Study of cellular immunity response of mB7-1 gene transfected mouse ovarian cancer cell line and its tumorigenecities in vivo

Objective:To investigate the cellular immunity response in vitro and the tumorigenecities in vivo of mB7 1 gene transfected murine ovarian cancer cell line.Methods:mB7 1 gene was transfected into the NuTu 19 cell line by retrovirus vector, and the expression of mB7 1 gene was confirmed by flow cytometry(FCM).NuTu 19/neo and NuTu 19/mB7 1 cells were injected subcutaneously into syngeneic Fischer 344 rats respectively,and their tumorigenecities were recorded.Proliferation indices of lymphocyte were assayed after syngenieic mixed tumor lymphocyte cultures(MTLCs).The lysis activity of CTL toward tumor cells was determined using methyl thiazolyl tetrazolium(MTT) assay.Results:Successful transfection of mB7 1 gene into NuTu 19 cell line was comfirmed with FCM.In vitro study showed that there was no obvious changes in cell growth of gene transfected cell line,compared with the cell line NuTu 19.NuTu 19/mB7 1 cells could induce more effective proliferation of effector lymphocytes(P<0.05). The lysis activity of CTL activated by NuTu 19/mB7 1 was stronger than that of NuTu 19/neo (P<0.01).Tumor sizes were smaller in the NuTu 19/mB7 1 receptance syngeneic Fischer 344 rats compared with those in the control group.Conclusion:mB7 1 genetically modified ovarian cancer cells could induce the cellular immunity response in vitro and the tumorigenecitiy of NuTu 19 cells was decreased after inoculation with the experimental vaccine.

Identification of an immune-related gene-based signature to predict prognosis of patients with gastric cancer

BACKGROUND Gastric cancer(GC)is the most commonly diagnosed malignancy worldwide.Increasing evidence suggests that it is necessary to further explore genetic and immunological characteristics of GC.AIM To construct an immune-related gene(IRG)signature for accurately predicting the prognosis of patients with GC.METHODS Differentially expressed genes(DEGs)between 375 gastric cancer tissues and 32 normal adjacent tissues were obtained from The Cancer Genome Atlas(TCGA)GDC data portal.Then,differentially expressed IRGs from the ImmPort database were identified for GC.Cox univariate survival analysis was used to screen survival-related IRGs.Differentially expressed survival-related IRGs were considered as hub IRGs.Genetic mutations of hub IRGs were analyzed.Then,hub IRGs were selected to conduct a prognostic signature.Receiver operating characteristic(ROC)curve analysis was used to evaluate the prognostic performance of the signature.The correlation of the signature with clinical features and tumor-infiltrating immune cells was analyzed.RESULTS Among all DEGs,70 hub IRGs were obtained for GC.The deletions and amplifications were the two most common types of genetic mutations of hub IRGs.A prognostic signature was identified,consisting of ten hub IRGs(including S100A12,DEFB126,KAL1,APOH,CGB5,GRP,GLP2R,LGR6,PTGER3,and CTLA4).This prognostic signature could accurately distinguish patients into highand low-risk groups,and overall survival analysis showed that high risk patients had shortened survival time than low risk patients(P<0.0001).The area under curve of the ROC of the signature was 0.761,suggesting that the prognostic signature had a high sensitivity and accuracy.Multivariate regression analysis demonstrated that the prognostic signature could become an independent prognostic predictor for GC after adjustment for other clinical features.Furthermore,we found that the prognostic signature was significantly correlated with macrophage infiltration.CONCLUSION Our study proposed an immune-related prognostic signature for GC,which could help develop treatment strategies for patients with GC in the future.

Killing effect of TNF-related apoptosis inducing ligand regulated by tetracycline on gastric cancer cell line NCI-N87

AIM To clone the cDNA fragment of human TRAIL (TNFrelated apoptosis inducing ligand) into a tetracyclineregulated gene expression system, the RevTet-On system, transduce expression vectors into a gastric carcinoma cell line-NCl-N87 and examine the effects of controlled expression of TRAIL in vitro on the gastric carcinoma cells.METHODS The full-length cDNA of TRAIL was inserted into a vector under the control of the tetracyclineresponsive element (TRE) to obtain the plasmid pRevTRETRAIL, which was transfected into a packaging cell line PT67. In addition, vector pRev-Tet- On and pRevTRE were also transfected into PT67 separately. After hygromycin and G418 selection, the viral titer was determined. The medium containing retroviral vectors was collected and used to transduce a gastric carcinoma cell line NCI-N87.The resulting cell line NCI-N87-Tet-On-TRE-TRAIL and a control cell line, NCI-N87-Tet-On-TRE, were established.TRAIL expression in the cell line was induced by incubating cells with doxycycline (Dox), which is a tetracycline analogue. The killing effect on gastric carcinoma cells was analyzed after induction.RESULTS The recombinant plasmid pRev-TRE-TRAIL was constructed. After hygromycin or G418 selection, the producer cell lines PT67-TRE, PT67-TRE-TRAIL and PT67TetOn were obtained, with titers of about 108CFU@ L-1 By transducing NCI-N87 cells with retroviral vectors from these cell lines, stable cell lines NCI-N87-Tet-On-TRETRAIL (NN3T) and control cell line NCI-N87-Tet-On-TRE (NN2T) were established. The growth curves of the selected cell lines were the same with the wild type NCIN87. When Dox was added, cell death was obvious in the test groups (29% -77%), whereas no difference was observed in control and wild type cell lines. With the addition of a medium from the test group, human leukemia cell line Jurkat was activated till death (83%), indicating the secretion of active TRAIL proteins from the test cells to the medium.CONCLUSION With the use of the RevTet-On system, a regulated expression system for TRAIL was constructed.Using this system, the selected killing effect of TRAIL on gastric carcinoma cell line NCI-N87 could be observed.

信迪利单抗联合白蛋白紫杉醇对晚期胃癌血清肿瘤标志物水平及免疫功能的影响

目的探究信迪利单抗联合白蛋白紫杉醇对晚期胃癌血清肿瘤标志物水平及免疫功能的影响。方法前瞻性选取2021年6月至2023年2月在长治医学院附属和平医院接受治疗的89例晚期胃癌患者为研究对象,按照随机数字表法分为对照组(n=44)和观察组(n=45)。对照组患者采取白蛋白紫杉醇治疗,观察组患者采取信迪利单抗联合白蛋白紫杉醇进行治疗。比较两组治疗效果、肿瘤标志物[糖类抗原(CA)125、CA19-9、癌胚抗原、甲胎蛋白及组织多肽特异性抗原(TPS)]水平、免疫功能指标(CD3^(+)、CD4^(+)、CD8^(+)、CD4^(+)/CD8^(+))水平,记录患者治疗期间发生的不良反应。结果治疗后,观察组患者的客观缓解率及疾病控制率分别为55.56%、77.78%,均高于对照组(34.09%、47.73%),差异均有统计学意义(P<0.05)。治疗后,两组患者的各肿瘤标志物水平均较治疗前降低,且观察组CA125、CA19-9、癌胚抗原、甲胎蛋白、TPS水平分别为(16.77±2.77)U/mL、(15.33±3.44)U/mL、(1.22±0.21)U/mL、(14.13±2.87)μg/L、(22.78±4.26)U/L,均低于对照组[(22.31±3.33)U/mL、(24.65±4.32)U/mL、(3.88±0.66)U/mL、(23.87±3.45)μg/L、(32.15±5.67)U/L],差异均有统计学意义(P<0.05)。治疗后,两组CD3^(+)、CD4^(+)水平及CD4^(+)/CD8^(+)值均较治疗前升高,CD8^(+)水平较治疗前降低,且观察组CD3^(+)、CD4^(+)水平及CD4^(+)/CD8^(+)值分别为(69.55±4.15)%、(42.26±4.54)%、2.01±0.15,均高于对照组[(60.34±3.44)%、(32.48±3.56)%、1.48±0.41],CD8^(+)水平为(21.06±2.09)%,低于对照组[(22.45±2.12)%],差异均有统计学意义(P<0.05)。两组不良反应发生方面比较,差异无统计学意义(P>0.05)。结论对晚期胃癌患者采用信迪利单抗联合白蛋白紫杉醇治疗,治疗效果显著,患者血清肿瘤标志物水平得到显著降低,免疫功能改善显著,且具有较好的安全性。

Hyperbaric oxygen treatment on keloid tumor immune gene expression

Background:Hyperbaric oxygen treatment(HBOT)has been demonstrated to influence the keloid recurrence rate after surgery and to relieve keloid symptoms and other pathological processes in keloids.To explore the mechanism of the effect of HBOT on keloids,tumor immune gene expression and immune cell infiltration were studied in this work.Methods:From February 2021 to April 2021,HBOT was carried out on keloid patients four times before surgery.Keloid tissue samples were collected and divided into an HBOT group(keloid with HBOT before surgery[HK]group,n=6)and a non-HBOT group(K group,n=6).Tumor gene expression was analyzed with an Oncomine Immune Response Research Assay kit.Data were mined with R package.The differentially expressed genes between the groups were compared.Hub genes between the groups were determined and verified with Quantitative Real-time PCR.Immune cell infiltration was analyzed based on CIBERSORT deconvolution algorithm analysis of gene expression and verified with immunohistochemistry(IHC).Results:Inflammatory cell infiltration was reduced in the HK group.There were 178 upregulated genes and 217 downregulated genes.Ten hub genes were identified,including Integrin Subunit Alpha M(ITGAM),interleukin(IL)-4,IL-6,IL-2,Protein Tyrosine Phosphatase Receptor Type C(PTPRC),CD86,transforming growth factor(TGF),CD80,CTLA4,and IL-10.CD80,ITGAM,IL-4,and PTPRC with significantly downregulated expression were identified.IL-10 and IL-2 were upregulated in the HK group but without a significant difference.Infiltration differences of CD8 lymphocyte T cells,CD4 lymphocyte T-activated memory cells,and dendritic resting cells were identified with gene CIBERSORT deconvolution algorithm analysis.Infiltration levels of CD4 lymphocyte T cell in the HK group were significantly higher than those of the K group in IHC verification.Conclusion:HBOT affected tumor gene expression and immune cell infiltration in keloids.CD4 lymphocyte T cell,especially activated memory CD4+T,might be the key regulatory immune cell,and its related gene expression needs further study.

胃腺癌进展及预后关键基因与免疫浸润分析

目的基于肿瘤研究公共数据库TCGA和GTEx的样本RNASeq数据信息鉴定在胃腺癌中异常表达且与预后相关的关键基因,并探讨其与免疫细胞浸润的相关性,旨在寻找新的诊治靶点。方法应用GEPIA 2.0可视化肿瘤数据分析平台,对公共数据库中胃腺癌及癌旁组织的差异表达基因和预后相关基因进行分析,并筛选出胃腺癌中差异表达并与预后相关的基因,使用FunRich软件进行GO和KEGG富集分析,通过TIMER 2.0和TISIDB分析关键基因表达与免疫细胞浸润的相关性。结果共筛选到4644个差异表达基因,包括3746个上调基因和898个下调基因。与胃腺癌总生存期(OS)和无病生存期(DFS)相关的前500个基因中GFRA3、APBB1、ABCA8、PLCXD3、FAM153B、CLRN3、CD300LG及ASF1B 8个基因差异表达。经过进一步验证确定ABCA8、PLCXD3、CLRN3、CD300LG及ASF1B 5个关键基因,ABCA8、PLCXD3和CD300LG在胃腺癌组织低表达,CLRN3和ASF1B在胃腺癌组织高表达(P<0.01);在胃腺癌中ABCA8、PLCXD3和CD300LG低表达患者OS和DFS较长,CLRN3和ASF1B高表达患者OS和DFS较长(P<0.05)。相关性分析发现以上5个关键基因表达与免疫细胞浸润水平、丰度具有相关性(P<0.01),且预后风险模型单因素Cox回归分析显示以上5个关键基因是胃腺癌预后的相关风险因素(P<0.05,P<0.01)。结论ABCA8、PLCXD3、CLRN3、CD300LG和ASF1B 5个基因在胃腺癌中异常表达,且与预后相关;其表达与免疫细胞浸润具有相关性,可作为预后风险模型的相关风险因素。

胃癌肿瘤突变负荷相关差异基因的研究

目的:筛选影响胃癌肿瘤突变负荷(TMB)高低的关键差异基因,为胃癌的诊治提供新思路。方法:本研究数据均下载自癌症和肿瘤基因图谱(TCGA)数据库,并通过R软件进行数据分析。我们对高、低TMB组进行差异基因分析、基因本体(GO)富集分析、京都基因与基因组百科全书(KEGG)信号转导途径分析和蛋白质-蛋白质相互作用(PPI)网络构建。结果:本研究在高低负荷差异基因分析总共发现491个差异基因。GO富集分析主要集中在生物过程(BP)、细胞成分(CC)和分子功能(MF)3个方面。KEGG信号转导通路分析主要包括26个信号转导通路。PPI分析共筛选出了7个最为关键的Hub基因。结论:发现影响胃癌的肿瘤突变负荷高低的最为关键Hub差异基因MYH11、TAC1、ALB、DMD、FGFR1、IGF1和AOX1,其中AOX1为首次发现,这有望为胃癌治疗方式的选择提供新思路。

Caveolin-1在胃癌组织中的表达及临床生物学意义

背景与目的:Caveolin-1作为候选抑癌基因,在多种肿瘤均有异常表达。本研究探讨Caveolin-1在非癌胃粘膜、肠上皮化生、异型增生和胃癌组织以及胃癌细胞系MGC803和BGC823的表达特点。方法:采用冰冻组织微阵列的免疫组织化学(immunohistochemistry,IHC)染色,在完全相同的实验条件下检测56例胃癌及29例非癌粘膜、11例肠上皮化生、7例异型增生组织中Caveolin-1的表达状况,及其与胃癌的临床病理分期、淋巴结转移、Lauren分型及组织学分型的关系。Westernblot和RT-PCR法检测胃癌及相应癌旁组织与MGC-803和BGC-823细胞中Caveolin-1蛋白和RNA的表达情况。结果:免疫组化结果显示,Caveolin-1在非癌胃粘膜、肠上皮化生、异型增生和胃癌中的阳性率分别为86.2%(25/29)、81.8%(9/11)、57.1%(4/7)、17.9%(10/56);胃癌与其它各组间的阳性率有显著性差异(P<0.05)。进展期胃癌的Caveolin-1阳性率(16.0%)低于早期胃癌(33.3%),但无统计学意义(P>0.05)。弥漫型胃癌的阳性率(7.0%)明显低于肠型胃癌(26.9%,P<0.05)。有淋巴结转移的胃癌Caveolin-1阳性率(9.7%)低于无淋巴结转移(31.8%,P<0.05)。Westernblot及RT-PCR结果显示,胃癌组织和相应非癌胃粘膜组织中Caveolin-1的表达无显著性差异,但MGC-803和BGC-823细胞中Caveolin-1表达?

胃癌及癌前病变中抑癌基因p16表达的意义

目的探讨p16基因在胃癌发生、发展中的重要作用以及在胃癌早期诊断中的意义。方法取胃粘膜由正常经癌前病变到癌不同阶段的手术切除标本,运用原位分子杂交技术检测其中p16基因mRNA的表达。结果 p16基因mRNA在各型胃粘膜中阳性率分别为:正常胃粘膜94.1％,轻、中、重度异型增生分别为93.9％,80.0％,31.3％;小肠型肠化为69.5％,结肠型肠化为21.4％;胃癌为25.0％;其中正常胃粘膜,轻、中度异型增生,小肠型肠化间差异无显著性(P>0.05);结肠型肠化,重度异型增生、胃癌间差异无显著性(P>0.05),而两组间的差异则有显著性(P<0.01)。结论 P16基因的变异在胃癌的发生、发展中起重要作用;重度异型增生、结肠型肠化是一种重要的癌前病变;对异型增生和肠化胃粘膜进行p16基因mRNA检测有助于胃癌的早期诊断和治疗。

mB7-1转染大鼠卵巢癌细胞诱导细胞免疫应答及体内致瘤性研究

为探讨转染mB7 1基因大鼠卵巢癌细胞系后体外诱导细胞免疫应答及其致瘤性 ,我们以逆转录病毒为载体将mB7 1基因转染入大鼠低分化卵巢上皮癌细胞株NuTu 19中 ,流式细胞仪检测mB7 1的表达。用同源淋巴细胞肿瘤细胞混合培养实验 (MTLCs)测定淋巴细胞增殖指数 ,MTT法检测细胞毒淋巴细胞的增殖及其杀瘤活性。将NuTu 19/Neu、NuTu 19/mB7 1细胞分别接种于同源大鼠Fischer344皮下 ,观察mB7 1修饰的肿瘤细胞在动物体内的致瘤性。结果显示 ,mB7 1基因成功地转染入大鼠低分化卵巢上皮癌细胞株NuTu 19,流式细胞仪测定mB7 1基因呈阳性表达。NuTu 19/mB7 1体外刺激脾淋巴细胞增殖能力明显高于对照组细胞 (P <0 0 5 )。NuTu 19/mB7 1体外诱导的CTL较对照组对NuTu 19细胞的杀伤率显著增高 (P<0 0 1)。两种细胞体外增殖能力无明显改变 ,NuTu 19/mB7 1组在Fischer344大鼠体内成瘤时间明显晚于对照组 (P <0 0 5 )。由此可见 ,mB7 1基因修饰的鼠卵巢癌细胞可体外诱导细胞免疫应答 ,接种动物后 。

树突状细胞诱导抗胃癌移植瘤免疫

目的 研究树突状细胞 (DC)体外诱导的抗肿瘤免疫能否抑制裸鼠移植瘤生长并预防其发生。方法 联合应用粒 /巨噬细胞集落刺激因子 (GM -CSF)及白介素 4(IL 4)直接从胃癌患者外周血中培养出DC ;以SGC -790 1细胞的肿瘤抗原粗提物刺激DC使其激活同源的T淋巴细胞产生细胞毒性T淋巴细胞 (CTL) ;建立裸鼠SGC -790 1细胞移植瘤模型 ;DC诱导的CTL治疗裸鼠SGC -790 1细胞移植瘤 ,观察移植瘤生长 ;DC诱导的CTL预防性治疗裸鼠 ,观察随后接种的SGC -790 1细胞移植瘤发生情况。结果 DC诱导的CTL不仅能抑制裸鼠移植瘤生长并能预防其发生。结论 经肿瘤抗原激活的DC作为一新概念上的抗肿瘤疫苗有可能在治疗肿瘤及预防其术后复发和转移中发挥重要作用。

p53、Rb、p16抑癌基因在胃粘膜上皮异型增生病变中表达的意义

目的 :研究p53、Rb和p163个抑癌基因在正常胃粘膜→异型增生粘膜→胃癌发展过程中的表达状态及相互关系 方法 :收集胃手术及胃镜活检标本共 60例 ,镜下观察胃粘膜并选取不同部位、不同程度的异型增生腺体病灶共 99处。应用免疫组化检测p53、Rb、p16蛋白在 99个异型增生病灶、35例胃腺癌和 32例正常胃粘膜中的表达情况 应用聚合酶链反应单链构象多态性分析技术 (PCR -SSCP)对 35例胃腺癌进行p16基因点突变检测 结果 :p53阳性率在轻、中、重度异型增生病灶中分别为 7.5%、35.1%、59.1% ,胃癌组为 68.6% ,正常对照组中无表达 Rb蛋白阳性率在轻、中、重度异型增生病灶中分别为 80 .0 %、89.3%、81.8% ,胃癌组为 57.1% ,正常对照组为90 6% .p16蛋白在正常胃粘膜、异型增生粘膜和胃癌中普遍表达 ,3组间阳性率比较无显著性差异 p16基因PCR -SSCP分析示只有 1例低分化腺癌出现异常单链泳动带 不同类型 (隐窝型、腺瘤型、再生型 )异型增生病灶组间、位于癌旁和良性病变旁的异型增生病灶组间p53、Rb和p16蛋白的阳性率无显著性差异 高 -中分化、低分化、未分化胃癌组间p53、Rb和p16蛋白的阳性率无显著性差异 结论 :突变型p53蛋白的积聚在胃粘膜异型增生阶段已经开始 ,随着异型增生程度的加重逐渐增加 ,重?

抑癌基因MTS1/p16的缺失与胃癌生物学特性的关系

目的:研究MTS1/p16基因在胃癌中失活的发生率及其临床意义。方法:采用PCR方法对35例胃癌标本MTS1/p16基因第2外显子进行扩增研究。结果:胃癌中MTS1/p16基因的缺失率为40.0％,明显高于正常胃粘膜组织。MTS1/p16基因的缺失与胃癌的组织学类型和分化程度无关,但与淋巴结转移以及临床病理分期有密切关系,随着癌转移的发生或疾病向晚期发展,MTS1/p16基因的缺失率明显提高。结论:MTS1/p16基因的缺失是胃癌细胞发生发展中重要的分子事件,可以作为胃癌恶性进展的一个标志因素。对MTS1/p16基因的检测,将可能有助于临床提高胃癌的基因诊断水平,并且为胃癌恶性程度的判断以及患者预后的评估提供有力的参考指标。

胃类癌33例诊治分析

目的探讨胃类癌的诊断和治疗方法。方法对1987至2007年期间收治并经病理证实的33例胃类癌患者的临床资料进行回顾性分析。结果33例患者中Ⅰ型类癌8例,Ⅲ型类癌25例。病变位于胃体10例,胃底贲门18例,胃窦5例。临床症状无特异性。全组共31例行手术治疗,1例行内镜下黏膜切除,1例行肝转移灶放疗。25例患者获得随访,时间5～173个月。总体1,3,5年生存率分别为76.2%、47.1%和25.0%。其中Ⅰ型患者全部存活。Ⅲ型患者中9例死亡,其1,3,5年生存率分别为66.7%、30.8%和18.2%。结论胃镜是早期发现胃类癌的重要手段,免疫组化有助于提高类癌的诊断率。胃类癌的治疗以及预后与准确的分型密切相关。

p16和nm23-H1基因表达与胃癌及淋巴结转移的关系

目的:进一步评估p16和nm23鄄H1基因表达在胃癌中的作用,尤其是与肿瘤淋巴结转移的关系。方法:采用免疫组织化学方法,检测p16和nm23鄄H1基因蛋白在46例原发性胃癌及69枚转移淋巴结中的表达,探索二者表达与胃癌临床病理特征的关系,评估二者作为预测肿瘤转移和临床预后的生物学指标的可能性。结果:p16和nm23鄄H1基因蛋白表达在原发性胃癌中的阳性率分别为17.4%和28.3%,在转移淋巴结中的阳性率分别为15.9%和14.5%。p16表达在伴和不伴淋巴结转移的原发性胃癌中差异显著(P<0.05)。nm23鄄H1在乳头状腺癌中呈明显高表达。p16和nm23鄄H1蛋白表达在原发性胃癌及转移淋巴结中无相关关系。结论:胃癌组织和转移淋巴结中存在p16和nm23鄄H1基因蛋白频繁缺失;p16阳性表达的病人淋巴结转移机会降低;nm23鄄H1基因蛋白表达率在乳头状腺癌中明显高于其他组织学类型;nm23鄄H1表达与胃癌淋巴结转移无关;p16和nm23鄄H1表达调节似无关联,同时检测p16和nm23鄄H1表达不比单纯检测p16表达具有更高预测胃癌淋巴结转移的价值。

胃癌转移的免疫分子和基因研究进展

近年来肿瘤免疫学的研究发展较快。综述了有关胃癌浸润转移的免疫分子和基因的研究现状 ,提出了新的免疫学诊断指标在预测肿瘤转移方面的应用 。

荷胃癌小鼠自杀基因治疗后宿主免疫功能观察

目的研究自杀基因疗法是否能诱导肿瘤宿主产生免疫应答。 方法将小鼠前胃癌细胞株 (MFC)接种于小鼠背部制作肿瘤模型 ,以单纯疱疹病毒胸苷激酶基因 (HSV -tk)和大肠杆菌胞嘧啶脱氧基酶基因 (CD)注射于瘤体内 ,并于瘤体周围注射mIL - 2和mGM -CSFDNA及向腹腔内注射前药 9-丙氧乌苷 (GVC)和 5 -Fc。治疗第5周取肿瘤消退小鼠的脾细胞测自然杀伤细胞 (NK)和LAK活性 ;第 12周取PBL和脾细胞测NK和细胞毒细胞(CTL)活性。 结果在TK/CD基因联合细胞因子及前药治疗后第 5周 ,实验组脾细胞NK和LAK活性均明显高于荷瘤鼠对照 (P <0 .0 1)。 12周后 ,实验组PBL的NK和CTL活性均显著高于正常对照组 (P =0 .0 5 ,P <0 .0 1) ;而脾细胞NK活性与对照组无显著差异 (P >0 .0 5 )。 结论自杀基因疗法能较长时间提高宿主抗肿瘤免疫反应。

胃癌患者血清VEGF水平与T淋巴细胞嗜银蛋白含量的相关性及临床意义

目的检测胃癌患者血清血管内皮生长因子(VEGF)水平和外周血T淋巴细胞嗜银蛋白(Ag-NORs)含量,探讨二者间的相关性及临床意义。方法采用酶联免疫双抗夹心法(ELISA)定量检测74例胃癌患者和32例健康人血清VEGF水平,同时检测外周血T淋巴细胞嗜银蛋白含量并进行相关分析。结果胃癌患者血清VEGF水平(383.33±121.67)pg/ml明显高于健康人(142.86±71.29)pg/ml(P<0.01),其水平与肿瘤大小、浸润深度、分化程度、有无转移、TNM分期有关(P<0.05或0.01);而胃癌患者外周血T淋巴细胞Ag-NORs含量(4.30±1.21)%明显低于健康人(7.65±1.38)%(P<0.01),其含量与胃癌分化程度、有无转移、TNM分期有关(P<0.05或0.01)。胃癌患者血清VEGF水平与外周血T淋巴细胞嗜银蛋白含量呈显著负相关(r=-0.659,P<0.01)。结论胃癌患者血清VEGF水平和外周血T淋巴细胞Ag-NORs含量是评估胃癌恶性生物学行为的重要指标。二者密切相关,对评价机体肿瘤免疫功能具有重要的临床参考价值。

CD137L基因转染对小鼠体内卵巢癌细胞生长的影响及其免疫调节研究

目的:初步探讨转染CD137L基因的小鼠卵巢癌细胞株OVHM细胞体内的致瘤性及其对免疫功能的影响。方法:以脂质体为载体将重组质粒pcDNA3.1和pcD-NA3.1/CD137LL分别导入OVHM细胞中,经HygroB筛选,用RT-PCR法检测目的基因的表达,筛选建立高表达的细胞株。计数法绘制体外培养细胞生长曲线。将转染前后的肿瘤细胞,分别接种于B6C3F1小鼠的腹部皮下观察其致瘤性。LDH法测定其CTL细胞的杀伤活性。结果:与野生型OVHM细胞相比,所建立的OVHM/CD137L细胞株可高表达CD137L。转染前后肿瘤细胞的体外生长速度无明显变化。OVHM/CD137L细胞在小鼠体内的致瘤性较OVHM/pcDNA3.1细胞和野生型OVHM细胞明显降低。接种OVHM/CD137L细胞小鼠的CTL细胞杀伤活性明显增强。结论:转染CD137L基因后OVHM细胞在小鼠体内的致瘤性显著降低,且诱导产生了明显的抗肿瘤免疫。CD137L有望成为卵巢癌基因治疗的候选基因。

诱导性表达人巨细胞病毒UL138蛋白的胃癌细胞株构建及其效应评价

目的:构建可诱导表达人巨细胞病毒(HCMV)UL138蛋白的稳定遗传胃癌细胞系,并评价UL138对胃癌细胞的生物学效应。方法:将p CMV-tet3G质粒转染BGC-823胃癌细胞,通过G418及荧光素酶活性检测筛选BGCTet-on细胞株。再将含有UL138外源基因的p TRE3G-UL138质粒转染稳定BGCTet-on细胞株,通过G418及潮霉素双抗筛选,获得诱导性表达UL138基因的胃癌细胞(BGC-UL138_(Tet-on))。强力霉素(DOX)诱导后,采用Western blot验证UL138蛋白表达,采用流式细胞术及CCK8试剂盒检测UL138蛋白表达对胃癌细胞的生长周期及增殖的影响。构建荷瘤裸鼠模型,体内验证UL138蛋白表达对胃癌细胞增殖的影响。结果:成功构建了DOX诱导性表达的、携带UL138基因的BGC-UL138_(Tet-on)稳转细胞株。UL138蛋白表达可显著抑制胃癌细胞增殖;流式细胞术检测显示UL138蛋白表达阻滞BGC-823细胞周期在G_1期。荷瘤裸鼠模型体内实验发现,DOX腹腔注射诱导UL138蛋白表达后,BGC-UL138_(Tet-on)移植瘤体积缩小甚至消失。结论:成功构建可诱导表达UL138蛋白的胃癌细胞株,进一步证实HCMV基因UL138的表达可在体内和体外抑制胃癌细胞的增殖,细胞实验提示细胞周期阻断在G_1期。