Expression of E-selectin, integrinβ_1 and immunoglobulin superfamily member in human gastric carcinoma cells and its clinicopathologic significance

AIM： To study the expression levels of E-selectin, integrin β1 and immunoglobulin supperfamily memberintercellular adhesion molecule-1 （ICAM-1） in human gastric carcinoma cells, and to explore the relationship between these three kinds of cell adhesion molecules and gastric carcinoma. METHODS： The serum contents of E-selectin, integrin β1 and ICAM-1 were detected by enzyme-linked immunosorbent assay （ELISA）, in 47 healthy individuals （control group） and in 57 patients with gastric carcinoma （gastric carcinoma group） respectively prior to operation and 7 d after operation. RESULTS： The serum E-selectin, ECAM-1 and integrin β1 were found to be expressed in both control and gastric carcinoma groups. However, they were highly expressed in patients with gastric carcinoma patients before operation or with unresectable tumours. The expression levels of ICAM-1 and integrin β1 were significantly higher in gastric carcinoma patients than in controls （P 〈 0.01）. A comparison of the E-selectin levels between the two groups showed statistically insignificant differnce （P = 0.64）. In addition, the expression levels were all decreased substantially in the postoperative patients subjected to radical resection of the tumours, indicating that the high level expressions of these compounds might be the important factor for predicting the prognosis of these patients. CONCLUSION： Serum E-selectin, ICAM-1 and integrin β1 expression levels are probably related to the metastasis and relapse of gastric cancer.

Low intensity ultrasound-induced apoptosis in human gastric carcinoma cells

AIM： To investigate the low intensity ultrasound （US）- induced apoptosis in human gastric carcinoma cells and its potential mechanism and to suggest a new therapeutic approach to gastric carcinoma. METHODS： Human SGC-7901 gastric carcinoma cells were cultured in vitro and irradiated by low intensity US for 10 min at different intensities with different incubation times after irradiation. Morphologic changes were examined under microscope with trypan blue staining and then the percentage of early apoptotic cells was detected by flow cytometry （FCM） with double staining of fiuorescein isothiocyanate （FITC）- Annexin V/propidium iodide （PI）. Two-dimensional electrophoresis （2DE） was used to get the protein profile and some proteins differently expressed after US irradiation were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry （MALDI-TOF-MS）. Functional analysis was performed to investigate the mechanism of US-induced cell apoptosis. RESULTS： The percentage of apoptotic cells increased about 10% after US irradiation （12.0 W/cm^2, 12 h culture）, The percentage of early apoptosis and secondary necrosis in the US-irradiated cells increased with the increased US intensity. Moreover, apoptotic cells increased with the increased culture time after US irradiation and reached its maximum at about 12 h.Several new proteins appeared after US irradiation and were up or down regulated more than 2 times. Some heat shock proteins （HSPs） were found to be associated with the signal process simulating the apoptosis of cells. CONCLUSION： Low intensity US could induce apoptosis in human gastric carcinoma cells. US-induced apoptosis is related to US intensity/culture time. US-induced apoptosis may be caspases-dependent and endoplasmic reticulum （ER） stress-triggered apoptosis may also contribute to it. Proteomic experimental system is useful in finding the protein alteration in carcinoma cells after US irradiation, helping to develop a new cancer therapy.

Inhibitory effect of Fuzheng Yiliuyin in combination with chemotherapeutics on human gastric carcinoma cell strain

AIM： To study the inhibitory effects of Fuzheng Yiliuyin （Decoction for Suppressing Tumors by Strengthening the Body Resistance） in combination with chemotherapeutics on human gastric carcinoma cell strain. METHODS： Fuzheng Yiliuyin （ZY） combined with various kinds of chemotherapeutics was put into two kinds of cultivated human gastric carcinoma cell strains, then its inhibitory effects on human gastric carcinoma cell strains were determind by the MTT method. Flow cytorneter was used to assay the apoptosis rate, and the ultrastructure of gastric carcinoma cells was observed under transmission electron microscope. RESULTS： Obvious apoptosis was seen in gastric carcinoma cells after treatment with ZY for 72 h. ZY and chemical drugs had synergistic inhibition effects on the cultivated gastric carcinoma cells, but the effects were different on various cell strains. The inhibitory effects of ZY could be strengthened by cytotoxic action and apoptosis. ZY combined with tluorouracil, etoposide and cisplatin （EFP） chemotherapeutics had better inhibitory effects on SGC-7901, while ZY combined with EFP or with DDP chemotherapeutics had better inhibitory effects than other drugs on MGC-803. CONCLUSION： ZY induces apoptosis and inhibits the growth of gastric carcinoma cells. ZY has the synergistic function of chemotherapeutics.

INTERACTIONS BETWEEN THE HUMAN GASTRIC CARCINOMA CELL AND THE HUMAN VASCULAR ENDOTHELIAL CELL

Objective To definite the interactions between the human gastric carcinoma cell and the human vascular endothelial cell during the establishment and maintenance of the tumor vascular system and the tumor hematogenous metastasis.Methods We prepared the conditioned mediums of each cell so as to study the effect of the conditioned medium on itself or others by MTT colorimetry. The comprehensive effect of interactions between two cells was determined by stratified transfilter co culture or direct contact co culture.Results The conditioned medium of human gastric carcinoma cell can stimulate the proliferation of the human vascular endothelial cell, but the CM of HVEC can inhibit the growth of HGCC. Both kinds of cells can inhibit the growth of itself. The ultimate comprehensive effect of the interactions between two kinds of cells was increase of total cell numbers.Conclusion There exist the complicated interactions between the human gastric carcinoma cell and the human vascular endothelial cell during the tumor angiogenesis and the tumor hematogenous metastasis. The ultimate comprehensive effect of the interactions is increase of total cells numbers and tumor volume.

Growth-inhibiting and Apoptosis-inducing Effects of Tanshinone ⅡA on Human Gastric Carcinoma Cells

To explore the effects of Tanshinone Ⅱ A on the proliferation, apoptosis and gene expression of p53 and bcl-2 in human gastric carcinoma MKN-45 cells. Cell count and MTT assay were used to study the proliferation-inhibiting effect of Tanshinone Ⅱ A on MKN-45 cells. The effect of Tanshinone Ⅱ A on the cell cycle and apoptosis of MKN-45 cells were examined by propidium iodide （PI） staining and flow cytometry. Semi-quantitative RT-PCR was used to further verify the ex- pression of p53 and bcl-2 gene after exposure to Tanshinone Ⅱ A in MKN-45 cells. The results showed that Tanshinone Ⅱ A significantly inhibited the growth and proliferation of MKN-45 cells in a dose- and time-dependent manner （P〈0.05）. Tanshinone Ⅱ A arrested MKN-45 cells in G2/M phase which led to an obvious accumulation of G2/M phase cells while decreased number of Go/G1 phase cells. This resulted in apoptosis of MKN-45 cells and the apoptosis rate was as high as 43.91% after treatment with 2.0 lag/mL Tanshinone Ⅱ A for 96 h. It was also found that Tanshinone Ⅱ A up-regulated expression of p53 gene and down-regulated expression of bcl-2 gene. The cytostatic and antiproliferative effect of Tanshinone Ⅱ A makes it a promising anticancer agent for the treatment of gastric carcinoma.

Paris Saponin Ⅰ Induces G_2/M Cell Cycle Arrest and Apoptosis in Human Gastric Carcinoma SGC7901 Cells

The aim of this study was to investigate the effect of Paris saponin I （PSI ） on human gastric carcinoma cell growth and apoptosis and to explore the potential mechanisms. The proliferation of SGC7901 cells was monitored by the MTT cell viability assay, while the nuclear morphology of apoptotic cells was assessed by Hoechst 33258 staining. Flow cytometry was performed to analyze the cell cycle progression of propidium iodide （PI）-stained SGC7901 cells and the apoptotic rate of annexin V/PI-stained ceils. Western blotting was used to examine the expression of several cell cycle proteins, including cyclin 131 and Cdkl, and the apoptosis-regulated proteins Bcl-2, Bax, cytochrome c, procaspase-9, and procaspase-3. The MTT assay demonstrated that PSI could induce significant dose- and time-dependent inhibition of SGC7901 cell proliferation. Marked morphological changes, including condensation of chromatin, nuclear fragmentation and apoptotic bodies were clearly shown on Hoechst 33258 staining. PS I treatment also resulted in the disruption of the cell cycle at Gz/M and the induction of apoptosis. Following PSI treatment, the cell cycle-related proteins cyclin B 1 and Cdkl were down- regulated. Expression of the pro-apoptotic protein Bax was increased, while anti-apoptotic protein Bcl-2 decreased. PSI treatment resulted in elevated cytoplasmic cytochrome c and activation of the apoptotic proteases caspase-9 and caspase-3. These data indicate that PS I acts as an inhibitor of proliferation in SGC7901 cells by inducing cell cycle arrest and mitochondria-dependent apoptosis. PSI is a potential therapeutic agent against human gastric carcinoma.

DIFFERENCES IN EXPRESSION AND REGULATION BETWEEN TRANSFORMED CELLS OF THE HUMAN GASTRIC CARCINOMA ONCOGENE Ha-ras AND THE UNTRANSFORMED PARENT CELLS

An Ha-ras oncogene was isolated from a cell line of gastric carcinoma called BGE-823 in order to elucidate genetic control and the influence of DNA sequences. The oncogene was cloned and identified as a single nucleotide substitution of thymine for guanine in the 12th codon through the sequencing of its first axon. We compared the differences of expression and regulation between the transformed Ha-ras cells and untransformed parent cells. Data indicated that the expression of Ha-ras in the transformed cells was five-fold higher than in the untransformed cells and that the Ha-ras gene in the former was hypersensitive toward DNase I. In addition, a nuclear protein of 35 kilodaltons bound strongly to the 2.5 Kb fragment located upstream of the 6.6 Kb Ha-ras gene and contained a CC rich region. These results suggest that there might be another mechanism of activation for the ras gene besides point mutation.

Human papillomavirus DNA and P16～(INK4A) expression in concurrent esophageal and gastric cardia cancers

AIM:To investigate the relationship between human papillomavirus (HPV) infection and concurrent esophagus and gastric cardia cancer from the same patient (CC) and examine the significance of P16 INK4A protein expression.METHODS:Polymerase chain reaction was used to detect the presence of HPV type16 (HPV16).The expression of P16 INK4A protein was detected using immunohistochemistry.RESULTS:Among the CC specimens,HPV16-DNA was found in eight cases of esophageal squamous cell carcinoma (ESCC) and five cases of gastric cardia adenocarcinoma (GCA),respectively (47% vs 29%),and two of both ESCC and GCA.P16 INK4A was highly expressed in both ESCC and GCA.In the HPV-associated positive CC,higher P16 INK4A expression was observed in the GCA than in the ESCC (75% vs 25%,P < 0.05).CONCLUSION:HPV16 as a correlated risk factor may play an important role in the development of ESCC and GCA.P16 INK4A may be a screening index in the HPVassociated carcinoma of gastric cardia.

共轭亚油酸对人胃腺癌细胞的抑制作用

采用体外细胞培养方法，研究共轭亚油酸（ＣＬＡ）对人胃腺癌细胞（ＳＧＣ－７９０１）生长的影响。ＣＬＡ的剂量分别为２５、５０、１００和２００μｍ ｏｌ／Ｌ，以乙醇为溶剂对照。结果表明：ＣＬＡ 能抑制ＳＧＣ－７９０１细胞的增殖、细胞核分裂、集落形成和细胞ＤＮＡ的合成。

苦荞异槲皮苷对人胃癌细胞SGC-7901增殖及凋亡的影响

从苦荞中提取制备异槲皮苷,研究其对人胃癌细胞SGC-7901增殖、凋亡、迁移和细胞周期的影响。将异槲皮苷作用于人胃癌SGC-7901细胞和人肾上皮细胞系293T,通过噻唑蓝法检测异槲皮苷对其增殖的影响;4’,6-二脒基-2-苯基吲哚(4’,6-diamino-2-phenyl indole,DAPI)荧光染色法观察细胞核的形态学变化;划痕擦伤迁移实验检测异槲皮苷对SGC-7901细胞迁移能力的影响;流式细胞术检测异槲皮苷对SGC-7901细胞凋亡及细胞周期的影响。结果表明:苦荞异槲皮苷可以抑制SGC-7901细胞的增殖,并呈时间和剂量依赖性,当用100μmol/L异槲皮苷作用细胞48 h后,对SGC-7901细胞的增殖抑制率达到35.92%,而对人肾上皮细胞系293T的增殖抑制率仅为3.15%;DAPI荧光染色法观察异槲皮苷处理细胞后,染色体凝聚,有凋亡小体产生;划痕擦伤实验显示,异槲皮苷能抑制SGC-7901细胞的迁移;流式细胞术检测结果表明,异槲皮苷可使G1和S期细胞减少,G2/M期细胞增多,且细胞凋亡率明显增加。综上所述,苦荞麦异槲皮苷能够诱导SGC-7901细胞发生凋亡,阻断细胞周期并抑制细胞增殖和迁移。

桉油精对补骨脂素体外抗肿瘤活性的增效作用研究

目的考察桉油精对补骨脂素体外抗肿瘤活性的增效作用。方法采用MTT法观察桉油精与补骨脂素对4种人胃癌细胞的生长抑制作用,计算IC50,选择对补骨脂素最为敏感的一种癌细胞,以确定桉油精与补骨脂素的最佳比例与浓度。结果补骨脂素作用于SGC-7901、MKN-45、BGC-823、MGC-803的IC50分别为36.23、66.26、36.9、40.27μg/mL,白蔻仁挥发油及桉油精单用无显著抗肿瘤作用,但其与补骨脂素联用,有显著的抑制增强作用。选用对补骨脂素最为敏感的SGC-7901进一步研究。结论桉油精对补骨脂素的体外抗肿瘤活性具有协同作用。桉油精与补骨脂素配伍的最佳抑制比例与剂量为2∶1,72∶36μg/mL。

40种香豆素类化合物对人胃癌细胞株BGC和人肝癌细胞株BEL-7402细胞生长抑制活性的筛选

目的:寻找中药中抗肿瘤活性化合物和先导化合物,为开发新药奠定基础。方法:采用人胃癌细胞株BGC和人肝癌细胞株BEL-7402体外培养法,鉴定受试的40种香豆素类化合物对其生长的抑制作用。结果:伞形花内酯刘寄奴酸酯、木桔素、环氧酸橙皮油素、酸橙皮油素、前胡酮、芸香苦素、牛防风素和异香柑内酯对人胃癌细胞株BGC细胞的生长有一定程度的抑制作用,且呈浓度-效应关系;伞形花内酯刘寄奴酸酯、异虎耳草素、前胡素和二氢山芹醇当归酸酯对人肝癌细胞株BEL-7402细胞的生长有一定程度的抑制作用,且呈浓度-效应关系。结论:香豆素类化合物是潜在的抗肿瘤药物。

维生素E琥珀酸酯抑制人胃癌细胞生长的研究

采用体外细胞培养的方法 ,观察维生素 E琥珀酸酯 (VES)对人胃癌细胞 (SGC- 790 1)生长的影响 ,并利用 MTT法比较 VES和α-生育酚对胃癌细胞生长的作用效果。结果表明 :VES(5、10和 2 0μg/ m l)对胃癌细胞生长有明显抑制作用 ,而 α-生育酚在同样剂量对肿瘤细胞生长无抑制作用。这提示 VES具有不同于维生素 E的抗癌机理。

共轭亚油酸对人胃腺癌细胞侵袭能力的影响及其作用机制

研究c9,t11-共轭亚油酸 (CLA)对人胃腺癌细胞 (SGC 790 1)侵袭能力的影响 ,探讨其抑制肿瘤转移的可能机理。用 0、2 5、5 0、10 0和 2 0 0 μmol LCLA处理细胞 2 4h后 ,分别用重组基底膜侵袭实验评价癌细胞侵袭能力 ;PAGE底物酶谱方法检测Ⅳ型胶原酶活性 ;RT PCR方法检测SGC 790 1细胞TIMP 1和TIMP 2mR NA表达。研究结果表明 :c9,t11 CLA处理后 ,SGC 790 1细胞侵袭重组基底膜的能力下降、SGC 790 1细胞培养上清中的Ⅳ型胶原酶活性降低、SGC 790 1细胞中TIMP 1和TIMP 2mRNA的表达增加。因此 ,c9,t11 CLA抑制SGC 790 1细胞侵袭重组基底膜 ,并且c9,t11 CLA的抗侵袭活性与降低肿瘤细胞培养上清中Ⅳ型胶原酶活性和诱导肿瘤细胞TIMP 1和TIMP

马钱子生物碱成分的体外抗肿瘤活性筛选

目的从中药中筛选有抗肿瘤活性的化合物。方法应用人鼻咽癌细胞株KB、人白血病细胞株HL-60、人胃癌细胞株BGC和人肝癌细胞株BEL-7402筛选6种马钱子生物碱类成分对其生长的抑制作用。结果士的宁对人鼻咽癌细胞株KB、人白血病细胞株HL-60和人胃癌细胞株BGC等细胞的生长有一定的抑制作用;伪士的宁对人鼻咽癌细胞株KB、人胃癌细胞株BGC和人肝癌细胞株BEL-7402等细胞的生长有一定的抑制作用;依卡金对人鼻咽癌细胞株KB和人胃癌细胞株BGC等细胞的生长有一定的抑制作用。结论士的宁、伪士的宁和依卡金对人鼻咽癌细胞株KB和人胃癌细胞株BGC等细胞的生长皆有抑制作用,呈现浓度-效应关系。

KDR反义寡核苷酸对人胃癌细胞的作用

目的 用 KDR反义寡核苷酸作用于人胃腺癌细胞 ,观察对细胞增殖及超微结构的影响 ,检测作用后 KDR基因的表达 ,并测定细胞分泌 VEGF的能力 .方法 MTT法测定细胞的增殖能力 ,电镜下观察细胞的超微结构 .原位杂交及免疫组化法检测 KDR m RNA及蛋白的表达 .ELISA法测细胞培养液中人 VEGF的含量 .结果 KDR ASODN可明显的抑制人胃腺癌细胞增殖 .增殖抑制率最高可达 55.4% .当剂量为 2 0μmol· L-1 时还可诱发细胞凋亡 .KDR ASODN能明显抑制细胞内 KDR m RNA和蛋白的表达 .胃癌细胞还能分泌一定量的人 VEGF:(92± 1 .7) ng· L-1 .结论 胃癌细胞能分泌一定量的人 VEGF.KDR ASODN能抑制细胞内 KDR基因的表达 ,并显著抑制胃癌细胞的增殖 .在一定剂量下还可诱发其凋亡 .说明 VEGF及受体 KDR在人胃腺癌细胞的增殖。

44种生物碱类化合物对人胃癌细胞株BGC和人肝癌细胞株BEL-7402细胞增殖抑制活性的筛选

目的:从中药中筛选有抗肿瘤活性的化合物。方法:应用人胃癌细胞株BGC和人肝癌细胞株BEL-7402筛选44种生物碱类化合物对其增殖的抑制作用。结果:伊波加因、哈尔明碱、黄连碱、小檗碱、延胡索碱、小檗胺、吐根碱、马兜铃酸I、番茄碱苷、澳洲茄次碱、茄次碱、金鸡宁、罂粟碱和酒石酸麦角胺对人胃癌细胞株BGC细胞的增殖有一定程度的抑制作用,且呈浓度-效应关系;最强者为黄连碱、延胡索碱、小檗胺、吐根碱、番茄碱苷、澳洲茄次碱和金鸡宁,其次为伊波加因、哈尔明碱、小檗碱、马兜铃酸I、茄次碱、罂粟碱和酒石酸麦角胺。蛇根精、γ-崖椒碱、黄连碱、小檗碱、延胡索碱、吐根碱、马兜铃酸I、茄次碱和喜树碱对人肝癌细胞株BEL-7402细胞的增殖有一定程度的抑制作用,且呈浓度-效应关系;最强者为吐根碱、小檗碱、延胡索碱和喜树碱,其次为黄连碱、马兜铃酸I和茄次碱。结论:上述生物碱类化合物可能是含有该生物碱、有抗肿瘤活性中药的有效成分,亦可能成为开发抗肿瘤新药的候选药物。

金雀异黄素诱导人胃癌细胞凋亡的机制研究

采用 4’ ,6 二乙酰基 2 苯基吲哚染色、琼脂糖凝胶电泳实验观察了金雀异黄素 (Gen)对体外培养的人胃癌SGC 790 1细胞的凋亡诱导作用 ,并采用WesternBlot法检测Gen对人胃癌细胞p5 3、Caspase 3蛋白的表达情况。结果显示 ,Gen诱导了胃癌细胞凋亡 ,并抑制突变型p5 3蛋白表达 ,增加Caspase 3蛋白表达。结果提示 ,Gen抑制突变型p5 3蛋白表达 ,增加Caspase 3蛋白表达 。

桂皮醛诱导人胃癌BGC-823细胞凋亡及相关分子机制的探讨

目的:探讨桂皮醛对人胃癌BGC-823细胞增殖、凋亡及其相关分子机制。方法:采用MTT法检测桂皮醛对人胃癌BGC-823细胞增殖的影响;采用流式细胞仪检测桂皮醛诱导的凋亡,Hoechst 33342染色法观察凋亡形态的变化;RT-PCR法检测桂皮醛对人胃癌BGC-823细胞相关凋亡基因的影响;Western Bolt方法检测桂皮醛对人胃癌BGC-823细胞相关凋亡蛋白的影响。结果:与对照组相比,桂皮醛有抑制人胃癌BGC-823细胞增殖的作用(P<0.01);桂皮醛能诱导凋亡的产生;桂皮醛能下调凋亡相关基因Bcl-2、Bcl-xL、Survivin,上调凋亡相关基因Bax、Bak;桂皮醛能下调凋亡相关蛋白Bcl-2、Procaspase-3,上调凋亡相关蛋白Bax。结论:桂皮醛对人胃癌细胞BGC-823的增殖具有抑制作用并能诱导凋亡,可能与内源性凋亡途径的激活有关。