Expression of tumor necrosis factor related apoptosis inducing ligand receptor in glioblastoma

BACKGROUND： Receptors for tumor necrosis factor related apoptosis inducing ligand （TRAIL） include death receptor 4, death receptor 5, decoy receptor 1, and decoy receptor 2. Activation of death receptor 4 and 5 selectively kills tumor cells. OBJECTIVE： To detect TRAIL receptor expression in glioblastoma by immunohistochemistry and RT-PCR, and to compare this expression to that in normal brain tissue. DESIGN： Observational analysis. SETTING： Department of Pathology, the First Affiliated Hospital of Zhengzhou University; Henan Tumor Pathology Key Laboratory. PARTICIPANTS： Twenty-five patients （17 males and 8 females） who received glioblastoma resection were selected from the Fifth Affiliated Hospital of Zhengzhou University, between September 2003 to June 2004. All glioblastoma samples were diagnosed pathologically. Twenty patients （12 males and 8 females） with craniocerebral injury who received normal brain tissue resection were selected in the same time period. There were no significant differences in sex and age between glioblastoma patients or between craniocerebral injury patients （P 〉 0.05）. All patients and appropriate relatives provided informed consent, and this study was approved by the local research ethics committee. METHODS： Polyclonal antibody against TRAIL receptors and an immunohistochemical kit （batch number： 200502） were purchased from Boster Company, Wuhan. Immunohistochemistry： Expression of death receptor 4, death receptor 5, decoy receptor l, and decoy receptor 2 were observed in both glioblastoma and normal brain tissue. The experiment was performed according to the kit instructions, and positive staining was brown-yellow. Assessment： There were no positive signals （-）; weakly positive signals, positive cells 〈 25% （＋）; weakly positive signals, positive cells 25%-50% （＋＋）; strongly positive signals, positive cells 50%-75% （＋＋＋）; strongly positive signals, positive cells 〉 75% （＋＋＋＋）. Evaluation： Expression levels of TRAIL receptors were estimated in both normal brain tissue and glioblastoma. Expression of decoy receptor 1 and decoy receptor 2 mRNA in glioblastoma were detected by reverse transcription polymerase chain reaction, and expression of decoy receptor in glioblastoma was estimated. MAIN OUTCOME MEASURES： Comparison of death receptor and decoy receptor protein expression between glioblastoma and normal brain tissue; decoy receptor mRNA expression in glioblastoma. RESULTS： Death receptor protein expression was strongly positive （＋＋＋） in glioblastoma, while it was weakly positive （＋, ＋＋） in normal brain tissue. Therefore, expression rate of death receptor protein in the glioblastoma was significantly higher than that in the normal brain tissue （.~ 2 = 18.48, 23.03, P 〈 0.01）. Decoy receptor protein expression in the glioblastoma was significantly lower than that in the normal brain tissue （ x2 = 6.65, 18.76, P 〈 0.01）. The level of decoy receptor mRNA expression in glioblastoma was significantly higher than those of protein expression （ x 2 = 9.82, 10.09, P〈 0.01）. CONCLUSION： High expression of death receptor and low expression of decoy receptor are frequently observed in glioblastoma, suggesting that TRAIL receptor genes show an anti-tumor and expressive response during the initiation and development of the tumor. There are significant differences in decoy receptor expression between normal brain tissue and glioblastoma, suggesting that the restricted expression of decoy receptor in glioblastoma is regulated at the post-transcriptional level.

Antitumor effect of tumor necrosis factor-related apoptosis inducing ligand combined with mevastatin on a human glioma cell line SWO-38

BACKGROUND： Previous studies have reported that statins are less toxic to the human body and have greater antitumor activity; however, few studies have addressed the antitumor effect of statins combined with tumor necrosis factor-related apoptosis inducing ligand （TRAIL）. OBJECTIVE： To explore the effect of TRAIL combined with mevastatin on the proliferation and apoptotic cell death of a human glioma cell line SWO-38, and to study its mechanism of action. DESIGN, TIME AND SETTING： An in vitro control experiment was performed at the Central Laboratory of the Third Hospital Affiliated to Sun Yat-sen University, between January and April 2009. MATERIALS： The human SWO-38 cell line was provided by Cell Research, Department of Animal Experimental Center of Sun Yat-sen University; human recombinant soluble TRAIL by R＆D, USA; and mevastatin by Sigma, USA. METHODS： SWO-38 cells were separately incubated in TRAIL （100, 200, 300, 400, and 500 tJg/L） and mevastatin （5, 10, 20, 30, and 40 pmol/L） for 72 hours. In addition, SWO-38 cells were incubated in TRAIL （300 μg/L）, mevastatin （30 μmol/L）, and a solution containing both TRAIL and mevastatin for 12, 24, 48 and 72 hours. MAIN OUTCOME MEASURES： Cell proliferation was detected using methyl thiazolyl tetrazolium assay; cell apoptosis was observed using Hoechst 33258 staining and fluorescence microscopy and was measured using Annexin V/propidium iodide flow cytometry; TRAIL R1/DR4 and TRAIL R2/DR5 protein expressions levels were measured using indirect immunofluorescence staining combined with flow cytometry in the recombinant soluble TRAIL （rsTRAIL, 300 tJg/L）, mevastatin （30 IJmol/L） and combination groups; TRAIL R1/DR4 and TRAIL R2/DR5 mRNA expression was detected using real-time polymerase chain reaction. RESULTS： rsTRAIL, mevastatin and their combination inhibited tumor proliferation in a time- and dose-dependent manner. The proliferation inhibitory rate and apoptosis rate of human SWO-38 cells in the combined group were significantly greater than the rsTRAIL or mevastatin alone group （P 〈 0.01）. TRAIL R1/DR4 and TRAIL R2/DR5 protein and mRNA expressions were increased in the combination group compared with mevastatin or rsTRAIL alone after 72 hours （P 〈 0.01）. CONCLUSION： Both rsTRAIL and mevastatin inhibit the proliferation and apoptosis of the human glioma cell line SWO-38, while their combination enhances the anti-tumor effect. The mechanism of action possibly correlates to the upregulation of TRAIL R1/DR4 and TRAIL R2/DR5 mRNA expression by mevastatin, thereby enhancing the cell sensitivity to rsTRAIL.

Tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis in glioma U87 cells

Studies have shown that tumor necrosis factor-related apoptosis-inducing ligand（TRAIL）exhibits strong induction of apoptosis in human glioma cells.It remains unclear whether the mitochondrion pathway,an important apoptosis signaling pathway,is involved in TRAIL-induced glioma cell apoptosis.In the present study,in vitro cultured human glioma U87 cells were treated with human recombinant soluble TRAIL.Apoptosis of glioma U87 cells,mitochondrial transmembrane potential（Δψm）,cytoplasmic cytochrome c concentration and changes in caspase-3,-8 and-9 activity following human recombinant soluble TRAIL treatment were investigated to determine the mechanism of glioma U87 cell apoptosis induced by TRAIL.Additionally,blocking caspase-8resulted in TRAIL-induced mitochondrion pathway activation,suggesting that TRAIL,through activating caspase-8,initiated a series of mitochondrial events and resulted in apoptosis of glioma U87 cells.

TNF related apoptosis-inducing ligand and its receptors in ocular tumors

Most of the ocular tumors have poor prognosis, and they remain a difficult problem in the area of ophthalmology. With the rapid development of molecular biology and immunologic techniques and the deep research on ocular tumor related genes, it becomes possible to diagnose and treat malignant tumors from the molecular level. The tumor necrosis factor related apoptosis-inducing ligand (TRAIL), a member of the tumor necrosis factor (TNF) super family, is a promising candidate, either alone or in combination with established cancer therapies, since it can initiate apoptosis through the activation of their death receptors. The ability of TRAIL to selectively induce apoptosis of transformed, virus-infected or tumor cells but not normal cells promotes the development of TRAIL-based cancer therapy. Here, we will review TRAIL and its receptors' structure, function, mechanism of action and application in ocular tumors therapy.

Brain edema and tumor necrosis factor-like weak inducer of apoptosis in rats with cerebral ischemia

BACKGROUND： Recent studies have demonstrated that tumor necrosis factor-like weak inducer of apoptosis （TWEAK） participates in brain edema. However, it is unclear whether blood-brain barrier （BBB） disruption is associated with TWEAK during the process of brain edema OBJECTIVE： To investigate the effects of TWEAK on BBB permeability in brain edema. DESIGN, TIME AND SETTING： An immunohistochemical observation, randomized, controlled animal experiment was performed at the Laboratory of Neurosurgical Anatomy, Xiangya Medical College, Central South University ＆ Central Laboratory, Third Xiangya Hospital, Central South University between January 2006 and December 2007. MATERIALS： A total of 48 adult Wistar rats were randomly divided into three groups： normal control （n = 8）, sham-operated （n = 8）, and ischemia/reperfusion （n = 32）. Rats from the ischemia/reperfusion group were randomly assigned to four subgroups according to different time points, i.e., 2 hours of ischemia followed by 6 hours （n = 8）, 12 hours （n = 8）, 1 day （n = 8）, or 12 days （n = 8） of reperfusion. METHODS： Focal cerebral ischemia/reperfusion injury was induced by middle cerebral artery occlusion （MCAO） using the suture method in rats from the ischemia/reperfusion group. Thread was introduced at a depth of 17-19 mm. Rats in the sham-operated group were subjected to experimental procedures similar to the ischemia/reperfusion group; however, the introducing depth of thread was 10 mm. The normal control group was not given any intervention. MAIN OUTCOME MEASURES： TWEAK expression was examined by immunohistochemistry; brain water content on the ischemic side was calculated as the ratio of dry to wet tissue weight; BBB permeability was measured by Evans blue extravasation. RESULTS： A total of eight rats died prior to and after surgery and an additional eight rats were randomly entered into the study. Thus 48 rats were included in the final analysis. In the ischemia/reperfusion group, TWEAK-positive cells were present in the ischemic penumbra surrounding the lamellar necrotic region in the fight cerebral hemisphere at 6 hours reperfusion and increased thereafter; by 2 days reperfusion they had reached a peak level, which was significantly higher than the sham-operated and normal control groups （P 〈 0.05）. At 6 hours reperfusion, both brain water content and Evans blue extravasation showed the same tendency for change as TWEAK expression. Pearson correlation analysis results revealed that the degree of TWEAK expression was positively correlated with brain water content （r = 0.892, P 〈 0.05）. CONCLUSION： The present results confirmed that TWEAK was involved in BBB disruption and participated in brain edema following cerebral ischemia.

Induction of apoptosis in osteogenic sarcoma cells by combination of tumor necrosis factor-related apoptosis inducing ligand and chemotherapeutic agents

Background Osteosarcoma is one of the most common primary malignant tumors of bone with poor prognosis. TNF-related apoptosis inducing ligand （TRAIL） is a member of the tumor necrosis factor （TNF） cytokine family. TRAIL induces apoptosis in various tumor cell lines but is not found to be cytotoxic to many normal cell types in vitro. We investigated the cytotoxic activity of TRAIL and chemotherapeutic agents, including methotrexate （MTX）, doxorubicin （DOX） and cisplatin （CDDP）, on established osteosarcoma cell line-OS-732. Methods OS-732 cells were incubated with chemotherapeutic agents MTX,DOX and CDDP at various peak plasma concentrations（PPC）, 0.1PPC,1PPC and 10PPC, alone or with 100 ng/ml of TRAIL for 24 hours or 48 hours. MTT was used to evaluate the cytotoxic activity of different agents on OS-732. The apoptosis proportion was assayed by flow cytometry. Cellular morphologic changes were observed by phase contrast microscope, scan electron microscope, and transmission electron microscope. Results The inhibitory rate was （24.438±3.414）% with TRAIL of 100 ng/ml for 24 hours. The cells were responsive to DOX and CDDP with a dose-effect relationship （P〈0.05）. In OS-732 cells, DOX and CDDP cooperated synergistically with TRAIL when incubated the cells with them for 24 hours （the combined inhibitory rate is （58.360±2.146）% and （54.101±-2.721）%, respectively）. TRAIL alone or drugs alone induced the apoptosis rate was less than 25% （P〈0.05）. However, the combination of TRAIL and MTX did not present synergistic effects on OS-732 cells （P〉0.05, compared with TRAIL alone）. Conclusions Osteosarcoma OS-732 cells were not responsive to TRAIL-induced apoptosis. DOX and CDDP sensitize osteosarcoma OS-732 cells to TRAIL-induced apoptosis. The combination of TRAIL and MTX presented no synergistic effects on killing OS-732 cells.

可溶性肿瘤坏死因子样凋亡弱诱导因子在斑秃病人血清及皮损中的表达及临床意义

目的检测可溶性肿瘤坏死因子样凋亡弱诱导因子(sTWEAK)在斑秃病人血清及皮损中的表达水平,并探讨其在斑秃发病过程中的作用及临床意义。方法以2017年10月至2018年11月在首都医科大学附属北京友谊医院确诊并治疗的斑秃病人70例为斑秃组,依据病情分为轻症组(44例)、重症组(26例)。同期该院因色素痣或皮肤良性肿瘤进行头皮手术的人群65例作为对照组。比较各组sTWEAK及相关细胞因子白细胞介素(IL)-4、IL-12、γ干扰素(IFN-γ)蛋白及mRNA表达水平;采用Pearson法分析两指标间相关性。多元线性回归分析各指标与sTWEAK或脱发严重度工具(SALT)评分的关系。结果斑秃组血清及皮损中sTWEAK表达水平[(196.73±32.48)ng/L,(2.35±0.62)ng/L]高于对照组[(121.75±19.62)ng/L,(1.49±0.45)ng/L](P<0.05)。斑秃组血清及皮损中细胞因子IL-12、IFN-γ表达水平高于对照组(P<0.05),IL-4表达水平比较差异无统计学意义(P>0.05)。斑秃病人血清及皮损中sTWEAK表达水平与IL-12及IFN-γ呈正相关,与IL-4表达水平无明显相关性。轻症组血清及皮损中sTWEAK[(173.25±26.84)ng/L,(2.04±0.57)ng/L]、IL-12及IFN-γ表达水平低于重症组[(236.47±42.02)ng/L,(2.87±0.70)ng/L](P<0.05);轻症组、重症组血清及皮损中IL-4表达水平差异无统计学意义(P>0.05);斑秃病人血清、皮损处sTWEAK、IL-12、IFN-γ与SALT评分均呈正相关(P<0.05)。多元线性回归分析显示,IL-12、IFN-γ与血清和皮损处sTWEAK呈显著正相关;血清及皮损处sTWEAK、IL-12、IFN-γ与SALT评分存在正相关性(P<0.05)。结论斑秃病人血清及皮损中sTWEAK表达水平增加,与IL-12及IFN-γ呈正相关,sTWEAK、IL-12及IFN-γ均与斑秃病情严重程度密切相关,sTWEAK可能参与斑秃炎症反应过程。

血清sTWEAK、Netrin-1联合APACHEⅡ评分对重型颅脑损伤患者术后预后不良的预测价值

目的探讨血清可溶性肿瘤坏死因子样凋亡弱诱导因子(sTWEAK)、神经轴突导向因子-1(Netrin-1)联合急性生理学与慢性健康状况评分系统Ⅱ(APACHEⅡ)评分对重型颅脑损伤患者术后预后不良的预测价值。方法选取2020年6月至2022年6月该院收治的120例重型颅脑损伤患者,根据术后30 d预后情况分为预后良好组和预后不良组。对比两组血清sTWEAK、Netrin-1水平及APACHEⅡ评分。采用单因素和多因素Logistic回归分析重型颅脑损伤患者术后预后不良的影响因素,并据以构建血清sTWEAK、Netrin-1及APACHEⅡ评分联合应用的预测模型,受试者工作特征(ROC)曲线分析血清sTWEAK、Netrin-1水平及APACHEⅡ评分对重型颅脑损伤患者术后预后不良的预测价值。结果预后不良组的重症监护室居住时间长于预后良好组,白蛋白水平、入院时格拉斯哥昏迷评分法评分和血清Netrin-1水平低于预后良好组,多发脑挫裂伤占比、机械通气占比、入院时APACHEⅡ评分和血清sTWEAK、血清肌酐、血尿素氮水平均高于预后良好组,差异有统计学意义(P<0.05)。多因素Logistic回归分析结果显示,多发脑挫裂伤、Netrin-1水平降低、入院时APACHEⅡ评分升高、sTWEAK水平升高为重型颅脑损伤患者术后预后不良的危险因素(P<0.05)。ROC曲线分析结果显示,血清sTWEAK、Netrin-1及APACHEⅡ评分3个指标单独及联合应用时曲线下面积及其95%CI分别为0.742(0.552~0.925)、0.731(0.488~0.963)、0.714(0.502~0.911)、0.882(0.795~0.947)。结论血清sTWEAK、Netrin-1联合APACHEⅡ评分对重型颅脑损伤患者术后预后不良具有较好的预测价值,可为临床治疗方案的制订提供参考。

自发性急性脑出血患者血浆sCD163/sTWEAK比值与预后的关系

目的探究自发性急性脑出血(ACH)患者血浆可溶性CD163(sCD163)/可溶性肿瘤坏死因子样凋亡弱诱导因子(sTWEAK)比值与预后的关系。方法纳入ACH患者90例作为病例组,根据格拉斯哥预后评分将病例组分为预后不良组(38例)和预后良好组(52例);另选取同期体检健康者45例为对照组。酶联免疫吸附试验检测血浆sCD163、sTWEAK水平并计算sCD163/sTWEAK比值。分析血浆sCD163、sTWEAK水平及sCD163/sTWEAK比值与临床资料的相关性;Logistic回归分析ACH患者预后不良的影响因素;受试者工作特征(ROC)曲线分析sCD163/sTWEAK比值对ACH患者预后不良的预测价值。结果病例组血浆sCD163、sTWEAK水平及sCD163/sTWEAK比值均显著高于对照组;预后良好组上述指标均低于预后不良组(P<0.05)。预后良好组血肿体积、美国国立卫生研究院卒中量表(NIHSS)评分、高血压及幕下出血比例均低于预后不良组,低密度脂蛋白胆固醇(LDL-C)高于预后不良组(P<0.05)。相关性分析表明,血浆sCD163、sTWEAK水平及sCD163/sTWEAK比值与出血部位、血肿体积、NIHSS评分、白细胞计数、血小板计数、中性粒细胞/淋巴细胞比值(NLR)呈正相关(P<0.05)。Logistic回归分析显示,sCD163/sTWEAK比值、出血部位、血肿体积、NIHSS评分为ACH患者预后不良的影响因素(P<0.05)。ROC曲线结果表明,sCD163/sTWEAK比值评估ACH患者预后不良的AUC为0.850,敏感度和特异度分别为86.84%和69.23%。结论sCD163/sTWEAK比值在ACH患者血浆中水平较高,并与预后不良有关,该值对此类患者的预后有一定预测价值。

针刀调控线粒体途径软骨细胞凋亡防治大鼠膝骨关节炎

背景:针刀治疗膝骨关节炎的疗效确切,但其作用机制并不十分明确。目的:基于破骨细胞相关受体-肿瘤坏死因子相关的凋亡诱导配体-骨保护素(OSCAR-TRAIL-OPG)途径分析针刀对膝骨关节炎大鼠膝关节软骨细胞凋亡的影响。方法:采用随机数字表法将27只SD大鼠分为正常组(9只)、模型组(9只)、针刀组(9只),正常组大鼠常规饲养,不进行任何处理;模型组、针刀组采用膝关节内注射木瓜蛋白酶建立左后肢膝骨关节炎模型,造模成功后给予针刀组大鼠针刀干预,1次/周,共3次。干预结束后进行相关检测。结果与结论:①与正常组相比,模型组大鼠Lequesne MG行为学评分升高(P<0.01);与模型组相比,针刀组大鼠Lequesne MG行为学评分降低(P<0.01)。②苏木精-伊红染色显示与正常组相比,模型组大鼠膝关节软骨表面磨损且不平整,软骨细胞肿胀、破裂且数量减少,细胞排列杂乱;针刀组大鼠膝关节软骨表面较为平整,软骨细胞数量较多且排列较规整,结构基本清晰。③免疫组化染色显示与正常组相比,模型组大鼠膝关节软骨组织中OSCAR、TRAIL阳性表达增加(P<0.01),OPG阳性表达减少(P<0.01);与模型组相比,针刀组大鼠膝关节软骨组织中OSCAR、TRAIL阳性表达减少(P<0.01),OPG阳性表达增加(P<0.01)。④TUNEL染色显示与正常组相比,模型组软骨细胞凋亡数量增加(P<0.01);与模型组相比,针刀组软骨细胞凋亡数量减少(P<0.01)。⑤RT-qPCR与Western blot检测显示与正常组相比,模型组大鼠关节软骨组织中OSCAR、TRAIL、Bax表达升高(P<0.01),OPG、Bcl-2表达降低(P<0.01);与模型组相比,针刀组大鼠关节软骨组织中OSCAR、TRAIL、Bax表达降低(P<0.01),OPG、Bcl-2表达升高(P<0.01)。⑥针刀干预可减轻膝骨关节炎大鼠关节软骨组织损伤,该作用可能与OSCAR-TRAIL-OPG通路阻断线粒体途径凋亡信号释放有关。

Apoptosis and Expression of Protein TRAIL in Granulosa Cells of Rats with Polycystic Ovarian Syndrome

The relationship between apoptosis of granulosa cells and follicle development arrest in polycystic ovarian syndrome (PCOS) rats, and the contribution of tumor necrosis factor related apoptosis inducing ligand (TRAIL) in apoptosis of granulosa cells were explored. By using sodium prasterone sulfate rat PCOS model was induced. The apoptosis of granulosa cells in ovaries of rats was observed by TdT-mediated dUTP-biotin nick end-labeling (TUNEL), and the expression of TRAIL protein and mRNA in granulosa cells was detected by using immunhistochemical staining and reverse transcription polymerase chain reaction (RT-PCR) respectively. The apoptotic rate and the expression of protein TRAIL in granulosa cells were significantly higher in antral follicles from the PCOS rats than in those from the control rats (P<0.01, P<0.05). There was no significant difference in apoptotic rate and the expression of TRAIL protein in granulosa cells of preantral follicles between the PCOS rats and the control rats (P>0.05). No apoptosis and the expression of TRAIL protein in granulosa cells of primordial follicles were found in the two groups. The expression of TRAIL mRNA was significantly stronger in granulosa cells from the PCOS rats than in those from the con- trol rats (P<0.01). It was suggested that the apoptotic rate in granulosa cells was significantly higher in antral follicle from the PCOS rats than in those from the control rats. TRAIL played a role in regu- lating the apoptosis of granulosa cells in PCOS rats.

T2DM患者血清sTWEAK、sCD40L水平与糖尿病微血管并发症的相关性分析

目的 探讨2型糖尿病(T2DM)患者血清可溶性肿瘤坏死因子(TNF)样凋亡弱诱导因子(sTWEAK)、可溶性CD40配体(sCD40L)水平与糖尿病微血管并发症(DMA)的相关性。方法 选取2021年3月-2023年3月航天中心医院收治的150例T2DM患者为研究对象,根据患者是否合并DMA分为DMA组(n=69)及T2DM组(n=81)。检测并比较两组患者血清sTWEAK、sCD40L水平及临床资料,采用Logistic回归分析影响T2DM并发微血管病变的危险因素,绘制受试者工作特征曲线(ROC)分析sTWEAK、sCD40L对T2DM并发微血管病变的预测价值。结果 DMA组患者血清sTWEAK、sCD40L水平均高于T2DM组(P<0.05);DMA组的病程、FPG及HbA1c、sTWEAK、sCD40L水平均高于T2DM,差异均有统计学意义(P<0.05);Logistic分析结果显示病程长、HbA1c高水平、sTWEAK高水平及sCD40L高水平均是导致T2DM并发微血管病变的独立危险因素(P<0.05)。ROC结果显示,血清sTWEAK、sCD40L单独及联合预测T2DM并发微血管病变的AUC(95%CI)分别为0.714(0.635~0.785)、0.744(0.666~0.812)、0.859(0.792~0.910),二者联合的预测效能优于单独检测(Z=2.626、2.395,P<0.05)。结论 血清sTWEAK、 sCD40L在DMA患者中均异常升高,与DMA关系密切,sTWEAK、sCD40L联合对于T2DM并发微血管病变的预测价值较高。

连翘苷通过调控CTRP3表达对高糖诱导的人视网膜血管内皮细胞损伤的影响及其机制研究

目的比较不同剂量连翘苷(PHN)对高糖(HG)诱导的人视网膜血管内皮细胞损伤的影响,并分析其对补体C1q/肿瘤坏死因子相关蛋白3(CTRP3)表达的调控作用及可能的作用机制。方法采用HG培养人视网膜血管内皮细胞并建立细胞损伤模型(HG组)。HG+PHN-L组、HG+PHN-M组、HG+PHN-H组人视网膜血管内皮细胞分别用1μmol·L^(-1)、10μmol·L^(-1)、100μmol·L^(-1)PHN处理后进行HG诱导。HG+pcDNA组、HG+pcDNA-CTRP3组分别用pcDNA、pcDNA-CTRP3转染至人视网膜血管内皮细胞后进行HG诱导。HG+PHN-H+sh-NC组、HG+PHN-H+sh-CTRP3组分别用sh-NC、sh-CTRP3转染后,用100μmol·L^(-1)PHN处理HG诱导的人视网膜血管内皮细胞。检测细胞丙二醛(MDA)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)水平。采用流式细胞术检测细胞凋亡率。采用实时荧光定量聚合酶链反应(qRT-PCR)、Western blot分别检测B淋巴细胞瘤-2(Bcl-2)、Bcl-2相关蛋白(Bax)、CTRP3蛋白表达水平。结果与Con组比较,HG组细胞凋亡率、MDA水平、Bax蛋白水平均升高,SOD、GSH-Px、Bcl-2蛋白、CTRP3 mRNA及蛋白水平均降低(均为P<0.05)。与HG组比较,HG+PHN-L组、HG+PHN-M组、HG+PHN-H组细胞凋亡率、MDA水平、Bax蛋白水平均降低,且HG+PHN-H组<HG+PHN-M组<HG+PHN-L组(均为P<0.05),SOD、GSH-Px、Bcl-2蛋白、CTRP3 mRNA及蛋白水平均升高,且HG+PHN-H组>HG+PHN-M组>HG+PHN-L组(均为P<0.05)。与HG+pcDNA组比较,HG+pcDNA-CTRP3组细胞凋亡率、MDA水平、Bax蛋白水平均降低,SOD、GSH-Px、CTRP3蛋白、Bcl-2蛋白水平均升高(均为P<0.05)。与HG+PHN-H+sh-NC组比较,HG+PHN-H+sh-CTRP3组细胞凋亡率、MDA水平、Bax蛋白水平均升高,SOD水平、GSH-Px水平、CTRP3蛋白、Bcl-2蛋白水平均降低(均为P<0.05)。结论PHN可减轻HG诱导的人视网膜血管内皮细胞损伤,其作用机制可能与上调CTRP3表达有关。

血清Gal-3、TWEAK对精神分裂症的诊断价值及与精神症状严重程度的关系

目的研究血清半乳糖凝集素3(Gal-3)、肿瘤坏死因子样弱凋亡诱导因子(TWEAK)对精神分裂症的诊断价值及与精神症状严重程度的关系。方法选取2019年3月至2021年3月在该院诊治的96例精神分裂症患者作为病例组,以同期60例健康体检者为对照组,进行回顾性分析。采用酶联免疫吸附试验检测血清Gal-3、TWEAK水平。采用Pearson相关分析血清Gal-3、TWEAK水平与临床指标的相关性;采用多因素Logistic回归分析影响精神分裂症发生的因素;采用受试者工作特征(ROC)曲线分析血清Gal-3、TWEAK单项及联合检测对精神分裂症的诊断价值。结果观察组PANSS评分总分、阳性症状评分、阴性症状评分、一般精神病理学症状评分及血清Gal-3、TWEAK水平均高于对照组,差异均有统计学意义(P<0.001)。血清Gal-3、TWEAK水平与PANSS评分总分、阳性症状评分、阴性症状评分、一般精神病理学症状评分呈正相关(P<0.001)。PANSS评分总分、阳性症状评分、一般精神病理学症状评分升高及血清Gal-3、TWEAK水平升高均是精神分裂症发生的独立危险因素(P<0.001)。血清Gal-3、TWEAK联合检测诊断精神分裂症的曲线下面积为0.870(95%CI:0.831~0.919),明显高于血清Gal-3、TWEAK单项检测诊断精神分裂症的AUC[0.812(95%CI:0.769~0.847)、0.820(95%CI:0.771~0.852)],差异均有统计学意义(Z=4.345,P<0.001;Z=4.010,P=0.002)。结论精神分裂症患者血清Gal-3、TWEAK水平升高,二者与精神症状严重程度有关,血清Gal-3、TWEAK联合检测对精神分裂症具有较高的诊断价值。

前列腺癌患者血清TWEAK和SREBP-1水平表达与临床病理特征及无进展生存预后的关系研究

目的 研究前列腺癌(prostate cancer,PC)患者血清肿瘤坏死因子样弱凋亡诱导因子(tumor necrosis factor like weak inducer of apoptosis,TWEAK)、固醇调节元件结合蛋白1(sterol regulatory element-binding protein 1,SREBP-1)表达与临床病理特征及无进展生存预后的关系。方法 选取2018年1月~2020年1月武汉市东西湖区人民医院行PC根治术的94例PC患者为PC组,以同期50例前列腺增生(benign prostatic hyperplasia,BPH)患者为BPH组,以同期体检的50例健康人为对照组。酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)检测血清TWEAK和SREBP-1表达水平。Kaplan-Meier生存分析比较血清TWEAK和SREBP-1对PC患者无进展生存预后的影响。多因素COX回归分析影响PC患者无进展生存预后的因素。结果PC组患者血清TWEAK(77.14±15.46 ng/L),SREBP-1(334.14±33.81 ng/L)高于BPH组(38.69±10.58 ng/L,201.69±28.74 ng/L)和对照组(36.26±10.27 ng/L,189.51±27.65 ng/L),差异具有统计学意义(t=23.752,25.249;34.636,37.821,均P<0.05)。PC患者血清TWEAK与SREBP-1表达呈显著正相关(r=0.668,P=0.001)。Gleason评分> 7分、TNM分期Ⅲ期及术前前列腺特异抗原(prostate specific antigen,PSA)水平≥20 ng/ml的PC患者血清TWEAK,SREBP-1水平高于Gleason评分≤7分,TNM分期Ⅰ~Ⅱ期及术前PSA水平<20 ng/ml,差异具有统计学意义(t=8.465~16.597,均P<0.05)。TWEAK高表达组和低表达组三年总体无进展生存率分别为60.42%(29/48)和86.96%(40/46),SREBP-1高表达组和低表达组三年总体无进展生存率分别为57.78%(26/45)和87.76%(43/49);TWEAK高表达组、SREBP-1高表达组三年累积无进展生存率低于TWEAK低表达组、SREBP-1低表达组,差异具有统计学意义(Log-rankχ2=8.125,9.547,P=0.004,0.002)。TNM分期Ⅲ期(OR=1.448,P <0.001)、Gleason评分> 7分(OR=1.401,P <0.001)、术前PSA≥20 ng/ml (OR=1.353,P <0.001)及血清TWEAK (OR=1.338,P <0.001)和SREBP-1 (OR=1.293,P <0.001)是影响PC患者无进展生存预后的独立危险因素。结论 PC患者血清TWEAK和SREBP-1升高,两者与PC临床病理特征相关,是评估无进展生存预后的血清标志物。

2型糖尿病患者血清PTX3、sTWEAK水平与非酒精性脂肪性肝病的关系研究

目的探讨2型糖尿病(T2DM)患者血清正五聚蛋白3(PTX3)、可溶性肿瘤坏死因子样凋亡弱诱导因子(sTWEAK)水平与非酒精性脂肪性肝病(NAFLD)的关系。方法选取北部战区总医院152例新诊断为T2DM的患者为研究对象,根据是否合并NAFLD将患者分为NAFLD组(92例)和非NAFLD组(60例);根据肝脏超声检查结果,将NAFLD患者分为轻度组、中度组和重度组。另选取35例健康人作为对照(对照组)。采用酶联免疫吸附试验检测血清PTX3、sTWEAK水平。采用Pearson相关分析T2DM患者PTX3和sTWEAK水平与总胆固醇(TC)、甘油三酯(TG)、低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)、空腹血糖(FPG)、糖化血红蛋白(HbA1c)、胰岛素抵抗指数(HOMA-IR)的关系。采用多因素Logistic回归分析T2DM合并NAFLD的影响因素;采用受试者工作特征(ROC)曲线分析PTX3、sTWEAK对NAFLD的预测价值,计算曲线下面积(AUC)。比较轻度组、中度组、重度组血清PTX3、sTWEAK水平。结果与对照组比较,NAFLD组和非NAFLD组血清PTX3、sTWEAK水平较高(P<0.05)。与非NAFLD组比较,NAFLD组血清PTX3、sTWEAK水平较高(P<0.05)。体质量指数(BMI,OR=3.387)、TG(OR=1.958)、HOMA-IR(OR=3.040)、PTX3(OR=4.836)、sTWEAK(OR=4.133)是T2DM合并NAFLD的影响因素(P<0.05)。T2DM患者血清PTX3水平分别与BMI、LDL-C、FPG、HbA1c、HOMA-IR呈正相关(P<0.05),而与HDL-C呈负相关(P<0.05)。血清sTWEAK水平分别与LDL-C、FPG、HOMA-IR呈正相关(P<0.05)。血清PTX3、sTWEAK预测T2DM患者发生NAFLD的AUC分别为0.873和0.821,二者联合可将AUC提高至0.915。轻度组、中度组、重度组血清PTX3和sTWEAK水平比较,差异有统计学意义(P<0.05),病情越重,患者血清PTX3和sTWEAK水平越高。结论PTX3、sTWEAK是T2DM患者发生NAFLD的影响因素,并且血清PTX3、sTWEAK水平越高,NAFLD病情越重。

COMBINATION OF γ-INTERFERON WITH TRAIL AND CISPLATIN OR ETOPOSIDE INDUCES APOPTOSIS IN HUMAN NEUROBLASTOMA CELL LINE SH-SY5Y

Objective To study the effect of γ-interferon (IFNγ), tumor necrosis factor related apoptosis inducing ligand (TRAIL), and cisplatin or etoposide induced apoptosis in human neuroblastoma cell line SH-SY5Y and its possible molecular mechanisms.Methods The expressions of Caspase 8 mRNA and protein were detected with RT-PCR and Western blot analysis. The effects of IFNγ, TRAIL, IFNγ+TRAIL, IFNγ+Caspase 8 inhibitor+TRAIL, IFNγ+cisplatin+TRAIL, and IFNγ+etoposide+TRAIL on the growth and apoptosis of SH-SY5Y cells were detected with the methods of MTT and flow cytometry. The relative Caspase 8 activity was measured with colorimetric assay. Results Caspase 8 was undetectable in SH-SY5Y cells but an increased expression of Caspase 8 mRNA and protein was found after treatment with IFNγ. SH-SY5Y cells themselves were not sensitive to TRAIL, but those expressing Caspase 8 after treatment with IFNγ were. The killing effect of TRAIL on SH-SY5Y cells expressing Caspase 8 was depressed by Caspase 8 inhibitor. Cisplatin and etoposide could enhance the sensitivity of TRAIL on SH-SY5Y cells. The relative Caspase 8 activity of SH-SY5Y cells in IFNγ+TRAIL group was significantly higher than those of control group, IFNγ group, TRAIL group, and inhibitor group (P<0.01). There was no significant difference among IFNγ+TRAIL group, IFNγ+cisplatin+TRAIL group, and IFNγ+etoposide+TRAIL group.Conclusions IFNγ could sensitize SH-SY5Y cells to TRAIL-induced apoptosis and this may be realized by the upregulation of Caspase 8. Cisplatin and etoposide could enhance the killing effect of TRAIL on SH-SY5Y cells.

Downregulation of cyclooxygenase-1 is involved in gastric mucosal apoptosis via death signaling in portal hypertensive rats

门高血压(PHT ) gastropathy 是肝肝硬化的经常的复杂并发症，带之一从肝硬化死亡引起。Apoptosis 广泛地被认为从坏死的房间死亡是房间死亡和一个不同实体的一个活跃精力依赖者模式。胃的 mucosal apoptosis 是否涉及 PHT gastropathy，是不清楚的。通过 cyclooxygenase (艇长) 生产的前列腺素(PG ) 被认为从损害和 apoptosis 在胃肠的 mucosa 的保护起一个关键作用。然而，在 PHT gastropathy 的艇长的角色仍然不清楚地被理解。这研究的目的是调查(1 ) 胃的 mucosal apoptosis 是否涉及 PHT gastropathy，(2 ) 艇长的 downregulation 贡献这 apoptosis。在这研究，当 mucosal 增长在 PHT 老鼠被禁止时，我们证明胃的 mucosal apoptosis 显著地被增加。胃的 mucosal COX-1 显著地在 mRNA 和蛋白质层次被压制，并且 PGE2 在 PHT 老鼠被减少。进一步， PGE2 处理在 PHT 老鼠压制了胃的 mucosal apoptosis。然而，胃的 mucosal COX-2 层次没在假冒操作老鼠和 PHT 老鼠之间不同。肿瘤坏死因素的胃的 mucosal 层次 --(TNF-) 并且船边交货 ligand，然而并非 TNF 相关的导致 apoptosis ligand，被增加，并且激活的 caspase-8 和 caspase-3 层次是在 PHT 老鼠的 upregulated。到 cytosol 的从线粒体的细胞色素 c 的版本没在 PHT 老鼠被观察。我们的数据显示 COX-1 的 downregulation 经由死亡涉及胃的 mucosal apoptosis 调停信号的类型 -- 我在 PHT 老鼠的房间死亡。

TRAIL-induced apoptosis of hepatocellular carcinoma cells is augmented by targeted therapies

AIM:To analyze the effect of chemotherapeutic drugs and specific kinase inhibitors,in combination with the death receptor ligand tumor necrosis factor-related apoptosis inducing ligand(TRAIL),on overcoming TRAIL resistance in hepatocellular carcinoma(HCC)and to study the efficacy of agonistic TRAIL antibodies,as well as the commitment of antiapoptotic BCL-2 proteins, in TRAIL-induced apoptosis. METHODS:Surface expression of TRAIL receptors (TRAIL-R1-4)and expression levels of the antiapoptotic BCL-2 proteins MCL-1 and BCL-xL were analyzed by flow cytometry and Western blotting,respectively. Knock-down of MCL-1 and BCL-xL was performed by transfecting specific small interfering RNAs.HCC cellswere treated with kinase inhibitors and chemotherapeutic drugs.Apoptosis induction and cell viability were analyzed via flow cytometry and 3-(4,5-Dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. RESULTS:TRAIL-R1 and-R2 were profoundly expressed on the HCC cell lines Huh7 and Hep-G2. However,treatment of Huh7 and Hep-G2 with TRAIL and agonistic antibodies only induced minor apoptosis rates.Apoptosis resistance towards TRAIL could be considerably reduced by adding the chemotherapeutic drugs 5-fluorouracil and doxorubicin as well as the kinase inhibitors LY294002[inhibition of phosphoinositol- 3-kinase(PI3K)],AG1478(epidermal growth factor receptor kinase),PD98059(MEK1),rapamycin(mam- malian target of rapamycin)and the multi-kinase inhibitor Sorafenib.Furthermore,the antiapoptotic BCL-2 proteins MCL-1 and BCL-xL play a major role in TRAIL resistance:knock-down by RNA interference increased TRAIL-induced apoptosis of HCC cells.Additionally, knock-down of MCL-1 and BCL-xL led to a significant sensitization of HCC cells towards inhibition of both c-Jun N-terminal kinase and PI3K.CONCLUSION:Our data identify the blockage of survival kinases,combination with chemotherapeutic drugs and targeting of antiapoptotic BCL-2 proteins as promising ways to overcome TRAIL resistance in HCC.

EGFR inhibitors sensitize non-small cell lung cancer cells to TRAIL-induced apoptosis

Apoptosis induced by tumor necrosis factor-related apoptosis-inducing ligand(TRAIL) can be regulated by the epidermal growth factor(EGF) signaling pathway.In this study,recombinant adenoviral vectors that encode TRAIL gene from the hTERT/RGD promoter(AdTRAIL) was combined with drugs including gefitinib,elotinib,and cetuximab that inhibit EGFR and the EGF signaling pathway in non-small cell lung cancer(NSCLC) cell lines to investigate their antitumor activity.In vitro,compared to single reagent,AdTRAIL combined with EGFR inhibitors reduced proliferation and enhanced apoptosis in H460,A549,and SW1573 cell lines.Western blot results suggested that these effects were relative to up-regulation of pro-apoptosis protein BAX and down-regulation of p-AKT.In vivo,AdTRAIL combined with cetuximab resulted in a significant growth reduction in H460 xenografts without damage to the main organs of nude mice.Histological examination and TUNEL analyses of xenografts showed that cetuximab enhanced cell apoptosis induced by AdTRAIL.These results indicate that EGFR inhibitors enhanced AdTRAIL anti-tumor activity in NSCLC cell lines and that inhibiting the AKT pathway played an important role in this enhancement.