siRNA沉默FAM83A基因表达对非小细胞肺癌细胞增殖、迁移和侵袭的影响

目的:研究FAM83A基因对非小细胞肺癌细胞增殖、迁移、侵袭及肿瘤生长的影响。方法:运用qRT-PCR法检测非小细胞肺癌细胞H1299中FAM83A的表达;将blank组（未作任何处理）、si-control组（转染si-control）、si-FAM83A组（转染si-FAM83A）用脂质体法转染至H1299细胞;Western blot检测细胞中FAM83A、p16、p21、MMP-2、MMP-9的蛋白表达;MTT法检测细胞的增殖;Transwell小室检测细胞的迁移和侵袭;裸鼠成瘤实验检测肿瘤的生长。结果:沉默FAM83A后,H1299细胞的增殖、迁移和侵袭能力均得到显著下调,并且p16、p21的蛋白表达明显上调,MMP-2、MMP-9的蛋白表达明显下调。最重要的是,沉默FAM83A后的异种移植裸鼠体内的肿瘤生长能力得到明显抑制。结论:沉默FAM83A可抑制非小细胞肺癌细胞的增殖、迁移和侵袭,并抑制裸鼠体内肿瘤的生长。     

circTADA2A在肺腺癌组织和细胞中的表达及其对A549细胞迁移及侵袭影响的实验研究

目的:研究环状RNA TADA2A(circular RNA TADA2A,circTADA2A)在肺腺癌(lung adenocarcinoma,LAD)组织和细胞中的表达及其对A549细胞迁移及侵袭的影响.方法:在Gene Expression Omnibus(GEO)数据库中下载非编码RNA阵列分析文件GSE10...     

CaMKⅡ高表达通过EMT促进肺腺癌侵袭转移

  目的  探究钙/钙调蛋白依赖性蛋白激酶Ⅱ（calcium/calmodulin-dependent protein kinase Ⅱ,CaMKⅡ）在肺腺癌组织中的表达及其促进肺腺癌的侵袭、转移。  方法  通过免疫组织化学染色（immunohistochemistry,IHC）分析肺腺癌患者的石蜡组织标本CaMKⅡ表达与临床病理参数关系,同时对肺腺癌组织与其配对淋巴结转移病灶中的CaMKⅡ表达情况进行比较分析,收集2011年1月至2011年12月于天津医科大学肿瘤医院行手术治疗的113例肺腺癌患者及21例配对的原发病灶及淋巴结转移病灶的石蜡组织标本。将肺腺癌细胞系H1299及Calu3进行慢病毒转染,实验组转染高表达CaMKⅡ病毒,对照组转染阴性对照病毒,通过Transwell实验和划痕实验检测肺腺癌细胞侵袭、转移能力,通过Western blot 检测CaMKⅡ表达水平与上皮间充质转化（pithelial-mesenchymal transition,EMT）相关指标和表皮生长因子受体（epidermal growth factor receptor,EGFR）通路激活间的关系。  结果  CaMKⅡ高表达与肺腺癌TNM分期和淋巴结转移呈正相关,肺腺癌淋巴结转移病灶中的癌组织CaMKⅡ表达明显高于其原发病灶（P<0.05）。细胞实验表明,CaMKⅡ高表达的肺腺癌细胞,其穿膜细胞数目明显增多,伤口愈合能力增强; EGFR通路激活后p-CaMKⅡ水平增加,且CaMKⅡ促进了EMT相关蛋白及转录因子的表达。  结论  CaMKⅡ参与EGFR信号传导,促进EMT过程,增加肺腺癌的侵袭转移能力。      

LINC00668在肺鳞癌中高表达并促进肿瘤细胞的侵袭转移

背景 与目的由于缺乏有效的治疗措施,肺鳞癌(lung squamous cell carcinoma,LUSC)仍是制约非小细胞肺癌(non-small cell lung cancer,NSCLC)5年生存率的重要影响因素.长链非编码RNA 00668(LINC00668)参与调控多种肿瘤的发生发展,但其在LUSC中...     

KIF3A在不同病变胃组织及淋巴结转移灶中的表达及其与胃腺癌转移和预后的关系

目的 检测KIF3A在不同病变胃组织及淋巴结转移灶中的表达,探讨其与胃腺癌发生、转移、临床病理特征及预后的关系。方法 应用免疫组织化学技术检测不同胃组织及淋巴结转移灶中KIF3A的表达。采用SPSS23.0软件包对数据进行统计分析。结果 KIF3A在胃腺癌组织中的表达水平明显高于癌旁相对正常组织（P<0.01）,而淋巴结转移灶中的表达水平较相应原发灶中更高（P<0.01）;低级别、高级别上皮内瘤变中KIF3A的表达水平均明显高于慢性胃炎组织（P<0.01）;KIF3A的表达与进展期胃腺癌患者的TNM分期、淋巴结转移、脉管侵犯相关（P<0.05）;进展期胃腺癌患者中,KIF3A高表达者总体生存和无病生存时间较低表达者明显缩短（P<0.01）,且KIF3A在细胞质及细胞核都有表达者总体生存时间较仅细胞质表达者短（P<0.05）; Cox多因素分析显示,KIF3A是胃腺癌患者总体生存期与无病生存期的独立危险因素（P<0.05）。结论 KIF3A可能与胃黏膜上皮内瘤变的发生有关,其表达增强和核表达可能与进展期胃腺癌患者预后不良有关,是进展期胃腺癌患者的独立预后因素。     

lncRNA NKILA促进结直肠腺癌细胞的增殖和侵袭

目的:探讨lncRNA NKILA对结直肠腺癌细胞增殖和侵袭的影响。方法:对结直肠腺癌病人的肿瘤组织和癌旁组织进行lncRNA测序分析,以肿瘤组织表达明显增高的lncRNA NKILA作为研究对象。将结直肠腺癌细胞DLD-1分为3组,即未转染慢病毒的空白对照组,转染无序序列慢病毒的阴性对照组,以及转染沉默lncRNA NKILA序列的试验组。流式细胞术检测转染效率,qRT-PCR检测lncRNA表达。CCK-8实验和Transwell实验分别检测DLD-1细胞的增殖和侵袭。将三组细胞分别皮下注射到小鼠体内构建荷瘤小鼠模型,观察移植瘤体积以及生存时间。Western blot检测lncRNA NKILA对DLD-1细胞中NF-κB信号通路的影响。结果:相比癌旁组织,结直肠癌组织中的lncRNA NKILA表达显著增高。通过转染慢病毒,试验组DLD-1细胞中lncRNA NKILA表达明显降低（P＜0.01）。CCK-8检测结果显示,相比阴性对照组,试验组增殖率降低（P＜0.05）。Transwell实验结果显示,相比阴性对照组,试验组DLD-1细胞侵袭能力明显降低（P＜0.01）。试验组小鼠肿瘤体积较阴性对照组减小（P＜0.05或P＜0.001）,生存时间明显延长（中位生存期75 vs 58 d,P＜0.05）。lncRNA NKILA可以抑制IκBα的磷酸化和p65的核转位。结论:lncRNA NKILA可以促进结直肠腺癌细胞DLD-1的增殖和侵袭,这种作用与lncRNA NKILA抑制NF-κB信号通路相关。     

RAB5A对人肺腺癌细胞系侵袭转移作用的研究

目的：探讨RAB5A基因在人肺腺癌细胞系侵袭和转移中的作用。方法：采用体外重建基底膜侵袭实验和癌细胞粘附能力；癌细胞趋化性运动能力、癌细胞分泌明胶酶的能力及活性的测定，分析RAB5A正义真核表达载体转染后的AGZY83－a和反义RNA转染后的Anip973细胞的侵袭、转移能力的改变。结果：RAB5A正义表达载体PcDNA3.1-RAB5A转染的AGZY83－a重建基底膜侵袭能力明显增强，统计学意义显著（P＜0．0005），细胞趋化性运动能力显著增高（P＜0．0005），对基底膜成分的粘附能力增大（P＜0．05），以及明胶酶分泌及活性增强等一系列趋向Anip973的变化。PcDNA3-AntiRAB5A反义RNA表达载体对Anip973重建基底膜侵袭力明显下降（P＜0．0025），趋化性运动能力显著降低（P＜0．005），对基底膜成分粘附能力降低（P＜0．05），以及明胶酶分泌减少等一系列趋向AGZY83－a的变化。结论：RAB5A在肺腺癌侵袭轩移表型形成中发挥重要的作用。反义RNA可阻断RAB5A基因的翻译过程。     

miR-625-5p通过靶向调控PRKACA促进肺腺癌细胞的增殖和侵袭

背景 与目的:肺腺癌是非小细胞肺癌的一种亚型,虽在诊断和治疗方面已经取得了很大进展,但晚期肺腺癌临床预后和总生存仍较差.近年来多项研究表明,miRNA在多种癌症中发挥作用,并在细胞增殖、转移、炎症等生物学过程中发挥重要作用.探究miR-625-5p对肺腺癌细胞增殖和侵袭能力的影响及分子机制,旨在为后续肺腺癌的诊断和治疗...     

过表达CUL4A对肺腺癌A549细胞生物学功能的影响

目的:探究CUL4A对人肺腺癌细胞株A549生物学行为的影响。方法:通过慢病毒感染的方法,构建CUL4A过表达的A549细胞株,通过体外平板克隆和Transwell实验检测A549细胞株的增殖、迁移和侵袭能力。结果:通过RT-PCR和Western-blotting证实CUL4A过表达的A549细胞株构建成功。 与对照组的细胞相比,上调CUL4A的表达能够促进A549细胞的增殖、迁移和侵袭能力。结论:我们的研究结果强调了CUL4A在人肺腺癌细胞株A549中的重要性,并提示CUL4A不仅可能与肺癌的发生和转移有关,而且可能成为肺癌潜在的治疗靶点以及生物标志物。     

FAM83A and FAM83A-AS1 both play oncogenic roles in lung adenocarcinoma

Lung adenocarcinoma (LUAD) is the most common subtype of lung cancer. Nevertheless, the detailed molecular mechanisms of the progression of LUAD remain largely unknown. The present bioinformatics analysis reported that FAM83A and FAM83A-AS1 were upregulated in LUAD tissues and associated with prognosis in patients with LUAD. The purpose of the current study was to investigate the role of FAM83A and its antisense long non-coding (lnc)RNA FAM83A-AS1 in LUAD. Gene Expression Profiling Interactive Analysis was used to screen for potential oncogenes in LUAD and to analyze the clinical significance of FAM83A and FAM83A-AS1. Small interfering RNAs were constructed and transfected into LUAD cells to knock down the expression of FAM83A and FAM83A-AS1. EdU, Cell Counting Kit-8, Transwell and Matrigel assays were performed to detect the proliferation, migration and invasion of LUAD cells. The interaction between FAM83A-AS1, microRNA (miR)-495-3p and FAM83A was explored using a luciferase reporter assay. FAM83A and FAM83A-AS1 were both overexpressed in LUAD tissues compared with adjacent normal tissues. High expression of FAM83A and FAM83A-AS1 predicted worse survival and more advanced clinical stage. Knockdown of FAM83A or FAM83A-AS1 could inhibit the proliferation, migration and invasion of LUAD cells. Moreover, lncRNA FAM83A-AS1 regulated the expression of FAM83A by functioning as competing endogenous RNA for miR-495-3p. These results implicated that FAM83A and FAM83A-AS1 both played oncogenic roles in LUAD and FAM83A-AS1 could regulate the expression of FAM83A by sponging miR-495-3p. The study revealed a novel regulatory mechanism of tumor development in LUAD and FAM83A and FAM83A-AS1 may be novel biomarkers and therapeutic targets for LUAD.     

A non-secretory form of FAM3B promotes invasion and metastasis of human colon cancer cells by upregulating Slug expression

FAM3B mRNA has been predicted to have multiple splicing forms. Its secretory form PANDER is decreased in gastric cancers with high invasiveness and metastasis. Here we found that its non-secretory form FAM3B-258 was highly expressed in most colon cancer cell lines and colorectal adenocarcinoma tissues but not hepatocellular carcinoma, lung carcinoma and pancreatic adenocarcinoma cell lines. Elevation of FAM3B-258 was associated with poor cancer cell differentiation. Stable overexpression of FAM3B-258 in colon cancer cells downregulated adhesion proteins, upregulated Slug and Cdc42, promoted cell migration and invasion in vitro and metastasis in nude mice. Slug mediated FAM3B-258-induced downregulation of adhesion molecules, upregulation of Cdc42, and invasion of colon cancer cells. The expression of FAM3B-258 in human colorectal adenocarcinomas was positively correlated with Slug. These results suggest that FAM3B-258 promotes colon cancer cell invasion and metastasis through upregulation of Slug.     

STAMBP promotes lung adenocarcinoma metastasis by regulating the EGFR/MAPK signaling pathway

Tumor metastasis is a leading cause of death in lung adenocarcinoma (LUAD) patients, but the molecular events that regulate metastasis have not been completely elucidated. STAMBP is a deubiquitinating enzyme of the Jab1/MPN metalloenzyme family that regulates the stability of substrates in cells by specifically removing ubiquitin molecules. We found that STAMBP expression was increased in the cytoplasm of tumor cells from LUAD patients. The STAMBP level was closely associated with tumor size, lymph node invasion and neoplasm disease stage. A high STAMBP level predicted poor overall survival and disease-free survival in LUAD patients. STAMBP overexpression promoted cell migration and invasion, whereas STAMBP knockdown attenuated these processes in LUAD cells after epidermal growth factor treatment. Mechanistically, increased STAMBP expression promoted the stabilization of Epidermal growth factor receptor (EGFR), whereas STAMBP knockdown induced the degradation of EGFR. STAMBP may deubiquitinate EGFR by localizing in early endosomes and increase EGFR membrane localization in LUAD cells. The overexpression of STAMBP triggered the activation of MAPK signaling after epidermal growth factor treatment. In contrast, this activation was attenuated in STAMBP knockdown cells. Small molecule inhibitors of EGFR and MAPK signaling pathway may block STAMBP-induced cell mobility and invasion as well as ERK activation in cells. Importantly, STAMBP knockdown suppressed LUAD tumor growth and metastasis by regulating the EGFR-mediated ERK activation in a xenograft mouse model. Our findings identified STAMBP as a novel potential target for LUAD therapy.     

Upregulation of FAM83D affects the proliferation and invasion of hepatocellular carcinoma

The identification of potential oncogenes plays an important role in finding novel therapeutic targets for many cancers, including hepatocellular carcinoma (HCC), which is one of the most common cancers worldwide. In our previous research, using microarray technology, we found that FAM83D was overexpressed in HCCs. However, whether the overexpression of FAM83D contributes to hepatocarcinogenesis remains unclear. In this study, we found that FAM83D was significantly upregulated in 76.6% (167 of 218) of the HCC specimens at the mRNA level and in 69.44% (50 of 72) of the HCC specimens at the protein level compared with adjacent non-cancerous liver specimens, as indicated by RT-PCR and immunohistochemical staining, respectively. The FAM83DmRNA expression level was positively correlated with the level of alpha-fetoprotein (AFP) (≥100 ng/ml), the clinical TNM stage, the presence of a portal vein tumor thrombus (PVTT), disease-free survival (DFS) and the overall survival (OS) time of the HCC patients (P < 0.05). Knocking down FAM83D significantly promoted the growth of Huh7 and HepG2 cells, as demonstrated in an RNA interference assay. Moreover, the DNA methylation status of the FAM83D promoter was significantly reduced in the HCC specimens with overexpression of FAM83D gene. Our data suggest that the upregulation of FAM83D, a potential oncotarget gene, may be triggered by epigenetic events and can contribute to hepatocarcinogenesis.     

miR-148b在肺腺癌中的表达及其功能分析

目的 探讨微小RNA-148b（miR-148b）在肺腺癌组织和细胞株中的表达及临床意义。方法 采用实时定量PCR检测13例肺腺癌及癌旁组织中miR-148b的表达,并检测其在肺腺癌细胞株H1437、H1975、A549、PC14/B和正常人肺上皮细胞株HBE中的表达,选取肺腺癌细胞株miR-148b表达水平最低者分别转染miR-148b模拟物（mimics）和miR-148b control。通过MTT检测、低密度细胞集落形成实验、Transwell实验检测miR-148b对细胞增殖、集落形成和侵袭能力的影响。结果 miR-148b在肺腺癌和癌旁组织中的相对表达量分别为0.61±0.42和0.91±0.32（P＜0.05）;与正常肺上皮细胞株HBE（相对表达量设为1.00）比较,4种肺腺癌细胞株H1437、H1975、A549、PC14/B中miR-148b的表达量分别为0.42±0.08、0.38±0.02、0.29±0.03和0.21±0.04（P＜0.05）,选取PC14/B细胞株进行后续实验。MTT检测显示,实验第3天,转染miR-148b mimics的PC14/B细胞吸光值明显低于转染miR-148b control组（P＜0.05）;低密度细胞集落形成实验中,与转染了miR-148b control的PC14/B细胞（相对克隆数设为100）相比,转染miR-148b mimics的PC14/B细胞的相对克隆数为47±8,差异有统计学意义（P＜0.05）;Transwell实验显示,转染miR-148b mimics后穿过基底膜的相对PC14/B细胞数为82±22,与miR-148b control组（200±34）相比,差异有统计学意义（P＜0.05）。结论 miR-148b在肺腺癌组织和细胞中表达下调,且可抑制肺腺癌PC14/B细胞株的增殖、集落形成和侵袭能力。     

Effects of bone sialoprotein on pancreatic cancer cell growth, invasion and metastasis

Bone sialoprotein (BSP) is an acidic glycoprotein that plays an important role in cancer cell growth, migration and invasion. The expression, localization and possible function of BSP in chronic pancreatitis (CP) and pancreatic ductal adenocarcinoma (PDAC) were analyzed by QRT-PCR, laser capture microdissection, DNA microarray analysis, immunoblotting, radioimmunoassays and immunohistochemistry as well as cell growth, invasion, scattering, and adhesion assays. BSP mRNA was detected in 40.7% of normal, in 80% of CP and in 86.4% of PDAC samples. The median BSP mRNA levels were 6.1 and 0.9copies/microl cDNA in PDAC and CP tissues, respectively, and zero copies/microl cDNA in normal pancreatic tissues. BSP was weakly present in the cytoplasm of islet cells and ductal cells in 20% of normal pancreatic tissues. BSP was localized in the tubular complexes of both CP and PDAC, as well as in pancreatic cancer cells. Five out of 8 pancreatic cancer cell lines expressed BSP mRNA. Recombinant BSP (rBSP) inhibited Capan-1 and SU8686 pancreatic cancer cell growth, with a maximal effect of -46.4+/-12.0% in Capan-1 cells and -45.7+/-14.5% in SU8686 cells. rBSP decreased the invasion of SU8686 cells by -59.1+/-11.2% and of Capan-1 cells by -13.3+/-3.8% (P<0.05), whereas it did not affect scattering or adhesion of both cell lines. In conclusion, endogenous BSP expression levels in pancreatic cancer cells and low to absent BSP expression in the surrounding stromal tissue elements may indirectly act to enhance the proliferation and invasion of pancreatic cancer cells.     

THBS2在肺腺癌组织中的表达及其对肺腺癌细胞迁移、侵袭和上皮间质转化的影响

目的:研究血小板反应蛋白2（thrombospondin 2,THBS2）在肺腺癌组织中的表达及其对肺腺癌细胞H1299和A549迁移能力、侵袭能力、上皮间质转化（epithelial-mesenchymal transition,EMT）的影响,并探究分子机制。方法:运用TIMER和GEPIA数据库分析THBS2 mRNA在泛肿瘤和肺腺癌组织中的表达,利用UALCAN数据库分析THBS2 mRNA在I-IV期肺腺癌组织中的表达差异,通过HPA数据库检测THBS2蛋白在肺腺癌组织中的表达情况。通过免疫印迹实验（Western blot）检测THBS2在人正常支气管上皮细胞（BEAS-2B）和肺腺癌细胞（H1299、A549）中的表达情况。应用质粒转染技术在肺腺癌细胞H1299和A549中分别过表达和敲低THBS2。细胞划痕实验和侵袭实验（Transwell）检测细胞迁移和侵袭能力。Western blot验证THBS2过表达和敲低效率,同时检测EMT相关蛋白和MMP2的表达水平。结果:TIMER数据库与GEPIA数据库检索结果显示THBS2 mRNA在肺腺癌组织中高表达（P＜0.001）。UALCAN数据库分析证明,与正常肺组织相比,THBS2 mRNA在I-IV期肺腺癌组织中均显著表达（P＜0.001）。HPA数据库结果显示,THBS2蛋白在肺腺癌组织中呈现中高等强度表达。细胞划痕实验、Transwell实验和Western blot结果表明,THBS2在人正常支气管上皮细胞和肺腺癌细胞中的表达有显著差异（P＜0.01）;过表达THBS2可增加肺腺癌细胞H1299和A549迁移与侵袭能力（P＜0.01）,EMT相关蛋白E-cadherin表达水平减少,而N-cadherin和Vimentin表达水平增加（P＜0.01）;相反的,敲低THBS2可抑制肺腺癌细胞H1299和A549迁移与侵袭能力（P＜0.01）,EMT相关蛋白E-cadherin表达水平增加,而N-cadherin和Vimentin表达水平减少（P＜0.01）。结论:THBS2在肺腺癌组织中高表达;THBS2在肺腺癌细胞（H1299、A549）中的表达均显著高于人正常支气管上皮细胞（BEAS-2B）;THBS2在肺腺癌细胞H1299和A549中的过表达和敲低影响迁移、侵袭和EMT的过程,可作为肺腺癌治疗的新型潜在靶点。