Correlation of tumor necrosis factor receptor superfamily 13B variation with sporadic intracranial aneurysm and clinical characteristics in Han Chinese populations

BACKGROUND： Inflammatory reaction correlates with sporadic intracranial aneurysm （IA）. Variation of tumor necrosis factor receptor superfamily 13B （TNFRSF13B）, an inflammatory mediator receptor, may associate with IA. OBJECTIVE： To explore the relationship between TNFRSF13B gene and sporadic IA, as well as the clinical characteristics of sporadic IA. DESIGN, TIME AND SETTING： Case-control study of genetic association was performed at the Experimental Technology Center of China Medical University from November 2006 to January 2008. PARTICIPANTS： A total of 367 patients with IA, confirmed by three-dimensional computed tomography angiography, magnetic resonance angiography, digital subtraction angiography, and neuro surgery, were admitted to the Department of Neurosurgery, First Affiliated Hospital of China Medical University from 2006 to 2007, and were selected as the case group. All patients were Han, with no family history of IA. In addition, a total of 396 non-lA patients were selected as control subjects. METHODS： Peripheral vein blood was harvested to extract whole blood genomic DNA. Genotyping and TNFRSF13B single nucleotide polymorphism （SNP） rs11078355 G〉A allele polymorphisms were determined by polymerase chain reaction-restriction fragment length polymorphism. The relationship of TNFRSF13B SNP rs11078355 G〉A polymorphisms to IA and IA clinical characteristics were analyzed using the chi-square and two-sided test. MAIN OUTCOME MEASURES： TNFRSF13B SNP rs11078355 G〉A genotype distribution. RESULTS： In the IA patients, TNFRSF13B SNP rs11078355 G〉A genotype frequency was significantly increased （X2 = 16.306, odds ratio = 1.881,95% confidence interval = 1.382 2.560, P 〈 0.001）. In IA patients aged 〉 65 years, the frequency of TNFRSF13B SNP rs11078355 GA ＋ AA genotype was significantly greater than the GG genotype （X2 = 26.604, odds ratio = 5.248, 95% confidence interval = 2.662 10.345, P 〈 0.001）. CONCLUSION： The TNFRSF13B gene may associate with sporadic IA in Han Chinese populations In elderly patients, allele A may be an independent risk factor for IA, in addition to senile diseases, such as hypertension and diabetes mellitus.

Tumor necrosis family receptor superfamily member 9/tumor necrosis factor receptor-associated f

Hepatitis C virus(HCV)infection is an excellent immunological model for understanding the mechanisms developed by non-cytopathic viruses and tumors to evade the adaptative immune response.The antigen-specific cytotoxic T cell response is essential for keeping HCV under control,but during persistent infection,these cells become exhausted or even deleted.The exhaustion process is progressive and depends on the infection duration and level of antigenemia.During high antigenic load and long duration of infection,T cells become extremely exhausted and ultimately disappear due to apoptosis.The development of exhaustion involves the impairment of positive co-stimulation induced by regulatory cytokines,such as transforming growth factor beta 1.This cytokine downregulates tumor necrosis factor receptor(TNFR)-associated factor 1(TRAF1),the signal transducer of the T cell co-stimulatory molecule TNFR superfamily member 9(known as 4-1BB).This impairment correlates with the low reactivity of T cells and an exhaustion phenotype.Treatment with interleukin-7 in vitro restores TRAF1 expression and rescues T cell effector function.The process of TRAF1 loss and its in vitro recovery is hierarchical,and more affected by severe disease progression.In conclusion,TRAF1 dynamics on T cells define a new pathogenic model that describes some aspects of the natural history of HCV,and sheds light on novel immunotherapy strategies for chronic viral infections and cancer.

新生儿GBS感染所致化脓性脑膜炎中血清维生素D和炎性细胞因子的表达及意义

目的检测新生儿B族链球菌(group B streptococcus,GBS)感染所致化脓性脑膜炎(purulent meningitis,PM)血清中维生素D、白细胞介素(interleukin,IL)-6、IL-10、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)和C反应蛋白(c-reactive protein,CRP)的表达水平,并探讨其临床价值。方法选取2017年5月至2020年5月在秦皇岛市第一医院出生的59例GBS感染的PM新生儿纳入观察组,同期59例非GBS感染的PM新生儿(晚发败血症)纳入对照组。检测所有受试者血清维生素D、CRP、IL-6、IL-10和TNF-α水平,并进行Pearson相关性分析;利用受试者操作特征(receiver operating characteristic,ROC)曲线分析血清维生素D和炎性细胞因子对新生儿GBS感染所致PM的诊断价值。统计学方法采用t检验、χ^(2)检验和Pearson相关性分析。结果观察组与对照组孕产妇胎膜早破[47.5%(28/59)与5.1%(3/59),χ^(2)=27.345]、产时窒息[52.5%(31/59)与18.6%(11/59),χ^(2)=14.787]和产褥感染[(44.1%(26/59)与(22.0%(13/59)),χ^(2)=6.473]的发生率比较,观察组明显高于对照组(P<0.05)。观察组血清维生素D水平显著低于对照组[(13.3±2.1)μg/L与(21.1±5.0)μg/L,t=11.345],IL-6[(87.1±14.5)μg/L与(63.9±11.9)μg/L,t=9.507]、IL-10[(49.6±15.2)μg/L与(29.3±10.0)μg/L,t=8.596]、TNF-α[(76.8±19.0)μg/L与(50.0±10.8)μg/L,t=9.410]和CRP[(21.5±5.0)μg/L与(13.7±3.7)μg/L,t=9.702]水平显著高于对照组(P值均<0.05)。Pearson相关性分析结果显示,观察组血清维生素D水平分别与IL-6、IL-10、TNF-α和CRP水平呈负相关(r=-0.662、-0.644、-0.564、-0.643,P<0.05);血清维生素D、IL-6、IL-10、TNF-α和CRP单独诊断GBS感染新生儿PM的曲线下面积(area under the curve,AUC)分别为0.831(95%CI:0.757~0.904)、0.887(95%CI:0.830~0.944)、0.859(95%CI:0.793~0.925)、0.888(95%CI:0.821~0.955)、0.879(95%CI:0.820~0.938),5项联合检测的AUC为0.991(95%CI:0.978~1.000)。结论GBS感染所致的PM新生儿血清中维生素D水平降低,炎性细胞因子水平增加,对于GBS感染所致的PM具有一定的辅助诊断价值。

Effect of Wenhua Juanbi Recipe(温化蠲痹方) on Expression of Receptor Activator of Nuclear Factor Kappa B Ligand,Osteoprotegerin,and Tumor Necrosis Factor Receptor Superfamily Member 14 in Rats with Collagen-Induced Arthritis

Objective： To study the effect of Wenhua Juanbi Recipe（温化蠲痹方, WJR） on expression of receptor activator of nuclear factor kappa B ligand（RANKL）, osteoprotegerin（OPG）, and tumor necrosis factor receptor superfamily member 14（TNFRSF14, also known as LIGHT） in rats with collagen-induced arthritis（CIA）. Methods： CIA rats were generated by subcutaneous injection of bovine collagen type-Ⅱ at the tail base. Sixty CIA rats were randomly assigned（10 animals/group） to： model, methotrexate（MTX）-treated（0.78 mg/kg body weight）, and WJR-treated（22.9 g/kg） groups. Healthy normal rats（n=10） were used as the normal control. Treatments or saline were administered once daily by oral gavage. Rats were sacrificed at day 28 post-treatment and knee synovium and peripheral blood serum were collected. Toe swelling degree and expression of RANKL, OPG, and LIGHT were determined by Western blot and immunohistochemistry. Results： Compared with the normal group, toe swelling degree was significantly increased in the model group（P〈0.01）. After treatment, toe swelling degree decreased significantly in the WJR and MTX groups compared with the model group（P〈0.01）. Compared with the normal group, expression of RANKL and LIGHT were significantly increased and OPG significantly decreased in peripheral blood and synovium of the model group（P〈0.01）. Conversely, RANKL and LIGHT expression were significantly reduced and OPG increased in the WJR and MTX groups compared with the model group（P〈0.01）. No statistically significant difference existed between WJR and MTX groups. Conclusion： WJR likely acts by reducing RANKL expression and increasing OPG expression, thus inhibiting RANKL/RANK interaction and reducing LIGHT expression, thereby inhibiting osteoclast formation/activation to block bone erosion.

Tumor necrosis factor receptor superfamily member 9 is upregulated in the endothelium and tumor cells in melanoma brain metastasis

Aim:The cytokine receptor tumor necrosis factor receptor superfamily member 9(TNFRSF9)is mainly considered to be a co-stimulatory activation marker in hematopoietic cells.Several preclinical models have shown a dramatic beneficial effect of treatment approaches targeting TNFRSF9 with agonistic antibodies.However,preliminary clinical phase I/II studies were stopped after the occurrence of several severe deleterious side effects.In a previous study,it was demonstrated that TNFRSF9 was strongly expressed by reactive astrocytes in primary central nervous system(CNS)tumors,but was largely absent from tumor or inflammatory cells.The aim of the present study was to address the cellular source of TNFRSF9 expression in the setting of human melanoma brain metastasis,a highly immunogenic tumor with a prominent tropism to the CNS.Methods:Melanoma brain metastasis was analyzed in a cohort of 78 patients by immunohistochemistry for TNFRSF9 and its expression was correlated with clinicopathological parameters including sex,age,survival,tumor size,number of tumor spots,and BRAF V600E expression status.Results:Tumor necrosis factor receptor superfamily member 9 was frequently expressed independently on both melanoma and endothelial cells.In addition,TNFRSF9 was also present on smooth muscle cells of larger vessels and on a subset of lymphomonocytic tumor infiltrates.No association between TNFRSF9 expression and patient survival or other clinicopathological parameters was seen.Of note,several cases showed a gradual increase in TNFRSF9 expression on tumor cells with increasing distance from blood vessels,an observation that might be linked to hypoxia-driven TNFRSF9 expression in tumor cells.Conclusion:The findings indicate that the cellular source of TNFRSF9 in melanoma brain metastasis largely exceeds the lymphomonocytic pool,and therefore further careful(re-)assessment of potential TNFRSF9 functions in cell types other than hematopoietic cells is needed.Furthermore,the hypothesis of hypoxia-driven TNFRSF9 expression in brain metastasis melanoma cells requires further functional testing.

TRAF5通过调控IL-17A/NF-κB信号通路减轻氧-糖剥夺/再灌注引起的心肌细胞炎症和细胞凋亡

目的 探讨肿瘤坏死因子受体相关因子5(tumor necrosis factor receptor-associated factor 5,TRAF5)对氧-葡萄糖剥夺/再灌注(oxygen-glucose deprivation/reperfusion,OGD/R)引起的心肌细胞炎症和细胞凋亡的影响及相关分子机制。方法 体外培养大鼠心肌细胞系H9c2细胞,构建OGD/R诱导的H9c2细胞模型,以脂质体转染法将TRAF5过表达质粒(pcDNA-TRAF5)和空载质粒(pcDNA-NC)转染至OGD/R诱导的H9c2细胞。采用实时定量聚合酶链反应(quantitative real time polymerase chain reaction,qRT-PCR)法检测细胞中TRAF5基因表达,CCK-8法测定细胞活力,试剂盒法检测细胞中乳酸脱氢酶(lactate dehydrogenase,LDH)、心肌肌钙蛋白T(cardiac troponin T,cTnT)和炎性因子水平,流式细胞术检测细胞凋亡,Western blot法检测细胞中相关蛋白表达。结果 与对照组比较,OGD/R组H9c2细胞中TRAF5 mRNA和蛋白表达量降低,细胞活力降低,细胞中LDH、cTnT、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-6(interleukin-6,IL-6)、白细胞介素-1β(interleukin-1β,IL-1β)、单核细胞趋化蛋白-1(monocyte chemoattractant protein-1,MCP-1)浓度升高,细胞凋亡率升高,细胞中B淋巴细胞瘤2(B-cell lymphoma-2,Bcl-2)蛋白表达量降低,Bcl-2相关蛋白(B-cell-lymphoma-2 related protein,Bax)、切割型半胱氨酸天冬氨酸蛋白水解酶-3(cleaved cysteine aspartate proteolytic enzyme-3,cleaved caspase-3)、白细胞介素-17A(interleukin-17A,IL-17A)蛋白表达量及磷酸化核转录因子-κB(phosphorylated nuclear factor-κB,p-NF-κB)/NF-κB比值升高,差异均有统计学意义(P<0.05)。与OGD/R组比较,OGD/R+pcDNA-TRAF5组H9c2细胞中TRAF5 mRNA和蛋白表达量升高,细胞活力升高,细胞中LDH、c TnT、TNF-α、IL-6、IL-1β、MCP-1浓度降低,细胞凋亡率降低,细胞中Bcl-2蛋白表达量升高,Bax、cleaved caspase-3、IL-17A蛋白表达量及p-NF-κB/NF-κB比值降低,差异均有统计学意义(P<0.05)。结论 TRAF5可减轻OGD/R引起的心肌细胞炎症和细胞凋亡,其作用机制可能与调控IL-17A/NF-κB信号通路有关。

Nuclear factor kB activity in patients with acute severe cholangitis

瞄准：在外部血决定 NF-kB 活动在有严重类型(ACST ) 和相互关联的尖锐胆管炎的病人的单音的原子房间(PBMC ) 有胆汁的道感染和临床的结果的严厉的 NF-kB 激活的度。方法：有 ACST 的二十个病人被划分成幸存者组(13 个盒子) 并且非幸存者组(7 个盒子) 。经历选任的 gastrectomy 或腹股沟的脱肠修理的另外的十个病人作为控制组被选择。外部血样品手术后地被花 24 个小时。PBMC 被密度坡度 centrifugation 分开，然后原子的蛋白质从 PBMC 被孤立，并且 Electrophoretic 活动性移动试金(EMSA ) 使用了坚定。结果被扫描一个简历图象分析系统的 densitometer 确定并且表示了同样相对的光密度(杆) 。在有 ACST 和健康控制题目的病人的血浆的 TNF-alpha， IL-6，和 IL-10 的层次被使用连接酶的免疫分析(ELISA ) 决定。结果：NF-kB 活动是 5.02 +/- 1.03 在里面非幸存者组， 0.51 在幸存者组织的 2.98 +/- 和 1.06 +/- 0.34 在控制组。在三个组(P【0.05 ) 有统计差别。在血浆的 TNF-alpha 和 IL-6 的层次是( 498 +/- 53 ) ng.L ( -1)and ( 587 +/- 64 ) ng.L ( -1)in 非幸存者组，( 284 +/- 32 ) ng.L (-1)并且( 318 +/- 49 ) ng.L ( -1)in 幸存者组并且( 89 +/- 11 ) ng.L (-1)并且( 102 +/-13 ) ng.L ( -1)in 控制组。有 ACST 的所有病人增加了 TNF-alpha 和 IL-6 的层次，它是控制组的比那些大的许多褶层，并且有一条证据在那些显著地高级非幸存者组比那在幸存者组(P【0.05 ) 。在血浆的 IL-10 的层次是(378+/-32 ) ng.L (-1),(384+/-37 ) ng.L (-1) 并且(68+/-11 ) ng.L (在三个组的 -1) 分别地。当与控制相比组(P【0.05 ) ，而是 IL-10 层次不是显著地更高级的在时，所有病人也增加了 IL-10 的层次非幸存者比在幸存者(P】0.05 ) 。结论：在在有 ACST 的病人的 PBMC 的 NF-kB 活动显著地增加并且 NF-kB 激活的度与胆汁的道感染和临床的结果的严厉被相关。

Role of Toll-like receptor 4 in inflammatory reactions of hippocampal neurons

Lipopolysaccharide stimulates Toll-like receptor 4 on immune cells to produce immune mediators. Toll-like receptor 4 is also expressed by non-immune cells, which can be stimulated by lipopolysaccharide. However, whether Toll-like receptor 4 is expressed by primary cultured hippocampal neurons and its specific role in lipopolysaccharide-induced neuroinflammation is currently undefined, in this study, Toll-like receptor 4 antibody blocking was used to analyze the Toll-like receptor 4 signaling pathway and changes in inflammation of lipopolysaccharide stimulated hippocampal neurons. Immunofluorescence showed that Toll-like receptor 4 protein was mainly located in the membrane of hippocampal neurons. Quantitative reverse transcription-PCR and western blot assay showed that after stimulation of lipopolysaccharide, the mRNA and protein levels of Toll-like receptor 4 and the mRNA levels of interleukin-ll3 and tumor necrosis factor-（] were significantly increased. In addition, there was increased phosphorylation and degradation of kappa B a inhibitor in the cytosol and increased nuclear factor-KB p65 expression in the nuclei. Pretreatment with Toll-like receptor 4 antibody could almost completely block this increase. These experimental findings indicate that lipopolysaccharide participates in neuroinflammation by stimulating Toll-like receptor 4/nuclear factor-KB pathway in hippocampal neurons, which may be both ＂passive victims＂ and ＂activators＂ of neuroinflammation.

TNFRSF10B在直肠癌中的表达及临床意义

目的探讨肿瘤坏死因子受体超家族成员(TNFRSF)10B在直肠癌组织中的表达及临床意义,以期为直肠癌预后判断提供理论依据。方法选取2013年9月至2017年7月在唐山市中医医院进行手术治疗的直肠癌患者87例为研究对象,采用实时荧光定量PCR和免疫组织化学染色法检测患者直肠癌组织及癌旁组织中TNFRSF10B mRNA及蛋白的表达情况,分析TNFRSF10B蛋白表达与直肠癌患者临床病理特征及预后的关系。结果癌旁组织中TNFRSF10B mRNA的表达水平明显低于直肠癌组织中TNFRSF10B mRNA的表达水平,差异有统计学意义(P<0.05)。癌旁组织中TNFRSF10B蛋白的高表达率为13.79%,明显低于直肠癌组织中TNFRSF10B蛋白的高表达率(54.02%),差异有统计学意义(P<0.05)。TNFRSF10B蛋白表达与直肠癌患者临床分期、肿瘤分化程度、淋巴结转移、远处转移有关(P<0.05)。TNFRSF10B蛋白高表达、临床分期Ⅲ+Ⅳ期、淋巴结转移是影响直肠癌患者预后的独立危险因素(P<0.05)。结论TNFRSF10B在直肠癌组织中高表达,与直肠癌的发生、发展相关,对直肠癌患者预后评估有一定的指导意义。

基于TLR4/MyD88/NF-kB信号通路探讨追风透骨胶囊减缓兔膝骨关节炎模型软骨退变的作用机制

目的 观察追风透骨胶囊对兔膝骨关节炎(knee osteoarthritis, KOA)模型关节软骨退变的干预作用,并基于Toll样受体4(Toll like receptor 4, TLR4)/髓细胞分化初级反应蛋白88(myeloid differentiation primary response protein 88, MyD88)/核因子kappa-B(nuclear factor kappa-B, NF-κB)信号通路探讨追风透骨胶囊的可能作用机制。方法 从50只6月龄雄性新西兰兔中随机选取10只作为正常组,其余40只为KOA造模组,用改良Videman造模法制备兔KOA模型。模型验证后,从正常组中随机选取6只作为空白组(A组),KOA造模组抽取30只随机分为模型组(B组)、硫酸氨基葡萄糖组(C组)、追风透骨胶囊低剂量组(D组)、追风透骨胶囊中剂量组(E组)、追风透骨胶囊高剂量组(F组),每组6只。A、B组予蒸馏水灌胃,C、D、E、F组予相应药物灌胃,疗程均为6周。给药结束后,取造模侧膝关节软骨,予大体及HE染色观察,并用Pelletier评分及Mankin评分评估;以Western blot、RT-PCR法检测软骨中TLR4、My D88、NF-κB蛋白及其mRNA的表达;免疫组化法检测白细胞介素-1β(interleukin-1β, IL-1β)、白细胞介素-6(interleukin-6, IL-6)、肿瘤坏死因子-α(tumor necrosis factor-α, TNF-α)的含量。结果 与A组比较,B组膝关节软骨破坏明显,Pelletier评分及Mankin评分均升高(P<0.01),软骨中TLR4、MyD88、NF-κB蛋白及其mRNA表达水平均上调(P<0.01),IL-1β、IL-6、TNF-α含量均升高(P<0.01)。与B组比较,C、D、E、F组的软骨损伤程度轻,软骨Mankin评分均降低(P<0.01),软骨中TLR4、MyD88、NF-κB蛋白及m RNA表达水平均下调(P<0.01),IL-1β、IL-6、TNF-α含量均降低(P<0.01),此外,F组软骨Pelletier评分较B组降低明显(P<0.05)。结论 追风透骨胶囊能延缓改良Videman造模法诱导的兔KOA模型关节软骨退变,其作用机制可能与下调软骨中TLR4、MyD88、NF-κB蛋白及m RNA的表达,抑制TLR4/MyD88/NF-κB信号通路的活化,降低IL-1β、IL-6、TNF-α在软骨中的含量,从而减轻炎症反应相关。

Decoy receptor 3: Its role as biomarker for chronic inflammatory diseases

Members of the tumor-necrosis factor-α(TNF-α) and TNF-α receptor(TNFR) superfamilies of proteins(TNFSF and TNFRSF, respectively) play important roles in the function of the immune system. Decoy receptor 3(Dc R3, TNFRSF6b) is a decoy receptor that binds to three TNFSF ligands, Fas L, LIGHT and TL1 A. Association to these ligands competes with the corresponding functional receptors and blocks downstream signaling, leading to immunomodulatory effects, including the prevention of apoptosis. Dc R3 lacks a transmembrane region and exists only as a secreted protein, which is detectable in biological fluids. Recent studies have shown that Dc R3 is upregulated and may be pathogenetically implicated in several and diverse chronic inflammatory diseases. The strongest associations have been described for rheumatological diseases, mainly systemic lupus erythematosus and rheumatoid arthritis, inflammatory bowel disease, and serious infectious conditions, including systemic inflammatory response syndrome. In the majority of these conditions, Dc R3 m RNA and protein expression is elevated both at the target tissues as well as in the systemic circulation. Dc R3 concentration in the serum is untraceable in the majority of healthy individuals but can be detected in patients with various inflammatory diseases. In mostsuch cases, soluble Dc R3 correlates with disease severity, as patients with severe forms of disease have significantly higher levels than patients with milder or no activity. In addition, effective anti-inflammatory treatment leads to the disappearance of soluble Dc R3 from the circulation. Taken together, current evidence suggests that serum Dc R3 may become a useful biomarker for chronic inflammatory disorders, as it is upregulated in response to inflammatory stimuli, and may serve both as a prognostic marker for disease severity and as a surrogate indicator of response to treatment.

柚皮素对过敏性鼻炎模型大鼠鼻黏膜组织TLR4/NF-κB/TNF-α信号通路的影响

目的探究柚皮素对过敏性鼻炎(AR)模型大鼠鼻黏膜组织Toll样受体4(TLR4)/核因子κB(NF-κB)/肿瘤坏死因子α(TNF-α)信号通路的影响。方法建立AR大鼠模型,48只大鼠以随机数字表法平均分为4组:模型组、柚皮素低(100 mg/kg)剂量组、柚皮素高(200 mg/kg)剂量组、柚皮素高(200 mg/kg)剂量+脂多糖(LPS)(TLR4通路激活剂,0.4 mg/kg)组,另取12只SD大鼠设为对照组。以药物分组干预治疗后,观察大鼠鼻炎症状并进行评分;苏木精-伊红(HE)染色检测大鼠鼻黏膜组织病理形态改变;试剂盒测定大鼠血清IgE及炎性因子白细胞介素(IL)-17、IL-18水平;免疫组织化学染色检测大鼠鼻黏膜组织CD19、CD23表达;免疫印迹法检测大鼠鼻黏膜组织TLR4/NF-κB/TNF-α通路蛋白表达。结果与对照组相比,模型组大鼠鼻黏膜组织发生严重病理损伤,鼻炎症状评分、血清IgE、IL-17及IL-18水平、鼻黏膜组织CD19、CD23阳性细胞比例、鼻黏膜组织TLR4、核内NF-κB p65、TNF-α蛋白表达水平显著升高(P<0.05);与模型组相比,柚皮素干预组大鼠鼻黏膜组织病理损伤均减轻,鼻炎症状评分、血清IgE、IL-17及IL-18水平、鼻黏膜组织CD19、CD23阳性细胞比例、鼻黏膜组织TLR4、核内NF-κB p65、TNF-α蛋白表达水平均降低,且柚皮素高剂量组大鼠各指标改变程度更强;与柚皮素高剂量组相比,柚皮素高剂量+LPS组大鼠鼻黏膜组织病理损伤加重,鼻炎症状评分、血清IgE、IL-17及IL-18水平、鼻黏膜组织CD19、CD23阳性细胞比例、鼻黏膜组织TLR4、核内NF-κB p65、TNF-α蛋白表达水平显著升高(P<0.05)。结论柚皮素可通过抑制TLR4/NF-κB/TNF-α通路表达而减轻超敏反应及炎症,从而缓解鼻黏膜组织损伤,改善AR大鼠症状。

蟾蜍TNFRSF11b、TNFRSF9基因克隆及生物信息学分析

目的:克隆中华蟾蜍肿瘤坏死因子受体超家族(TNFRSF)BbgTNFRSF11b、BbgTNFRSF9基因的全长序列,并分析其序列特征。方法:根据转录组测序所得的TNFRSF11b、TNFRSF9基因片段设计特异性引物,以中华蟾蜍蟾皮为材料,利用逆转录聚合酶链式反应(PCR)技术获得BbgTNFRSF11b、BbgTNFRSF9基因全长互补脱氧核糖核酸(cDNA)序列,并采用生物信息学手段分析其序列特征,通过实时荧光定量PCR方法检测BbgTNFRSF11b、BbgTNFRSF9基因在中华蟾蜍7种组织/器官中的表达情况。结果:克隆获得蟾蜍BbgTNFRSF11b基因的全长cDNA序列为1233bp,编码410个氨基酸,理论相对分子质量为46.97kDa,等电点为8.64,不存在跨膜区及信号肽,为亲水性蛋白,具有多个磷酸化位点。BbgTNFRSF9基因的全长cDNA序列为829bp,编码247个氨基酸,理论相对分子质量为67.874kDa,理论等电点为5.12,存在信号肽,为疏水性蛋白,具有多个磷酸化位点,进化树分析发现BbgTNFRSF11b、BbgTNFRSF9基因与其他动物TNFRSF11b、TNFRSF9基因相似度不高,表明该基因虽有特殊结构域但不同物种间差异较大。组织表达分析显示,BbgTNFRSF11b、BbgTNFRSF9基因在耳后腺中表达显著高于其他部位。结论:成功获得蟾蜍BbgTNFRSF11b、BbgTNFRSF9基因序列,掌握其序列特征,为后续深入研究该蛋白的功能提供参考。

探究芩柏加减方通过NF-κB/TNF-α信号通路对溃疡性结肠炎的保护机制

目的通过网络药理学和动物实验探讨芩柏加减方通过核因子κB(nuclear factorκB,NF-κB)/肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)信号通路对溃疡性结肠炎(ulcerative colitis,UC)的保护机制。方法将48只SPF级SD大鼠利用2,4,6-三硝基苯磺酸(trinitrobenzene sulfonic acid,TNBS)建立UC模型,造模成功后随机分为模型组(以生理盐水灌肠)、奥沙拉嗪组(以200 mg/kg奥沙拉嗪灌肠)、芩柏加减方低剂量组(以1.43 g/kg芩柏加减方灌肠)和芩柏加减方高剂量组(以2.86 g/kg芩柏加减方灌肠),每组12只,每天灌肠2次,连续14 d,另取12只正常饲养的SD大鼠作为正常对照组。于给药第1、7、14天每组随机取4只大鼠进行麻醉采血并取结肠组织。采用免疫组织化学法检测结肠组织中核因子κB亚基p65亲和肽(nuclear factorκB p65,NF-κB p65)、TNF-α、白细胞介素-10(interleukin-10,IL-10)、半胱氨酸天冬氨酸蛋白酶-3(Caspase-3)的表达来评价芩柏加减方对UC大鼠的影响,同时采用TUNEL荧光法检测评价大鼠结肠组织细胞调亡情况;利用ELISA法评价血清中细胞因子NF-κB p65、TNF-α、IL-10、Caspase-3的水平差异变化情况。结果通过网络药理学富集分析发现芩柏加减方中5种主要活性成分与UC成明显相关性且与NF-κB p65/TNF-α信号通路密切相关。经UC造模发现:与正常对照组比较,模型组大鼠结肠组织中TNF-α、Caspase-3和NF-κB p65水平均升高(P<0.05),IL-10水平显著降低(P<0.05)。HE染色结果显示,正常对照组大鼠结肠腺体排列整齐,未见炎性浸润;模型组的结肠腺体排列紊乱,肌底膜断裂且伴有明显的炎性浸润,凋亡指数相比正常对照组均升高(P<0.05);奥沙拉嗪组及芩柏加减方低、高剂量组随着给药时间的延长,结肠病理损伤均得到不同程度的缓解。经检测给药14 d后,血清中TNF-α、Caspase-3、IL-10和NF-κB p65水平结果与组织病理检测结果高度一致性。相比模型组,奥沙拉嗪组、芩柏加减方高剂量组大鼠血清中TNF-α、Caspase-3和NF-κB p65含量明显降低(P<0.05),IL-10水平显著升高(P<0.05)。结论芩柏加减方对TNBS诱导的UC模型具有一定保护作用,能降低细胞凋亡指数,可能是通过调节NF-κB/TNF-α信号通路来降低炎症因子分泌、减轻机体炎症反应、提高抗炎因子表达、促进黏膜愈合来实现。

针灸调控TNF-α/TLR4/NF-κB改善顺铂小鼠肝损伤的作用机制

目的通过观察针灸对顺铂(Cisplatin,DDP)所致肝损伤小鼠与肝细胞炎症反应相关的肿瘤坏死因子-α(TNF-α)、Toll样受体4(TLR4)和核因子-κB(NF-κB)表达含量的变化,以及血清中谷丙转氨酶(ALT)、谷草转氨酶(AST)和谷氨酸脱氢酶(GLDH)活性变化,并结合小鼠肝脏病理变化,阐释针灸对DDP引起肝损伤小鼠的作用机制。方法选用SPF雄性KM小鼠,随机分为4组,按照10 mL·kg^(-1)空白组腹腔注射0.9%NaCl溶液,其余3组按照体质量腹腔注射同剂量DDP溶液,24 h后模型成功。干预组采用“大椎”“肝俞”“肾俞”“足三里”,分别给予针刺和艾灸干预,其余两组陪同固定不干预,每天1次,持续5天。断食1天后取材,生化检测血清中ALT、AST、GLDH含量,Western blot测定肝脏中TNF-α、TLR4、NF-κB蛋白表达含量。结果与空白组比,模型组血清中ALT、AST、GLDH活性增高,肝脏中TNF-α、TLR4、NF-κB蛋白表达升高,均有统计学意义(P<0.05)。与模型组比,针刺组和艾灸组ALT、AST、GLDH活性和TNF-α、TLR4、NF-κB蛋白表达均降低,均有统计学意义(P<0.05)。与针刺组比,艾灸组血清中ALT、AST活性降低,GLDH活性升高,TNF-α蛋白表达降低,但均无统计学意义(P>0.05),TLR4、NF-κB蛋白表达降低,有统计学意义(P<0.05)。结论针灸可能通过抑制TNF-α、TLR4的过度表达,从而降低NF-κB过度表达,减少炎症因子的过度释放,从而减轻DDP小鼠肝脏炎症的发生,降低DDP对肝脏产生的毒副作用,对肝脏起到有效的保护作用。

山楂原花青素下调Traf6/NF-κB信号通路诱导肝癌细胞凋亡

目的:探讨山楂原花青素(hawthorn procyanidins,HPC)诱导肝癌细胞Huh-7增殖和促进其凋亡的机制。方法:体外常规培养Huh-7细胞株和过表达Traf6基因Huh-7细胞株,配制0、5、10、20、40、80μg/mL不同质量浓度的HPC,采用细胞计数8(cell counting kit-8,CCK-8)试剂盒检测HPC对Huh-7细胞和过表达Traf6基因Huh-7细胞增殖的抑制作用,并选40μg/mL作为HPC实验质量浓度;利用实时定量聚合酶链反应检测Huh-7细胞中Traf6、NF-κB?p65、Bax和Caspase-3的mRNA表达水平;利用蛋白质免疫印迹法(Western blot)检测Huh-7细胞中Traf6、NF-κB p65、Bax和Caspase-3蛋白表达水平;利用Annexin V-FITC/PI凋亡检测试剂盒检测HPC对Huh-7细胞凋亡的影响。结果:HPC质量浓度大于20μg/mL时可以明显抑制Huh-7细胞增殖,并随HPC质量浓度升高Huh-7细胞的相对存活率显著降低(P<0.05);Traf6蛋白过表达可以促进Huh-7细胞的增殖。与空白对照组比较,40μg/mL的HPC可以显著抑制肿瘤坏死因子受体相关因子6(tumor necrosis factor receptor associated factor 6,Traf 6)和核转录因子-κB(nuclear factor kappa B,NF-κB) p65 mRNA和蛋白表达(P<0.05),显著增强Bax和Caspase-3 mRNA和蛋白表达(P<0.05),显著促进Huh-7细胞的凋亡(P<0.05)。结论:HPC可以通过下调Traf6/NF-κB信号通路诱导肝癌细胞凋亡。

包虫抗原B通过肿瘤坏死因子受体2调控巨噬细胞极化对小鼠免疫性血小板减少症的改善作用

目的:探讨包虫抗原B (HA-B)对小鼠免疫性血小板减少症(ITP)的改善作用,阐明其相关作用机制。方法:将野生型(WT)和肿瘤坏死因子受体2 (TNFR2)基因敲除(TNFR2^(-/-))的C57/B6小鼠分为WT对照组、WT-ITP模型组、WT-HA-B组、TNFR2^(-/-)对照组、TNFR2^(-/-)ITP模型组和TNFR2^(-/-)HA-B组,检测各组小鼠体质量、脏器指数和血常规指标,采用流式细胞术检测各组小鼠外周血中M2巨噬细胞百分率,酶联免疫吸附试验(ELISA)法检测各组小鼠血清中精氨酸酶1(Arg1)、白细胞介素10 (IL-10)、诱导型一氧化氮合成酶(iNOS)和白细胞介素6 (IL-6)水平,采用实时荧光定量PCR (RT-qPCR)法检测各组小鼠骨髓来源巨噬细胞(BMDM)中Arg1、IL-10、iNOS和IL-6表达水平。分别将WT对照组和TNFR2^(-/-)对照组小鼠BMDM诱导为M2巨噬细胞(WT M2组和TNFR2^(-/-)M2组),加入HA-B (WT M2+HA-B组和TNFR2^(-/-)M2+HA-B组),采用RT-qPCR法检测各组细胞中Arg1和IL-10mRNA表达水平。结果:分别与WT对照组和TNFR2^(-/-)对照组比较,WT-ITP模型组和TNFR2^(-/-)ITP模型组小鼠体质量降低(P<0.05),脾脏和胸腺指数升高(P<0.05),血小板和红细胞数量减少(P<0.05),血红蛋白水平降低(P<0.05),白细胞数量增加(P<0.05),凝血时间延长(P<0.05);外周血中M2巨噬细胞百分率降低(P<0.05),血清中Arg1和IL-10水平降低(P<0.05),iNOS和IL-6水平升高(P<0.05);BMDM中Arg1和IL-10 mRNA表达水平降低(P<0.05),iNOS和IL-6 mRNA表达水平升高(P<0.05)。与WT-ITP模型组比较,WT-HA-B组小鼠体质量增加(P<0.05),脾脏和胸腺指数降低(P<0.05),血小板和红细胞数量增加(P<0.05),血红蛋白水平升高(P<0.05),白细胞数量减少(P<0.05),凝血时间缩短(P<0.05);外周血中M2巨噬细胞百分率升高(P<0.05),血清中Arg1和IL-10水平升高(P<0.05),iNOS和IL-6水平降低(P<0.05);BMDM中Arg1和IL-10 mRNA表达水平升高(P<0.05),iNOS和IL-6 mRNA表达水平降低(P<0.05)。与WT-HA-B组比较,TNFR2^(-/-)HA-B组小鼠体质量降低(P<0.05),脾脏和胸腺指数升高(P<0.05),血小板和红细胞数量减少(P<0.05),血红蛋白水平降低(P<0.05),白细胞数量增加(P<0.05),凝血时间延长(P<0.05);外周血中M2巨噬细胞百分率降低(P<0.05),血清中Arg1和IL-10水平降低(P<0.05),iNOS和IL-6水平升高(P<0.05);BMDM中Arg1和IL-10mRNA表达水平降低(P<0.05),iNOS和IL-6 mRNA表达水平升高(P<0.05)。与WT M2组比较,WT M2+HA-B组M2巨噬细胞中Arg1和IL-10 mRNA表达水平升高(P<0.05);与WT M2+HA-B组比较,TNFR2^(-/-)M2+HA-B组M2巨噬细胞中Arg1和IL-10 mRNA表达水平降低(P<0.05)。结论:HA-B可通过TNFR2促进巨噬细胞M2极化,进而发挥治疗ITP的作用。

肿瘤坏死因子受体超家族成员TNFRSF19的研究进展

肿瘤坏死因子受体超家族(tumor necrosis factor receptor superfamily,TNFRSF)通过与肿瘤坏死因子超家族成员相互作用调节免疫应答,调控细胞生存、增殖和分化等。TNFRSF19是TNFRSF家族的新成员之一,主要表达于上皮细胞、毛囊和大脑组织细胞中。近年来相关研究表明,TNFRSF19在调节神经系统发育和维持干细胞干性中发挥重要生理功能。TNFRSF19在不同肿瘤中发挥截然相反的促癌或抑癌功能,其功能与肿瘤的组织来源密切相关。本文就TNFRSF19的生理功能及其在癌症中的最新研究进展进行简要综述。

儿童病毒性肺炎血清肿瘤坏死因子超家族成员14、甲壳质酶蛋白40、可溶性白细胞介素2受体水平变化与短期预后的相关性

目的探讨儿童病毒性肺炎血清肿瘤坏死因子超家族成员14(TNFSF14)、甲壳质酶蛋白40(YKL-40)、可溶性白细胞介素2受体(sIL-2R)水平变化与短期预后的关系。方法选取2019年1月—2020年6月在西安国际医学中心医院儿科接受治疗的病毒性肺炎患儿120例(观察组)。另选取这一时期在该院体检健康儿童80例(对照组)。采用酶联免疫吸附法检测血清TNFSF14、YKL-40、sIL-2R水平,并分析其与患者病情程度及预后的关系。采用受试者工作特征(ROC)曲线,分析TNFSF14、YKL-40、sIL-2R对病毒性肺炎患儿临床预后的诊断效能。结果观察组血清TNFSF14、YKL-40、sIL-2R水平分别均高于对照组,差异有统计学意义(P<0.05)。重度组血清TNFSF14、YKL-40、sIL-2R水平分别高于轻度组,差异有统计学意义(P<0.05)。预后不良组血清TNFSF14、YKL-40、sIL-2R水平分别高于预后良好组,差异有统计学意义(P<0.05)。Pearson相关性分析结果显示,血清TNFSF14、YKL-40、sIL-2R分别与急性生理学和慢性健康状况评价Ⅱ(APACHEⅡ)的评分呈正相关(P<0.05)。ROC曲线结果示:血清TNFSF14、YKL-40、sIL-2R联合检测预测患儿不良预后的AUC为0.921(95%CI:0.867~0.984,P<0.001),灵敏度和特异度分别为0.905和0.816。多因素logistic回归分析结果示:APACHEⅡ评分(95%CI:1.001~3.268,P=0.005)及血清CRP(95%CI:1.755~6.143,P=0.001)、TNFSF14(95%CI:1.427~5.619,P=0.001)、YKL-40(95%CI:1.109~3.525,P<0.001)、sIL-2R(95%CI:1.265~4.173,P=0.002)是病毒性肺炎患儿不良预后的独立危险因素。结论血清TNFSF14、YKL-40、sIL-2R水平变化与病毒性肺炎患儿的病情严重程度及临床预后密切相关,也是影响患儿不良预后的重要危险因素,且对评估患儿的临床结局有较高参考价值。

NF-κB抑制剂对烫伤脓毒症大鼠致炎/抗炎细胞因子表达的影响

目的 观察NF κB抑制剂对动物主要器官致炎 /抗炎细胞因子基因及蛋白表达的调节效应 ,并对其作用机制进行初步探讨。方法 34只动物随机分为正常对照组 (n=6 )、烫伤 (2 0 %TBSAⅢ度 )对照组 (n=6 )、烫伤后金葡菌感染组 (n=12 )和NF κB抑制剂二硫氨基甲酸酞吡咯烷 (PDTC)拮抗组 (n =10 ) ,检测动物肝、肾、肺组织中TNF α、IL 10基因及蛋白表达的改变。结果 烫伤脓毒症组 0 5～2h肝、肺、肾组织中TNF αmRNA表达迅速增强 ,同时各组织TNF α蛋白水平亦显著升高。PDTC早期干预对肺脏TNF αmRNA及蛋白水平影响不明显 ,但肝、肾组织其表达量在 2h均被显著抑制 (P <0 0 5或 0 0 1)。烫伤合并金葡菌感染后 2h大鼠肝、肺、肾组织IL 10mRNA与蛋白水平均显著升高 (P <0 0 5或 0 0 1) ,早期给予PDTC对肺、肾组织各时间点IL 10mRNA与蛋白表达均无明显影响 ,肝组织仅 2h呈现一定程度地下降。结论 NF κB抑制剂在有效拮抗G+ 菌脓毒症TNF α等致炎介质的同时 ,对机体并存的抗炎细胞因子反应机制具有保护作用。