抗粘附肽YIGSR的衍生物对Lewis肺癌侵袭、转移能力的抑制作用

采用高转移肿瘤细胞对基底膜成分的体外粘附、侵袭模型和Lewis肺癌细胞自发肺转移模型 ,研究了YIGSR均叉、杂叉聚合肽的抗肿瘤侵袭、转移活性。实验结果 ,RGD和YIGSR的杂叉肽和YIGSR的均叉肽对PG细胞较对照的YIGSR肽表现了更明显的粘附抑制作用 ,并降低了PG细胞的体外侵袭能力 ,降低了移值Lewis肺癌细胞肺转移小鼠的肺重量或肺转移瘤结节数 ,但对小鼠原发部位移植瘤重量无明显影响 ;YIGSR聚合肽的以上作用有一定的剂量依赖性。     

威麦宁抗小鼠Lewis肺癌转移作用及分子机制的研究

目的: 探讨威麦宁抗小鼠Lewis肺癌转移作用及其分子机制。方法: 接种Lewis肺癌细胞复制移植性肺癌转移模型,用药后计数肺表面转移灶的数目;免疫组化法检测瘤组织中血管内皮细胞CD34的表达情况、并计数微血管密度(MVD)和瘤细胞表面E-cadherin蛋白表达率;RT-PCR法检测瘤细胞E-cadherin mRNA的表达水平。结果: NS(生理盐水组)、WL(威麦宁低剂量组)、WH(威麦宁高剂量组)、CTX(环磷酰胺组)各组肺转移灶发生率依次为100%、90%、60%和40%。瘤组织内MVD计数, WH组MVD明显低于NS组(P<0.01)。WH组瘤细胞膜表面E-cadherin蛋白表达率及E-cadherin mRNA水平明显高于NS组(P<0.01)。 结论: 威麦宁可能通过提高瘤细胞E-cadherin mRNA及E-cadherin蛋白的表达;减少瘤组织微血管形成等机制,发挥抗Lewis肺癌荷瘤小鼠移植性肺转移作用。     

树突状细胞诱导的CIK细胞对肺癌生长的抑制作用

目的探讨树突状细胞诱导的CIK细胞对肺癌移植瘤细胞生长的抑制作用。方法建立肺癌皮下移植瘤模型;分离患者外周血单个核细胞,培养成熟的DC及CIK细胞;用反复冻融肺癌组织的方法制备肿瘤细胞裂解物并负载DC,诱导DC-CIK细胞;进行肺癌移植瘤内注射,比较两组裸鼠移植瘤体积及重量,观察其抑瘤情况。结果 DC-CIK细胞对肺癌皮下移植瘤细胞的抑制率为70.3%,抑制作用明显强于DC或CIK细胞的抑制作用(P0.05)。结论树突状细胞诱导的CIK细胞能够有效抑制肺癌移植瘤生长。     

益气活血解毒方对小鼠Lewis肺癌生长转移及血管生成的作用

目的：研究益气活血解毒方对小鼠Lewis肺癌生长及自发性肺转移的作用并探讨其作用机制。方法:复制小鼠Lewis肺癌模型并随机分组,腹腔注射给药 10 d 后，观察抑瘤率，HE染色观察瘤组织病理学变化，免疫组化法检测瘤组织增殖细胞核抗原（PCNA）的表达；给药 20 d 后，观察肺转移瘤结节数、瘤组织微血管密度（MVD），免疫组化法和ELISA法分别检测瘤组织及血清血管内皮生长因子（VEGF）的表达。结果:益气活血解毒方能减少小鼠Lewis肺癌肿瘤重量和肺转移灶数，高低剂量(24 mg·kg^-1·d^-1,12 mg·kg^-1·d^-1)抑瘤率分别为48.29%、37.26%，肺转移结节数分别为1.67、3.50，对照组为6.44；并能抑制肿瘤细胞PCNA的表达，降低肿瘤组织微血管密度,抑制肿瘤细胞及血清中VEGF的表达。结论:益气活血解毒方能抑制小鼠Lewis肺癌的生长与转移,其机制可能与抑制VEGF表达,减少肿瘤血管生成有关。     

葡萄糖转运蛋白1抑制剂BAY-876对肺癌细胞增殖、迁移、侵袭和肿瘤生长的影响

目的 探讨BAY-876对肺癌细胞增殖、迁移、侵袭和肿瘤生长的影响及可能机制。方法 利用CCK-8实验检测BAY-876对肺癌细胞增殖的影响,利用Transwell实验分析BAY-876对肺癌细胞迁移和侵袭的影响,利用荷瘤小鼠模型检测BAY-876对肿瘤生长的影响,免疫组织化学法检测各组移植瘤组织中ITGβ1蛋白的表达水平。结果 BAY-876抑制A549细胞的增殖、迁移和侵袭,同时抑制裸鼠移植瘤的生长,明显缩小肿瘤体积;BAY-876处理后移植瘤组织中ITGβ1蛋白表达降低;过表达ITGβ1逆转了BAY-876对A549细胞迁移和侵袭的抑制作用。结论 BAY-876可能通过抑制ITGβ1蛋白的表达抑制肺癌细胞增殖、迁移、侵袭和肿瘤生长。     

c-Met基因调控肺腺癌A549细胞放射敏感性

目的探讨上调和下调肺腺癌细胞株A549细胞中c-Met基因表达水平后,该细胞鼠移植瘤放射敏感性的变化情况。方法利用电穿孔法将c-Met siRNA和重组表达载体pc DNA3.1-c-Met分别转染入肺癌A549细胞后采用G418筛选得到稳定转染的肺癌细胞系。转染48 h后用RT-PCR法和Western blot法分别检测细胞中c-Met基因和蛋白表达水平,采用CCK-8法检测细胞增殖活性;克隆形成实验研究分别上调和下调c-Met基因表达水平对肺腺癌细胞株A549放射敏感性的影响;TUNEL法检测细胞凋亡影响,移植瘤实验检测基因沉默和激进c-Met基因表达水平并联合放射射线照射对肺腺癌细胞生长的抑制作用。结果 c-Met下调组中肺癌A549细胞内c-Met基因和蛋白表达水平明显降低,而上调组中的表达水平则得到了明显升高。c-Met下调组肺癌A549细胞放射敏感性升高,c-Met上调组肺癌A549细胞放射敏感性降低(P0.05)。下调组中肺癌A549细胞的增殖活性受到抑制而细胞凋亡率显著增加(P0.05),上调组肺癌细胞的增殖能力明显增强而细胞凋亡率明显降低(P0.05);下调组裸鼠移植瘤的平均体积明显小于对照组,而上调组裸鼠移植瘤的瘤体平均体积则显著大于对照组(P0.05)。结论基因沉默c-Met的基因表达水平能显著降低肺癌细胞的增殖活性,抑制人肺癌A549细胞裸鼠移植瘤的生长并明显提高移植瘤瘤体的放射敏感性。     

藏药红景天甙诱导DC成熟并增强T细胞抑制肺癌的实验研究

观察红景天甙对Lewis肺癌移植瘤生长的作用,初步探讨其增强肿瘤免疫的机制。建立小鼠Lewis肺癌移植瘤模型,随机分为红景天甙(sachalin rhodiola rhizome,SRR)组,紫杉醇(taxelol,TAX)组和PBS组,计算抑瘤率并观察小鼠生存期;同时取SRR治疗荷瘤小鼠肿瘤引流DC,观察SRR体外处理后对DC表面分子、吞噬功能及T细胞杀伤功能的影响。在治疗第23天,SRR组肿瘤体积明显小于PBS组(P<0.05),与TAX组无显著性差异(P>0.05);SRR组小鼠生存期较PBS组显著延长(P<0.05),与TAX组无显著差异(P>0.05);SRR处理的树突细胞(dendritic cell,DC)表面分子MHCII、CD80表达显著上升,吞噬功能显著下降,由DC诱导的活化T细胞对肿瘤细胞的杀伤效应显著高于对照组(P<0.05)。SRR有效促进DC成熟、增强T细胞功能,同时可以抑制肿瘤生长。     

生存素-D53A突变体对人肺腺癌SPC-A1移植瘤的治疗作用

为了研究生存素基因负性突变体生存素-D53A(SVV-D53A)对人肺腺癌SPC-A1移植瘤的治疗作用,并探讨其作用机制,我们在体外实验中使用SVV-D53A质粒转染人肺腺癌SPC-A1细胞,蛋白免疫印迹检测生存素突变体表达,同时通过流式细胞检测细胞凋亡情况。在体内实验中,裸鼠皮下接种人肺腺癌SPC-A1细胞建立肺癌移植瘤模型,使用阳离子脂质体包裹SVV-D53A质粒进行治疗。治疗结束后免疫组化检测各组肿瘤标本细胞增殖,末端脱氧核苷酸转移酶脱氧尿苷三磷酸切口末端标记(TUNEL)检测肿瘤细胞凋亡。与对照组相比,SVV-D53A质粒在体外显著诱导SPC-A1细胞凋亡,SVV-D53A组裸鼠皮下移植瘤体积减小,细胞增殖减弱,大量肿瘤细胞发生凋亡。研究结果表明SVV-D53A的表达在体外和体内均能够显著抑制人肺腺癌SPC-A1细胞的生长,其作用机制主要为诱导肿瘤细胞凋亡。     

芦荟苷对非小细胞肺癌的增殖和抗转移作用

目的：探讨芦荟苷对非小细胞肺癌的增殖和抗转移作用的调控及其机制。方法：qPCR检测不同浓度的芦荟苷对非小细胞肺癌的细胞活性影响情况;分析芦荟苷对非小细胞肺癌细胞增殖行为的影响情况;流式细胞术实验检测芦荟苷对非小细胞肺癌细胞凋亡行为和细胞周期的影响;Transwell侵袭实验和划痕愈合实验检测芦荟苷对非小细胞肺癌细胞侵袭和迁移能力的影响,裸鼠体外成瘤实验检测芦荟苷对非小细胞肺癌细胞成瘤能力的影响;Western blot检测增殖相关蛋白的表达情况。结果：100 μmol/L的芦荟苷对非小细胞肺癌的细胞活性的抑制作用最佳;而随着时间的进展,芦荟苷对非小细胞肺癌细胞的增殖能力有直接抑制作用,可以在一定程度上干扰非小细胞肺癌细胞的生长进程;而合适浓度芦荟苷对非小细胞肺癌细胞侵袭和迁移能力有直接的抑制作用,同时可以抑制体外成瘤能力,干扰增殖蛋白的表达。结论：芦荟苷可以抑制非小细胞肺癌细胞的增殖和侵袭行为,对其转移有一定的抑制作用。     

黄芪、党参提取物增强紫杉醇抑制肿瘤血管生成和转移的实验研究

目的:观察黄芪、党参提取物对紫杉醇抑制Lewis肺癌血管生成和肿瘤转移的增强作用,为中医药抗肿瘤应用提供实验依据.方法:C57BL/6小鼠右肋处皮下接种Lewis肺癌,6 h后随机分为实验组、紫杉醇组和对照组.实验组接受紫杉醇联合参芪注射液治疗,紫杉醇组接受同等剂量的紫杉醇治疗,对照组用等体积的生理盐水代替.观察指标:移植瘤体积,肿瘤组织内微血管度和肺脏转移瘤数量,小鼠存活时间.结果:与紫杉醇治疗相比,紫杉醇联合参芪可以明显减少移植瘤内的微血管密度(10.1±4.4 vs 16.8±7.3,P＜0.05)和肺脏转移瘤个数(13.4±4.3 vs 18.4±3.9,P＜0.05),明显延长小鼠的存活时间(43.3±8.4 d vs 36.3±6.6 d,P＜0.05). 结论:黄芪、党参提取物对紫杉醇抑制肿瘤血管生成和移植肿瘤转移有一定的增强作用.     

Syngeneic lymph-node-targeting model of green fluorescent protein-expressing Lewis lung carcinoma

The Lewis lung tumor has been extensively studied in both syngeneic and allogeneic mouse models. However, its metastatic potential and mechanism are poorly understood. The aim of the present study was to develop a highly metastatic lymph-node targeting, imageable model of the Lewis lung carcinoma in a syngeneic host. We report here a syngeneic model of the Lewis lung carcinoma in which the carcinoma cells are labeled with green fluorescent protein (GFP). The tumor cells were transplanted in the dorsal side of the ear of C57-B16 mice in order to give the tumor cells access to the lymphatic system. This model of the Lewis lung carcinoma extensively metastasized to numerous lymph nodes throughout the body of the animal as well as visceral organs, as visualized by fluorescence microscopy using the bright GFP signal. Twenty-one different metastatic sites, including lymph nodes throughout the body, were identified among the cohort of transplanted animals. The data demonstrate a predilection of the Lewis lung carcinoma for lymphatic pathways for metastasis throughout the animal body. The concomitant macrometastases to the visceral organs observed in this model may be remetastasis from the lymph nodes. This model of the Lewis lung carcinoma should be very useful in defining cellular trafficking and targeting mechanisms of metastasis, in particular those involving lymphatic pathways.     

慢病毒介导Gag-caspase-8对小鼠乳腺癌细胞移植瘤的抑制作用*

目的：探讨慢病毒介导Gag-caspase-8 对小鼠乳腺癌4T1 细胞BALB/C移植瘤的抑制效果。方法：PEI 转染 法将Gag-caspase-8 转入293T 细胞包装,并收集病毒颗粒。BALB/C小鼠建立4T1 乳腺癌移植瘤模型,按照数字 表法随机分为对照组（ 注射等量PBS）、Gag-caspase-8 治疗组（ 注射1×106/mL Gag-caspase-8）和Gag 组（ 注射 1×106/mL Gag）。在种植细胞第10 天开始经瘤内注射Gag-caspase-8 进行抑瘤实验,每只0.1 mL,每3 d 给药1 次, 共计3 次,观察小鼠移植瘤的生长情况;抑瘤实验第12 天处死小鼠,采用免疫荧光法检测移植瘤组织caspase-8 蛋白表达。结果：抑瘤实验12 d 时,对照组、Gag 组移植瘤体积明显高于Gag-caspase-8 组,Gag-caspase-8 组 抑瘤率明显高于对照组,差异有统计学意义。Gag-caspase-8 组Caspase-8 蛋白表达高于对照组和Gag 组,Gagcaspase- 8 组肺部转移结节数明显少于对照组,差异有统计学意义。结论：Gag-caspase-8 慢病毒颗粒对小鼠乳腺 癌荷瘤生长及肺转移有抑制作用。     

Syngeneic lymph-node-targeting model of green fluorescent protein-expressing Lewis lung carcinoma

The Lewis lung tumor has been extensively studied in both syngeneic and allogeneic mouse models. However, its metastatic potential and mechanism are poorly understood. The aim of the present study was to develop a highly metastatic lymph-node targeting, imageable model of the Lewis lung carcinoma in a syngeneic host. We report here a syngeneic model of the Lewis lung carcinoma in which the carcinoma cells are labeled with green fluorescent protein (GFP). The tumor cells were transplanted in the dorsal side of the ear of C57-B16 mice in order to give the tumor cells access to the lymphatic system. This model of the Lewis lung carcinoma extensively metastasized to numerous lymph nodes throughout the body of the animal as well as visceral organs, as visualized by fluorescence microscopy using the bright GFP signal. Twenty-one different metastatic sites, including lymph nodes throughout the body, were identified among the cohort of transplanted animals. The data demonstrate a predilection of the Lewis lung carcinoma for lymphatic pathways for metastasis throughout the animal body. The concomitant macrometastases to the visceral organs observed in this model may be remetastasis from the lymph nodes. This model of the Lewis lung carcinoma should be very useful in defining cellular trafficking and targeting mechanisms of metastasis, in particular those involving lymphatic pathways.This revised version was published online in August 2005 with a corrected cover date.     

A highly metastatic Lewis lung carcinoma orthotopic green fluorescent protein model

The Lewis lung carcinoma has been widely used for many important studies. However, the subcutaneous transplant or orthotopic cell-suspension injection models have not allowed the expression of its full metastatic potential. A powerful new highly metastatic model of the widely-used Lewis lung carcinoma is reported here using surgical orthotopic implantation (SOI) of tumor fragments and enhanced green fluorescent protein (GFP) transduction of the tumor cells. To achieve this goal, we first developed in vitro a stable high-expression GFP transductant of the Lewis lung carcinoma with the pLEIN retroviral expression vector containing the enhanced Aequorea victoria GFP gene. Stable high-level expression of GFP was found maintained in vivo in subcutaneously-growing Lewis lung tumors. The in vivo GFP-expressing tumors were harvested and implanted as tissue fragments by SOI in the right lung of additional nude mice. This model resulted in rapid orthotopic growth and extensive metastasis visualized by GFP-expression. 100% of the animals had metastases on the ipsilateral diaphragmatic surface, contralateral diaphragmatic surface, contralateral lung parenchima, and in mediastinal lymph nodes. Heart metastases were visualized in 40%, and brain metastases were visualized in 30% of the SOI animals. Mice developed signs of respiratory distress between 10–15 days post-tumor implantation and were sacrificed. The use of GFP-transduced Lewis lung carcinoma transplanted by SOI reveals for the first time the high malignancy of this tumor and provides an important useful model for metastasis, angiogenesis and therapeutic studies.     

复方紫杉醇注射液对 C57BL/6小鼠Lewis肺癌的作用研究

目的:研究复方紫杉醇注射液对C57BL/6小鼠Lewis肺癌的作用及其机制。方法:60只C57BL/6小鼠按体重随机分为5组,每组12只,移植皮下接种Lewis细胞。在接种后24小时,对照组给予生理盐水;实验组分别给予紫杉醇注射液大、小两种剂量,复方紫杉醇注射液大、小两种剂量,各组均腹腔注射,连续5天。停药后第6天脱颈处死动物,计算抑瘤率;用HE染色光镜观察细胞形态学的变化;用流式细胞仪(FCM)检测研究复方紫杉醇注射液诱导肿瘤细胞凋亡以及对肿瘤细胞周期的影响。结果:复方紫杉醇注射液大、小剂量组对荷瘤鼠Lewis肺癌有明显的抑制作用,且各剂量组均可诱导Lewis细胞凋亡,与对照组比较有显著性差异(P<0.05);而对细胞周期的影响不太明显。结论:复方紫杉醇注射液对移植于C57BL/6小鼠的Lewis肺癌具有明显的抑制作用,其机制可能与诱导肿瘤细胞凋亡有关。     

硫修饰脂质体包裹VEGF反义寡核苷酸对肺癌血管生成及转移的抑制作用

目的：探讨硫修饰脂质体包裹的VEGF反义寡核苷酸对肺癌血管生成和转移的抑制作用。方法：将硫修饰脂质体包裹的VEGF反义寡核苷酸（ASODN）、正义寡核苷酸（SODN）、错义寡核苷酸（MODN）加入培养的Lewis肺癌细胞中，采用免疫组织化学方法检测肺癌细胞中VEGF蛋白表达，观察经ODN处理的条件培养基对牛主动脉内皮细胞增殖的影响。另外，复制小鼠Lems肺癌模型40只，随机分为对照组、VEGFASODN组、VEGFSODN组及VEGFMSODN组，每组10只，接种肿瘤细胞24小时内，给予脂质体、VEGFASODN、VEGFSODN、VEGFMSODN皮下注射，每周2次，连续4周。检测皮下肿瘤的变化和肺转移率，并用免疫组织化学方法检测肿瘤组织微血管密度（MVD），超声检测肿瘤组织Ps、砌变化。结果：ASODN能够下调肺癌细胞中VEGF蛋白的表达，经ASODN处理的条件培养基能显著抑制牛主动脉内皮细胞的增殖。ASODN组小鼠肿瘤生长及肺转移受到显著抑制，MVD、PS、RI与其它各组相比有明显差异（P〈0．01）。结论：硫修饰脂质体包裹的VEGF反义寡核苷酸能够显著抑制肺癌血管生成，进而抑制肿瘤生长及转移。     

Enhancing T lymphocytes from tumor-bearing mice suppress host resistance to a syngeneic tumor

The cell-mediated immune response of animals to a lethal syngeneic tumor was investigated by inoculating C57BL mice with Lewis lung carcinoma (3LL) cells T lymphocytes, obtained from the enlarged spleens of the tumor-bearing mice were found to be cytotoxic to 3LL target cells in vitro. However, we found that such spleen cells enhanced tumor growth in vivo when mice were injected with a mixture of spleen cells and tumor cells. Removal of T lymphocytes by treatment of the spleen cells with anti-Θ serum plus complement reduced the enhancement of tumor growth. Hence, the tumor enhancing cells, like the cytotoxic cells, appeared to be T lymphocytes. Removal of T lymphocytes from normal mice by adult thymectomy before tumor inoculation led to a reduction in the number of tumor metastases. Thus, enhancing T lymphocytes appear to exist in normal as well as in tumor-bearing mice. Investigation of this mechanism of tumor enhancement suggested that the enhancing T lymphocytes act as suppressor T cells inhibiting natural immune resistance to tumor growth.     

Experimental metastasis is suppressed in MMP-9-deficient mice

Matrix metalloproteinases (MMPs) are thought to play a key role in tumor invasion and metastasis. The role of MMP-9 (gelatinase B) in tumor metastasis was examined in MMP-9-deficient mice produced by gene targeting using embryonic stem cells. MMP-9-deficient mice develop normally and are fertile. In these mice, the number of metastatic colonies of B16-BL6 melanoma cells or Lewis lung carcinoma cells that were implanted intravenously fell by 45% for B16-BL6 melanoma and 59% for Lewis lung carcinoma (p=0.03 and p=0.0043, respectively). Gelatin zymography showed that both tumor cell lines did not secrete MMP-9 by themselves but the host cells surrounding the tumor cells secrete MMP-9 in vivo. These results indicated that host-derived MMP-9 plays an important role in the process of tumor metastasis.     

Inhibitory effects of bovine lactoferrin on colon carcinoma 26 lung metastasis in mice

In order to determine the effects of the multifunctional iron-binding glycoprotein, lactoferrin (LF), and related compounds on tumor growth and metastasis, bovine LF (bLF), and bLF hydrolysate and lactoferricin (bLFcin), active products generated by acid-pepsin hydrolysis were administered orally to BALB/c mice bearing subcutaneous (s.c.) implants of the highly metastatic colon carcinoma 26 (Co 26Lu). bLF and the bLF hydrolysate demonstrated significant inhibition of lung metastatic colony formation from s.c. implanted tumors without appreciable effects on tumor growth. bLFcin displayed a tendency for inhibition of lung metastasis. On the other hand, bLF did not exert marked anti-metastatic activity in athymic nude mice bearing Co 26Lu, though bLF had a tendency to inhibit the lung metastatic colony formation associated with anti-asialoGM1 antibody (Ab) treatment. AsialoGM1+ and CD8+ cells in white blood cells were increased after treatment with bLF. In vitro, the viability of Co 26Lu-F55 cells was markedly decreased when co-cultured with white blood cells from mice administrated bLF p.o., but recovered on treatment with anti-asialoGM1 Ab or anti-CD8 mAb and complement. The results suggest bLF and related compounds might find application as tools in the control of metastasis and that asialoGM1+ and CD8+ cells in the blood are important for their inhibitory effects.