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@#[摘 要] 目的：探讨桥接整合因子1（BIN1）在上皮性卵巢癌（EOC）组织中的表达及其临床意义，以及BIN1对EOC细胞A2780增殖、迁移和侵袭的影响。方法：收集2017年7月至2018年1月河北医科大学第四医院手术切除的67例EOC患者的肿瘤组织及同期因其他妇科疾病手术切除的30例非肿瘤患者的卵巢组织（正常对照组）标本。用免疫组织化学染色法检测EOC组织和非肿瘤卵巢组织中BIN1蛋白的表达水平，χ2检验分析BIN1表达与患者临床病理特征之间的关联，Kaplan-Meier法分析BIN1表达与患者的无病生存期（DFS）和总生存期（OS）之间的关系。用qPCR和WB法检测EOC细胞SKOV3、A2780和人卵巢上皮细胞IOSE80中BIN1 mRNA和蛋白的表达水平。利用基因转染技术将BIN1质粒CMV-MCS-GFP-SV40-Neomycin-BIN1和空载体质粒CMV-MCS-GFP-SV40-Neomycin分别转染到A2780细胞以构建过表达BIN1细胞及其对照，用qPCR和WB法分别检测转染细胞中BIN1 mRNA和蛋白的表达水平，CCK-8、划痕愈合和Transwell实验分别检测过表达BIN1对A2780细胞增殖、迁移和侵袭的影响。结果：EOC组织中BIN1阳性表达率显著低于正常卵巢组织（P<0.01）。BIN1表达与EOC患者较晚的术后病理分期、较差的组织学分级、淋巴结转移及腹膜转移存在正向关联（均P<0.05）；BIN1低表达组患者的DFS和OS均短于BIN1高表达组患者（均P<0.05）。SKOV3和A2780细胞中BIN1 mRNA和蛋白的表达水平均显著低于IOSE80细胞（均P<0.01）；过表达BIN1显著抑制A2780细胞的增殖、迁移和侵袭（P<0.05或P<0.01）。结论：BIN1在EOC组织和细胞中呈低表达状态，与患者的不良预后有关；过表达BIN1可降低EOC细胞A2780的增殖、迁移和侵袭能力。
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@#[摘 要] 目的：探讨PD-1抗体联合化疗对比抗血管生成药物联合化疗在晚期驱动基因阴性肺腺癌一线治疗中的疗效和安全性。方法：收集2018年3月至2021年8月河北医科大学第四医院收治的141例不可手术切除的ⅢB/ⅢC和Ⅳ期驱动基因阴性肺腺癌患者，回顾性分析PD-1抗体联合化疗对比抗血管生成药物联合化疗在一线治疗中的疗效与安全性。主要研究终点为无进展生存期（PFS），次要终点为客观缓解率（ORR）、疾病控制率（DCR）和不良反应。结果：141例患者均纳入生存分析，中位随访时间为13.0个月（95% CI：12.0～14.0）。PD-1抗体联合化疗组（A组）和抗血管生成药物联合化疗组（B组）的ORR分别为33.33%和27.38%，DCR分别为98.25%和89.29%，差异均无统计学意义。A组和B组的中位PFS分别为8.4个月（95% CI: 7.3～9.9）和6.9个月（95% CI: 6.1～7.7），差异无统计学意义。亚组分析结果显示，ⅢB/ⅢC期、肝或脑转移患者中，A组中位PFS较B组均延长（均P<0.01）。A组和B组不良反应发生率分别为26.32%和14.29%，多数为1~2级。结论：PD-1抗体联合化疗对比抗血管生成药物联合化疗一线治疗晚期驱动基因阴性肺腺癌疗效相当，不良反应可耐受，可成为晚期驱动基因阴性肺腺癌标准一线治疗。
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@#[摘 要] 目的：探讨ERBB2.1转导蛋白反义RNA1（transducer of ERBB2.1 antisense RNA 1，TOB1-AS1）在上皮性卵巢癌（epithelial ovarian cancer，EOC）组织中的表达情况及其临床意义，初步探讨TOB1-AS1对EOC细胞体外增殖、迁移和侵袭的影响。方法：使用TCGA数据库对EOC组织中TOB1-AS1表达情况进行分析；收集2017年7月至2018年1月在河北医科大学第四医院妇科行肿瘤切除并经病理检查证实为EOC的67例患者的肿瘤组织，收集同期因其他妇科疾病接受手术的30例患者的非肿瘤卵巢组织作为对照。采用qPCR法检测EOC组织和非肿瘤卵巢组织中TOB1-AS1的表达水平，χ2检验分析TOB1-AS1的表达与不同临床病理特征之间的相关性，Kaplan-Meier和Cox比例风险回归模型分析患者生存及预后的潜在影响因素。CCK-8实验、划痕实验和Transwell实验分别检测敲低TOB1-AS1表达对EOC细胞SKOV3和A2780增殖、迁移和侵袭的影响。结果：TCGA数据库中资料和qPCR检测结果均显示，在EOC组织中TOB1-AS1的表达水平显著高于非肿瘤卵巢组织（均P<0. 01）。TOB1-AS1的高表达与EOC患者较晚的FIGO分期、较差的组织分级、淋巴结转移及腹膜转移有关（均P<0.05）。Kaplan-Meier生存分析结果显示，TOB1-AS1高表达组患者术后DFS和OS均短于TOB1-AS1低表达组（均P<0.05）。Cox比例风险回归模型分析结果显示，FIGO分期、淋巴结转移、腹膜转移及TOB1-AS1表达是EOC患者预后的独立影响因素（均P<0.05）。TOB1-AS1在EOC细胞系SKOV3、A2780中的表达水平也显著高于正常卵巢上皮细胞系IOSE80（均P<0.01）。细胞功能实验结果显示，敲低TOB1-AS1可抑制SKOV3和A2780细胞的增殖、迁移和侵袭（均P<0.05）。结论：TOB1-AS1在EOC组织中高表达，与患者的不良预后显著相关。TOB1-AS1可能通过促进EOC细胞SKOV3、A2780的增殖、迁移和侵袭来影响EOC的恶性进展。
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@#[Abstract] Objective: To investigate the effect of long non-coding RNA DiGeorge syndrome critical region gene 5 (DGCR5) on proliferation, invasion and migration of esophageal squamous cell carcinoma (ESCC) TE1 cells and its mechanism. Methods: qPCR was used to detect the expression level of DGCR5 in ESCC cell lines (TE1, Yes-2, KYSE150 and Eca9706). TE1 cells were transfected with siRNA-DGCR5(si-DGCR5) and negative control (si-NC) plasmids, respectively. CCK-8, Wound healing and Transwell assay were used to detect the proliferation, migration and invasion of TE1 cells before and after DGCR5 knockdown. The relationship between DGCR5 expression and epidermal growth factor receptor (EGFR) in ESCC tissues was analyzed by GEPIA database. The mRNA and protein expressions of EGFR in ESCC cell line were examined by qPCR and Western blotting (WB). WB was further used to detect the expression of EGFR protein in TE1 cells before and after DGCR5 knockdown. Results: lncRNA DGCR5 was highly expressed in ESCC cell lines (all P<0.01). qPCR confirmed that the expression of DGCR5 in TE1 cells of si-DGCR5 group was significantly lower than that of si-NC group (P<0.01). The proliferation, migration and invasion ability of TE1 cells in si-DGCR5 group were significantly lower than those in si-NC group (all P<0.01). GEPIA database showed that the expression of DGCR5 was positively correlated with EGFR in ESCC tissues (P<0.01). WB showed that the protein level of EGFR in TE1 cells of si-DGCR5 group decreased significantly (P<0.01). Conclusion: lncRNA DGCR5 is highly expressed in ESCC cells, and promotes the proliferation, invasion and migration of TE1 cells possibly by up-regulating EGFR expression.
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@#[Abstract] Objective：To study the expression of long non-coding RNA(lncRNA) titin antisense RNA1 (TTN-AS1) in lung adenocarcinoma (LUAD) tissues, and explore its relationship with clinicopathologic characteristics and prognosis of LUAD patients. Methods: The TTN-AS1 expression in LUAD data set was analyzed using TCGAdatabase. 52 pairs of tumor tissues and matched para-carcinoma tissues from LUAD patients, who underwent surgical resection and were later pathologically conformed in Fourth Hospital of Hebei Medical University between Jan. 2014 and Jan. 2015, were used in this study. qPCR was performed to detect TTN-AS1 expression in the specimens. Then, the correlations between TTN-AS1 expression and clinicopathologic characteristics were analyzed. Survival analysis was used to determine the significance of TTN-AS1 expression for predicting the prognosis of LUAD patients. Results: TCGAdatabase analysis and qPCR results showed that TTN-AS1 expression in LUAD tissues was significantly higher than that in normal lung and para-carcinoma tissues (both P<0.01). TTN-AS1 expression in LUAD tissues was significantly correlated with the TNM stage and lymph node metastasis (P<0.05), but not correlated with gender, age, tumor invasion range (P>0.05). Kaplan-Meier univariate analysis result demonstrated that the patients with high TTN-AS1 expression had shorter post-operative disease-free survival (DFS) and overall survival (OS) than those patients with low TTN-AS1 expression (all P<0.01). Cox proportional hazard regression model result demonstrated that wider tumor invasion range, positive lymph node metastasis and high TTN-AS1 expression were significantly correlated with shorter postoperative DFS and OS (P<0.05). Conclusion: TTN-AS1 was highly expressed in LUAD tissues, and closely correlated with TNM stage and lymph node metastasis of LUAD patients (all P<0.05). High expression of TTN-AS1 is significantly correlated with shorter DFS and OS, indicating that TTN-AS1 may be a biomarker for predicting poor prognosis of LUAD patients.
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@#[Abstract] Objective: To investigate the changes in malignant biological behaviors and expression of programmed cell death-ligand 1 (PD-L1) in esophageal squamous cell carcinoma (ESCC) YES-2 cell line after cis-dichlorodiammine platinum (CDDP) induction (YES-2/CDDP-R). Methods: YES-2 cells were treated with CDDP from low concentration to high concentration (0.25-2.0 μg/ml) with intermittent impact (15-25 days per concentration) to establish ESCC CDDP-resistant cell line YES-2/CDDP-R. The morphological change of YES-2/CDDP-R cells was observed under the inverted microscope. Methyl thiazolyl tetrazolium (MTT) was used to detect cell sensitivity to CDDP. Wound healing assay was used to detect cell migration ability. qPCR and Western blotting were used to detect mRNA and protein expressions of PD-L1. Results: After CDDP gradien ttreatment for9 months,YES-2/CDDP-R cells were successfully established. The morphology of the YES-2/CDDP-R cells showed uneven size, intracellular vacuoles and significantly increased black particles along with the appearance of huge cells. The IC50 of CDDP for YES-2/CDDP-R cells was significantly higher than that for parental cells, indicating decreased sensitivity to CDDP (P<0.05). Compared to theYES-2 cells, the proliferation and migration of YES-2/CDDP-R cells were significantly increased (P<0.05 or P<0.01), and the mRNA and protein expressions of PD-L1 were significantly up-regulated (all P<0.001). Conclusion: YES-2 cells with CDDP resistance (YES-2/CDDP-R) were successfully established. The sensitivity of YES-2/CDDP-R cells to CDDP was significantly reduced while the abilities of cell proliferation and migration were enhanced. The up-regulation of PD-L1 in YES-2/CDDP-R cells suggests that CDDP-resistance could promote immune escape by inducing PD-L1 up-regulation.
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@# Objective: To study the expression of miR-142-5p in lung adenocarcinoma tissues, and to explore its effect on proliferation, invasion, migration and epithelieal-mesenchymal transition (EMT) of H1650 cells and the potential mechanisms. Methods:Atotal of 107 pairs of lung adenocarcinoma tissues and corresponding para-cancerous tissues from patients, who underwent tumor resection and were pathologically confirmed at the Department of Thoracic Surgery, the Fourth Hospital of Hebei Medical University between Jan. 2014 and Jan. 2015, were collected for this study; in addition, human lung adenocarcinoma cell lines (H1650, HCC827, A549, H1975, PC9) and human bronchial epithelial BEAS-2B cells were also used in this study. qPCR was used to detect the expression of miR-142-5p in lung adenocarcinoma tissues and cell lines. The correlation between expression of miR-142-5p and clinical features was analyzed.After transfection with miR-142-5p mimics or miR-negative control (miR-NC) plasmid, the proliferation, invasion and migration of H1650 cells were detected with CCK-8, Transwell invasion assay and Wound healing assay, respectively. The bioinforamtics tool was used to predict the target genes of miR-142-5p, and Luciferase reporter gene assay was performed to validate the regulation of miR-142-5p on target gene. Western blotting (WB) was used to detect the expressions of cyclin-dependent kinase 5 (CDK5) and EMTrelated protein. Results: Compared to Para-cancerous tissues and BEAS-2B cells, the expression of miR-142-5p was lower in lung adenocarcinoma tissues and cell lines (all P<0.01). Of the 107 cases of lung adenocarcinoma tissues, 61 cases (57.01%) showed decreased miR-142-5 expression, which was correlated with the TNM stage and lymph node metastasis (both P<0.01). Transfection of miR-142-5p mimics significantly up-regulated the expression of miR-142-5p and decreased the proliferation, invasion and migration of H1650 cells (all P<0.05 or P<0.01). Bioinformatics showed that CDK5 was a target gene of miR-142-5p. Luciferase reporter gene assay and WB validated that miR-142-5p could significantly down-regulate CDK5 expression in H1650 cells, up-regulate the expression of E-cadherin and down-regulate the expressions of N-cadherin, Twist and Snail in H1650 cells (all P<0.01). Conclusion: miR-142-5p is low expressed in lung adenocarcinoma tissues and cell lines; it suppresses the EMT process to inhibit, invasion and migration of H1650 cells via down-regulating the expression of CDK5.
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@# Objective: To investigate the effects of long non-coding RNA nuclear enriched abundant transcript 1 (lncRNA NEAT1) on the proliferation of lung adenocarcinoma PC-9 cells and to explore its mechanism. Methods: qPCR was used to detect the expression level of lncRNA NEAT1 in human lung adenocarcinoma PC-9 cells and human embryonic lung diploid 2BS cells. The sequence of small interfering RNA(siRNA) targeting lncRNANEAT1 gene was designed and synthesized, and then transfected into PC-9 cells by liposome method. The expression level of NEAT1 in PC-9 cells before and after transfection was detected by qPCR. MTT and flow cytometry were used to detect the effect of lncRNANEAT1 knockdown on proliferation and cell cycle distribution of PC-9 cells, respectively. WB assay was used to detect the expressions of DNA damage-related proteins, namely, double-stranded DNA breaks (DSBs) biomarker γ-H2AX and ataxia-telangiectasia mutated (ATM), before and after transfection. Results: Compared with 2BS cells, lncRNA NEAT1 was highly expressed in PC-9 cells (P<0.05). The PC-9 cells with lncRNA NEAT1 knock-down were successfully established. After being transfected with siRNA for 12 h, the proliferation of PC-9 cells in siNEAT1 group and siNEAT2 group significantly decreased as compared with the blank control group and the empty transfection group (P<0.05). In the interference groups, cell cycle was arrested in G1 phase ([88.97±2.64]%, [88.15±1.48]% vs [84.5±1.72]%, P<0.05) and G2/M phase ([8.35±2.02]%, [8.11± 1.36]% vs [4.28±1.28]%, P<0.05). The expression levels of DNA damage-related proteinsATM and γ-H2AX in the interference groups were significantly increased (all P<0.05). Conclusion: lncRNA NEAT1 is highly expressed in lung adenocarcinoma PC-9 cells. lncRNA NEAT1 inhibits DNA damage and causes cell cycle at G1/M phase switch, and thus promotes the proliferation of lung adenocarcinoma cells.
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@# Objective: To investigate the effect and mechanism of bridging intergrator-1 (BIN1) on expression of programmed death-ligand 1 (PD-L1) in non-small cell lung cancer (NSCLC) A549 cells. Methods: Quantitative real-time polymerase chain reaction (qRTPCR) and Western blotting were used to detect the mRNA and protein expression of BIN1 and PD-L1 in A549 cells and normal human embryo lung fibroblast 2BS cells, respectively. Eukaryotic expression plasmid CMV-MCS-GFP-SV40-Neomycin-BIN1 containing human full length BIN1 gene sequence was transfected into A549 cells via cationic liposomes by using gene transfection technology (as BIN1+group); c-MYC-siRNAwas used to knockdown the expression of c-MYC inA549 cells through RNAinterference technique (as cMYC-siRNA group). The transfection efficiencies were verified by qRT-PCR and Western blotting, the effects of BIN1 over-expression and c-MYC knock-down on the expression of c-MYC and PD-L1 in A549 cells were detected as well. Results: Comparing with 2BS cells, the expression of BIN1 was down-regulated in A549 cells at both mRNA and protein levels, while the expression of PD-L1 was up-regulated (all P<0.05). The expression of BIN1 was increased at both mRNA and protein level in BIN1+ group, while the expression of PD-L1 was decreased significantly after B1N1 transfection (all P<0.05). After transfection of c-MYC-siRNA into A549 cells, the expression of c-MYC and PD-L1 in c-MYC-siRNAgroup was down-regulated significantly (all P<0.01). Conclusion: Over-expression of BIN1 could reduce the expression of PD-L1 by inactivating the c-MYC pathway, thereby inhibiting the immune escape ofA549 cells.