的表达较正常黏膜明显降低;miR-187*在结肠癌细胞株中的表达较正常结直肠黏膜组织明显下调;而B细胞特异性莫洛尼小鼠白血病病毒整合位点1（BMI-1） mRNA在结肠癌细胞株中表达较正常黏膜组织中明显增高;转染miR-187*模拟物后,miR-187*表达明显增加;同时miR-187*的高表达可以显著抑制BMI-1 mRNA的水平;miR-187*模拟物转染组和阴性对照组相比,细胞增殖活力明显受抑制（P＜0.05）,同时增殖细胞核抗原表达亦明显降低;细胞周期检测结果显示,miR-187*模拟物转染组G2/M期细胞增多,和阴性对照组间差异有统计学意义。结论：miR-187*在人结肠癌细胞株中表达下调;上调miR-187*表达可抑制结肠癌细胞增殖活性,影响结肠癌细胞周期;miR-187*可能通过抑制BMI-1 在结肠癌中发挥抑癌作用。     

Heterogeneity of Tie2 expression in tumor microcirculation: influence of cancer type, implantation site, and response to therapy

To evaluate the expression of the Tie2/Tek tyrosine kinase receptor in tumor blood vessels, we examined Tie2lacZ(+)/RAG1(-) mice. There was considerable heterogeneity (Tie2-negative, Tie2-positive, or Tie2-composite blood vessels) in subcutaneous xenografts of human colorectal carcinoma (HCT116; 97.5% Tie2-positive vessels) versus human melanoma (WM115; 75.9% Tie2-positive vessels). Similar patterns of Tie2 expression occurred in abdominal metastases derived from the same cell lines. Immunostaining for endothelial markers and Tie2 revealed that endogenous protein levels corresponded with transgene activity. Endothelial cells were confirmed to be of mouse origin through triple immunofluorescence staining with mouse antiserum to human nuclei, isolectin GS-IB(4), and anti-Tie2. Similar Tie2 heterogeneity was observed in clinical specimens from a variety of human cancers, including malignant melanoma and colorectal carcinoma. We also examined the effect of Tek-Delta Fc anti-angiogenic therapy on tumor growth and Tie2 expression patterns in HCT116 and WM115 subcutaneous xenografts. Tek-Delta induced extensive tumor regression in HCT116 tumors and concomitant reductions in Tie2-expressing blood vessels. However, no significant responses were seen in Tek-Delta-treated WM115 tumors. Thus, vascular heterogeneity of Tie2 expression is cancer-type specific, suggesting that the tumor microenvironment and/or direct cancer cell interactions influence Tie2 endothelial expression.     

慢病毒介导的RNA干扰沉默Slug基因对结肠癌HCT116细胞增殖和凋亡的影响

目的:探讨利用RNA干扰(RNA interference,RNAi)技术沉默Slug基因,观察对结肠癌HCT116细胞增殖和周期的影响。方法:构建Slug基因特异性siRNA慢病毒载体,感染结肠癌HCT116细胞,设立空白对照组、阴性对照组及SlugsiRNA三组,应用Real-time PCR和Western blot方法分别从基因和蛋白质水平检测各组干扰质粒对Slug基因的干扰效果,MTT法检测Slug基因在siRNA作用下的细胞增殖率,流式细胞仪检测细胞凋亡周期变化情况。结果:转染Slug siRNA后,结肠癌HCT116细胞中Slug基因mRNA和蛋白表达明显受到抑制(P<0.05);MTT检测,干扰组细胞增殖水平明显低于阴性对照组;流式细胞仪检测细胞G1期细胞百分比(52.3±0.6)高于阴性对照组(45.1±0.3,P<0.05)。结论:Slug siRNA能明显下调靶基因Slug的表达,在体外可抑制结肠癌HCT116细胞的生长并促进其凋亡。     

错配修复基因hMLH1在雌激素诱导结肠癌细胞凋亡中的作用

 摘 要 目的：探讨错配修复基因hMLH1在雌激素诱导结肠癌细胞凋亡中的作用。方法：将含野生型hMLH1-cDNA的质粒转入hMLH1缺陷的人结肠癌细胞株HCT116,在不同浓度的雌激素干预下,分别以流式细胞仪和活细胞计数试剂盒（cell counting kit-8）法检测转染后细胞的凋亡率和活性。结果：与空载组相比,转染hMLH1后雌激素能明显诱导HCT116细胞活性降低,且凋亡率增加。结论:hMLH1参与雌激素诱导的结肠癌细胞株HCT116凋亡。     

Potent anti-tumor effects of EGFR-targeted hybrid peptide on mice bearing liver metastases

In this study, we investigated the therapeutic efficacy of EGFR2R-lytic hybrid peptide for the treatment of liver metastasis from colon carcinoma. The cytotoxic activity of the hybrid peptide against luciferase-expressing human colon cancer (HCT-116-luc) cells was determined by the WST-8 assay. The experimental mouse model of liver metastases was generated by splenic injection of HCT-116-luc cells. The hybrid peptide was intravenously injected into mice the day after cell implantation at a dose of 5 mg/kg and this was repeated on alternate days for a total of 7 doses. Saline-treated mice were used as controls. Tumor growth and therapeutic responses were monitored by an IVIS imaging system. It was shown that the hybrid peptide exhibited potent cytotoxic activity against HCT-116-luc cells and the liver metastases were significantly reduced after intravenous injections of hybrid peptide compared with controls. Furthermore, Kaplan–Meier analysis showed that hybrid peptide-treated mice had significantly longer survival than controls. In addition, bright-field and ex vivo imaging of liver tissue revealed that mice treated with the hybrid peptide had significantly fewer tumors compared with controls. These results demonstrated that the EGFR2R-lytic hybrid peptide is a potential treatment option for patients with colorectal cancer metastases in the liver.     

DNA methyltransferase 1 is essential for initiation of the colon cancers

DNA methyltransferase 1 (Dnmt1) is essential for the maintenance of hematopoietic and somatic stem cells in mice; however, its roles in human cancer stem-like cells (CSCs)/cancer-initiating cells (CICs) are still elusive. In the present study, we investigated DNMT1 functions in the maintenance of human colon CSCs/CICs using the human colon cancer cell line HCT116 (HCT116 w/t) and its DNMT1 knockout cell line (DNMT1^−/−). The rates of CSCs/CICs were evaluated by side population (SP) analysis, ALDEFLUOR assay and expression of CD44 and CD24. SP, ALDEFLUOR-positive (ALDEFLUOR⁺) and CD44-positive and CD24-positive (CD44⁺CD24⁺) cell rates were lower in DNMT1^−/− cells than in HCT116 w/t cells. Since CSCs/CICs have higher tumor-initiating ability than that of non-CSCs/CICs, the tumor-initiating abilities were addressed by injecting immune deficient (NOD/SCID) mice. DNMT1^−/− cells showed less tumor-initiating ability than did HCT116 w/t cells, whereas the growing rate of DNMT1^−/− cells showed no significant difference from that of HCT116 cells both in vitro and in vivo. Similar results were obtained for cells in which DNMT1 had been transiently knocked-down using gene-specific siRNAs. Taken together, these results indicate that DNMT1 is essential for maintenance of colon CSCs/CICs and that short-term suppression of DNMT1 might be sufficient to disrupt CSCs/CICs.     

Preclinical study of treatment response in HCT-116 cells and xenografts with (1) H-decoupled (31) P MRS

The topoisomerase I inhibitor, irinotecan, and its active metabolite SN‐38 have been shown to induce G₂/M cell cycle arrest without significant cell death in human colon carcinoma cells (HCT‐116). Subsequent treatment of these G₂/M‐arrested cells with the cyclin‐dependent kinase inhibitor, flavopiridol, induced these cells to undergo apoptosis. The goal of this study was to develop a noninvasive metabolic biomarker for early tumor response and target inhibition of irinotecan followed by flavopiridol treatment in a longitudinal study. A total of eleven mice bearing HCT‐116 xenografts were separated into two cohorts where one cohort was administered saline and the other treated with a sequential course of irinotecan followed by flavopiridol. Each mouse xenograft was longitudinally monitored with proton (¹H)‐decoupled phosphorus (³¹P) magnetic resonance spectroscopy (MRS) before and after treatment. A statistically significant decrease in phosphocholine (p = 0.0004) and inorganic phosphate (p = 0.0103) levels were observed in HCT‐116 xenografts following treatment, which were evidenced within twenty‐four hours of treatment completion. Also, a significant growth delay was found in treated xenografts. To discern the underlying mechanism for the treatment response of the xenografts, in vitro HCT‐116 cell cultures were investigated with enzymatic assays, cell cycle analysis, and apoptotic assays. Flavopiridol had a direct effect on choline kinase as measured by a 67% reduction in the phosphorylation of choline to phosphocholine. Cells treated with SN‐38 alone underwent 83 ± 5% G₂/M cell cycle arrest compared to untreated cells. In cells, flavopiridol alone induced 5 ± 1% apoptosis while the sequential treatment (SN‐38 then flavopiridol) resulted in 39 ± 10% apoptosis. In vivo ¹H‐decoupled ³¹P MRS indirectly measures choline kinase activity. The decrease in phosphocholine may be a potential indicator of early tumor response to the sequential treatment of irinotecan followed by flavopiridol in noninvasive and/or longitudinal studies. Copyright © 2011 John Wiley & Sons, Ltd.     

Fentanyl inhibits proliferation and invasion of colorectal cancer via β-catenin

Background and aim: Fentanyl is widely used for relieving pain and narcotizing in cancer patients. However, there are few published reports regarding the effects of fentanyl on tumor control and treatment. Here we investigated the effects of fentanyl on tumor growth and cell invasion in the human colorectal carcinoma (HCT116) cells. Methods: Nude mice xenografts of HCT116 cells were established to assess the inhibition effect on tumor growth by fentanyl. MTT and Transwell were employed to determine the cell survival rate and cell invasion, respectively. MicroRNAs and mRNAs expression were quantified by real-time PCR. β-catenin and matrix metalloproteinases (MMP-2 and MMP-9) expression were assayed by western blotting. β-Catenin-specific small interfering RNA (Si-β-catenin) and miR-182 mimics were transfected in cells to investigate the mechanism underlying the effects of fentanyl on the colorectal tumor and HCT116 cells. Results: Treatment with fentanyl inhibited the tumor growth and HCT116 cells invasion. Fentanyl also downregulated the expression of β-catenin and miR-182 in both xenograft tumors and HCT116 cells, and decreased the protein level of MMP-9 in HCT116 cells. Downregulation of β-Catenin resulted in the decrease of miR-182 expression in colorectal cells. In addition, the overexpression of miR-182 reversed the effect of fentanyl on MMP-9 expression and cell invasion of HCT116 cells. Conclusions: The current study demonstrated that the inhibition of tumor growth and cell invasion in colorectal cancer by fentanyl is probably due to downregulation of miR-182 and MMP-9 expression by β-catenin.     

黄芩苷对CA46细胞裸鼠异种移植瘤的作用及机制

目的: 研究黄芩苷抗CA46细胞裸鼠异种移植瘤的作用并探讨其机制。方法: 构建CA46细胞裸鼠异种移植瘤模型,将荷瘤裸鼠随机分为5组:阴性对照组、15 mg/kg黄芩苷组、30 mg/kg黄芩苷组、60 mg/kg黄芩苷组和4 mg/kg依托泊甙 (VP-16) 阳性对照组。采用腹腔注射方式给药,12 d后处死部分裸鼠,对剥取的瘤块称重计算抑瘤率,并进行透射电镜、病理学和Western blotting检测;观察各组未处死的裸鼠直至全部死亡,研究黄芩苷对荷瘤裸鼠生存时间的影响。结果: 黄芩苷明显抑制了CA46细胞裸鼠异种移植瘤的生长,引起肿瘤细胞的凋亡、坏死以及p-Akt、NF-κB、mTOR和p-mTOR蛋白表达的下调;随着黄芩苷剂量的增加,黄芩苷治疗组荷瘤裸鼠的生存时间明显延长。结论: 黄芩苷可抑制CA46细胞裸鼠异种移植瘤的生长,诱导肿瘤细胞的凋亡,并延长荷瘤裸鼠的生存时间,其机制可能与下调PI3K/Akt/NF-κB和PI3K/Akt/mTOR信号通路有关。     

CD40配体对结肠癌细胞株的体外抑制作用

研究免疫共刺激分子CD40在结肠癌细胞株的表达及重组可溶性人CD40配体(rshCD40L)对结肠癌细胞株的体外抑制作用。采用流式细胞仪分别检测4株结肠癌细胞(HCT116、Caco-2、SW48和COLO-320)CD40分子的表达,同时利用流式细胞仪检测γ干扰素(IFN-γ)对结肠癌细胞株CD40表达的影响。MTT法检测rshCD40L对结肠癌细胞的抑制作用,TUNEL法检测rshCD40L对结肠细胞的促凋亡作用,同时检测联合rshCD40L和IFN-γ抗结肠癌作用。结果流式细胞仪检测有3株结肠癌细胞(HCT116、Caco-2和SW48)表达CD40分子,IFN-γ能增强CD40+结肠癌细胞CD40分子的表达。rshCD40L能明显抑制CD40+结肠癌细胞的增殖作用(抑制率分别为HCT116:33.5%±5.5%;Caco-2:21.5%±3.6%;SW48:30.1%±4.2%),而对CD40-结肠癌细胞株(COLO-320)无明显抑制作用(P<0.05)。rshCD40L能诱导CD40+结肠癌细胞的凋亡作用(凋亡细胞百分比为HCT116:25.6%±4.52%;Caco-2:15.71%±2.27%;SW48:18.0%±3.7%),而对CD40-结肠癌细胞株(COLO-320)无诱导凋亡作用(P<0.05)。另外,IFN-γ能明显增强rshCD40L对结肠癌细胞的抑制增殖和促凋亡作用。重组人CD40配体与IFN-γ联合可显著诱导CD40阳性的结肠癌细胞凋亡,抑制其增殖,具有潜在的抗肿瘤作用。