桥接整合因子1 通过c-MYC途径抑制非小细胞肺癌A549 细胞中PD-L1的表达

目的：研究人非小细胞肺癌（non-small cell lung cancer,NSCLC）A549 细胞中桥接整合因子-1（bridging intergrator-1,BIN1）对程序性死亡受体-配体1（programmed death-ligand 1,PD-L1）表达的影响及其机制。方法：采用qRT-PCR和Western blotting方法检测A549 细胞和正常人胚肺成纤维细胞2BS中BIN1 与PD-L1 基因和蛋白表达情况,通过基因转染技术、利用阳离子脂质体将含有人全长BIN1 基因序列的真核表达质粒CMV-MCS-GFP-SV40-Neomycin-BIN1 转染到A549 细胞中（BIN1+组）,构建过表达BIN1 的细胞株,采用RNA干扰技术将干扰骨髓细胞瘤病毒癌基因（cellular-myelocytomatosis viral oncogene ,c-MYC）的c-MYC-siRNA转染到A549 细胞中（c-MYC-siRNA组）以敲低c-MYC基因表达,通过qRT-PCR和Western blotting 方法验证转染效果及过表达BIN1 基因或敲低c-MYC基因对A549 细胞中c-MYC 和PD-L1 表达的影响。结果：与2BS 细胞相比,A549 细胞中BIN1 基因和蛋白均呈低表达状态,而PD-L1 呈高表达状态（均P<0.05）。将携带BIN1 基因的真核表达质粒转染到A549 细胞后,BIN1 基因和蛋白表达水平较对照组显著升高（P<0.05）,而PD-L1 表达显著降低（P<0.05）。c-MYC-siRNA 转染到A549 细胞后,细胞内c-MYC表达显著降低（P < 0.01）,PD-L1 表达明显下调（P<0.01）。结论：BIN1 过表达可以通过失活c-MYC通路降低PDL1表达,从而抑制A549细胞的免疫逃逸。     

桥接整合因子1 去甲基化诱导细胞周期阻滞抑制食管鳞状细胞癌EC109细胞增殖

目的：分析桥接整合因子1（bridging integrator-1, Bin1）去甲基化对Bin1 基因表达和食管鳞状细胞癌EC109 细胞增殖能力的影响,并初步探讨其可能的作用机制。方法：甲基化特异性PCR（methylation specific polymerase chain reaction, MSP）法检测去甲基化药物5-氮杂-2''脱氧胞苷（5-Aza-2''-deoxycytidine, 5-Aza-dc）处理后EC109 细胞Bin1 启动子区域的甲基化状态,用qPCR、Western blotting 和MTT法分别检测单独5-Aza-dc 和5-Aza-dc 加转染Bin1 基因干扰片段（Bin1 siRNA）处理对EC109 细胞Bin1 mRNA及其蛋白表达和细胞增殖能力的影响,流式细胞术和Western blotting 检测5-Aza-dc 处理后EC109 细胞周期和细胞周期相关蛋白（Cyclin D1 与CDK4）表达的变化。结果：去甲基化药物5-Aza-dc 处理后,EC109 细胞Bin1 基因启动子区域发生去甲基化。5-Aza-dc 处理的Bin1 去甲基化EC109 细胞Bin1 mRNA和蛋白表达明显上调（均P<0.05）,细胞增殖能力明显下降（P<0.05）,细胞阻滞在G0/G1 期,表现为S 期细胞比例显著减少,细胞周期相关蛋白Cyclin D、CDK4 表达均明显下调（均P<0.05）。Bin1 去甲基化EC109 细胞转染Bin1 siRNA后,Bin1 mRNA和蛋白表达明显下调,细胞增殖能力增强（均P<0.05）。结论：Bin1 基因启动子区域在EC109 细胞中呈完全甲基化状态,5-Aza-dc 去甲基化可使食管鳞癌细胞EC109 细胞Bin1 表达升高,通过降低细胞周期相关蛋白表达诱导细胞周期阻滞抑制EC109 细胞增殖。证实表观遗传学变化可能与食管癌细胞恶性增殖有关,可为食管癌治疗提供新的思路。     

细胞周期蛋白D1在非小细胞肺癌中表达的临床意义

目的 探讨非小细胞肺癌（ＮＳＣＬＣ）中细胞周期蛋白Ｄ１（ＣｙｃｌｉｎＤ１）的表达及其临床意义。方法 用鼠抗人ＣｙｃｌｉｎＤ１单克隆抗体（ＤＣＳ／６）对８９例石蜡包埋的原发性ＮＳＣＣＬ组织进行免疫组化染色，并与２０例肺炎性病变比较。结果 ＣｙｃｌｉｎＤ２在肺鳞癌、腺癌、大细胞癌中都有较高的表达，其阳性率分别为６０．５％、６１．０％、４０．０％；而在肺炎性病变中，在肺鳞癌、腺癌中，肿块直径≥３ｃｍ、有     

乏氧诱导因子-1α在非小细胞肺癌中的表达研究

目的　探讨乏氧诱导因子 1α(HIF 1α)在非小细胞肺癌中的表达水平及其临床意义。方法　采用免疫组织化学技术检测HIF 1α在 68例非小细胞肺癌标本中的表达。结果　HIF 1α在非小细胞肺癌中的阳性表达率为 5 7.3 5 %( 3 9/ 68)。腺癌中HIF 1α阳性表达率为 5 4.76%( 2 3 / 42 ) ,鳞癌为 61.5 4%( 16/ 2 6) (P>0 .0 5 ) ;中高分化的肿瘤标本中HIF 1α阳性表达率 ( 74.2 9%,2 6/ 3 5 )显著高于低分化的肿瘤标本 3 9.3 9%( 13 / 3 3 ) (P <0 .0 5 ) ;Ⅰ～Ⅱ期HIF 1α的阳性表达率 ( 86.96%,2 0 / 2 3 )显著高于Ⅲ～Ⅳ期 42 .2 2 %( 19/ 45 ) (P<0 .0 5 )。HIF 1α的表达与患者性别、年龄及有无淋巴结转移无明显关系。结论　HIF 1α在非小细胞肺癌中有着较高的表达率 ,且与肿瘤的分化程度和临床分期有一定的关系 ,提示在非小细胞肺癌的治疗中应该重视HIF 1α导致的放化疗抗拒     

非小细胞肺癌中细胞周期蛋白D1与E的作用及相互关系

目的 :研究细胞周期蛋白D1与E在非小细胞肺癌 (NSCLC)发生、发展中的作用及相互关系。方法 :免疫组化方法检测 87例NSCLC肿瘤组织中细胞周期蛋白D1、E的表达及PCNA估计增殖指数 (PI) ,并将上述结果与临床病理及预后资料进行对比分析。结果 :87例NSCLC中细胞周期蛋白D1、E阳性率分别为 4 4 .8% (39/87)、4 8.3% (42 /87) ,细胞周期蛋白D1阳性组的PI值显著高于阴性组 (P〈0 .0 5 ) ,细胞周期蛋白E阳性组PI值与阴性组无显著差异 (P〉0 .0 5 ) ;细胞周期蛋白D1阳性组肿瘤直径、淋巴结转移率和生存率均与阴性组有显著差异(P〈0 .0 1、〈0 .0 5、〈0 .0 1) ,细胞周期蛋白E阳性组淋巴结转移率、临床分期和生存率均与阴性组有显著差异 (P〈0 .0 5、〈0 .0 5、〈0 .0 1) ;细胞周期蛋白D1阳性组中细胞周期蛋白E的阳性率显著高于其阴性组 (P〈0 .0 5 ) ;细胞周期蛋白D1与细胞周期蛋白E双阳性组的PI值、肿瘤直径、淋巴结转移率显著高于非双阳性组 (P值均〈0 .0 5 ) ,生存率显著低于非双阳性组 (P〈0 .0 1)。结论 :细胞周期蛋白D1、细胞周期蛋白E均参与NSCLC的发生、发展 ,并影响其预后 ,但两者在其中所起作用不同 :细胞周期蛋白D1是调节NSCLC增殖的主要因素 ,细胞周期蛋白E主要与NSCLC进展有关 ;细胞周期蛋白D1可促     

细胞周期蛋白D_1在非小细胞肺癌中表达的临床意义

目的　探讨非小细胞肺癌 (NSCLC)中细胞周期蛋白D1(CyclinD1)的表达及其临床意义。方法　用鼠抗人CyclinD1单克隆抗体(DCS/6 )对 89例石蜡包埋的原发性NSCLC组织进行免疫组化染色 ,并与 2 0例肺炎性病变比较。结果　CyclinD1在肺鳞癌、腺癌、大细胞癌中都有较高的表达 ,其阳性率分别为 6 0 .5 %、6 1.0 %、40 .0 % ;而在肺炎性病变中 ,则全部阴性。在肺鳞癌、腺癌中 ,肿块直径≥ 3cm、有淋巴结转移或远处转移者 ,其CyclinD1阳性率较高 (P <0 .0 5 ) ,且与临床分期有关 ,Ⅲ～Ⅳ期癌中的阳性率高于Ⅰ～Ⅱ期癌 (P <0 .0 5 )。结论　CyclinD1的过表达与NSCLC的发生、发展和预后等密切相关 ,对NSCLC的诊断、治疗有一定的指导意义 ,可作为一个诊断及预后指标。     

乏氧诱导因子-1α在非小细胞肺癌中的表达研究

目的探讨乏氧诱导因子-1α(HIF-1α)在非小细胞肺癌中的表达水平及其临床意义。方法采用免疫组织化学技术检测乏氧诱导因子-1α在68例非小细胞肺癌标本中的表达。结果(1)HIF-1α在非小细胞肺癌中的总的阳性表达率为57.35%(39/68);(2)腺癌中HIF-1α的阳性表达率为54.76%(23/42),鳞癌中HIF-1α的阳性表达率61.54%(16/26),腺癌与鳞癌之间HIF-1α的阳性表达率无显著差异(P>0.05);中高分化的肿瘤标本中HIF-1α的阳性表达率为74.28%(26/35)高于低分化的肿瘤标本中HIF—1α的阳性表达率39.39%(13/33)(P<0.05);(3)HIF-1α的表达与性别、年龄及有无淋巴结转移和临床分期无关。结论HIF-1α在非小细胞肺癌中有着较高的表达率,且与肿瘤的分化程度有一定的关系。     

基因通过 NF-κB途径抑制非小细胞肺癌A549细胞的迁移和侵袭能力

目的:研究桥接整合因子1(bridging intergrator-1,Bin1)基因过表达对非小细胞肺癌细胞株A549细胞迁移和侵袭能力的影响,并初步探讨其作用机制.方法:通过基因转染技术,利用阳离子脂质体将含有人全长Bin1基因序列的真核表达质粒CMV-MCS-GFP-SV40-Neomycin-Bin1转染到A549细胞株,分别设置空白对照组及空质粒转染组,利用RT-PCR和West-em blotting分别检测各处理组细胞中Bin1基因和蛋白表达水平.通过细胞划痕实验、Transwell侵袭实验分别检测Bin1过表达对A549细胞迁移、侵袭能力的影响;Western blotting实验检测Bin1过表达对A549细胞内NF-κB磷酸化水平和迁移相关蛋白E-钙黏着蛋白、N-钙黏着蛋白、MMP-9表达水平的影响.结果:与空白对照组和空质粒转染组相比,Bin1转染组A549细胞中Bin1基因和蛋白表达水平均明显升高(P＜0.05);Bin1转染组细胞迁移、侵袭能力均较空白质粒组和空白对照组明显下降[穿膜细胞数:(50.50±3.15) vs (124.00±4.25),(130.00±4.37)个;均P＜0.05];与空白转染组和空白对照组相比,Bin1转染组细胞内NF-κB表达水平明显上调(P＜0.05)而p-NF-κB表达明显下调(P＜0.05),N-钙黏着蛋白、MPP-9明显下调(P＜0.05),E-钙黏着蛋白明显上调(P＜0.05).结论:Bin1过表达可以抑制A549细胞的迁移及侵袭能力,其机制可能与NF-κB途径的失活及细胞迁移侵袭相关蛋白表达变化有关.     

乏氧诱导因子-1α在非小细胞肺癌中的表达研究(英文)

目的探讨乏氧诱导因子-1α(HIF-1α)在非小细胞肺癌中的表达水平及其临床意义。方法采用免疫组织化学技术检测乏氧诱导因子-1α在68例非小细胞肺癌标本中的表达。结果 (1)HIF-1α在非小细胞肺癌中的总的阳性表达率为57.35%(39/68);(2)腺癌中HIF-1α的阳性表达率为54.76%(23/42), 鳞癌中 HIF-1α的阳性表达率61.54%(16/26),腺癌与鳞癌之间 HIF-1α的阳性表达率无显著差异(P>0.05);中高分化的肿瘤标本中 HIF-1α的阳性表达率为74.28%(26/35)高于低分化的肿瘤标本中 HIF-1α的阳性表达率39.39%(13/33)(P<0.05);(3)HIF-1α的表达与性别、年龄及有无淋巴结转移和临床分期无关。结论 HIF-1α在非小细胞肺癌中有着较高的表达率,且与肿瘤的分化程度有一定的关系。     

PD-1信号通路在非小细胞肺癌中的研究进展

摘 要：近年来,对于程序性死亡因子-1（programmed death-1,PD-1）与肿瘤免疫逃逸关系的研究已越来越深入,尤其是对以PD-1信号通路为靶点的临床肿瘤治疗研究取得了令人瞩目的进步。全文对PD-1及其信号通路的理论基础、临床前证据及以PD-1通路为靶点在NSCLC中治疗研究进展进行综述,并对其应用前景进行展望。     

LIF promotes tumorigenesis and metastasis of breast cancer through the AKT-mTOR pathway

Leukemia inhibitory factor (LIF) is a multi-functional cytokine protein. The role of LIF in tumorigenesis is not well-understood. Here, we found that LIF promotes tumorigenesis and metastasis of breast cancer. LIF promotes cell proliferation and anchorage-independent growth of breast cancer cells in vitro, and the growth of xenograft breast tumors in vivo. LIF also promotes invasion and migration of breast cancer cells in vitro and metastasis of breast cancer in vivo. We found that LIF activates the AKT-mTOR signaling pathway to promote tumorigenesis and metastasis of breast cancer. Inhibiting the AKT activity can largely block the activation of the mTOR pathway by LIF, suggesting that LIF activates the mTOR pathway through AKT. Inhibiting the AKT activity as well as inhibiting the mTOR activity largely block the promoting effect of LIF on tumorigenesis and metastasis. Furthermore, overexpression of LIF is significantly associated with a poorer relapse free survival in breast cancer patients. Taken together, our data strongly suggest that LIF plays an important role in the tumorigenesis and metastasis of breast cancer, and could be an important prognostic marker for breast cancer.     

Myosin X对肺癌H1975细胞系放射敏感性调节及机制探讨

目的:探讨Myosin X对肺癌H1975细胞系放射敏感性调节及相关机制。方法:蛋白免疫印迹法检测肺癌细胞系中Myosin X表达量以获取实验对象。运用CRISPR/Cas9技术建立敲除Myosin X的H1975细胞系(KO组)和感染对照病毒的H1975细胞系(NC组),并进行敲除效率验证。克隆形成实验及多靶单击模型...     

Oncoprotein HBXIP promotes tumorigenesis through MAPK/ERK pathway activation in non-small cell lung cancer

Objective:The oncoprotein,hepatitis B X-interacting protein(HBXIP),has been reported to play an important role in human malignancies.However,its functions in non-small cell lung cancer(NSCLC)are poorly understood.The goal of the present study was to identify the role of HBXIP in the regulation of NSCLC development.Methods:The level of HBXIP expression in NSCLC tissue was assessed by immunohistochemical and Western blot analyses,and its relationships with clinicopathological features and outcomes were statistically evaluated.The effects of HBXIP on NSCLC cell progression were assessed through cell viability,colony formation,and flow cytometry analyses in vitro.The mechanism by which HBXIP regulated the MAPK pathway was studied by Western blot,immunofluorescence,and immunoprecipitation assays.In addition,in vivo experiments were performed to evaluate the progression of NSCLC and ERK signaling pathway activation after HBXIP knockdown.Results:HBXIP was overexpressed in human NSCLC and was correlated with the invasiveness of NSCLC.The high expression of HBXIP in NSCLC was significantly correlated with gender(P=0.033),N stage(P=0.002),and tumor-node-metastasis stage(P=0.008).In vitro experiments using an NSCLC cell line revealed that HBXIP knockdown resulted in the suppression of cell proliferation and colony formation,which was consistent with the enhanced cell cycle arrest in G1 phase.The results of a mechanistic investigation suggested that binding of HBXIP to MEK1 protein promoted MAPK/ERK signaling pathway activation in NSCLC by preventing the proteasome-mediated degradation of MEK1.In addition,the results obtained using in vivo subcutaneous tumor xenografts confirmed that HBXIP deficiency decreased MEK1 protein levels and NSCLC tumor growth.Conclusions:Taken together,our results showed that the HBXIP-MEK interaction promoted oncogenesis via the MAPK/ERK pathway,which may serve as a novel therapeutic target for cancers in which MAPK/ERK signaling is a dominant feature.     

Rapamycin induces p53-independent apoptosis through the mitochondrial pathway in non-small cell lung cancer cells

The mammalian target of rapamycin (mTOR) is a key kinase acting downstream of growth factor receptor PI3K and AKT signaling, leading to processes resulting in increased cell size and proliferation through translation control. Rapamycin, a specific inhibitor of mTOR, results predominately in G1 cell cycle arrest through translation control and occasionally, cell type-dependent apoptosis by an unknown mechanism. In this study, we investigated the effect and mechanism of action of rapamycin on non-small cell lung cancer (NSCLC) cell lines with p53 mutations. Cell proliferation was evaluated by modified MTT assay. The apoptotic effect of rapamycin was measured by caspase-3 activation and flow cytometric analysis of Annexin V binding. The expression of Bcl-2 and the release of cytochrome?c from mitochondria were evaluated by western blotting. We found that rapamycin induced apoptosis in NSCLC cell lines with p53 mutations. Western blot analysis demonstrated that rapamycin downregulates the expression levels of Bcl-2, which leads to increased cytochrome c release from mitochondria and subsequent activation of caspase cascades. These findings suggest that rapamycin induces p53-independent apoptosis through downregulation of Bcl-2 and the mitochondrial pathway in NSCLC cell lines as a novel antitumor mechanism.     

CDO1通过PI3K/AKT信号通路调控胃癌细胞增殖及细胞周期

目的:探讨半胱氨酸双加氧酶1(CDO1)对胃癌细胞增殖、细胞周期的调控机制。方法:用脂质体法将si-NC组（转染si-NC）、si-CDO1组（转染si-CDO1）、pcDNA组（转染pcDNA）、pcDNA-CDO1组（转染pcDNA-CDO1）、pcDNA-CDO1+DMSO组（转染pcDNA-CDO1并用DMSO处理）、pcDNA-CDO1+IGF-1组（转染pcDNA-CDO1并用IGF-1处理）转染至AKG细胞。用实时荧光定量逆转录聚合酶链反应（qRT-PCR）、免疫印迹（Western blot）、细胞计数试剂盒（CCK-8）、流式细胞术检测细胞CDO1、PI3K、Akt、p-Akt蛋白的表达、细胞增殖、细胞周期。结果:与人胃黏膜上皮细胞GES-1相比,胃腺癌细胞AKG中CDO1的表达明显降低（P<0.05）;与si-NC组相比,si-CDO1组AKG细胞的增殖明显上调,细胞发生明显的S期、G2/M期阻滞,过表达CDO1则具有相反的作用。重要的是,敲减CDO1可上调PI3K/AKT信号通路关键基因PI3K、p-Akt的表达,而过表达CDO1具有相反的作用。激活PI3K/AKT信号通路后,过表达CDO1对胃癌细胞的增殖、细胞周期的调控作用可被部分逆转。结论:CDO1可抑制胃癌细胞的增殖,调控细胞周期,其机制与抑制PI3K/AKT信号通路的活性有关,将为胃癌的治疗提供参考。     

Overexpression of TYRO3 indicates poor prognosis and induces gastric cancer progression via AKT-mTOR pathway

Gastric cancer (GC) is among of the leading causes of cancer mortality worldwide. This is because many patients are diagnosed with advanced GC and postoperative radiotherapy and chemotherapy have also exhibited limited effects on GC. TYRO3 has been considered carcinogenic and a potential therapeutic target for GC. However, TYRO3 function and mechanism in GC remains elusive. The study results indicated that TYRO3 was aberrantly elevated in GC tissues and predicted poor prognosis. TYRO3 is closely associated with clinicopathological indicators in GC tissues such as lymph node metastasis, venous invasion, neural invasion, and the tumor-node-metastasis stage. In addition, TYRO3 expression levels are closely related to the AKT-mTOR pathway in GC tissues. Moreover, the oncogenic role of TYRO3 was determined through in vitro and in vivo functional assays, and knockdown of the TYRO3 expression level in GC cell lines can effectively suppress the AKT-mTOR pathway and inhibit tumor cell proliferation and migration. In conclusion, this study provides a theoretical basis for establishing the potential association and regulatory mechanism between TYRO3 and AKT-mTOR and offers a new strategy for GC-targeted therapy.     

KCTD10抑制非小细胞肺癌中Hippo通路活性

目的:检测KCTD10在非小细胞肺癌中的表达水平及其对Hippo通路的影响。方法:采用免疫组化方法检测KCTD10在非小细胞肺癌组织标本中的表达;采用Western blot检测YAP、P-YAP、P-LATS1、MMP7、CyclinE在非小细胞肺癌细胞中的表达;采用集落形成实验和Transwell实验检测细胞增殖和迁移。结果:KCTD10在非小细胞肺癌组织及细胞系中高表达;KCTD10过表达促进A549细胞增殖和迁移;过表达KCTD10的A549细胞系YAP的表达总量未见明显变化,但抑制了P-YAP、P-LATS1的表达,促进了MMP7、CyclinE的表达。结论:KCTD10抑制了非小细胞肺癌中Hippo通路活性;KCTD10可能作为非小细胞肺癌防治的一个潜在靶向分子。     

晚期非小细胞肺癌化疗周期的新争议

2003年美国临床肿瘤协会(ASCO)年会中Socinski教授对颇有争议的晚期、转移性非小细胞肺癌(NSCLC)的化疗周期有一篇精彩的综述，文章中报告了针对ⅢB/Ⅳ期NSLC的3项Ⅲ期临床试验结果，其中2项试验是比较一线常规治疗(一组为3个和4个周期化疗，另一组为6个或6个以上周期)的结果。观察指标包括：短期缓解率、症状缓解率、生活质量、生存率。这2项试验结果发现长化疗周期组(≥6     

Inhibition of RalA signaling pathway in treatment of non-small cell lung cancer

Non-small cell lung cancer (NSCLC) is the most common type of lung cancer and relatively resistant to chemotherapy. The most prevalent molecular abnormality in NSCLC is the overactivation of K-Ras proto-oncogene; therefore, elucidating down-stream Ras signaling in NSCLC is significantly important in developing novel therapies against this malignancy. Our work indicates that RalA, an important effector of Ras, is activated in NSCLC cell lines. While RalA was also overactivated in fetal human broncho-epithelial cells, RalBP1 (Ral binding protein-1), an important down-stream effector of RalA, was expressed at higher levels in cancer cell lines. Aurora kinase-A (AKA), an upstream activator of RalA, was also found to be active only in malignant cells. The outcome of inhibition of RalA (by gene specific silencing using a lentivirus) on the malignant phenotype of A549 cells was also studied. While proliferation and invasiveness of A549 cells were reduced upon silencing RalA, apoptosis and necrosis were elevated in such conditions. Additionally, the in vivo tumorigenesis of A549 cells was reduced upon partial inhibition of RalA and AKA using pharmacological inhibitors. Finally, we were interested in evaluating the level of active RalA in the fraction of NSCLC cells expressing cancer stem cell markers. For this purpose cells with increased expression of CD44 were separated from A549 cells and compared with cells with low level of expression of this marker and an unsorted population. A significant enhancement of RalA activation in high CD44+ cells was found as potential evidence for involvement of RalA signaling in initiation of the neoplastic procedure and an important contributor for tumor maintenance in NSCLC. Further studies can reveal therapeutic, preventive and diagnostic value of RalA pathway in this deadly disease.