Silencing profilin-1 inhibits gastric cancer progression via integrin β1/focal adhesion kinase pathway modulation

AIM:To investigate the role of profilin-1(PFN1)in gastric cancer and the underlying mechanisms.METHODS:Immunohistochemical analysis,quan-titative real-time polymerase chain reaction(q RTPCR)and Western blot were performed to detect PFN1expression in clinical gastric carcinoma and adjacent tissues,and the association of PFN1 expression with patient clinicopathological characteristics was analyzed.PFN1 was knocked down to investigate the role of this protein in cell proliferation and metastasis in the SGC-7901 cell line.To explore the underlying mechanisms,the expression of integrinβ1 and the activity of focal adhesion kinase(FAK)and the downstream proteins extracellular-regulated kinase(ERK)1/2,P38 mitogen-activated protein kinase(MAPK),phosphatidylinositol 3-kinase(PI3K),AKT and mammalian target of rapamycin(m TOR)were measured through Western blot or q RT-PCR analysis.Fibronectin(FN),a ligand of integrinβ1,was used to verify the correlation between alterations in the integrinβ1/FAK pathway and changes in tumor cell aggressiveness upon PFN1 perturbation.RESULTS:Immunohistochemical,Western blot and q RT-PCR analyses revealed that PFN1 expression was higher at both the protein and m RNA levels in gastric carcinoma tissues compared with the adjacent tissues.In addition,high PFN1 expression(53/75,70.4%)was correlated with tumor infiltration,lymph node metastasis and TNM stage in gastric cancer,but not with gender,age,location,tumor size,or histological differentiation.In vitro experiments showed that PFN1knockdown inhibited the proliferation of SGC-7901cells through the induction G0/G1 arrest.Silencing PFN1 inhibited cell migration and invasion and downregulated the expression of matrix metalloproteinase(MMP)-2 and MMP9.Moreover,silencing PFN1 reduced the expression of integrinβ1 at the protein level and inhibited the activity of FAK,and the downstream effectors ERK1/2,P38MAPK,PI3K,AKT and m TOR.FN-promoted cell proliferation and metastasis via the integrinβ1/FAK pathway was ameliorated by PFN1silencing.CONCLUSION:These findings suggest that PFN1 plays a critical role in gastric carcinoma progression,and these effects are likely mediated through the integrinβ1/FAK pathway.

Effect of NHE1 antisense gene transfection on the biological behavior of SGC-7901 human gastric carcinoma cells

AIM: To study the effect of type 1 Na+/H+ exchanger (NHE1 ) antisense human gene transfection on the biological behavior of gastric carcinoma cell line SGC-7901. METHODS: Antisense NHE1 eukaryotic expression on vector pcDNA3.1 was constructed by recombinant DNA technique and transfected into gastric carcinoma cell line SGC-7901 with DOTAP liposome transfection method. Morphological changes of cells were observed with optic and electron microscopes. Changes in cell proliferative capacity, apoptosis, intracellular pH (pHi), cell cycle, clone formation in two-layer soft agar, and tumorigenicity in nude mice were examined. RESULTS: Antisense eukaryotic expressing vectors were successfully constructed and transfected into SGC-7901. The transfectant obtained named 7901 -antisense (7901-AS) stablely produced antisense NHE1. There was a significant difference between the pHi of 7901-AS cells (6.77 ± 0.05) and that of 7901-zeo cells and SGC-7901 cells (7.24 ± 0.03 and 7.26 ± 0.03, P < 0.01). Compared with SGC-7901 and 7901-zeo cells, 7901-AS cells mostly showed cell proliferation inhibition, G1/G0 phase arrest, increased cell apoptotic rate, recovery of contact inhibition, and density contact. The tumorigenicity in nude mice and cloning efficiency in the two-layer soft agar were clearly inhibited. CONCLUSION: NHE1 antisense gene significantly restrains the malignant behavior of human gastric carcinoma cells, suppresses cell growth and induces cell apoptosis, and partially reverses the malignant phenotypes of SGC-7901 . These results suggest a potential role for human tumor gene therapy.

ITGA1 polymorphisms and haplotypes are associated with gastric cancer risk in a Korean population

AIM:To evaluate the association between the geneticpolymorphisms and haplotypes of the ITGA1 gene and the risk of gastric cancer.METHODS:The study subjects were 477 age-and sex-matched case-control pairs.Genotyping was performed for 15 single nucleotide polymorphisms(SNPs)in ITGA1.The associations between gastric cancer and these SNPs and haplotypes were analyzed with multivariate conditional logistic regression models.Multiple testing corrections were carried out following methodology for controlling the false discovery rate.Gene-based association tests were performed using the versatile gene-based association study(VEGAS)method.RESULTS:In the codominant model,the ORs for SNPs rs2432143(1.517;95%CI:1.144-2.011)and rs2447867(1.258;95%CI:1.051-1.505)were statistically significant.In the dominant model,polymorphisms of rs1862610 and rs2447867 were found to be significant risk factors,with ORs of 1.337(95%CI:1.029-1.737)and 1.412(95%CI:1.061-1.881),respectively.In the recessive model,only the rs2432143 polymorphism was significant(OR=1.559,95%CI:1.150-2.114).The C-C type of ITGA1 haplotype block 2 was a significant protective factor against gastric cancer in the both codominant model(OR=0.602,95%CI:0.212-0.709,P=0.021)and the dominant model(OR=0.653,95%CI:0.483-0.884).The ITGA1 gene showed a significant gene-based association with gastric cancer in the VEGAS test.In the dominant model,the A-T type of ITGA1 haplotype block 2 was a significant risk factor(OR=1.341,95%CI:1.034-1.741).SNP rs2447867 might be related to the severity of gastric epithelial injury due to inflammation and,thus,to the risk of developing gastric cancer.CONCLUSION:ITGA1 gene SNPs rs1862610,rs2432143,and rs2447867 and the ITGA1 haplotype block that includes SNPs rs1862610 and rs2432143 were significantly associated with gastric cancer.

Integrin αvβ6 and matrix metalloproteinase 9 correlate with survival in gastric cancer

AIM: To investigate the expression of integrin αvβ6 and matrix metalloproteinase 9 (MMP-9), their association with prognostic factors and to assess their predictive role in gastric cancer patients.METHODS: Immunohistochemistry was used to determine the expressions of integrin αvβ6 and MMP-9 in 126 specimens from patients with primary gastric carcinoma. Associations between immunohistochemical staining and various clinic pathologic variables of tissue specimens were evaluated by the χ2 test and Fisher’s exact test. Expression correlation of αvβ6 and MMP-9 was assessed using bivariate correlation analysis. The patients were followed-up every 3 mo in the first two years and at least every 6 mo afterwards, with a median follow-up of 56 mo (ranging from 2 mo to 94 mo). Four different combinations of αvβ6 and MMP-9 levels (that is, both markers positive, both markers negative, αvβ6 positive with MMP-9 negative, and αvβ6 negative with MMP-9 positive) were evaluated for their relative effect on survival. The difference in survival curves was evaluated with a log-rank test. Survival analysis was conducted using the Kaplan-Meier survival and Cox proportional hazards model analysis.RESULTS: The expressions of integrin αvβ6 and MMP-9 were investigated in 126 cases, among which 34.92% were positive for αvβ6 expression, and 42.06% for MMP-9 expression. The expression of αvβ6 was associated with Lauren type, differentiation, N stage, and TNM stage (the P values were 0.006, 0.038, 0.016, and 0.002, respectively). While MMP-9 expression was associated with differentiation, T stage, N stage, and TNM stage (the P values were 0.039, 0.014, 0.033, and 0.008, respectively). The positive correlation between αvβ6 and MMP-9 in gastric cancer was confirmed by a correlation analysis. The Kaplan-Meier survival analysis showed that patients with expression of αvβ6 or MMP-9 alone died earlier than those with negative expression and that patients who were both αvβ6 and MMP-9 positive had a shorter overall survival than those with the opposite pattern (both αvβ6 and MMP-9 negative) (P = 0.000). A Cox model indicated that positive expression of αvβ6 and MMP-9, diffuse Lauren type, as well as a senior grade of N stage, M stage, and TNM stage were predictors of a poor prognosis in univariate analysis. Only αvβ6 and MMP-9 retained their significance when adjustments were made for other known prognostic factors in multivariate analysis (RR = 2.632, P = 0.003 and RR = 1.813, P = 0.007).CONCLUSION: The expression of αvβ6 and MMP-9 are closely correlated, and the combinational pattern of αvβ6 and MMP-9 can serve as a more effective prognostic index for gastric cancer patients.

Effects of angiopoietin-1 on attachment and metastasis of human gastric cancer cell line BGC-823

AIM： To evaluate the effects of angiopoietin-1 （Ang-1） on adhesion of gastric cancer cell line BGC-823 and expression of integrin β1, CD44V6, urokinase-type plasminogen activator （uPA） and matrix metalloproteinase-2 （MMP-2）. METHODS： BGC-823 cells were transfected transiently with adenovirus-Ang-1 （Ad-Ang-1）. Cells transfected transiently with adenovirus-green fluorescent protein （Ad-GFP） and untransfected cells were used as a negative and blank control group, respectively. The cell adhesion rate between cell and extracellular matrix （ECM） was determined by cell adhesion assay. To investigate whether Ang-1 could reinforce gastric carcinoma metastasis, we performed migration and invasion assays in BGC-823 cells. The mRNA and protein expression of integrin β1, CD44V6, uPA and MMP-2 were detected by reverse transcription polymerase chain reaction and Western blotting, respectively. The expression of integrin β1 and CD44V6 was measured by immunohistochemistry. RESULTS： BGC-823 cells were transfected successfully. The adhesion rate increased significantly in the Ad-Ang-1 group （P 〈 0.05）. The Ad-Ang-1-transfected group had a significant increase in migration and invasion compared with that of the mock-transfected and Ad-GFP groups. The mRNA and protein expression of integrin β1, CD44V6, uPA and MMP-2 in the Ad- Ang-1 group was higher than that in the Ad-GFP and blank control groups （P 〈 0.05）. Compared with mocktransfected and Ad-GFP groups, integrin 131 and CD44V6 expression intensity greatly increased （P 〈 0.05）. CONCLUSION： Transfection of Ang-1 into human gastric cancer cell line BGC-823 can significantly increase expression of integrin β1 and CD44V6, by which cell adhesion and metastasis to the ECM are promoted.

A novel long non-coding RNA NFIA-AS1 is down-regulated in gastric cancer and inhibits proliferation of gastric cancer cells

Gastric cancer is one of the most common malignant gastrointestinal tumors whose morbidity and mortality account for the second and third place respectively in malignant tumors in China.As an important participant in tumor biology,the abnormal expression of long non-coding RNA(lncRNAs)in cancer cells is closely related to the occurrence and development of tumors and plays the role of oncogenes or tumor suppressor genes.In this study,we identified a novel lncRNA NFIA antisense RNA 1(NFIA-AS1)and explored its role and clinical significance in gastric cancer.Real-time quantitative PCR was performed to detect the expression of NFIA-AS1 in tumor tissues and corresponding normal tissues from 42 pairs of gastric cancer samples.The lower expression of NFIA-AS1 was significantly associated with larger tumor size,lower histological grade,and advanced TNM stage.Kaplan-meier analysis showed that NFIA-AS1 expression could be used as an independent predictor of overall survival.We also demonstrated that overexpression of NFIA-AS1 significantly inhibited the proliferation of gastric cancer cells through affecting p16 levels.In conclusion,our results suggest that the lncRNA NFIA-AS1 may play the role of tumor suppressor gene,and serve as a biomarker for prognosis or progression of gastric cancer.

E-选择素、整合素β_1、免疫球蛋白超家族成员在人胃癌细胞中的表达及其意义

目的了解E-选择素(E-selectin)、整合素β(1Integrinβ1)、免疫球蛋白超家族成员(ICAM-1)在人胃癌细胞中的表达水平,探讨这3种细胞粘附分子与胃癌的关系。方法采用NothernBlotting法和ELISA法检测胃癌细胞、正常胃上皮细胞和人脐静脉内皮细胞上E-selectin、Integrinβ1及ICAM-1的表达水平并进行比较。结果胃癌细胞、正常胃上皮细胞和血管内皮细胞均有E-selectin、Integrinβ1及ICAM-1的表达。但胃癌细胞3种粘附分子表达水平均高于正常胃上皮细胞和人脐静脉内皮细胞,差别均有显著性意义(P<0.05)。结论E-selectin、Integrinβ1、ICAM-1可能与胃癌细胞转移有关。

硫酸右旋糖苷对人胃癌MKN1细胞的黏附以及整合素β_1表达影响的研究

目的 通过观察细胞的黏附过程和整合素 β1的cDNA的表达 ,研究硫酸右旋糖苷 (DS)抑制人胃癌MKN1细胞株的黏附过程的机制。方法 用位相差显微镜对MKN1细胞在培养盘上的附着过程及形态变化进行了观察。用半定量RT -PCR法对整合素β1的cDNA的表达水平进行了测定。结果 DS抑制了MKN1细胞在培养盘上的附着 ,培养 4 8小时后仍有部分细胞处于游离状态。在DS治疗组 ,黏附细胞和游离细胞内整合素 β1的cDNA的表达水平分别减少到非治疗细胞的 74 %和 38%。结论 DS对人胃癌细胞株MKN1的黏附过程的抑制与对整合素 β1cDNA表达的抑制有关。

Galectin-1和integrin β1在胃癌中表达及其临床意义

目的:探讨galectin-1和integrinβ1在胃癌中表达及其临床意义。方法:采用免疫组织化学技术检测胃癌和癌旁胃组织中galectin-1和integrinβ1的表达,分析其表达与胃癌临床指标的相关性。结果:Galectin-1在胃癌组织中的阳性表达率为60.00%,高于癌旁胃黏膜的13.75%,差异有统计学意义(P<0.05)。Integrinβ1在胃癌组织中的阳性表达率为57.50%,高于癌旁胃黏膜的16.25%,差异有统计学意义(P<0.05)。Galectin-1表达与胃癌的淋巴结转移、临床分期和浸润深度相关。Integrinβ1表达与胃癌的淋巴结转移和浸润深度相关。Galectin-1和integrinβ1在胃癌中的表达呈正相关。结论:Galectin-1和integrinβ1与胃癌发生和恶性演进有关,二者表达存在相关性。

血清lncRNAs ABHD11-AS1、miR-1231表达与胃癌根治术后复发转移及远期预后的相关性

目的探究血清长链非编码RNAα/β水解酶域11反义RNA1(lncRNAs ABHD11-AS1)、微小RNA-1231(miR-1231)表达与胃癌根治术后复发转移及远期预后的相关性。方法选取2018年7月至2020年7月进行胃癌根治术治疗的147例患者(胃癌组),根据复发转移情况,将患者分为复发转移组48例和未复发转移组99例,根据3年预后情况,分为生存组40例和死亡组107例,选取同期体检的健康人员132例作为对照组,采用实时荧光定量PCR法测定血清lncRNAs ABHD11-AS1、miR-1231表达水平,采用Pearson法分析血清lncRNAs ABHD11-AS1、miR-1231的相关性,采用Logistic回归分析影响胃癌根治术后复发转移的因素。结果与对照组比较,胃癌组lncRNAs ABHD11-AS1表达水平显著升高(P<0.05),miR-1231表达水平显著降低(P<0.05)。血清lncRNAs ABHD11-AS1和miR-1231表达呈负相关(r=-0.691,P<0.05)。复发转移组肿瘤直径>5 cm、TNM分期Ⅲ+Ⅳ期、分化程度低分化、浸润深度T3+T4、有淋巴结转移的患者比例及lncRNAs ABHD11-AS1表达水平显著高于未复发转移组(P<0.05),miR-1231表达水平显著低于未复发转移患者,差异有统计学意义(P<0.05)。肿瘤直径、浸润深度、lncRNAs ABHD11-AS1是胃癌根治术后复发转移的独立危险因素,miR-1231是保护因素(P<0.05)。死亡组lncRNAs ABHD11-AS1表达水平显著高于生存组(P<0.05),miR-1231水平显著低于生存组(P<0.05)。结论胃癌患者血清中lncRNAs ABHD11-AS1表达水平升高,miR-1231表达水平降低,与胃癌根治术后复发转移及远期预后密切相关,可能作为胃癌根治术后复发转移及远期预后的标志物。

胃癌组织中lncRNA PTPRG-AS1、miR-599表达与临床病理特征及预后的关系

目的分析胃癌组织中长链非编码RNA(long non-coding RNA,lncRNA)蛋白酪氨酸磷酸酶受体G基因反义链1(protein tyrosine phosphatase receptor G gene antisense chain 1,PTPRG-AS1)和miR-599表达水平,探讨其与临床病理特征及预后的关系。方法选取华中科技大学同济医学院附属梨园医院2016年1月至2020年2月收治的106例胃癌患者为研究对象。检测胃癌组织及瘤旁组织中lncRNA PTPRG-AS1和miR-599水平。结果胃癌组织中lncRNA PTPRG-AS1水平显著高于瘤旁组织,miR-599水平显著低于瘤旁组织(P<0.05)。lncRNA PTPRG-AS1与miR-599水平呈负相关(r=-0.485,P<0.05)。lncRNA PTPRG-AS1、miR-599均与TNM分期、浸润深度、分化程度、淋巴结转移相关(P<0.05)。lncRNA PTPRG-AS1低表达组、miR-599高表达组生存率显著升高(χ^(2)=8.206、6.881,P<0.05)。lncRNA PTPRG-AS1、miR-599表达是影响胃癌患者不良预后的危险因素(P<0.05)。结论胃癌组织中lncRNA PTPRG-AS1和miR-599表达水平与患者临床病理特征及预后密切相关。

LEF1-AS1通过Wnt/β-catenin信号通路对胃癌细胞迁移侵袭的影响

目的探讨淋巴增强结合因子1反义RNA 1(LEF1-AS1)通过调控Wnt/β-连环蛋白(β-catenin)信号通路对胃癌细胞增殖、迁移和侵袭的影响。方法实时荧光定量PCR(qPCR)检测胃癌细胞和正常胃黏膜细胞GES-1中LEF1-AS1的表达情况。BGC-823细胞分为阴性对照组、LEF1-AS1干扰组和LEF1-AS1干扰+Wnt激动剂组。CCK-8法检测细胞活力,划痕实验和Transwell实验检测细胞迁移和侵袭能力;qPCR和Western blot检测Wnt1、β-catenin和Survivin表达。结果LEF1-AS1在胃癌细胞中的相对表达量高于GES-1细胞(P<0.05)。与阴性对照组比较,LEF1-AS1干扰组BGC-823细胞的增殖和迁移侵袭能力均下降(P<0.05)。LEF1-AS1干扰组Wnt1和β-catenin表达水平低于阴性对照组(P<0.05)。LEF1-AS1干扰+Wnt激动剂组BGC-823细胞的增殖和迁移侵袭能力均强于LEF1-AS1干扰组(P<0.05),且β-catenin和Survivin水平较LEF1-AS1干扰组升高(P<0.05)。结论LEF1-AS1沉默通过抑制Wnt/β-catenin信号通路活化来负调控胃癌细胞的生物学行为,该因子有作为胃癌治疗靶点的潜能。

长链非编码RNA脑源性神经营养因子-反义物和信号素3B-反义物1在胃癌病人中的表达及联合超声检查在胃癌诊断中的应用

目的探究长链非编码RNA(lncRNA)脑源性神经营养因子-反义物(BDNF-AS)和信号素3B-反义物1(SEMA3B-AS1)在胃癌病人中的表达及联合超声检查在胃癌诊断中的应用价值。方法2021年1月~2023年2月收治的118例胃癌病人为胃癌组,同期113例胃部良性病变者为良性病变组。实时荧光定量PCR(qRT-PCR)检测血清BDNF-AS和SEMA3B-AS1表达水平,并以平均值分为BDNF-AS高表达组(55例)和BDNF-AS低表达组(63例)、SEMA3B-AS1高表达组(57例)和SEMA3B-AS1低表达组(61例);Kappa检验分析超声诊断与临床病理诊断的一致性;受试者工作特征曲线(ROC)分析血清BDNF-AS和SEMA3B-AS1联合超声对胃癌的诊断价值。结果与良性病变组比较,胃癌组血清BDNF-AS和SEMA3B-AS1水平均明显下降(t=10.205,t=9.590,P<0.05);BDNF-AS和SEMA3B-AS1在肿瘤直径≥3 cm、浸润深度较深、分化程度较低、发生淋巴结转移的胃癌病人中表达水平明显较低(P<0.05);Kappa检验结果显示,超声诊断与临床病理诊断的一致性较高(Kappa值=0.723,P<0.05);ROC结果显示,血清BDNF-AS、SEMA3B-AS1水平和超声诊断胃癌的AUC分别为0.848、0.835、0.861,三者联合诊断的AUC(0.949)显著大于血清BDNF-AS单独诊断的AUC(Z=4.713,P=0.000)、血清SEMA3B-AS1水平单独诊断的AUC(Z=4.112,P=0.0015)和超声诊断的AUC(Z=3.350,P=0.0008),联合诊断的敏感度、特异度优于三者单独诊断。结论血清BDNF-AS和SEMA3B-AS1联合超声检查在胃癌的诊断上具有较高的应用价值。

β1-整合素反义寡核苷酸抑制胃癌细胞系SGC7901粘附侵袭的实验研究

目的:观察β1-整合素反义寡核苷酸对高侵袭性人胃癌细胞SGC7901粘附侵袭的抑制作用。方法:以脂质体为载体将硫代磷酸修饰的反义寡核苷酸转染SGC7901细胞,免疫化学方法观察转染后以Biotin标记的Inte鄄grinβ1asODN在细胞内的分布;RT-PCR及流式细胞术分别测定Integrinβ1mRNA和蛋白表达变化;MTT法和Boy鄄den小室模型分别测定其粘附力和侵袭力。结果:Integrinβ1asODN转染SGC7901后1h,胞浆及胞核内可见均匀分布的浅棕色颗粒,随着时间的延长,颗粒颜色不断加深;RT-PCR结果显示489bp处条带不同程度变浅;Integrinβ1蛋白表达曲线明显左移;粘附和侵袭实验证明,Integrinβ1asODN转染的SGC7901细胞的粘附力及侵袭力明显下降(P<0.001),抑制率分别为54%和76%,明显优于随机序列(P<0.001)。结论:Integrinβ1asODN转染SGC7901可降低其mRNA和蛋白表达水平,从而抑制其对细胞外基质(ECM)的粘附和侵袭。

LncRNA ZFAS1靶向微小RNA-150-5p对胃癌细胞侵袭和迁移的影响

目的探讨长链非编码RNA ZNFX1反义RNA 1(ZFAS1)靶向调控微小RNA-150-5p(miR-150-5p)表达及对胃癌细胞侵袭和迁移的影响。方法采用实时定量PCR(QPCR)检测正常胃黏膜上皮细胞GES-1及不同胃癌细胞(SGC-7901、HGC-27、MGC-803和BGC-823)的ZFAS1水平。脂质体法向BGC-823细胞转染针对ZFAS1的小干扰RNA片段si-ZFAS1(干扰组)或无关序列si-NC(NC组)并设未转染的细胞为对照组。QPCR、MTT法、Transwell小室实验和划痕实验测定BGC-823细胞的ZFAS1水平、增殖、侵袭和迁移能力的变化,QPCR和Western blotting检测Bcl-2和基质金属蛋白酶(MMP)-9的水平;双荧光素酶报告基因实验验证ZFAS1对miR-150-5p的靶向调控作用。结果胃癌细胞的ZFAS1水平均高于GES-1细胞(P<0.05),后续干扰实验选取ZFAS1水平最高的BGC-823细胞。干扰组的ZFAS1水平为0.269±0.074,低于对照组的1.171±0.263和NC组的1.088±0.142(P<0.05)。与NC组和对照组相比,干扰组BGC-823细胞的增殖活力下降(P<0.05);干扰组的划痕愈合率和穿膜细胞数目分别为(51.509±5.348)%和(145.662±12.328)个,低于NC组的(82.317±7.459)%和(341.342±21.814)个及对照组的(80.770±6.163)%和(339.234±17.255)个(P<0.05);干扰组的Bcl-2和MMP-9水平均低于其余两组(P<0.05)。对照组和NC组上述指标的差异无统计学意义(P>0.05)。miR-150-5p模拟物能够抑制含有其结合位点的ZFAS1野生型质粒的荧光素酶活性(P<0.05),不影响ZFAS1突变型质粒的荧光素酶活性(P>0.05)。结论ZFAS1在胃癌细胞中高表达,下调其水平可抑制BGC-823细胞的增殖和迁移侵袭能力并发挥类似癌基因的作用,可能与靶向调控miR-150-5p有关,在胃癌靶向治疗中有一定应用前景。

促血管生成素-1对人胃癌细胞黏附和转移作用的研究

目的:探讨促血管生成素-1(Ang-1)对人胃癌细胞中整合素β1(integrin β1)和CD44V6的作用,研究其对肿瘤细胞黏附和转移的影响及可能的作用机制。方法:利用腺病毒为载体,将已构建成功的、含Ang-1基因的重组腺病毒(Ad-Ang-1)瞬时转染人胃癌细胞株BGC-823,转染含绿色荧光蛋白基因的重组腺病毒(Ad-GFP)作为阴性对照,未转染的正常细胞作为空白对照,用细胞黏附法检测转染前后细胞与细胞外基质黏附率的改变,再分别用半定量RT-PCR和Western blot方法对3组细胞中整合素β1和CD44V6的mRNA和蛋白表达水平进行分析。结果:细胞黏附实验显示,转染Ad-Ang-1后,细胞与细胞外基质之间的黏附率明显增强(P<0.01);RT-PCR、Western blot结果显示整合素β1、CD44V6 mRNA和蛋白的表达在Ad-Ang-1组明显高于空白对照组及Ad-GFP组。结论:转染Ang-1能明显提高人胃癌细胞BGC-823中整合素β1、CD44V6 mRNA、蛋白的表达,从而促进肿瘤细胞的黏附和转移。

硫酸右旋糖苷对人胃癌细胞腹腔种植转移和整合素β1表达的影响

目的探讨硫酸右旋糖苷(DS)对人胃癌细胞腹腔种植转移整合素β1表达的影响。方法培养BGC-823细胞,构建裸鼠动物模型。将90只裸鼠,随机分为对照组(40只)与实验组(50只),对照组与实验组裸鼠在注射肿瘤细胞同时分别注射生理盐水与硫酸右旋糖苷。裸鼠剖腹行HE染色,观察癌结节的腹腔种植数量;应用免疫组化和RT-PCR方法检测整合素β1mRNA和蛋白表达。结果剖腹观察的瘤结节数经HE染色证实,实验组明显少于对照组的瘤结节数(P<0.01)。免疫组化和RT-PCR结果显示,实验组整合素β1的表达量明显小于对照组(P<0.01)。结论硫酸右旋糖苷可能通过下调整合素β1的表达抑制人胃癌细胞的腹腔种植转移,预防性使用大分子硫酸右旋糖苷能够抑制胃癌细胞的腹腔种植转移。

NHE-1反义基因对胃癌细胞的酸化作用及其意义

目的探讨Na+-H+交换泵-1(Na+-H+exchanger1,NHE-1)反义基因转染对人胃癌细胞内pH值(intracellu-larpH,pHi)的调节作用及其在肿瘤基因治疗中的价值。方法构建人NHE-1基因反义真核表达载体,采用脂质体法将其转染至SGC-7901胃癌细胞中,采用荧光探针检测转染前后细胞内pH值的变化以及对细胞增殖和凋亡的影响。结果转染反义基因的细胞pHi明显降低,增殖能力降低,细胞凋亡率增加。结论NHE-1基因的表达在肿瘤细胞pHi及增殖和凋亡调节中起重要作用。

长链非编码RNA AFAP1-AS1在胃癌组织中的表达及其临床意义

目的:探讨长链非编码RNA(long non-coding RNA,lncRNA)肌动蛋白纤维相关蛋白-1反义RNA1(actin filamentassociated protein 1-antisense RNA1,AFAP1-AS1 RNA1)在胃癌组织中的表达情况及其临床意义。方法:采用Real-time PCR方法检测2010年1月至2014年12月宜昌市第二人民医院及三峡大学仁和医院手术切除的274例胃癌组织及相应癌旁正常胃组织中AFAP1-AS1的表达,并分析AFAP1-AS1表达量与临床及病理特征的关系。结果:AFAP1-AS1在胃癌组织比癌旁正常组织显著高表达(P<0.01)。AFAP1-AS1在早期胃癌组织中平均表达量明显低于进展期胃癌(P<0.01)。在不同病理类型胃癌中,AFAP1-AS1的表达量没有差异(P>0.05)。AFAP1-AS1表达量和胃癌淋巴侵袭或远处转移密切相关,在伴有淋巴结侵袭或远处转移的胃癌组织中,AFAP1-AS1明显高表达(P<0.01)。结论:AFAP1-AS1可能是胃癌发生发展的一个重要因素;有望成为新的胃癌预后标志物,用于胃癌的临床诊断和预后判断。

Tiam 1反义寡核苷酸转染对胃癌细胞体内外侵袭转移能力的影响

背景与目的：作为细胞骨架结构调节因子，T淋巴瘤侵袭转移诱导因子1（Tiam 1）与胃肠道肿瘤侵袭转移密切相关。本研究应用反义寡核苷酸（ASODN）转染技术，特异抑制胃癌细胞中Tiam 1表达，进而观察对胃癌细胞体内、外侵袭转移能力的影响。方法：采用层粘连蛋白黏附法，由胃癌MKN-45细胞株（M0）中筛选获得高侵袭转移亚株（MH）。以脂质体介导将Tiam 1 ASODN转染至MH细胞中，并应用逆转录聚合酶链反应（RT-PCR）及定量细胞ELISA技术分别检测Tiam 1 mRNA和蛋白的表达。应用Boyden小室及裸鼠接种法分别观察ASODN转染后MH细胞在体内、外侵袭转移能力方面的改变。结果：应用0．43μmol／L Tiam 1 ASODN转染能特异性抑制胃癌MH细胞中Tiam 1 mRNA和蛋白表达（0．162±0．018，0．982±0．119），与脂质体转染组（0．789±0．054，1．237±0．108）、正义寡核苷酸（SODN）-脂质体转染组（0．754±0．039，1．234±0．103）、未转染组（0．801±0．065，1．290±0．182）相比，分别约为后三者的20．2％-21．5％、76．1％-79．6％（P〈0．05）。ASODN转染组细胞的体外侵袭、移行能力（10±3、21±9）较SODN转染组（27±10、56±13）、未转染组（26±6、53±12）明显受抑制（P〈0．05），同时ASODN转染组细胞在裸鼠体内的肺转移率（1／5=20％）也较SODN转染组（4／5=80％）、未转染组（4／5=80％）显著下降（P〈0．05）。结论：特异性ASODN转染可有效抑制Tiam 1在胃癌细胞中的表达并削弱其体内、外侵袭转移能力。