抗Fas单克隆抗体诱导人胃癌细胞系SGC-7901细胞

目的探讨抗Ｆａｓ单克隆抗体诱导胃癌细胞凋亡的规律及在胃癌治疗中的意义．方法应用细胞形态观察、琼脂糖凝胶电泳、流式细胞光度术检测抗Ｆａｓ单克隆抗体对胃癌细胞ＳＧＣ７９０１增殖周期的影响以及对细胞杀伤作用的方式,并检测了ＳＧＣ７９０１细胞表面Ｂｃｌ２的表达情况．．结果抗Ｆａｓ单克隆抗体有阻滞细胞周期、通过诱发凋亡而抑制肿瘤细胞生长的作用．经抗Ｆａｓ单克隆抗体处理后,ＳＧＣ７９０１细胞表面Ｂｃｌ２蛋白表达无明显变化．．结论抗Ｆａｓ单克隆抗体可以诱导胃癌细胞系ＳＧＣ７９０１细胞凋亡．抗Ｆａｓ单克隆抗体诱导胃癌细胞凋亡与Ｂｃｌ２表达无关     

肝细胞提取物组分S4对肿瘤细胞体外增殖的影响

目的观察肝细胞提取物组分Ｓ４对８株肿瘤细胞增殖的影响．方法采用ＭＴＴ比色法研究不同浓度Ｓ４对８株肿瘤细胞不同时相增殖的影响．结果Ｓ４对７４０２，７７２１，７７０３，Ｈｅｐｅ肝癌细胞作用７２ｈ后ＩＣ５０（ｍｇ／Ｌ）分别为３９，４２，９５和６５．对胃腺癌细胞（ＳＧＣ７９０１）、回盲部腺癌细胞（ＨＣＴ８）、肺腺癌细胞（ＧＬＣ８２）作用７２ｈ的ＩＣ５０分别为１４３，９９和５４．对鼻咽癌细胞（ＣＮＥ２）４８ｈ未显示抑制作用．结论Ｓ４抑制７株肿瘤细胞呈现明显的量效与时效关系，在１ｍｇ／Ｌ～１０ｍｇ／Ｌ浓度范围内及２４ｈ～７２ｈ时限内，抑制作用随剂量增加、时间延长而增强     

重组反义c-myc 腺病毒对人胃癌细胞的体内及体外分子治疗

目的研究重组反义ｃｍｙｃ腺病毒对胃癌细胞的体内外生物学作用．方法采用ＬａｃＺ基因Ｘｇａｌ染色、ＭＴＴ,ＤＮＡ梯度降解试验、原位末端标记、流式细胞仪、ＰＣＲ分析、裸鼠致瘤性、裸鼠皮下移植瘤模型实验等方法,对反义ｃｍｙｃ重组腺病毒在人胃癌ＳＧＣ７９０１细胞系中的作用进行体内外研究．结果ＡｄＡＳｃｍｙｃ对ＳＧＣ７９０１细胞能产生明显的生长抑制作用,ＭＴＴ显示生长抑制率为４４１％．ＤＮＡ梯度降解试验、原位末端标记、流式细胞仪显示ＡｄＡＳｃｍｙｃ诱导了ＳＧＣ７９０１细胞凋亡．经ＡｄＡＳｃｍｙｃ处理的ＳＧＣ７９０１细胞裸鼠致瘤性消失．ＡｄＡＳｃｍｙｃ对裸鼠皮下移植瘤模型瘤内注射能有效降低肿瘤的生长速度,生长抑制率为６８９％．结论ＡｄＡＳｃｍｙｃ对胃癌细胞具有显著的体内外生长抑制及凋亡诱导作用     

树突状细胞体外诱导抗肝癌免疫

目的应用树突状细胞（ＤＣ）在体外诱导高效而特异抗肝癌免疫．方法自肝癌患者外周血中分离ＤＣ;以粒／巨噬细胞集落刺激因子（ＧＭＣＳＦ）及白介素４（ＩＬ４）联合刺激ＤＣ;以人肝癌细胞系ＨｅｐＧ２肿瘤细胞的肿瘤相关抗原（ＴＡＡ）激活ＤＣ;ＤＣ诱导自体Ｔ淋巴细胞增殖、分化为细胞毒性Ｔ细胞（ＣＴＬ）;检测ＣＴＬ及其上清液对ＨｅｐＧ２肿瘤细胞、ＬＯＶＯ肿瘤细胞及ＨＯＳ８６０３肿瘤细胞的细胞毒作用．结果经人肝癌细胞系ＨｅｐＧ２肿瘤细胞的ＴＡＡ激活并经ＧＭＣＳＦ及ＩＬ４联合刺激后,肝癌患者外周血ＤＣ能够诱导自体Ｔ淋巴细胞增殖分化为ＣＴＬ,该ＣＴＬ及其上清液对ＨｅｐＧ２肿瘤细胞均有高效而特异性的杀伤作用（杀伤率分别为９２％±１０５％和４１％±８９％）．结论肝癌患者外周血ＤＣ体外能够诱导高效而特异抗肝癌免疫．提示ＤＣ作为一新概念上的抗肿瘤疫苗可能在肿瘤治疗及预防中发挥重要作用．     

奥曲肽和表皮生长因子受体单克隆抗体诱导人胃癌细胞凋亡

目的：定量检测奥曲肽（ＳＭＳ）和表皮生长因子受体单克隆抗体（ＥＧＦＲＭＣＡｂ）对人胃癌细胞系 ＳＧＣ－７９０１的诱 导凋亡作用。方法：将不同浓度的ＳＭＳ和ＥＧＦＲＭｃＡｂ与胃癌细胞共同孵育，用流式细胞术分析其细胞凋亡情况。 结果：ＳＭＳ ５ × １０－５和ＥＧＦＲＭｃＡｂ１：１００与胃癌细胞共同孵育７２ ｈ，胃癌细胞凋亡率分别为７．９６％和７．４９％，与对 照组（１．１９％湘比，有显著差异ｐ＜０．０５）。结论：诱导细胞凋亡是胃肠肽类激素发挥其抗肿瘤作用的重要途径之一。     

胃癌浸润性淋巴细胞的细胞表型及杀伤活性

目的和方法：本实验用９例胃癌患者ＴＩＬ和ＩＬ２在体外共同孵育后，用流式细胞仪分析胃癌ＴＩＬ的细胞表型特征。结果：ＴＩＬ细胞表型特征是以ＣＤ３为主，ＣＤ４／ＣＤ８为０８３，ＮＫ细胞１６３±３６％。经白细胞介素２（ＩＬ２）激活２０天后，ＣＤ３减少，ＣＤ４／ＣＤ８为１８５，ＮＫ细胞数量增至４１３±１３７％，Ｐ＜００１。ＬＤＨ释放法测定激活后的ＴＩＬ对自体胃癌细胞杀伤率比培养初期高３７４倍，变比对同时培养的７９０１胃癌细胞杀伤率高１７５倍。对自体和异体胃癌细胞杀伤作用均显著高于ＬＡＫ细胞，且具有靶细胞特异性。结论：结果表明胃癌ＴＩＬ对胃癌细胞杀伤作用，可能与ＮＫ细胞数量增多，导致胃癌细胞凋亡有关     

顺铂鱼肝油酸钠伍治疗胃癌的实验研究

用ＭＴＴ法观察顺铂（ＣＤＤＰ０与鱼肝油酸钠（ＳＭ）配伍对人胃癌细胞ＳＧＣ－７９０１的体外杀伤效果，人胃癌细胞ＳＧＣ－７９０１被接种于裸鼠皮下以建立荷瘤裸鼠。ＣＤＤＰ每次３ｍｇ．ｋｇ＾－１．ｔｉｍｅ＾－１，ＳＭ：５０ｍｇ．ｋｇ＾－１．ｔｉｍｅ＾－１，容量：５０μＬ肿瘤组织中心注射。配伍组织肿瘤ＳＭ和ＣＤＤＰ交叉给药，两药间隔３０分钟，每３天１次，共３次，ＳＭ在体外对胃癌细胞有很强的杀伤作用，局部注射     

扶正抗癌冲剂对体外胃癌细胞的抑制作用研究

目的研究中药扶正抗癌冲剂对体外胃癌细胞有无直接抑制或杀伤作用．方法采用体外培养的ＭＫＮ４５，ＭＫＮ２８及ＳＧＣ７９０１三种胃癌细胞株，以扶正抗癌冲剂配制药液（１６１５ｍｇ／ｍｌ含量）的不同稀释度，对３种胃癌细胞体外作用观察，并设丝裂霉素阳性对照组及单纯培养液加生理盐水阴性对照组．结果扶正抗癌冲剂对３种胃癌细胞株均有一定抑制作用，对低分化ＭＫＮ４５胃癌细胞抑制作用最明显，在１∶２稀释度４８ｈ抑制作用最强，抑制率达７４７％，持续到１２０ｈ抑制率仍在７１０％，与正常对照组比较有显著差异（Ｐ＜００５）．在电镜下观察被抑制的ＭＫＮ４５胃癌细胞发生线粒体肿胀、内质网水肿、胞浆脂肪空泡样变性、核仁缩小等变化．结论扶正抗癌冲剂对ＭＫＮ４５低分化胃癌细胞有较强的抑制和杀伤作用，并能使胃癌细胞结构改变．     

腺病毒介导多基因对肝癌细胞凋亡的诱导作用

目的 研究腺病毒介导的多基因（ｐ５３、Ｂ７－１、ＧＭ－ＣＳＦ、ＩＬ－２）对肝癌细胞系调亡的诱导,及对肝癌细胞系裸鼠体内成瘤性改变的影响。方法 应用光镜、电镜和ＴＵＮＥＬ法,检测肝癌细胞系感染腺病毒介导的多基因后凋亡的发生,检测ＨｅｐＧ２细胞导入腺病毒多基因后裸鼠体内的成瘤性改变。结果 肝癌细胞系导入多基因后发生凋亡,并对化疗药物顺铂的敏感性增加,册时给予１０ｍｇ／Ｌ的顺铂,可使近３０％的肝癌细胞发     

免疫抑制酸性蛋白单克隆抗体MI2体外抗肿瘤作用的观察

作者采用ＭＴＴ比色法观察了本室自制免疫抑制酸性蛋白单克隆抗体ＭＩ＿２的体外抗肿瘤作用。结果表明：ＭＩ＿２在浓度为７．８１ｍｇ／Ｌ时即可明显抑制胃癌细胞株ＳＧＣ７９０１的生长（抑制率７．５％±２．４％Ｐ＜０．０１），并见剂量依赖性表现；在ＬＡＫ细胞与ＳＧＣ７９０１胃癌细胞的效靶比为１０：１时，加入１．９５ｍｇ／Ｌ的ＭＩ＿２即可明显增强ＬＡＫ细胞的细胞毒作用（增强效应为２０６．３％Ｐ＜０．０１），其作用也呈剂量依赖性表现。作者还发现ＭＩ＿２的这种抗肿瘤作用与补体无关。     

硫代修饰端粒酶反义核酸联合顺铂、阿霉素对胃癌细胞作用的研究

目的：探讨应用硫代修饰人端粒酶RNA（hTR)反义核酸后胃癌细胞对顺铂（DDP）和阿霉素（ADM）敏感性的变化。方法：采用脂质体将针对hTR模版区设计的13相碱基硫代磷酸修饰的反义寡核苷酸CAGTTAGGGTTAG导入胃癌细胞SGC7901，应用四甲基偶氮唑蓝（MTT）法，流式细胞仪和TRAP－PCR－ELISA法测定联合应用化疗药后对细胞增殖，凋亡和端粒酶活性的影响。结果：化疗药物ADM和DDP地端粒酶活性抑制作用不同，而且有明显依赖趋势。hTR反义PS－ODN可增加ADR和DDP抑制胃癌细胞系SGC7901端粒酶活性，诱导细胞凋亡和抑制细胞增殖的作用。结论：hTR反义PS－ODN在体外能增加胃癌细胞系SGC701对化疗药ADR、DDP敏感性，其机制可能与其抑制细胞端粒酶活性，诱导细胞凋亡有关。     

Multidrug resistance reversal in human gastric carcinoma cells by neferine

AIIVI: To investigate the reversal effect of neferine on multidrug resistance in human gastric carcinoma cell line. METHODS: Cells of a human gastric cancer cells line, SGC7901, and its vincristine (VCR) -resistant variant, SGC7901/VCR, were cultivated with or without neferine and/or VCR. The cytotoxic effect of VCR was evaluated by the MTT assay. Cell apoptosis induced by VCR was determined by flow cytometry(FCM). The expression of P-glycoprotein (P-gp) and a multidrug-resistance-associated protein (MRP) in cells was examined by immunofluorescence and FCM. RESULTS: Neferine at the concentration from 2.5 μmol/L to 10 μmol/L had no cytotoxicity to SGC7901 cells, and its variant SGC7901/VCR cells. The ICso of VCR against SGC7901 and SGC7901/VCR cells was 0.059 μg/mL and 2.32 μg/mL, respectively, indicating that SGC7901/VCR cells were 39 times more resistant to VCR than its parent SGC7 901 cells. After treatment with neferine at concentrations of 2.5, 5 and 10 μmol/L, the IC50 of VCR to SGC7901/VCR cell line decreased to 0.340, 0.128 and 0.053 μg/mL, respectively,thus, increased the chemosensitivity by 6.8-, 18.1- and 43.8-fold, respectively. SGC7901/VCR cells were apoptosis resistant to VCR. Neferine (2.5, 5 and 10 μmol/L) promoted the VCR-induced apoptosis of SGC7901/VCR cells in a dosedependent manner. The expressions of P-gp and MRP were strongly positive in SGC7901/VCR cells, which were significantly down-regulated after treatment with neferine (10 μmol/L)for 24 h. CONCLUSION: Neferine reverses multidrug resistance of human gastric carcinoma SGC7901/VCR cells, which may be associated with the down-regulations of P-gp and MRP expression in SGC701/VCR cells.     

miR-449a Regulates Proliferation and Chemosensitivity to Cisplatin by Targeting Cyclin D1 and BCL2 in SGC7901 Cells

Background

Recently, several miRNAs have been determined as tumor suppressors in various cancers, such as microRNA-449a. However, the exact molecular mechanisms underlying miR-449a regulated cell proliferation and chemosensitivity in gastric cancer cells have not been well documented.

Aim

The present study was designed to test whether miR-449a mediates cell proliferation and chemosensitivity in gastric cancer cells via regulating cyclin D1 and BCL2.

Methods

In vitro, the ability of cell proliferation and cell viability were measured by MTT assay; cell cycle and cell apoptosis was detected by FCM. qRT-PCR was used to measure the expression of miR-449a. Western blot and real-time PCR assays were used to detect the expression of cyclin D1 and BCL2 in gastric cancer cell line SGC7901.

Results

miR-449a expression was downregulated in gastric cancer cell line SGC7901 and human gastric cancer tissues, compared to the gastric epithelial cell line GES-1 and matched non-tumor associated tissues. Upregulation of miR-449a reduced the proliferation of SGC7901 cells. Ectopic expression of miR-449a decreased the percentage of S phase cells, increased the percentage of G1/G0 phase cells and increased the apoptosis induced by cisplatin. Moreover, miR-449a inhibited SGC7901 cells proliferation and enhanced cisplatin chemosensitivity by downregulating expression of BCL2 and cyclin D1, respectively, via directly targeting the 3′-untranslated regions of BCL2 and cyclin D1 mRNA.

Conclusions

This is the first report to provide evidence that miR-449a could modulate cell cycle and apoptosis through regulating cyclin D1 and BCL2 expression in SGC7901 cells.     

Alisol B acetate induces apoptosis of SGC7901 cells via mitochondrial and phosphatidylinositol 3-kinases/Akt signaling pathways

AIM： To examine the effect of alisol B acetate on the growth of human gastric cancer cell line SGC7901 and its possible mechanism of action.
METHODS： The cytotoxic effect of alisol B acetate on SGC7901 cells was measured by 3-（4,5-dimethylthiazol- 2-yl）-2,5-diphenyltetrazolium bromide （MI-I-） assay. Phase-contrast and electron microscopy were used to observe the morphological changes. Cell cycle and mitochondrial transmembrane potential （A~Pm） were determined by flow cytometry. Western blotting was used to detect the expression of apoptosis-regulated gene Bcl-2, Bax, Apaf-1, caspase-3, caspase-9, Akt, P-Akt and phosphatidylinositol 3-kinases （PI3K）.
RESULTS： Alisol B acetate inhibited the proliferation of SGC7901 cell line in a time- and dose-dependent manner. PI staining showed that alisol B acetate can change the cell cycle distribution of SGC7901, increase the proportion of cells in G0-G1 phase and decrease the proportion of S phase cells and G2-M phase cells. Alisol B acetate at a concentration of 30 pmol/L induced apoptosis after 24, 48 and 72 h incubation, with occurrence rates of apoptotic cells of 4.36%, 14.42% and 21.16%, respectively. Phase-contrast and electron microscopy revealed that the nuclear fragmentation and chromosomal condensed, cells shrank and attachment loss appeared in the SGC7901 treated with alisol B acetate. Apoptosis of SGC7901 cells was associated with cell cycle arrest, caspase-3 and caspase-9 activation, loss of mitochondrial membrane potential and up-regulation of the ratio of Bax/Bcl-2 and inhibition of the PI3K/Akt.
CONCLUSION： Alisol B acetate exhibits an antiproliferative effect in SGC7901 cells by inducing apoptosis. Apoptosis of SGC7901 cells involves mitochondria-caspase and PI3K/Akt dependent pathways.     

Periostin基因过表达对顺铂和5-氟尿嘧啶诱导人胃癌细胞凋亡的影响

背景：肿瘤细胞耐药性的产生是肿瘤化疗失败的主要原因之一。Periostin是一种基质特异性细胞黏附分子。既往研究发现,periostin除参与胃癌细胞的生长、侵袭、转移外,还能降低其对化疗药物的敏感性。目的：明确periostin基因过表达对顺铂和5-氟尿嘧啶（5．Fu）诱导人胃癌细胞凋亡的影响及其可能机制。方法：将表达人periostin全长序列的重组质粒稳定转染人胃癌细胞株SGC7901,同时设置空载体稳定转染组和未转染对照组,加入不同浓度顺铂或5-Fu处理24h,以流式细胞术检测细胞凋亡。以蛋白质印迹法检测经5μmol／L顺铂或10μmol／L5-Fu处理的SGC7901细胞的凋亡相关蛋白表达。结果：顺铂和5-Fu均能剂量依赖性地诱导SGC7901细胞凋亡,促进细胞色素c由线粒体释放至胞质,同时easpase-3激活,easpase-3底物PARP剪切增强。periostin稳定转染组SGC7901细胞对顺铂和5-Fu诱导的细胞凋亡具有显著抗性,细胞凋亡率显著低于经相同剂量化疗药物处理的空载体稳定转染组（P〈0．05）,胞质细胞色素C、cleavedcaspase-3、cleavedPARP蛋白表达亦明显下调。结论：过表达periostin基因可增强SGC7901细胞对顺铂和5-Fu诱导的细胞凋亡的抵抗能力,其抗凋亡活性与抑制线粒体依赖性凋亡途径有关。     

Induction of apoptosis by arsenic trioxide and hydroxy camptothecin in gastriccancer cells in vitro

AIM To study the effects of arsenic trioxide and HCPT on different degrees of differentiated gastric cancer cells (SGC-7901, MKN-45, MKN-28)with respect to both cytotoxicity and induction of apoptosis in vitro. ~ODS The cytotoxicity of As2O3 and HCPT on gastric cancer cells was determined by MTTassay. Morphologic changes of apoptosis of gastric cancer cells were observed by light microscopy and transmission electron microscopy. Apoptosis and cell cycle changes of gastric cancer cells induced by HCPT and As2O3 were investigated by TUNEL method and flow cytometry. RESULTS As2O3 and HCPT had remarkable cytotoxic effects on different degrees of differentiated gastric cancer cells. The IC50 of As2O3 on well differentiated gastric cancer cell MKN-28, moderately differentiated gastric cancer cell SGC-7901, and poorly differentiated gastric cancer cell MKN-28 were 8. 91 μmol/L, 10. 57 μmol/L, and 11.65 μmol/L, respectively. The IC50 of HCPT on MKN-28, SGC-7901, and MKN-45 were 9. 35 rg/L, 10. 21 rg/L, and 12. 63 mg/L respectively after 48 h treatment. After 12 h of exposure to both drugs, gastric cancer cells exhibited morphologic features of apoptosis, including cell shrinkage, nuclear condensation,and formation of apoptotic bodies. A typical subdiploid peak before G0/G1 phase was observed by flow cytometry. The apoptotic rates of SGC7901, MKN-45, and MKN-28 were 13. 84%, 22.52%, and 9. 68%, respectively after 48 h exposure to 10 μmol/L As2O3. The apoptotic rates of SGC-7901, MKN-45, and MKN-28 were 21.88%, 12.35%, and 30. 26%, respectively after 48 h exposure to 10 mg/L HCPT. The apoptotic indice were 7% - 15% as assessed by TUNEL method. The effect of As2O3 on SGC-7901 showed remarkable cell cycle specificity, which induced cell death in G1 phase, and blocked G2/M phase. HCPT also showed a remarkable cell cycle specificity, by inducing cell death and apoptosis in G1 phase and arrest of proliferation at S phase. CONCLUSION AS2O3 and HCPT exhibit significant cytotoxicity on gastric cancer cells by induction of apoptosis. As2O3 and HCPT might have a promising prospect in the treatment of gastric cancer, which needs to be further studied.     

NS-398对人胃癌细胞株增殖及COX-2表达的影响

目的 体外观察选择性环氧化酶2(COX-2)抑制剂NS-398对人胃癌细胞株SGC7901细胞增殖及COX-2表达的影响。方法 采用噻唑蓝(MTT)法观察NS-398对SGC7901细胞增殖的影响，流式细胞仪(FCM)研究NS-398对SGC790l细胞凋亡的作用．免疫细胞化学观察COX-2蛋白的表达。结果 体外NS-398能减少SGC790l细胞株COX-2的表达．对SGC7901有细胞毒作用．可增加细胞凋亡率。结论 体外NS-398对SGC7901细胞增殖有抑制作用。可能与抑制COX-2表达及诱导细胞凋亡有关。     

海洋微生物次级代谢产物1403-B的抗胃癌活性

目的筛选能被海洋微生物次级代谢产物1403-B显著抑制的肿瘤细胞,初步探讨其机制,为临床抗肿瘤提供候选药物。方法用2.5、5.0、10、20、40μmol/L 1403-B分别处理鼻咽癌细胞株(3种:HONE-1、CNE-2、5-8F),肝癌细胞株(2种:HepG2、Bel7402),胃癌细胞株(SGC7901)48 h后,MTT法检测细胞增殖的情况;荧光显微镜观察细胞形态学变化;用Annexin V/PI双染法分析其凋亡情况,Western印迹法检测caspase-3的切割。结果 MTT实验显示,1403-B呈浓度依赖性抑制多种肿瘤细胞的增殖(P<0.05),其中对胃癌细胞SGC7901的IC50最低,为(4.12±0.57)μmol/L。1403-B作用SGC7901细胞24 h后,出现细胞膜、细胞核皱缩,染色质浓集和形成包裹细胞核片段的凋亡小体等凋亡现象,且呈剂量依赖性增加。Annexin V/PI双染法显示,随着1403-B浓度增加,SGC7901凋亡率升高。1403-B促进SGC7901细胞caspase-3切割。结论海洋药物1403-B能够通过诱导SGC7901凋亡而显著抑制其增殖,有望成为一种抗胃癌的新药物。