LATS1 Promotes B-ALL Tumorigenesis by Regulating YAP1 Phosphorylation and Subcellular Localization

Objective YAP1 plays a dual role as an oncogene and tumor suppressor gene in several tumors;differentiating between these roles may depend on the YAP1 phosphorylation pattern.The specific function of YAP1 in B cell acute lymphoblastic leukemia(B-ALL),however,is currently unclear.Thus,in the present study,the role of YAP1 in B-ALL was investigated using relevant cell lines and patient datasets.Methods The effects of shRNA-mediated knockdown on YAP1 and LATS1 levels in the NALM6 and MOLT-4 cell lines were examined using Western blotting,quantitative real-time polymerase chain reaction,flow cytometry,immunostaining,and nude mouse subcutaneous tumorigenesis experiments.Gene expression levels of Hippo pathway-related molecules before and after verteporfin(VP)treatment were compared using RNA-Seq to identify significant Hippo pathway-related genes in NALM6 cells.Results Patients with ALL showing high YAP1 expression and low YAP1-Ser127 phosphorylation levels had worse prognoses than those with low YAP1 protein expression and high YAP1-Ser127 phosphorylation levels.YAP1-Ser127 phosphorylation levels were lower in NALM6 cells than in MOLT-4 and control cells;YAP1 was distributed in the nuclei in NALM6 cells.Knockdown of YAP1 inhibited MOLT-4 and NALM6 cell proliferation and arrested the NALM6 cell cycle in the G0/G1 phase.Before and after VP treatment,the expression of the upstream gene LATS1 was upregulated;its overexpression promoted YAP1-Ser127 phosphorylation.Further,YAP1 was distributed in the plasma.Conclusion LATS1 may downregulate YAP1-Ser127 phosphorylation and maintain B-ALL cell function;thus,VP,which targets this axis,may serve as a new therapeutic method for improving the outcomes for B-ALL patients.

AU-rich element-binding proteins in colorectal cancer

Trans-acting factors controlling mRNA fate are critical for the post-transcriptional regulation of inflammation-related genes, as well as for oncogene and tumor suppressor expression in human cancers. Among them, a group of RNA-binding proteins called "Adenylate-Uridylate-rich elements binding proteins"(AUBPs)control mRNA stability or translation through their binding to AU-rich elements enriched in the 3'UTRs of inflammation-and cancer-associated mRNA transcripts. AUBPs play a central role in the recruitment of target mRNAs into small cytoplasmic foci called Processing-bodies and stress granules(also known as P-body/SG). Alterations in the expression and activities of AUBPs and Pbody/SG assembly have been observed to occur with colorectal cancer(CRC)progression, indicating the significant role AUBP-dependent post-transcriptional regulation plays in controlling gene expression during CRC tumorigenesis.Accordingly, these alterations contribute to the pathological expression of many early-response genes involved in prostaglandin biosynthesis and inflammation,along with key oncogenic pathways. In this review, we summarize the current role of these proteins in CRC development. CRC remains a major cause of cancer mortality worldwide and, therefore, targeting these AUBPs to restore efficient post-transcriptional regulation of gene expression may represent an appealing therapeutic strategy.

Aberrant expression of genes and proteins in pterygium and their implications in the pathogenesis

Pterygium is a common ocular surface disease induced by a variety of factors. The exact pathogenesis of pterygium remains unclear. Numbers of genes and proteins are discovered in pterygium and they function differently in the occurrence and development of this disease. We searched the Web of Science and PubMed throughout history for literatures about the subject. The keywords we used contain pterygium, gene, protein, angiogenesis, fibrosis, proliferation, inflammation, pathogenesis and therapy. In this review, we summarize the aberrant expression of a range of genes and proteins in pterygium compared with normal conjunctiva or cornea, including growth factors, matrix metalloproteinases and tissue inhibitors of mefalloproteinases, interleukins, tumor suppressor genes, proliferation related proteins, apoptosis related proteins, cell adhesion molecules, extracellular matrix proteins, heat shock proteins and tight junction proteins. We illustrate their possible mechanisms in the pathogenesis of pterygium as well as the related intervention based on them for pterygium therapy.

Alteration of tumor suppressor gene p16 and Rb in gastric cancinogesis

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Science Letters:IGFBP7 plays a potential tumor suppressor role against colorectal carcinogenesis with its expression associated with DNA hypomethylation of exon 1

Insulin-like growth factor binding-protein-7 (IGFBP7) was obtained from our previous colonic adenocarcinoma (CRC) and normal mucosa suppression subtraction hybridization (SSH) cDNA libraries. By RT-PCR and immunohistochemistry, we found that IGFBP7 was overexpressed in CRC tissue compared to normal tissue. However, our in vitro experiments performed in 10 CRC cell lines showed that IGFBP7 expressed only in SW480 and Caco2 cell lines, which implied an underlying reversible regulatory mechanism. Using methylation-specific PCR (MSP) and bisulfite sodium PCR (BSP), we found that its expression was associated with DNA hypomethylation of exon1. This was further supported by the in vitro study which showed restored IGFBP7 expression after demethylation agent 5-aza-2’-deoxycytidine treatment. Correlation analysis between IGFBP7 expression and prognosis indicated that overexpression of IGFBP7 in CRC tissue correlated with favourable survival. Investigation of the func-tional role of IGFBP7 through transfection studies showed that IGFBP7 protein could inhibit growth rate, decrease colony for-mation activity, and induce apoptosis in RKO and SW620 cells, suggesting it a potential tumor suppressor protein in colorectal carcinogenesis. In conclusion, our study clearly demonstrated that IGFBP7 plays a potential tumor suppressor role against colo-rectal carcinogenesis and its expression is associated with DNA hypomethylation of exon 1.

Function of apoptosis and expression of the proteins Bcl-2,p53 and C-myc in the development of gastric cancer

INTRODUCTIONIn China ,the incidence and mortality of gastric cancer rank the second among all cancers. Recent development of cancer [1-20].The aim of this study was investigat the insight of apoptosis and bcl-2, p53 and C-myc protein expression in the development of gastric cancer .

Altered expression of nuclear matrix proteins in etoposide induced apoptosis in HL-60 cells

The events of cell death and the expression of nuclear matrix protein (NMP) have been investigated in a promyelocytic leukemic cell line HL-60 induced with etoposide. By means of TUNEL assay, the nuclei displayed a characteristic morphology change, and the amount of apoptotic cells increased early and reached maximun about 39% after treatment with etoposide for 2 h. Nucleosomal DNA fragmentation was observed after treatment for 4 h. The morphological change of HL-60 cells, thus, occurred earlier than the appearance of DNA ladder. Total nuclear matrix proteins were analyzed by 2-dimensional gel electrophoresis. Differential expression of 59 nuclear matrix proteins was found in 4 h etoposide treated cells. Western blotting was then performed on three nuclear matrix acssociated proteins, PML, HSC70 and NuMA. The expression of the suppressor PML protein and heat shock protein HSC70 were significantly upregulated after etoposide treatment, while NuMA, a nuclear mitotic apparatus protein, was down regulated. These results demonstrate that significant biochemical alterations in nuclear matrix proteins take place during the apoptotic process.

Prokaryotic expression, purification of a novel candidate tumor suppressor gene FUS1 and characterization of its polyclonal antibodies

FUS1 is a novel candidate tumor suppressor gene identified in human chromosome 3p21.3. Its expression showed significantly reduction or even loss in lung cancer and other types of cancers. In order to further investigate the biological function of FUS1 protein, FUS1 cDNA from MRC-5 cells was amplified by RT-PCR and cloned into prokaryotic expression vector pQE-30. The recombinant expression plasmids were transformed into M15 strain and grown at 20℃ or 37℃. SDS–PAGE analysis revealed that the accumulation of the recombinant protein FUS1 (rFUS1) in inclusion body forms reached maxium amount when induced with 0.5 mM IPTG for 5 h at 37℃. The inclusion bodies were solubilized in 2M urea and purified by a 6 ×His tagged affinity column under denaturing condition. The purified rFUS1 was identified by electrospray ionization-mass spectrometry (ESI-MS) and tested for purity by HPLC chromatography. The purified rFUS1 proteins were then used to immunize rabbits to obtain anti-human FUS1 polyclonal antibodies, which were suitable to detect both the recombinant exogenous FUS1 and the endogenous FUS1 from tissues and cells by western blot and immunohistochemistry, Available purified rFUS1 proteins and self-prepared polyclonal antibodies against FUS1 may provide effective tools for further studies on biological function and application of FUS1.

虎杖苷调节Akt/MDM2/p53信号通路对胆囊癌细胞增殖、迁移和细胞周期的影响

目的探讨虎杖苷(PD)调节蛋白激酶B/原癌基因MDM2/抑癌基因p53信号通路对胆囊癌细胞增殖、迁移和细胞周期的影响。方法以人胆囊癌细胞株(GBC-SD)为研究对象,体外培养人胆囊癌细胞株(GBC-SD),使用浓度为10~160 mmol/L的虎杖苷处理细胞24、48、72 h,采用CCK-8法检测细胞的增殖能力,确定最佳实验浓度。将GBC-SD细胞分为对照组(Control组)、虎杖苷低、中、高浓度组(PD-L组、PD-M组、PD-H组)、虎杖苷+Akt激活剂组(PD+SC79组),Transwell小室法评价细胞的迁移能力,Hoechst染色观察细胞的凋亡,流式细胞术检测细胞周期与细胞凋亡,Western blot检测Akt、MDM2、p53磷酸化水平,建立荷瘤小鼠模型评价虎杖苷对胆囊癌肿瘤生长的影响。结果浓度为10~160 mmol/L的虎杖苷处理细胞24 h,可显著抑制GBC-SD细胞的增殖活性,选择10、20、40 mmol/L的虎杖苷进行后续实验;与Control组比较,PD-L组、PD-M组、PD-H组GBC-SD细胞的迁移数、细胞凋亡率、G2/M期细胞比例及S期细胞比例、P-Akt、P-MDM2蛋白表达显著降低,G0/G1期细胞比例、P-p53蛋白表达显著升高,且呈浓度依赖性(P<0.05);与PD-H组比较,PD+SC79组GBC-SD细胞的迁移数、细胞凋亡率、G2/M期细胞比例及S期细胞比例、P-Akt、P-MDM2蛋白表达显著升高,G0/G1期细胞比例、P-p53蛋白表达显著降低(P<0.05);虎杖苷干预治疗后,小鼠移植瘤的生长速度显著降低(P<0.05)。结论虎杖苷可以通过调节Akt/MDM2/p53信号通路使细胞周期阻滞,抑制胆囊癌细胞增殖、迁移。

CTNNB1、TP53蛋白在儿童肝母细胞瘤组织中的表达及与病理特征、预后的关系

目的 探讨钙黏蛋白相关蛋白(cadherin associated protein beta 1,CTNNB1)、肿瘤抑制因子P53(tumor suppressor factor P53,TP53)蛋白在儿童肝母细胞瘤(hepatoblastoma, HB)组织中的表达及与病理特征、预后的关系。方法 选取2017-06/2020-06月作者医院收治的72例HB患儿为研究对象,手术留取癌组织标本及对应的癌旁组织。连续随访3年,记录患儿的预后生存情况,并计算总生存率(overall survival, OS)。采用免疫组织化学法检测CTNNB1、TP53蛋白在HB患儿癌组织及癌旁组织中的表达情况;采用χ~2检验分析CTNNB1、TP53蛋白表达与HB患儿临床病理特征的关系;采用多因素Cox回归分析探讨HB患儿预后的影响因素。结果 HB患儿癌组织中CTNNB1阳性表达率(76.39%)高于对照组(43.06%),TP53阴性表达率(70.83%)高于对照组(38.89%)(P均<0.05)。初诊甲胎蛋白浓度≥100μg/L、POST-TEXT分期Ⅲ~Ⅳ期、肿瘤直径≥3 cm、有肿瘤侵袭或转移HB患儿的CTNNB1阳性表达率、TP53阴性表达率高于初诊甲胎蛋白<100μg/L、POST-TEXT分期Ⅰ~Ⅱ期、肿瘤直径<3 cm、无肿瘤侵袭或转移HB患儿(P均<0.05)。初诊甲胎蛋白≥100μg/L、POST-TEXT分期Ⅲ~Ⅳ期、有肿瘤侵袭或转移、CTNNB1阳性表达、TP53阴性表达的HB患儿3年OS低于初诊甲胎蛋白<100μg/L、POST-TEXT分期Ⅰ~Ⅱ期、无肿瘤侵袭或转移、CTNNB1阴性表达、TP53阳性表达的HB患儿(P均<0.05)。多因素Cox回归分析结果显示,POST-TEXT分期Ⅲ~Ⅳ期(HR=2.077,95%CI:1.423~3.032)、有肿瘤侵袭或转移(HR=2.291,95%CI:1.536~3.417)、CTNNB1阳性表达(HR=2.757,95%CI:1.781~4.268)、TP53阴性表达(HR=2.477,95%CI:1.635~3.753)是HB患儿预后的独立危险因素(P<0.05)。结论 HB患儿癌组织中CTNNB1主要呈阳性表达,而TP53主要呈阴性表达,二者与初诊甲胎蛋白、POST-TEXT分期、肿瘤直径、有肿瘤侵袭或转移密切相关,有望作为评估HB患儿预后的生物学指标。

铁死亡抑制蛋白1在肿瘤中的作用

铁死亡抑制蛋白1(FSP1)是铁死亡过程中的关键抑制因子,能够阻止细胞死亡,具有重要的生物学功能和潜在的临床应用价值。本研究详细探讨了FSP1的发现背景、基因定位与结构特性,以及其在抑制铁死亡和促进细胞凋亡中的双重作用。临床研究已经开发出针对FSP1的抑制剂,如铁死亡抑制蛋白1抑制剂(iFSP1)和相分离诱导型铁死亡抑制蛋白1抑制剂(icFSP1),未来的研究将进一步探讨FSP1的表达调控机制、与肿瘤免疫逃逸的关联以及其作为肿瘤预后和治疗反应监测指标的潜力。

三阴性乳腺癌EGFR、Ki-67、P53及CTC表达与预后的关系研究

目的探讨三阴性乳腺癌表皮生长因子受体(EGFR)、细胞核增殖相关抗原Ki-67(Ki-67)、P53及循环肿瘤细胞(CTC)表达与预后的关系。方法选取95例三阴性乳腺癌患者,免疫组化检测病理组织标本中EGFR、Ki-67、P53表达;所有患者接受8个周期化疗,采用膜滤过分离肿瘤细胞技术(ISET)检测化疗前后的CTC表达,并分析化疗前后CTC表达与化疗疗效的关系;分析CTC与EGFR、Ki-67、P53表达的关联性;随访患者无进展生存期(PFS),采用COX回归分析三阴性乳腺癌进展的危险因素。结果EGFR、Ki-67、P53阳性检出率分别为44.21%(42/95)、63.16%(60/95)、56.84%(54/95);化疗后患者的CTC阳性检出率(14.74%)低于化疗前(61.05%,P<0.05);化疗疗效与化疗后CTC阳性表达呈负相关(P<0.001);COX回归分析显示,临床分期为Ⅲ期、EGFR阳性、化疗后CTC阳性是三阴性乳腺癌进展的独立危险因素(P<0.05);不同临床分期患者PFS比较,Ⅰ期>Ⅱ期>Ⅲ期,EGFR阳性患者PFS短于阴性患者,化疗后CTC阳性患者PFS短于阴性患者(P<0.05)。结论化疗前EGFR阳性表达、化疗后CTC阳性表达与三阴性乳腺癌患者预后差有关,化疗后CTC阳性率越低患者化疗疗效越好。

组蛋白H2A去泛素化酶BAP1对恶性胶质瘤细胞发生发展的作用及临床应用价值研究

目的探索乳腺癌/卵巢癌易感基因1相关蛋白1(breast/ovarian cancer susceptibility gene 1 associated protein 1,BAP1)对人源恶性胶质瘤发生、发展的作用与BAP1作为恶性胶质瘤临床诊断标志物的可行性。方法基于基因表达综合数据库(gene expression omnibus,GEO)的子数据集GSE4290,GSE90598,分析BAP1在正常组织及胶质瘤组织中的差异性表达情况;受试者工作特征(receiver operating characteristic,ROC)曲线分析BAP1对恶性胶质瘤的早期诊断价值;选取自主收集的非配对28例恶性胶质瘤患者的原发灶组织、5例颅脑外伤患者内减压术切除的非瘤脑组织,采用实时荧光定量PCR(quantitative real-time polymerase chain reaction,qRT-PCR)检测BAP1的表达水平;利用靶向BAP1的特异性小干扰RNAs(small interfering RNAs,siRNAs)瞬时转染U251细胞系,进一步检测其干涉效率;基于流式细胞仪分析BAP1下调的U251细胞系,其细胞周期、凋亡的变化情况。结果生物信息学结果显示,BAP1在恶性胶质瘤组织中的表达水平均低于正常脑组织(GSE4290:1209±18.49 vs 1476±53.90;GSE90598:5.19±0.10 vs 5.65±0.21),差异具有统计学意义(t=5.115,2.267,均P<0.05)。ROC曲线显示,BAP1可高效区分恶性胶质瘤组织与正常脑组织(GSE4290:AUC=0.78;GSE90598:AUC=0.75,均P<0.05)。临床标本结果显示,BAP1在恶性胶质瘤原发灶组织中的表达水平显著低于非瘤脑组织(0.27±0.04 vs 1.06±0.07),差异具有统计学意义(t=10.22,P<0.001)。在U251细胞系中下调BAP1的表达,其细胞周期中S期细胞比例明显增多,由17.59%分别增至27.21%(siBAP1-1)和25.79%(siBAP1-2),差异具有统计学意义(t=6.576,6.642,均P<0.01),而细胞凋亡水平则有所下降,由10.17%分别降至2.70%(siBAP-1)和3.00%(siBAP-2),差异具有统计学意义(t=10.31,9.428,均P<0.01)。结论组蛋白H2A去泛素化酶BAP1能够通过抑制恶性胶质瘤细胞周期快速进展并促进其凋亡,进而发挥肿瘤抑癌基因的功能,可作为潜在的恶性胶质瘤临床诊断标志物。

Hp、RASAL2、CDH1及TP53对胃癌前病变与早期胃癌的鉴别诊断价值

目的探讨幽门螺杆菌(Hp)、RAS蛋白激活样因子2(RASAL2)、钙黏蛋白1(CDH1)及肿瘤抑制基因P53(TP53)对胃癌前病变与早期胃癌的鉴别诊断价值。方法选取2021年6月至2023年6月收治的早期胃癌52例,根据1:1选例原则另选取同期胃癌前病变52例、胃炎52例,分别纳入胃癌组、癌前组、胃炎组。比较3组及不同病理特征的早期胃癌患者Hp、RASAL2、CDH1、TP53阳性表达率,比较胃癌组Hp阳性、阴性患者RASAL2、CDH1、TP53阳性表达率,采用Spearman相关分析探讨各指标阳性表达与早期胃癌患者部分病理特征的相关性,采用受试者工作特征(ROC)曲线分析联合检测对早期胃癌及胃癌前病变的鉴别诊断价值。结果3组Hp、CDH1、TP53阳性表达率胃癌组>癌前组>胃炎组,RASAL2阳性表达率胃癌组<癌前组<胃炎组,差异有统计学意义(P<0.05,P<0.01)。胃癌组Hp阳性患者CDH1、TP53阳性表达率高于Hp阴性患者,RASAL2阳性表达率低于Hp阴性患者(P<0.05,P<0.01);早期胃癌患者Hp、RASAL2、CDH1、TP53阳性表达率在肿瘤浸润深度及淋巴结转移方面比较差异有统计学意义(P<0.05,P<0.01)。Hp、CDH1、TP53阳性表达与早期胃癌肿瘤浸润深度、淋巴结转移均呈正相关,RASAL2阳性表达与之呈负相关(P<0.01)。ROC曲线分析结果显示,Hp、RASAL2、CDH1、TP53联合诊断胃癌前病变的曲线下面积(AUC)为0.904(95%CI:0.846,0.945),联合诊断早期胃癌的AUC为0.894(95%CI:0.819,0.946)。结论Hp、CDH1、TP53在早期胃癌组织中阳性表达率较高,RASAL2阳性表达率较低,联合检测对胃癌前病变及早期胃癌具有一定鉴别诊断价值,可作为临床鉴别诊断胃癌前病变、早期胃癌的辅助指标。

组织中P53、人表皮生长因子受体2表达与乳腺癌患者临床病理特征的关系

目的:探讨组织中P53、人表皮生长因子受体2(HER2)表达与乳腺癌患者临床病理特征的关系.方法:选取2019年2月至2023年2月于本院采取手术治疗的60例乳腺癌患者作为研究对象.检测其组织中P53、HER2表达,分析其阳性表达与乳腺癌患者不同临床病理特征的相关性.结果:60例乳腺癌患者肿瘤组织中,P53、HER2阳性率分别为53.33%(32/60)和45%(27/60).Ⅲ期、组织低分化、发生淋巴结转移乳腺癌患者的P53、HER2阳性率,显著高于Ⅰ/Ⅱ期、组织中高分化、未发生淋巴结转移者,差异均有统计学意义(P<0.05).乳腺癌患者肿瘤组织中P53、HER2阳性表达,与肿瘤分期、淋巴结转移情况呈正相关关系(r>0,P<0.05),与组织分化程度呈负相关关系(r<0,P<0.05).结论:乳腺癌患者肿瘤组织中P53、HER2多呈阳性表达.肿瘤分期、组织分化程度、淋巴结转移情况,可能与P53、HER2表达有关.

Aberrant methylation of secreted protein acidic and rich in cysteine gene and its significance in gastric cancer

BACKGROUND Aberrant methylation in DNA regulatory regions could downregulate tumor suppressor genes without changing the sequences.However,our knowledge of secreted protein acidic and rich in cysteine(SPARC)and its aberrant methylation in gastric cancer(GC)is still inadequate.In the present research,we performed fundamental research to clarify the precise function of methylation on SPARC and its significance in GC.AIM To investigate promoter methylation and the effects of the SPARC gene in GC cells and tissues and to evaluate its clinical significance.METHODS Plasmids that overexpressed the SPARC gene were transfected into human GC BGC-823 cells;non-transfected cells were used as a control group(NC group).Quantitative real-time polymerase chain reaction and western blotting(WB)were then used to detect the expression of SPARC.Methylation-specific polymerase chain reaction was executed to analyze the gene promoter methylation status.Cell viability was measured by the cell counting kit-8 assay.The migration and invasion ability of cells were detected by scratch assays and transwell chamber assays,respectively.Cell cycle events and apoptosis were observed with a flow cytometer.RESULTS The expression of SPARC mRNA in GC tissues and cells was significantly lower and showed differing degrees of hypermethylation,respectively,than that in normal adjacent tissues and control cells.Treatment with 5-Aza-2’-deoxycytidine(5-Aza-Cdr)was able to restore the expression of SPARC and reverse promoter hypermethylation.Overexpression of the SPARC gene significantly inhibited proliferation,migration,and invasion of GC cells,while also causing cell cycle arrest and apoptosis;the NC group exhibited the opposite effects.CONCLUSION This study demonstrated that SPARC could function as a tumor suppressor and might be silenced by promoter hypermethylation.Furthermore,in GC cells,SPARC inhibited migration,invasion,and proliferation,caused cell cycle arrest at the G0/G1 phase,and promoted apoptosis.

Tumor suppressor p53 cross-talks with TRIM family proteins

p53 is a key tumor suppressor.As a transcription factor,p53 accumulates in cells in response to various stress signals and selectively transcribes its target genes to regulate a wide variety of cellular stress responses to exert its function in tumor suppression.In addition to tumor suppression,p53 is also involved in many other physiological and pathological processes,e.g.anti-infection,immune response,development,reproduction,neurodegeneration and aging.To maintain its proper function,p53 is under tight and delicate regulation through different mechanisms,particularly the posttranslational modifications.The tripartite motif(TRIM)family proteins are a large group of proteins characterized by the RING,B-Box and coiled-coil(RBCC)domains at the N-terminus.TRIM proteins play important roles in regulation of many fundamental biological processes,including cell proliferation and death,DNA repair,transcription,and immune response.Alterations of TRIM proteins have been linked to many diseases including cancer,infectious diseases,developmental disorders,and neurodegenera-tion.Interestingly,recent studies have revealed that many TRIM proteins are involved in the regulation of p53,and at the same time,many TRIM proteins are also regulated by p53.Here,we review the cross-talk between p53 and TRIM proteins,and its impact upon cellular biological processes as well as cancer and other diseases.

Potential effect of hepatitis C Virus non-structural protein 4B on liver carcinogenesis

Objective：To investigate the effect of hepatitis C virus non-structural protein 4B（HCV NS4B） on c-Myc, P53, ras gene expression＂ and apoptosis in hepatic cells and study the possible role that NS4B played in the carcinogenesis of heparoma. Methods： The recombinant plasmid（PCXN2-NS4B, PCXN2-P53） and the empty, vector were transfected or co-transfected into Chang liver cells with liposome. Screening was performed with G418. Plasmid mRNA was detected by RT-PCR. The pro rein expressions of c-Myc and ras genes were analyzed by immunocytochemistry. The expressions of wild-type P53 （wtp53） gene were detected by in situ hybridization. TUNEL（flow cytometry） was used for assessing the rate of apoptosis. Results：No expression of c-Myc gene was found in PCXN2 group. The expression of c-Myc gene in NS4B group was 21.3% ＋ 1.2%. The ex pression of ras gene in PCXN2 group was lower than that in NS4B group. Compared with PCXN2 group, the expression of P53 mRNA was not promoted or inhibited in NS4B group. But the expression of P53 mRNA in NS4B-P53 group was lower than that in P53 group. In PCXN2, NS4B, P53 and NS4B-P53 group, the rates of apoptosis were 17.02% ± 1.24%, 11.94% ± 2.24%, 25.84% ± 3.49% and 18.34% ± 1.55% respectively. Conclusion ：HCV NS4B induces the expression of c-Myc and ras gene. HCV NS4B may play a role in the inhibition of cell death through P53-dependent manner. Results from this study suggested that HCV NS4B might contribute to the viral carcinogenesis.

牛磺酸对胰腺癌细胞系BxPC-3和PANC-1细胞增殖、凋亡和迁移的影响

目的探讨牛磺酸(Tau)对胰腺癌导管细胞增殖、凋亡和迁移的影响。方法体外培养胰腺癌BxPC-3和PANC-1细胞,采用不同浓度Tau(0、10、20、40、80、160 mmol/L)进行预处理。采用CCK-8、细胞划痕实验、TUNEL法观察Tau对BxPC-3和PANC-1细胞增殖、迁移和凋亡的影响;实时荧光定量聚合酶链反应(qPCR)和蛋白质免疫印迹法检测Tau影响BxPC-3和PANC-1细胞中相关细胞凋亡及细胞周期分子mRNA和蛋白表达的情况。结果Tau可抑制BxPC-3和PANC-1的细胞增殖和迁移活性;同时,Tau能够促进BxPC-3和PANC-1细胞凋亡。与对照组相比,Tau处理后的BxPC-3细胞中相关凋亡因子P53、P21、Bcl-2关联X蛋白(BAX)表达水平呈上升趋势,而增殖细胞核抗原(PCNA)的表达降低;Tau处理后的PANC-1细胞中B淋巴细胞瘤-2蛋白(Bcl-2)、PCNA、细胞周期蛋白CyclinA2、CyclinB1、CyclinE、周期蛋白依赖性激酶CDK1、CDK2、CDK4、CDK6等的表达较对照组均下降,而相关凋亡蛋白P53、P21的表达水平升高。结论Tau可抑制胰腺癌细胞BxPC-3和PANC-1的增殖和迁移活性,并促进细胞凋亡,可能是通过影响相关细胞凋亡基因和细胞周期蛋白来抑制胰腺癌细胞的活性。

DOK3基因表达下调对原代结肠癌细胞生物学行为的影响及其可能机制

目的 分析对接蛋白3(DOK3)基因表达下调对原代结肠癌细胞增殖、侵袭/迁移能力的影响,并基于DOK3与G蛋白偶联受体84(GPR84)的相互作用探讨可能的作用机制。方法 培养原代结肠癌细胞HCT16,将其分为对照组、小干扰RNA(siRNA)-NC组、siRNA-DOK3组进行实验。分别将siRNA-NC、siRNA-DOK3转染至siRNA-NC组、siRNA-DOK3组,而仅将培养液加入对照组。培养24 h后通过实时荧光定量PCR评估转染效果,再使用CCK-8法、Transwell实验分别检测各组细胞的增殖情况、侵袭/迁移能力,采用蛋白免疫共沉淀实验检测HCT16细胞中DOK3与GPR84之间的相互作用关系。结果 与对照组比较,siRNA-DOK3组HCT16细胞中DOK3 mRNA表达量降低(P<0.05),而siRNA-NC组HCT16细胞中DOK3mRNA表达量差异无统计学意义(P>0.05)。培养48 h、72 h后,siRNA-DOK3组HCT16细胞的增殖活力较对照组增加(P<0.05)。与对照组相比,siRNA-DOK3组HCT16细胞的迁移和侵袭数量增加(P<0.05),而siRNA-NC组HCT16细胞的迁移和侵袭数量差异无统计学意义(P>0.05)。蛋白免疫共沉淀实验结果显示,在HCT16细胞中GPR84与DOK3相互结合。结论 DOK3基因可能作为抑癌基因,参与了结肠癌的发生与发展,且其可能通过与GPR84结合发挥抑癌作用。