Development and verification of a detection method for activity of neutralizing antibody in ELISA antibody positive serum of rats immunized with recombinant human interleukin-1 receptor antagonist / 中国生物制品学杂志

@#Objective To develop and verify a method for detecting the activity of neutralizing antibodies in ELISA antibody positive serum of rats immunized with recombinant human interleukin-1 receptor antagonist(rhIL-1Ra). Methods The SD rats were subcutaneously immunized with 3,20 and 100 mg/kg rhIL-1Ra injection respectively,10 rats in each group,half male and half female,twice a day at an interval of at least 4 h between each dose for 13 consecutive weeks. The blood samples were collected from the jugular vein of rats during the administration period and the recovery period. The serum samples were isolated and detected for the antibody titers by ELISA,and the samples positive for rhIL-1Ra antibody were purified by Protein A chromatographic column. Based on,D10G4·1 cells biological activity assay,a method for the detection of neutralizing antibody activity was developed and verified for the specificity,sensitivity and precision. The neutralizing antibody activity of rhIL-1Ra antibody positive serum determined by ELISA was detected by using the developed method.Results With the increase of doses,the serum antibody titers of rats in various dose groups gradually increased,and there were still antibodies in the recovery period,and the titer was still high. Rabbit anti-rhIL-1Ra monoclonal antibody showed obvious neutralizing effect on rIL-1Ra,while rabbit anti-rIFN-2b monoclonal antibody had no dose-effect relationship with rIL-1Ra. The sensitivity of the method was 171. 93 μg/mL;The CVs of precision verification were not more than 20%. The positive antibody sera detected by ELISA all had neutralizing effect on rhIL-1Ra injection,which was consistent with the results detected by ELISA. Conclusion The method developed in this study has good specificity and high sensitivity in the detection of serum neutralizing antibody activity in rats immunized with rhIL-1Ra,which can be used to detect the serum neutralizing antibody activity of animals with rhIL-1Ra repeated administration.

Construction,expression,purification and quality identification of kanamycin resistant strain of recombinant human interleukin-1 receptor antagonist / 中国生物制品学杂志

@#Objective To construct a recombinant human interleukin-1 receptor antagonist(rhIL-1Ra)strain of kanamycin resistance,express,purify and identify rhIL-1Ra protein in order to reduce the risks of β-lactam antibiotics.Methods The ampicillin-resistant rhIL-1Ra(A-rhIL-1Ra)plasmid and PET-28a plasmid were used as templates to prepare the linearized vector and kanamycin gene fragment by PCR.The kanamycin resistant rhIL-1Ra plasmid(K-rhIL-1Ra)was constructed by homologous recombination,which was transformed into E.coli BL21(DE3)to construct the recombinant engineered bacteria after correct sequencing.The engineered bacteria were induced by IPTG and then purified by CM Bestarose Fast Flow and DEAE Bestarose Fast Flow column chromatography.The purified products were collected,and then detected for the purity by 15% SDS-PAGE,size-exclusion high-performance liquid chromatography(SEC-HPLC),and reversed-phase high-performance liquid chromatography(RP-HPLC),determined for the relative molecular mass by tandem mass spectrometry,identified for the specificity by Western blot,measured for the biological activity by reporter gene assay,and compared for the related impurities by liquid chromatography-mass spectrometry(LC-MS)analysis.Results Colony PCR and sequencing results showed that K-rhIL-1Ra plasmid was constructed correctly.The expressed K-rhIL-1Ra protein had a relative molecular mass of about 17 000,mainly existing in soluble form,and the expression amount accounted for more than 30% of the total bacterial proteins.The purity of K-rhIL-1Ra protein purified by two steps was 97% and 99%,the monomer content was 99.33%and 100%,and the chromatographic purity was 91.86% and 96.96%,respectively.The mass spectral molecular mass was consistent with that of the standard(A-rhIL-1Ra protein),and the protein reacted specifically relative with mouse antihuman IL-1Ra monoclonal antibody.The biological activity of the purified K-rhIL-1Ra protein was 1.29 × 105U/mg,and the chemical modification types of related proteins were the same as those of the standard(A-rhIL-1Ra protein).Conclusion The K-rhIL-1Ra strain was successfully constructed,and the expressed and purified protein was in line with the characteristics and quality standards of rhIL-1Ra,which lays a foundation for the study of rhIL-1Ra strain change and comparability

New progress on clinical characteristics and related mechanisms of chronic ocular graft-versus-host disease / 器官移植

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) could effectively treat multiple hematological diseases. At present, with persistent improvement of transplantation techniques and rapid development of economy, more and more patients with hematological diseases are able to survive for a long time due to allo-HSCT treatment. Chronic ocular graft-versus-host disease (coGVHD) is the most common ocular complication after allo-HSCT, which is primarily manifested with refractory dry eye. In severe cases, it may cause imbalance of ocular surface homeostasis and limbal stem cell insufficiency, further leading to a series of complications that threaten the visual function and eye health, such as corneal perforation and symblepharon, etc. It is highly difficult to cure these symptoms. At present, relevant studies of clinical manifestations, diagnostic criteria, treatment specification and pathogenesis of coGVHD have been gradually deepened within the international community. However, related research and the establishment of clinical specification are still in the primary stage in China. In this article, research progress on clinical characteristics and related mechanisms of coGVHD was reviewed, aiming to deepen the understanding of this disease by ophthalmologists, especially specialists in corneal and ocular surface diseases, and provide novel ideas for subsequent in-depth research.

Expression and clinical significance of IL-1β and IL-1β receptor antagonist in persistent pulmonary hypertension of the newborns secondary to sepsis / 国际儿科学杂志

Objective:To explore the expression and clinical significance of IL-1β and IL-1β receptor antagonist(IL-1ra)in persistent pulmonary hypertension of the newborn(PPHN)secondary to sepsis.Methods:The newborns with sepsis were enrolled in the Department of Neonatal Intensive Care Unit(NICU)of Xi′an Children′s Hospital from January 2018 to November 2020.The newborns with sepsis were divided into two groups: the newborns without PPHN( n=108)were the control group and the newborns with PPHN( n=44)were the experimental group.Clinical data, laboratory examination and bedside echocardiography of all the newborns were collected to analyze the differences between the two groups.The expression levels of IL-1β and IL-1ra in neonatal plasma of the two groups were detected by enzym-linked immunosorbination(ELISA), and their roles in neonatal sepsis with PPHN were further analyzed.The risk factors of neonatal sepsis with PPHN were analyzed by multivariate Logistic regression, and the early prediction value of the risk factors for neonatal sepsis with PPHN were evaluated by the receiver operating characteristic(ROC)curve. Results:There were no significant differences in gestational age[(39.11±0.55)w vs(38.85±0.72)w], birth weight[(3.30±0.49)kg vs(3.24±0.55)kg]and proportions of males[60(55.6%)vs 30(68.2%)]between the two groups( P>0.05). The right ventricular diameter[(9.57±0.35)mm], pulmonary artery pressure[(51.36±5.91)mmHg]and the level of N-terminal brain natriuretic peptide(NT-proBNP)[(25436.83±12343.18)ng/L)]significantly increased in the experimental group than those in the control group[(8.77±0.41)mm, (31.24±5.11)mmHg, (11267.09±4405.48)ng/L, respectively, P<0.05]. Before treatment, the expression levels of plasma IL-1β[(31.24±5.25)ng/L]and IL-1ra[(41.94±10.13)ng/L]in the experimental group were significantly higher than those in the control group[(18.27±4.47)ng/L, (21.47±8.76)ng/L, respectively, P<0.05]. The expression levels of plasma IL-1β[(10.46±3.17)ng/L]and IL-1ra[(10.58±2.94)ng/L]in the experimental group after treatment were significantly lower than those before treatment[(31.24±5.25)ng/L, (41.94±10.13)ng/L , respectively, P<0.05]. Multivariate Logistic regression analysis showed that IL-1β and NT-proBNP were the independent risk factors for neonatal sepsis with PPHN( P<0.05). ROC curve analysis showed that IL-1β and NT-proBNP had the good predictive value for the occurrence of neonatal sepsis with PPHN( P<0.05). IL-1β combined with NT-proBNP has the better predictive value for neonatal sepsis with PPHN. Conclusion:IL-1β combined with NT-proBNP have the high predictive value for PPHN of the newborns secondary to sepsis.

Anti-interleukin-1 treatment among patients with familial Mediterranean fever resistant to colchicine treatment. Retrospective analysis

ABSTRACT BACKGROUND: Up to 5% of familial Mediterranean fever (FMF) cases are unresponsive to colchicine, through resistance, side effects and toxicity. Anakinra is an alternative treatment for FMF patients whose disease remains uncontrolled with colchicine. We aimed to evaluate anti-interleukin-1 treatment regarding clinical findings, laboratory parameters and quality of life (QoL) among FMF patients presenting resistance and toxicity towards colchicine. DESIGN AND SETTING: Descriptive observational study at the rheumatology clinic, Adnan Menderes University Medical School, Aydın, Turkey. METHODS: Among the patients included, age, sex, MEFV genotypes, acute-phase reactants, hepatic/renal function tests, average colchicine dose, disease duration, attack frequency, attack duration, disease severity, proteinuria, amyloidosis and QoL were evaluated. Colchicine resistance was defined as > 6 typical episodes/year or > 3 per 4-6 months. Kolmogorov-Smirnov, Friedman and two-way analysis of variance tests were used for statistical analyses. RESULTS: Between 2015 and 2017, 14 FMF patients receiving anakinra were enrolled. The mean colchicine dose was 1.7 ± 0.3 mg/day before use of anakinra. Ten patients were attack-free after treatment, while three showed reductions of at least 50% in attack frequency, attack duration and disease severity. Proteinuria levels in all patients with renal amyloidosis decreased after treatment. QoL among patients with renal amyloidosis differed significantly from QoL among non-amyloidosis patients. Mean visual analogue scale scores significantly improved in both groups after use of anakinra. CONCLUSIONS: Use of anakinra reduced attack frequency and proteinuria and acute-phase reactant levels, and improved QoL, with only a few uncomplicated side effects among colchicine-resistant or intolerant FMF patients. Injection-site reactions of severity insufficient to require discontinuation of treatment were seen.

Summary of molecular mechanisms of spontaneous remission in gouty inflammation / 中国药理学通报

Acute gout subsides spontaneously within a certain period of time after onset, a process known as spontaneous remission of gouty inflammation. Its pathophysiological mechanism is related to the dynamic interaction of various components in the immune system, i. e. through the phagocytosis of autoimmune cells, the negative regulation of inflammatory mediators such as NLRP3 inflammasome, 1L-1R and Toll-like receptors, the recruitment of neutrophil extracellular traps at the site of crystal deposition and the formation of local chronic granuloma, to alleviate the necrotizing inflammation mediated by monosodium urate crystals. This review summarizes the molecular mechanism of spontaneous remission of gouty inflammation.

Conditioned Medium from Tonsil-Derived Mesenchymal Stem Cells Relieves CCl₄-Induced Liver Fibrosis in Mice

BACKGROUND: The liver is an organ with remarkable regenerative capacity; however, once chronic fibrosis occurs, liver failure follows, with high mortality and morbidity rates. Continuous exposure to proinflammatory stimuli exaggerates the pathological process of liver failure; therefore, immune modulation is a potential strategy to treat liver fibrosis. Mesenchymal stem cells (MSCs) with tissue regenerative and immunomodulatory potential may support the development of therapeutics for liver fibrosis. METHODS: Here, we induced hepatic injury in mice by injecting carbon tetrachloride (CCl₄) and investigated the therapeutic potential of conditionedmedium from tonsil-derivedMSCs (T-MSCCM). In parallel, we used recombinant human IL-1Ra,which, as we have previously shown, is secreted exclusively from T-MSCs and resolves the fibrogenic activation of myoblasts. Hepatic inflammation and fibrosis were determined by histological analyses using H&E and Picro-Sirius Red staining. RESULTS: The results demonstrated that T-MSC CM treatment significantly reduced inflammation as well as fibrosis in the CCl₄-injured mouse liver. IL-1Ra injection showed effects similar to T-MSC CM treatment, suggesting that T-MSC CM may exert anti-inflammatory and anti-fibrotic effects via the endogenous production of IL-1Ra. The expression of genes involved in fibrosis was evaluated, and the results showed significant induction of alpha-1 type I collagen, transforming growth factor beta, and tissue inhibitor of metalloproteases 1 upon CCl₄ injection, whereas treatment with T-MSC CM or IL-1Ra downregulated their expression. CONCLUSION: Taken together, these data support the therapeutic potential of T-MSC CM and/or IL-1Ra for the alleviation of liver fibrosis, as well as in treating diseases involving organ fibrosis.

Association of interleukin 1 receptor antagonist intron 2 variable number of tandem repeats polymorphism with vitiligo susceptibility in Gujarat population

Background: Vitiligo is a multifactorial, polygenic, autoimmune skin disorder caused by selective destruction of melanocytes. Interleukin 1 receptor antagonist intron 2 polymorphism was found to be associated with various autoimmune disorders. Aims: We aimed to investigate the association of interleukin 1 receptor antagonist intron 2 variable number of tandem repeats polymorphism (rs2234663) with vitiligo to assess interleukin 1 receptor antagonist transcript levels and to perform possible genotype–phenotype correlation. Methods: Three hundred and seven vitiligo patients and 316 controls were enrolled in the study, genotyping of interleukin 1 receptor antagonist rs2234663 was performed by polymerase chain reaction, and relative gene expression of interleukin 1 receptor antagonist was carried out in peripheral blood mononuclear cells from patients (n = 36) and controls (n = 36) by real-time-PCR. Results: A significant difference was observed in the frequency of interleukin 1 receptor antagonist *A (1/2) genotype among patients with active and stable vitiligo (P = 0.0172). Interleukin 1 receptor antagonist*A (2/2) genotype and allele frequencies were significantly different between SV patients and controls (P = 0.0246 and P = 0.0046, respectively). Significant difference was also observed for interleukin 1 receptor antagonist*A2 (allele) in active and stable vitiligo patients (P = 0.0060). However, other comparisons did not show any significant difference in genotype and allele frequencies. Moreover, interleukin 1 receptor antagonist*A (3/2) genotype was observed only in patients whereas interleukin 1 receptor antagonist*A (5/2) was observed only in controls. Gene expression analysis showed no significant difference in interleukin 1 receptor antagonist transcript levels in patients compared to controls (P = 0.5962). Interestingly, genotype–phenotype correlation analysis revealed that individuals with IL1RN*A (2/2) exhibited higher interleukin 1 receptor antagonist expression compared to other major genotypes interleukin 1 receptor antagonist*A (1/2) (P = 0.01) and interleukin 1 receptor antagonist*A (1/1) (P = 0.03). Limitations: More case-control studies on interleukin 1 receptor antagonist rs2234663 polymorphism and gene expression from different ethnic populations are required to explore the impact of interleukin 1 receptor antagonist in vitiligo susceptibility. Conclusion: Interleukin 1 receptor antagonist*A2 might be a risk factor for progressive vitiligo.

Protective effect of interleukin-1 receptor antagonist on intestinal ischemia-reperfusion induced injury in rats / 中国药理学通报

Aim To explore the effects of interleukin-1 receptor antagonist (IL-1Ra) on intestinal ischemia-reperfusion (I/R) induced injury in rats. Methods Thirty-five male SD rats were randomly divided into sham operation group (S), model group (I/R), dif-ferent dosage drug groups(C1,C2,C3). Rat intesti-nal I/R model was established via clamping the superi-or mesenteric artery (SMA). After 1 h of ischemia, the arterial clamps were released for 1 h of reperfusion. 10,20,50 mg·kg-1of IL-Ra was injected via caudal vein 15min before reperfusion. Results After 2 h of I/R,compared with S group,I/R group rats exhibited severe damage on the intestinal mucosa, increase in MDA content, decrease in SOD activity, and signifi-cant release of TNF-α, IL-1β, IL-6. The results showed that, following the injection of IL-1Ra after clipping superior mesenteric artery, damage of the in-testinal mucosa was obviously relieved in different dos-age drug groups. Furthermore, there was different de-gree of relief on oxidative stress and inflammatory re-sponses. Conclusion IL-1Ra showed obvious protec-tive effect on intestinal I/R induced injury by relieving oxidative stress and inflammatory response,and it may potentially be used in the clinical treatment of intestinal I/R injury.

A Case of Schnitzler's Syndrome without Monoclonal Gammopathy-Associated Chronic Urticaria Treated with Anakinra

Chronic urticaria may often be associated with interleukin (IL)-1-mediated autoinflammatory disease, which should be suspected if systemic inflammation signs are present. Here, we report a case of Schnitzler's syndrome without monoclonal gammopathy treated successfully with the IL-1 receptor antagonist anakinra. A 69-year-old man suffered from a pruritic urticarial rash for 12 years. It became aggravated episodically and was accompanied by high fever, arthralgia, leukocytosis, and an elevated C-reactive protein and erythrocyte sedimentation rate. The episodes each lasted for over one week. Neutrophilic and eosinophilic inflammation was found on skin biopsy. However, serum and urine electrophoresis showed no evidence of monoclonal gammopathy. The cutaneous lesions were unresponsive to various kinds of anti-histamines, systemic glucocorticoids, colchicine, cyclosporine, dapsone, and methotrexate, which were administered over a span of 3 years immediately preceding successful treatment. A dramatic response, however, was observed after a daily administration of anakinra. This observation suggests that the correct diagnosis of this case is Schnitzler's syndrome without monoclonal gammopathy. For an adult patient with refractory chronic urticaria and systemic inflammation, Schnitzler's syndrome could be considered as a possible differential diagnosis. Although the typical form of Schnitzler's syndrome exhibits the presence of monoclonal gammopathy as a diagnostic criterion, monoclonal gammopathy may be absent in an atypical form. In such a situation, an IL-1 antagonist should be effective for the management of chronic urticaria.

The Allele 2 of the VNTR Polymorphism in the Gene That Encodes a Natural Inhibitor of IL-1β, IL-1RA Is Favorably Associated With Chronic Otitis Media

OBJECTIVES: Chronic otitis media (COM) is followed by irreversible tissue damage and destruction of the middle ear structures, with the possibility of complications under the maintenance of inflammation. Inflammatory mediators such as cytokines play a crucial role in the initial stage of inflammation. The aim of this study was to evaluate the association of the polymorphisms in two innate immunity/inflammation cascade genes from interleukin-1 (IL-1) gene cluster with COM with regard to cholesteatoma. METHODS: In the cross-sectional case-control study, DNA samples were collected from 189 patients with COM and 119 controls from a population of Serbia. The +3953 C/T (rs1143634), TaqI polymorphism in interleukin-1 beta (IL-1β) gene and 86 bp variable number tandem repeat (VNTR, rs2234663) polymorphism in the IL-1 receptor antagonist (IL-1RA) gene were analyzed by polymerase chain reaction. RESULTS: The IL-1β TaqI polymorphism was not significantly different in patients compared with the control group. The significant difference between patients and controls was observed for both, genotype and allele frequencies of IL-1RA VNTR polymorphism (chi-square P < 0.01). We found that carriers of IL-1RA allele 2 (odds ratio, 0.47; 95% confidence interval, 0.29 to 0.76; P=0.004) have a favorable association with COM, using multivariate logistic analysis that included both polymorphisms, age and sex. The IL-1RA allele frequency distribution was significantly different with regard to cholesteatoma. CONCLUSION: The carriers of allele 2 of VNTR IL-1RA polymorphism had a decreased odds ratio for COM, which is in agreement with findings in other inflammatory disease and its previous association with higher IL-1RA levels. Possible down-regulation of IL-1 mediated proinflammatory signaling pathways via IL-1RA in COM as well as results of our study should be further investigated and replicated.

The beneficial effect of mesenchymal stem cells transplantation in combination with interleukin-1 receptor antagonist on post-hepatectomy liver failure / 中华肝胆外科杂志

Objective To study the synergetic effect and possible mechanism of transplanting mesenchymal stem cells (MSCs) in combination with interleukin-1 receptor antagonist (IL-1Ra) on acute liver failure (ALF).Methods MSCs transplantation combined with IL-1Ra was used for a swine model of ALF induced by 85％ total hepatectomy.The living conditions,blood samples and survival time were recorded or collected for analysis of hepatic function.Liver injury histology was analyzed.Hepatic cell regeneration and apoptosis were studied by immunohistochemistry staining of Ki67 and TUNELassays respectively.The expression levels of AKT and NF-κB were analyzed by Western blotting.Results The difference on the survival time between the model group and combined therapy group was statistically significant (P ＜ 0.05).Combined therapy displayed improvement not only in the serum biochemical conditions but also in the serum inflammatory cytokines.Furthermore,the observed hepatic histopathological score was significantly less compared to model group.In addition,the combined therapy group significantly inhibited the liver cell apoptosis and increased hepatic cell regeneration.Finally,a significant increase in AKT expression and decrease of NF-κB expression (P ＜ 0.05) were observed,which was consistent with their important roles in liver regeneration.Conclusion The combined therapy displayed a synergistic effect on liver regeneration,by promoting restoration and reconstruction of ALF,through regulation of inflammation and apoptosis signaling network.

The beneficial effect of mesenchymal stem cells transplantation in combination with interleukin-1 receptor antagonist on post-hepatectomy liver failure / 中华肝胆外科杂志

Objective To study the synergetic effect and possible mechanism of transplanting mesenchymal stem cells (MSCs) in combination with interleukin-1 receptor antagonist (IL-1Ra) on acute liver failure (ALF).Methods MSCs transplantation combined with IL-1Ra was used for a swine model of ALF induced by 85％ total hepatectomy.The living conditions,blood samples and survival time were recorded or collected for analysis of hepatic function.Liver injury histology was analyzed.Hepatic cell regeneration and apoptosis were studied by immunohistochemistry staining of Ki67 and TUNELassays respectively.The expression levels of AKT and NF-κB were analyzed by Western blotting.Results The difference on the survival time between the model group and combined therapy group was statistically significant (P ＜ 0.05).Combined therapy displayed improvement not only in the serum biochemical conditions but also in the serum inflammatory cytokines.Furthermore,the observed hepatic histopathological score was significantly less compared to model group.In addition,the combined therapy group significantly inhibited the liver cell apoptosis and increased hepatic cell regeneration.Finally,a significant increase in AKT expression and decrease of NF-κB expression (P ＜ 0.05) were observed,which was consistent with their important roles in liver regeneration.Conclusion The combined therapy displayed a synergistic effect on liver regeneration,by promoting restoration and reconstruction of ALF,through regulation of inflammation and apoptosis signaling network.

Intranasal Administration of Interleukin-1 Receptor Antagonist in a Transient Focal Cerebral Ischemia Rat Model

The interleukin-1 receptor antagonist (IL-1RA) is a potential stroke treatment candidate. Intranasal delivery is a novel method thereby a therapeutic protein can be penetrated into the brain parenchyma by bypassing the blood-brain barrier. Thus, this study tested whether intranasal IL-1RA can provide neuroprotection and brain penetration in transient cerebral ischemia. In male Sprague-Dawley rats, focal cerebral ischemia was induced by middle cerebral artery occlusion (MCAO) for 1 h. The rats simultaneously received 50 mg/kg human IL-1RA through the intranasal (IN group) or intraperitoneal route (IP group). The other rats were given 0.5 mL/kg normal saline (EC group). Neurobehavioral function, infarct size, and the concentration of the administered human IL-1RA in the brain tissue were assessed. In addition, the cellular distribution of intranasal IL-1RA in the brain and its effect on proinflammatory cytokines expression were evaluated. Intranasal IL-1RA improved neurological deficit and reduced infarct size until 7 days after MCAO (p<0.05). The concentrations of the human IL-1RA in the brain tissue 24 h after MCAO were significantly greater in the IN group than in the IP group (p<0.05). The human IL-1RA was confirmed to be co-localized with neuron and microglia. Furthermore, the IN group had lower expression of interleukin-1β and tumor necrosis factor-α at 6 h after MCAO than the EC group (p<0.05). These results suggest that intranasal IL-1RA can reach the brain parenchyma more efficiently and provide superior neuroprotection in the transient focal cerebral ischemia.

Therapeutic effects of recombinant human interleukin-1 receptor antagonist on toluene-2,4-diisocyanate-induced guinea pig allergic rhinitis model / 国际药学研究杂志

Objective To evaluate the effects of the recombinant human interleukin-1 receptor antagonist(rhIL-1ra)on a tol?uene-2,4-diisocyanate(TDI)-induced guinea pig allergic rhinitis (AR)model. Methods An AR model was established via sensiti?zation and challenge of two-step procedure using TDI in guinea pigs. Normal animals were treated only with the olive oil(TDI vehicle). Sixty adult guinea pigs were randomly divided into six groups(n=10):normal group,model group(rhIL-1ra vehicle),positive con?trol group(budesonide,25.6μg/kg),rhIL-1ra treated groups(rhIL-1ra 50,100 and 200μg/kg,respectively). From day 8 after sensi?tization,animals of all the groups were treated respectively with the agents or vehicle once a day for 14 days. During the observation pe?riod,the index of clinic score was recorded for every animal. At day 14 of the dosing,guinea pigs were sacrificed 30 min after the last TDI challenge and observation. Blood samples were taken from the abdominal aorta to prepare the serum for detection of histamine , and the nasal mucosase were dissected for histamine detection and histopathological observation. Results Compared with the guinea pigs in normal group,those in the model group exerted the typical symptoms of AR. It was shown that rhIL-1ra could improve nasal symptoms and cause a significant decrease in the instances of nasal sneezing as well. In addition,rhIL-1ra significantly reduced the concentrations of histamine in the nasal mucosa and IgE in the blood compared with those in the model group(P<0.05). Moreover, the pathological results showed that less edema,vasodilation and inflammatory cell infiltration were found in the nasal mucosa after rhIL-1ra application. Budesonide also showed the above effects with no significant difference compared with rhIL-1ra. Conclusion A guinea pig allergic rhinitis model is successfully induced by TDI. The results indicated that rhIL-1ra(50-200μg/kg)is effective in im?proving allergic rhinitis. Our findings indicated that rhIL-1ra might serve as a potential new drug for allergic rhinitis therapy.

Evaluation of the Interleukin-1 Receptor Antagonist and Immunoregulatory Interleukin-10 in the Middle Ear in Chronic Otitis Media With Effusion in Children With and Without Atopy

OBJECTIVES: The role of pro-inflammatory cytokines in the course of chronic otitis media with effusion (COME) has been documented. However, there are fewer studies on the action of anti-inflammatory cytokines in the middle ear. We sought determine whether there is an association between COME and anti-inflammatory cytokines and whether there are any differences in the cytokine profile in COME children with and without atopy. METHODS: Eighty-four children were divided into 3 groups: 32 nonatopic children with COME (group NA), 31 atopic children with COME (group A), and 21 children without COME and without atopy (control group C). Specimens from the middle ear were collected and evaluated by enzyme-linked immunosorbent assay for the cytokines interleukin-1 receptor antagonist (IL-1Ra) and immunoregulatory IL-10. RESULTS: Significantly higher IL-10 concentrations were found in both nonatopic and atopic children with COME compared to controls. No significant differences in IL-1Ra levels were found between atopic and nonatopic children with COME and the control group. CONCLUSION: We found no differences in the levels of IL-1Ra in atopic and nonatopic children with COME compared to controls. However, we found elevated IL-10 levels in the middle ear effusions from children with COME, with or without atopy. These elevated immunoregulatory cytokine levels suggest a role for new immunomodulatory treatments to prevent disease progression in COME, regardless of atopy.

Therapeutic effects of recombinant human interleukin-1 receptor antagonist on toluene-2, 4-diisocyanate-induced guinea pig allergic rhinitis model / 国际药学研究杂志

Objective To evaluate the effects of the recombinant human interleukin-1 receptor antagonist(rhIL-1ra)on a toluene-2, 4-diisocyanate (TDI)-induced guinea pig allergic rhinitis (AR)model. Methods An AR model was established via sensitization and challenge of two-step procedure using TDI in guinea pigs. Normal animals were treated only with the olive oil(TDI vehicle). Sixty adult guinea pigs were randomly divided into six groups (n=10): normal group, model group (rhIL-1ra vehicle), positive control group (budesonide, 25.6 µg/kg), rhIL-1ra treated groups (rhIL-1ra 50, 100 and 200 µg/kg, respectively). From day 8 after sensitization, animals of all the groups were treated respectively with the agents or vehicle once a day for 14 days. During the observation period, the index of clinic score was recorded for every animal. At day 14 of the dosing, guinea pigs were sacrificed 30 min after the last TDI challenge and observation. Blood samples were taken from the abdominal aorta to prepare the serum for detection of histamine, and the nasal mucosase were dissected for histamine detection and histopathological observation. Results Compared with the guinea pigs in normal group, those in the model group exerted the typical symptoms of AR. It was shown that rhIL-1ra could improve nasal symptoms and cause a significant decrease in the instances of nasal sneezing as well. In addition, rhIL-1ra significantly reduced the concentrations of histamine in the nasal mucosa and IgE in the blood compared with those in the model group (P<0.05). Moreover, the pathological results showed that less edema, vasodilation and inflammatory cell infiltration were found in the nasal mucosa after rhIL-1ra application. Budesonide also showed the above effects with no significant difference compared with rhIL-1ra. Conclusion A guinea pig allergic rhinitis model is successfully induced by TDI. The results indicated that rhIL-1ra(50-200 µg/kg)is effective in improving allergic rhinitis. Our findings indicated that rhIL-1ra might serve as a potential new drug for allergic rhinitis therapy.

The effect of interleukin-1 receptor antagonist on metastasis through inhibiting HGF secretion in human colon cancer cell lines / 中华普通外科杂志

Objective The aim of this study was to investigate the co-operative role of HGF and IL-1ra in metastatic processes by interactions between colon cancer cells and stromal cells in their microenvironment.Methods Expression of IL-1α,HGF and c-Met mRNA and proteins were determined by RT-PCR and Western blot.The effect of HGF on metastatic potential was evaluated by proliferation,invasion,and angiogenesis assays using an in vitro system consisting of co-cultured tumor cells and stromal cells.Results IL-1α expression was closely correlated with metastatic potential,and cancer cell-derived IL-1α significantly promoted HGF expression by fibroblasts (P ＜ 0.01).HGF enhanced the migration and proliferation of human umbilical vein endothelial cells (HUVECs),and angiogenesis (P ＜ 0.01).The high liver-metastatic colon cancer cell line (HT-29),which secretes IL-1 α,significantly enhanced angiogenesis compared to the low liver-metastatic cell line (CaCo-2),which does not produce IL-1 α (P ＜ 0.01).IL-1 ra significantly inhibit migration,proliferation and angiogenesis (P ＜ 0.01).Conclusions Autocrine IL-1α and paracrine HGF enhance the metastatic potential of colon cancer cells;IL-1ra inhibit the metastatic potential of colon cancer cells by blocking IL-1α and HGF signaling pathways.

Association between IL-1RN-2018 gene polymorphism and IL-1Ra/IL-1βwith non-small cell lung cancer / 重庆医学

Objective To investigate the association between interleukin receptor antagonist (IL‐1Ra) encoded genes(IL‐1RN‐2018) polymorphisms and serum IL‐1Ra/IL‐1β with non‐small cell lung cancer (NSCLC ) .Methods Totally 85 cases of NSCLC were selected as the NSCLC group and 80 cases of healthy physical examination were selected as the control group from January 2012 to January 2014 .The IL‐1RN‐2018 T/C gene polymorphisms of the two groups were determined with the fluorescence quantitative PCR technique based on high resolution melting ,10% randomly selected samples were sequenced to prove the accura‐cy .The levels of IL‐1Ra and IL‐1βof two groups were determined with ELISA .Results (1)The onset risk of NSCLC in TC and TC+CC genotypes on IL‐1RN‐2018 site in the NSCLC group were increased by 2 .646 times and 2 .315 times respectively compared with TT genotype ,the difference had statistical significance(P0 .05) .(3)The serum IL‐1Ra and IL‐1βlev‐els in the NSCLC group were significantly higher than those in the control group ,but IL‐1Ra/IL‐1βin the NSCLC group was signif‐icantly lower (P0 .05) .Conclusion The C allele in IL‐1RN‐2018 site may increase the onset risk of NSCLC ;the reduction of serum IL‐1Ra/IL‐1βmay presage the risk of NSCLC .

Lesiones preneoplásicas gástricas en pacientes colombianos: asociación de polimorfismos genéticos interleucinas 1B-511, 1RN, 10-819, 10-1082, factor de necrosis tumoral-&alpha -308 y anticuerpos inmunoglobulina G hacia cagA de Helicobacter pylori / Pre-cancerous gastric lesions in Colombian patients: association with IL-1B-511, IL-1RN, IL-10-919, IL-10-1082, TNF-&alpha -308 genes polymorphisms, and anti-Helicobacter pylori cagA IgG antibodies

Objetivo: Evaluar la asociación de los polimorfismos de alguna de las citocinas más estudiadas en relación con el cáncer gástrico (IL-1B-511, IL-1RN intron-2-VNTR, TNF-α-308, IL-10-819 e IL-10- 1082) y la presencia de anticuerpos hacia la proteína cagA de Helicobacter pylori con las lesiones preneoplásicas gástricas en pacientes colombianos. Materiales y métodos: Se estudiaron 185 pacientes con lesiones preneoplásicas (gastritis atrófca, metaplasia intestinal y displasia), y 154 controles (gastritis no atrófica), provenientes de hospitales de una zona de riesgo alto y otra de riesgo bajo para cáncer gástrico. Se obtuvieron biopsias gástricas y muestras de sangre; la genotipificación de los polimorfismos se hizo por discriminación alélica usando PCR en tiempo real y por PCR convencional y electroforesis en agarosa (VNTR del intron 2 de IL-1RN); la serología de Helicobacter pylori y Helicobacter pylori cagA se determinó por ELISA. Se utilizó regresión logística multinomial en el análisis estadístico. Resultados: El genotipo IL-1B-511TT (odds ratio = 4,05; intervalo de confianza 95% 1,35-12,10) se asoció a metaplasia intestinal; no se observaron otras asociaciones entre los diferentes polimorfismos y las lesiones preneoplásicas. La infección por Helicobacter pylori cagA positivo se asoció a gastritis atrófica, metaplasia intestinal y displasia (OR = 2,66; 13,70; 40,29, respectivamente). Conclusión: Los resultados sugieren que entre los genotipos proinflamatorios el genotipo IL-1B-511TT estaría asociado a la metaplasia intestinal, y la serología de Helicobacter pylori cagA positivo sería un biomarcador útil para intervenir y prevenir la presencia de lesiones preneoplásicas. Se necesitan otros estudios con población colombiana que evalúen la asociación hallada de IL1B-511 con la metaplasia intestinal.

Objective: To evaluate the relationship of some of the most studied cytokines (IL-1B-511, IL-1RN intron-2-VNTR, TNF-a-308, IL-10-819, and IL-10-1082) with gastric cancer, as well as the presence of anti-Helicobacter pylori cagA IgG antibodies with pre-cancerous lesions in Colombian patients. Materials and methods: A study was conducted on 185 patients with pre-cancerous lesions (atrophic gastritis, intestinal metaplasia and dysplasia), and 154 controls (non-atrophic gastritis), seen in hospitals in a high risk area, and another in a low risk area, for gastric cancer. Gastric biopsy specimens and blood samples were obtained. The genotyping of the polymorphisms was performed by allelic discrimination using real-time PCR, conventional PCR, and agarose electrophoresis (VNTR of IL-1RN intron 2). The serology of Helicobacter pylori and Helicobacter pylori cagA was determined by ELISA. A multinomial logistic regression was used in the statistical analysis. Results: The IL-1B-511TT genotype was associated with intestinal metaplasia (OR=4.05; 95% CI; 1.35-12.10). No other relationships were observed between the different polymorphisms and preimg/revistas/rcc/cancerous lesions. Infection due to a positive Helicobacter pylori cagA was associated with atrophic gastritis, intestinal metaplasia and dysplasia (OR=2.66; 13.70; 40.29, respectively). Conclusion: The results suggest that, among the pro-inflammatory genotypes, the IL-1B-511TT would be associated with intestinal metaplasia, and that a positive Helicobacter pylori cagA serology could be a useful biomarker for the intervention and prevention of pre-cancerous lesions. Further studies are required in the Colombian population in order to evaluate the relationship found between IL1B-511 and intestinal metaplasia.