Study of tumor necrosis factor receptor in the inflammatory bowel disease

Ulcerative colitis(UC)and Crohn’s disease(CD)are part of Inflammatory Bowel Diseases(IBD)and have pathophysiological processes such as bowel necrosis and enteric neurons and enteric glial cells.In addition,the main inflammatory mediator is related to the tumor necrosis factor-alpha(TNF-α).TNF-αis a mediator of the intestinal inflammatory processes,thus being one of the main cytokines involved in the pathogenesis of IBD,however,its levels,when measured,are present in the serum of patients with IBD.In addition,TNF-αplays an important role in promoting inflammation,such as the production of interleukins(IL),for instance IL-1βand IL-6.There are two receptors for TNF as following:The tumor necrosis factor 1 receptor(TNFR1);and the tumor necrosis factor 2 receptor(TNFR2).They are involved in the pathogenesis of IBD and their receptors have been detected in IBD and their expression is correlated with disease activity.The soluble TNF form binds to the TNFR1 receptor with,and its activation results in a signaling cascade effects such as apoptosis,cell proliferation and cytokine secretion.In contrast,the transmembrane TNF form can bind both to TNFR1 and TNFR2.Recent studies have suggested that TNF-αis one of the main pro-inflammatory cytokines involved in the pathogenesis of IBD,since TNF levels are present in the serum of both patients with UC and CD.Intravenous and subcutaneous biologics targeting TNF-αhave revolutionized the treatment of IBD,thus becoming the best available agents to induce and maintain IBD remission.The application of antibodies aimed at neutralizing TNF-αin patients with IBD that induce a satisfactory clinical response in up to 60%of patients,and also induced long-term maintenance of disease remission in most patients.It has been suggested that anti-TNF-αagents inactivate the pro-inflammatory cytokine TNF-αby direct neutralization,i.e.,resulting in suppression of inflammation.However,anti-TNF-αantibodies perform more complex functions than a simple blockade.

Genes of tumor necrosis factors and their receptors and the primary open angle glaucoma in the population of Central Russia

AIM：To examine the association of genetic polymorphisms（-308）G/A TNFα,（＋250）A/G Ltα,（＋36）A/G TNFR1,（＋1663）A/G TNFR2 with the development of primary open angle glaucoma（POAG）among people in Central Russia.METHODS：The study sample included 443 individuals,of which 252 patients with POAG and 191 individuals in the control group.Genotyping of（-308）G/A TNFα,（＋250）A/G Ltα,（＋36）A/G TNFR1,（＋1663）A/G TNFR2 was performed using polymerase chain reaction.The distribution of alleles and genotypes of the studied DNA markers in the groups was examined by 2×2 contingency tables andχ2with the Yates’s correction for continuity and odds ratios（OR）with95%confidence intervals（CI）.RESULTS：Allele（-308）G TNFα（Р=0.01,OR=1.78,95%CI1.12-2.85）was identified as a risk factor for POAG.Homozygotes（-308）AA TNFαare at a lowest risk for development of the disease（Р=0.01,OR=0.0005）.The following combination of genetic variants of cytokines were associated with a reduced risk of POAG：（＋1663）A TNFR2 and（＋250）G Ltα（OR=0.34）CONCLUSION：Genetic polymorphisms（-308）G/A TNFα,（＋250）A/G Ltα,（＋1663）A/G TNFR2 associated with the development of POAG in the population of Central Russia.

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 factor alpha receptor 1 deficiency in hepatocytes does not protect from non-alcoholic steatohepatitis, but attenuates insulin resistance in mice

BACKGROUND End-stage liver disease caused by non-alcoholic steatohepatitis(NASH)is the second leading indication for liver transplantation.To date,only moderately effective pharmacotherapies exist to treat NASH.Understanding the pathogenesis of NASH is therefore crucial for the development of new therapies.The inflammatory cytokine tumor necrosis factor alpha(TNF-α)is important for the progression of liver disease.TNF signaling via TNF receptor 1(TNFR1)has been hypothesized to be important for the development of NASH and hepatocellular carcinoma in whole-body knockout animal models.AIM To investigate the role of TNFR1 signaling in hepatocytes for steatohepatitis development in a mouse model of diet-induced NASH.METHODS NASH was induced by a western-style fast-food diet in mice deficient for TNFR1 in hepatocytes(TNFR1ΔHEP)and their wild-type littermates(TNFR1fl/fl).Glucose tolerance was assessed after 18 wk and insulin resistance after 19 wk of feeding.After 20 wk mice were assessed for features of NASH and the metabolic syndrome such as liver weight,liver steatosis,liver fibrosis and markers of liver inflammation.RESULTS Obesity,liver injury,inflammation,steatosis and fibrosis was not different between TNFR1ΔHEP and TNFR1fl/fl mice.However,Tnfr1 deficiency in hepatocytes protected against glucose intolerance and insulin resistance.CONCLUSION Our results indicate that deficiency of TNFR1 signaling in hepatocytes does not protect from diet-induced NASH.However,improved insulin resistance in this model strengthens the role of the liver in glucose homeostasis.

Study on effect of imbalance of TRAF-6, IRAK-1 and NALP3 inflammatory factors in patients with gouty arthritis

Objective:To explore the effect of imbalance of tumor necrosis factor receptor related factor-6(TRAF-6),interleukin 1 receptor associated kinase-1(IRAK-1)and neutrophil alkaline phosphatase-3(NALP3)in patients with gouty arthritis.Methods:The retrospective experiment was conducted on 105 patients with gouty arthritis admitted to our hospital(47 patients with acute onset and 58 patients with remission,namely group A and group B);meanwhile,another 61 healthy volunteers were selected for control,namely group C.The enrolling of the three groups was dated from May 2017 to May 2018,and TRAF-6,IRAK-1 and NALP3 of all subjects were tested through real-time fluorescence quantification(RT-PCR),and the correlation between the three inflammatory factors and gouty arthritis was compared.Results:1)Through treatment,ESR,BUA and total addiment in group A and B were higher than those in group C,among which the three indicators in group A were higher than those in group B(P<0.05),while CRP was lower than that of group C,and the two indicators in group A were lower than those in group B(P<0.05).2)There was no significant difference in the relative expression of TRAF-6 mRNA between group A and group B before treatment(P>0.05),significantly lower than group C(P<0.05);the above indicators of group A and group B were improved to some extent after treatment,but group A was still lower than group B(P<0.05),and the degree of improvement of group A was also lower than that of group C(P<0.05),while the degree of improvement of group B was not significantly different from that of group C(P>0.05).3)The relative expression level of IRAK-1mRNA in group A and group B before treatment showed no significant difference(P>0.05),but was also lower than that in group C(P<0.05).The relative expression level of IRAK-1mRNA in group A and group B increased to some extent after treatment,with group A significantly lower than group C(P<0.05),and group B showed no significant difference compared with group C(P>0.05).4)The relative expression level of NALP-3 mRNA in group A and group B showed no significant difference(P>0.05)before treatment,significantly higher than that in group C(P<0.05);the relative expression of NALP-3 mRNA in group A was not significantly decreased(P>0.05)after treatment,while that in group B was significantly decreased after treatment(P<0.05),indicating significant different compared with group A and group C.5)There was no correlation between)TRAF-6,ESR,CRP and total addiment(P>0.05);IRAK-1 was negatively correlated with CRP,BUA and total addiment(P<0.05);NALP-3 was negatively correlated with ESR and CRP(P<0.05).Conclusion:TRAF-6,IRAK-1 and NALP-3 are all under abnormal expression in the developing of new gouty arthritis,acting as important participants in promoting the occurrence,development and outcome of illness states,so the intervening measures should be taken.

Moxibustion Treatment Restoring the Intestinal Epithelium Barrier in Rats with Crohn's Disease by Down-Regulating Tumor Necrosis Factor Alpha,Tumor Necrosis Factor Receptor 1,and Tumor Necrosis Factor Receptor 2

Objective： To investigate whether moxibustion regulates tumor necrosis factor alpha （TNF-α）, tumor necrosis factor receptor 1 （TNFR1）, and TNFR2 in the intestinal mucosa and to explore whether moxibustion could be used by means of this mechanism, to repair the intestinal epithelium barrier disruption in Crohn＇s disease （CD）. Methods： The CD rat models were established by tdnitrobenzene sulfonic acid （TNBs）, randomly divided into a model control （MC） group, an herb-partition moxibustion （HPM） group, a mild-warm moxibustion （MWM） group, and a salicylazosulfapyridine （SASP） group, and all were compared with a normal control （NC） group. The HPM and MWM groups were treated by moxibustion at Tianshu （ST25） and Qihai （RN6） for 14 days, and the SASP group obtained the SASP solution orally for the same pedod of time. The intestinal epithelium morphology and TNF-α, TNFR1, and TNFR2 contents were observed by the transmission electron microscopy and enzyme linked immunosorbent assay. Results： The sevedty of morphological changes in CD intestinal epithelium was obviously improved, and the levels of TNF-α, TNFR1, and TNFR2 in the intestinal mucosa all significantly decreased in the HPM and MWM groups. However, there were no significant differences between the HPM and MWM groups. Conclusion： The moxibustion therapies （HPM and MWM） could reduce intestinal inflammation and restore intestinal epithelium barrier disruption in CD, which might be due to down- regulating TNF- α, TNFR1, and TNFR2 in intestinal mucosa and improving intestinal epithelium morphology.

Knockout of the tumor necrosis factor a receptor 1 gene can up-regulate erythropoietin receptor during myocardial ischemia-reperfusion injury in mice

Background Tumor necrosis factor a receptor 1 （TNFaR1） plays an important role in the signal pathway of apoptosis. The objective of this study was to investigate the effects of TNFaR1 knockout on the up-regulation of erythropoietin receptor （Epo-R） and the coordinated anti-apoptosis functions during myocardial ischemia-reperfusion injury in mice. Methods The ischemia-reperfusion injury model for cardiomyocytes was performed by ligating the left circumflex branch artery of TNFaR1 knockout （P55/） C17 B6 mice, as well as wild-type （P55^＋/^＋） C17 B6 mice. Triphenyltetrazolium chloride （TTC） staining was performed to observe the damaged area of the heart. TUNEL staining and DNA fragmentation were used to identify apoptosis. Mitochondrial Bcl-2 and Bax as well as expression of Epo-R and its downstream genes （Jak-2, stat-5, Akt, lkB-a, HIF-1a） were measured by Western blotting. The gene knockout mice were assigned into those undergoing the apoptosis surgical model group （KO group）, and those subjected to sham operation （KOs group）. Similarly, wild-type mice were either exposed to the surgical model （WT group） or subject to a sham operation （WTs group）. Results The myocardial damage ratio of the wild-type group after the operation was significantly higher than that of the knockout group, （50.5±6.4）% vs （36.9±6.9）%, P 〈0.01. Similarly, TUNEL positive ratio of the wild-type group was significantly higher than that of the knockout group, （63.1±5.6）% vs （42.1±4.7）%, P〈0.01. The gray value ratios of Epo-R, Jak-2, stat-5, Akt, IkB-a, HIF-1 and mitochondrial Bcl-2 in the KO group were significantly higher than those of the WT group, P 〈0.05; however, mitochondrial Bax was significantly lower than that of the WT group significantly （P 〈0.05）. Conclusions Using the ischemia-reperfusion injury model in mice, cardiomyocytes of TNFaR1 knockouts exhibited anti-apoptotic characteristics. This information could be used to coordinate the prevention of myocardial apoptosis by up-regulating and activating the Epo-R pathway.

Gut region-specific TNFR expression:TNFR2 is more affected than TNFR1 in duodenal myenteric ganglia of diabetic rats

BACKGROUND Cytokines are essential in autoimmune inflammatory processes that accompany type 1 diabetes.Tumor necrosis factor alpha plays a key role among others in modulating enteric neuroinflammation,however,it has a dual role in cell degeneration or survival depending on different TNFRs.In general,TNFR1 is believed to trigger apoptosis,while TNFR2 promotes cell regeneration.The importance of the neuronal microenvironment has been recently highlighted in gut region-specific diabetic enteric neuropathy,however,the expression and alterations of different TNFRs in the gastrointestinal tract has not been reported.AIM To investigate the TNFR1 and TNFR2 expression in myenteric ganglia and their environment in different intestinal segments of diabetic rats.METHODS Ten weeks after the onset of hyperglycemia,gut segments were taken from the duodenum,ileum and colon of streptozotocin-induced(60 mg/body weight kg i.p.)diabetic(n=17),insulin-treated diabetic(n=15)and sex-and age-matched control(n=15)rats.Myenteric plexus whole-mount preparations were prepared from different gut regions for TNFR1/HuCD or TNFR2/HuCD double-labeling fluorescent immunohistochemistry.TNFR1 and TNFR2 expression was evaluated by post-embedding immunogold electron microscopy on ultrathin sections of myenteric ganglia.TNFRs levels were measured by enzyme-linked immunosorbent assay in muscle/myenteric plexus-containing(MUSCLE-MP)tissue homogenates from different gut segments and experimental conditions.RESULTS A distinct region-dependent TNFRs expression was detected in controls.The density of TNFR1-labeling gold particles was lowest,while TNFR2 density was highest in duodenal ganglia and a decreased TNFRs expression from proximal to distal segments was observed in MUSCLE-MP homogenates.In diabetics,the TNFR2 density was only significantly altered in the duodenum with decrease in the ganglia(0.32±0.02 vs 0.45±0.04,P<0.05),while no significant changes in TNFR1 density was observed.In diabetic MUSCLE-MP homogenates,both TNFRs levels significantly decreased in the duodenum(TNFR1:4.06±0.65 vs 20.32±3.1,P<0.001;TNFR2:11.72±0.39 vs 15.91±1.04,P<0.01),which markedly influenced the TNFR2/TNFR1 proportion in both the ganglia and their muscular environment.Insulin treatment had controversial effects on TNFR expression.CONCLUSION Our findings show diabetes-related region-dependent changes in TNFR expression and suggest that TNFR2 is more affected than TNFR1 in myenteric ganglia in the duodenum of type 1 diabetic rats.

Tumor necrosis factor alpha affect hydrocortisone expression in mice adrenal cortex cells mainly through tumor necrosis factor alpha-receptor 1

Background Tumor necrosis factor alpha （TNF-α） is important in promoting relative adrenal insufficiency （RAI） due to systemic inflammatory response syndrome （SIRS）. We identified the TNF-α receptor involved in the inhibition of adrenal corticotrophin （ACTH）-stimulated hydrocortisone release by studying the expression of TNF-α receptors in adrenal cortex Y1 cells and the effect of downregulating TNF receptors on ACTH-stimulated hydrocortisone release. Methods We used real-time PCR and immunocytochemistry to evaluate the expression of TNF receptors on Y1 cells. TNF-receptor 1 （TNF-R1） DNA fragments corresponding to the short hairpin RNA （shRNA）-sequences were synthesized and cloned into pcDNATM 6.2-GW/EmGFP expression vector. Knockdown efficiency of TNF-R1 expression was evaluated in miRNA transfected and mock-miRNA transfected Y1 cells by quantitative real-time PCR （Q-PCR）. Hydro- cortisone expression levels were determined in TNF-Rl-knockdown and control Y1 cells treated with TNF-α and ACTH. Results Mouse adrenal cortex Y1 cells were positive for type I TNF-R1, but not type II TNF-receptor （TNF-R2）. Blocking TNF-R1 expression resulted in loss of TNF-a-mediated inhibition of ACTH-stimulated hydrocortisone expression suggesting a role for the TNF-R1 related signaling pathway in ACTH-stimulated hydrocortisone synthesis. Conclusion The inhibitory effect of TNF-α on ACTH-stimulated hydrocortisone synthesis was mediated via TNF-R1 in adrenal cortex.

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.

Co-Inhibitors of Second Signal of Lymphocyte Response in Human Renal Transplants: PD-L2, GITR, and ILT-2/3/5 Positive Cells from Aspiration Biopsies Associate with Acute Rejection-Freedom

Following organ transplantation, the outcome of the encounter between an APC and a T lymphocyte is strongly dependent on the presence of costimulatory and co-inhibitory molecules, the former associated with allograft rejection and the latter with allograft acceptance. We evaluated the expression of PD-L2, GITR, ILT-2/3/5, and ILT-4 on graft-infiltrating cells procured by Fnab from human KTx under different immunosuppressive regimens. Methods: Fnab biopsies were performed on days 7 or 14 - 30 in stable KTx and on the day of acute rejection diagnosis. Cytopreparations were studied by the enzymatic avidin biotin complex staining. Results: Acute rejection group showed a significant down-regulated expression of PD-L2, GITR, and ILT-2/3/5 as compared to stable group, while for ILT-4 we did not find significant difference. Anti-IL2αR and rapamicyn treatment trend to down-regulate ILT-4 expression, although meaningless. A significantly positive correlation was observed between PD-L2 and GITR expression in Fnab. The PPV for acute rejection diagnosis for both PD-L2 and GITR was clearly above 0.8. Conclusions: Our findings point to an early entrance of cells expressing PD-L2, GITR and ILT-2/3/5 inside human KTx who are going to remain rejection-free. Both PD-L2 and GITR shared a high ability to rule-in and rule-out acute rejection.

Role of Osteoprotegerin and Receptor Activator of Nuclear Factor-kB Ligand in Bone Loss Related to Advanced Chronic Obstructive Pulmonary Disease

Background： Osteoporosis is a common complication of chronic obstructive pulmonary disease （COPD）. Recent clinical and biological researches have increasingly delineated the biomolecular pathways of bone metabolism regulation in COPD. We extended this work by examining the specific association and potential contribution of the osteoprotegerin （OPG）/receptor activator of nuclear factor-riB ligand （RANKL） axis to the pathogenesis of osteoporosis in advanced COPD. The aim of this study was to assess the relationships of serum OPG, RANKL, and tumor necrosis factor-alpha （TNF-a） with bone turnover in men with very severe COPD. Methods： Pulmonary function, T-score at the lumbar spine （LS） and femoral neck （FN）, serum OPG, RANKL, soluble receptor of tumor necrosis factor-alpha-1 and 11 （sTNFR-I, sTNFR-II）, osteocalcin （OC）, and β-CrossLaps （βCL） levels were measured in 45 men with very severe stage COPD and 36 male non-COPD volunteers. COPD patients and healthy controls were compared using all independent t-test and Mann-Whitney U-test. The Pearson coefficient was used to assess the relationships between variables. Results： OPG and OC were lower in male COPD patients than in control subjects whereas RANKL, serum ~CL, TN F-o~, and its receptors were higher. OPG directly correlated with forced expiratory volume in I s （FEVI） % predicted （r = 0.46, P 〈 0.005）, OC （r= 0.34, P 〈 0.05）, LS （I.= 0.56, P 〈 0.001 ）, and FN T-score （r= 0.47, P 〈 0.01 ）. In contrast, serum RANKL inversely associated with LS and FN T-score （r = -0.62, P 〈 0.001 and r = -0.48, P 〈 0.001 ） but directly correlated with [3CL （r = 0.48, P 〈 0.001 ）. In addition, OPG was inversely correlated with RANKL （r = -0.39, P 〈 0.01 ）, TNF-a （r = -0.56, P 〈 0.001 ）, and sTNFR-I （r = -0.40, P 〈 0.01 ）. Conelusion： Our results suggest that serum OPG and RANKL levels are inversely associated with bone loss in men with advanced stage COPD.

Bionic immunoactivator copresenting autophagy promoting and costimulatory molecules for synergistic cancer immunotherapy

Immunotherapy has great promise in improving malignant tumor treatment.However,the efficacy of existing strategies is often limited by the immunosuppressive environment.Here,we demonstrate an in situ bionic immunoactivator,PLT-Bec1/DTA-1,with possessed natural advantages of platelets for tumor recruitment and activation,on which DTA-1(CD357 monoclonal antibody)and Bec1 were tethered as combined immune boosters.PLT-Bec1/DTA-1,as a self-triggered release repository,can deliver the pre-tethered Bec1 and DTA-1 deeply through the secretion of platelet microparticles(PMPs),thereby cooperate tacitly and exhibit superiority in immune activation of dendritic cells(DCs)and T cells via autophagy inducibility,coupled with glucocorticoid-induced tumor necrosis factor receptor(GITR)-triggered T_(Reg) suppression,remodeled the immunosuppressive network of tumor microenvironment.PLT-Bec1/DTA-1 promoted antigen presentation and T cell proliferation,and alleviated the low activity state of bone marrow-derived dendritic cells(BMDCs)in tumor suppressive environment.PLT-Bec1/DTA-1 inhibited tumor recurrence(5-and 13-fold lower of control group in tumor volume)and CD8^(+)T/T_(Reg) ratio(6.3-and 8.8-fold vs.control group)in mouse tumor model after intravenous or subcutaneous administration.Also,PLT-Bec1/DTA-1 prevented tumor colonization in lung through in situ immune activation,and was slightly superior to the combined of Bec1 and PD-L1.Our findings highlight the promise of delivering immunostimulatory payloads via bionic carriers,eliciting automatic in situ activation of effector immune cells in tumor microenvironment for tumor eradication.All these results provide promising prospects into the application of immunoactivator in improving cancer synergistic immunotherapy to overcome the bottlenecks in clinic.

TNF-αTNFR1 Signaling is Required for the Full Expression of Acute and Chronic Itch in Mice via Peripheral and Central Mechanisms

Increasing evidence suggests that cytokines and chemokines play crucial roles in chronic itch. In the present study, we evaluated the roles of tumor necrosis factor-alpha （TNF-c0 and its receptors TNF receptor subtype-I （TNFR1） and TNFR2 in acute and chronic itch in mice. Compared to wild-type （WT） mice, TNFRl-knockout （TNFR1-KO） and TNFR1/R2 double-KO （DKO）, but not TNFR2-KO mice, exhibited reduced acute itch induced by compound 48/80 and chloroquine （CQ）. Application of the TNF-synthesis inhibitor thalidomide and the TNF-at antagonist etanercept dose-dependently suppressed acute itch. Intradermal injection of TNF-α was not sufficient to evoke scratching, but potentiated itch induced by compound 48/80, but not CQ. In addition, compound 48/80 induced TNF-α mRNA expression in the skin, while CQ induced its expression in the dorsal root ganglia （DRG） and spinal cord. Furthermore, chronic itch induced by dry skin was reduced by administration of thalidomide and etaner- cept and in TNFR1/R2 DKO mice. Dry skin induced TNF- expression in the skin, DRG, and spinal cord and TNFR1 expression only in the spinal cord. Thus, our findings suggest that TNF-c^-fNFR1 signaling is required for the full expression of acute and chronic itch via peripheral and central mechanisms, and targeting TNFR1 may be benefi- cial for chronic itch treatment.

Construction and expression of a Rev-dependent TNF-R1 expressing HIV-infected-cell injurious vectors

Background Rev is necessary for exporting unspliced and incompletely spliced intron containing HIV mRNAs and for HIV replication. The aim of this study is to develop a kind of selective suicide construct that can specifically and directly induce HIV infected cells into apoptosis based on the high affinity of Rev and Rev response element （RRE）. Methods Molecular-cloning technique was used to synthesis Rev dependent TNF-R1 expression construct pDM128-TNF-R1 （pT128） that contains RRE and TNFR1 gene. Restriction digestion, Polymerase Chain Reaction （PCR） and DNA sequencing were processed and the exactness and correctness of the inserted TNF-R1 gene in pT128 were confirmed repeatedly. The expression of pT128 co-transfected with different combination of other plasmids by calcium phosphate-DNA co-precipitation in Helas and by gene gun transfection in keratinocytes was further tested by flow-cytometry and cell counted under microscope. Results The new plasmid specifically expressed TNF-R1 in Helas when co-transfected with pRev but did not when without pRev. Indirect expression of TNF-R1 from pT128 was slower than the direct expression of that from Hu p60 TNFR1 in pDC302 （pT60） , but all those pT60 or pT128 transfected cells showed apoptosis at last while TNF-R1 was sufficiently expressed. Other kinds of Rev expression construct such as pAD8 and a chimeric HIV vaccine also can switched on the selective expression of pT128. Not only Rev-dependent expression in Helas, pT128 also normally expressed its TNF-R1 in keratinocytes. Co-transfected with prey or pAD8 that expressed Rev, pT128 expressed TNF-R1 and induced apoptosis of green fluorescent keratinocytes in skin explant. The number of green fluorescent keratinocytes co-transfected by pT128 plus prey or pAD8 was gradually outnumbered by that co-transfected by pT128 only. The difference was more significant after culturing for 72 hours. Conclusions Rev dependent pT128 is able to selectively induce apoptosis of HIV-infected or Rev-expressed target cells by expression of TNF-R1. The new strategy based on manipulation of the regulatory protein of HIV may be valuable in design of new HIV vaccine.

Identification and modulation of expression of a TNF receptor superfamily member 25 homologue in grass carp (Ctenopharyngodon idella)

The TNF receptor superfamily member 25(TNFRSF25)is part of the tumor necrosis factor receptor superfamily and contains a typical death domain.It is also known as DR3,TRAMP,LARD,WSL-1,Apo-3 and TR3,and has a vital role in regulating cell proliferation,differentiation and apoptosis.In this study,a homologue of the TNFRSF25 gene was identified in grass carp(Ctenopharyngodon idella).It encodes a transmembrane protein with an extracellular domain containing a cysteine-rich domain and an intracellular domain containing a death domain.It is an orthologue of fish TNFRSF1ALs and shares conserved gene synteny with human TNFRSF25.Expression studies showed that CiTNFRSF25 was constitutively expressed in the majority of fish tissues and can be modulated by interleukin 4/13B and infection by F.columnare.To our knowledge,this is the first report describing the existence of a TNFRSF25 homologue in teleost fish.