Leukotriene B4 Loaded in Microspheres Inhibits Osteoclast Differentiation and Activation

Abstract To investigate osteoclast formation in vivo and if leukotriene B4 (LTB4) loaded in microspheres (MS) could be used as a therapeutical strategy to promote a sustained delivery of the mediator and prevent osteoclast differentiation. Methods: In vivo, apical periodontitis was induced in mice to investigate osteoclast differentiation and signaling in absence of 5-lipoxygenase (5-LO). In vitro, LTB4-MS were prepared using an oil-in-water emulsion solvent extraction-evaporation process. Characterization and efficiency of LTB4 encapsulation were investigated. J774A.1 macrophages were cultured in the presence of monocyte colony-stimulating factor (M-CSF) and ligand for receptor activator of nuclear factor kappa B (RANKL) and then stimulated with LTB4-MS. Cytotoxicity, in vitro MS-LTB4 uptake, osteoclast formation and gene expression were measured. Results: We found that 5-LO negatively regulates osteoclastic formation in vivo during apical periodontitis development. In vitro, LTB4-MS were up-taken by macrophages and were not cytotoxic to the cells. LTB4-MS inhibited osteoclast formation and the synthesis of osteoclastogenic genes Acp5, Mmp9, Calcr and Ctsk. LTB4-MS inhibited differentiation of macrophages into an osteoclastic phenotype and cell activation under M-CSF and RANKL stimulus.

Resumo O objetivo deste trabalho foi Investigar a formação de osteoclastos in vivo e se o leucotrieno B4 (LTB4) incorporado em microesferas (MS) poderia ser usado como estratégia terapêutica para promover uma entrega sustentada do mediador e prevenir a diferenciação dos osteoclastos. Métodos: In vivo, a periodontite apical foi induzida em camundongos para investigar a diferenciação e sinalização de osteoclastos na ausência de 5-lipoxigenase (5-LO). In vitro, LTB4-MS foi preparado usando um processo de evaporação e extração de solvente de emulsão de óleo em água. A caracterização e a eficiência do encapsulamento do LTB4 foram investigadas. Macrófagos J774A.1 foram cultivados na presença de fator estimulador de colônia de monócitos (M-CSF) e ligante para o receptor ativador do fator nuclear kappa B (RANKL) e, então, estimulados com LTB4-MS. Citotoxicidade, captação in vitro de MS-LTB4, formação de osteoclastos e expressão gênica foram avaliadas. Resultados: A via 5-LO regula negativamente a formação de osteoclastos in vivo durante o desenvolvimento da periodontite apical. In vitro, LTB4-MS foram fagocitadas pelos macrófagos e não foram citotóxicos para as células. LTB4-MS inibiu a formação de osteoclastos e a síntese dos genes pró-osteoclastogênicos Acp5, Mmp9, Calcr e Ctsk. Conclusões: LTB4-MS inibiu a diferenciação de macrófagos em um fenótipo osteoclástico e a ativação celular sob estímulo de M-CSF e RANKL.

Correlation between hand-arm vibration disease and vasoactive factors / 环境与职业医学

Background Hand-arm vibration disease (HAVD) is a chronic progressive disease caused by long-term exposure to hand-transmitted vibration, but the mechanism by which vibration affects peripheral vascular function of fingers is not completely clear. Objective To study the association between vasoactive factors and HAVD, and to screen specific indicators for its early diagnosis and prevention. Methods Judgmental sampling method was used to select workers with (HAVD group) and without HAVD (vibration contact group), and non-hand-transmitted vibration operation workers (control group), with 60 workers in each group. The levels of leukotriene B4 (LTB4), vascular endothelial growth factor (VEGF), 5-hydroxy tryptamine (5-HT), interleukin-1β (IL-1β), and calcitonin gene-related peptide (CGRP) in plasma of the three groups were measured by enzyme-linked immunosorbent assay. The association between vasoactive factors and HAVD was analyzed using logistic regression, and the diagnostic HAVD indicators were screened by receiver operator characteristic (ROC) curve of a multivariate model indicator \begin{document}$ \widehat{Y} $\end{document}. Results The hand symptom rates of the HAVD group, the vibration contact group, and the control group were 26.7%, 66.7%, and 96.7% respectively, with a significant difference (P<0.05). The levels of LTB4, 5-HT, IL-1β, and CGRP in the HAVD group were the highest followed by the vibration contact group, and lowest levels were in the control group (P<0.05). There was no significant difference in the VEGF level among the three groups (P>0.05). The logistic regression results showed that higher levels of LTB4 (OR=1.048, 95%CI: 1.022-1.076), 5-HT (OR=1.011, 95%CI: 1.004-1.018), IL-1β (OR=1.148, 95%CI: 1.071-1.230), and CGRP (OR=1.055, 95%CI: 1.008-1.104) were associated with a higher risk of HAVD (P<0.05). The order of the potential indicators' area under the ROC curve from high to low was:\begin{document}$ \widehat{Y} $\end{document} (0.969) > IL-1β (0.907) > LTB4 (0.876) > 5-HT (0.858) > CGRP (0.836). Conclusion The expression levels of LTB4, 5-HT, IL-1β, and CGRP are altered with occupational exposure in hand-transmitted vibration operations and may be associated with HAVD; VEGF is not found to be associated with HAVD. The accuracy of early screening for HAVD can be improved by combining the monitoring of various biochemical indicators.

Prognostic implications and functional enrichment analysis of LTB4R in patients with acute myeloid leukemia / 南方医科大学学报

OBJECTIVE@#To explore the expression patterns, prognostic implications, and biological role of leukotriene B4 receptor (LTB4R) in patients with acute myeloid leukemia (AML).@*METHODS@#We collected the data of mRNA expression levels and clinical information of patients with AML from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) database for mRNA expression analyses, survival analyses, Cox regression analyses and correlation analyses using R studio to assess the expression patterns and prognostic value of LTB4R. The correlation of LTB4R expression levels with clinical characteristics of the patients were analyzed using UALCAN. The co-expressed genes LTB4R were screened from Linkedomics and subjected to functional enrichment analysis. A protein-protein interaction network was constructed using STRING. GSEA analyses of the differentially expressed genes (DEGs) were performed based on datasets from TCGA-LAML stratified by LTB4R expression level. We also collected peripheral blood mononuclear cells (PBMCs) from AML patients and healthy donors for examination of the mRNA expression levels of LTB4R and immune checkpoint genes using qRT-PCR. We also examined serum LTB4R protein levels in the patients using ELISA.@*RESULTS@#The mRNA expression level of LTB4R was significantly increased in AML patients (4.898±1.220 vs 2.252±0.215, P < 0.001), and an elevated LTB4R expression level was correlated with a poor overall survival (OS) of the patients (P=0.004, HR=1.74). LTB4R was identified as an independent prognostic factor for OS (P=0.019, HR=1.66) and was associated with FAB subtypes, cytogenetic risk, karyotype abnormalities and NPM1 mutations. The co- expressed genes of LTB4R were enriched in the functional pathways closely associated with AML leukemogenesis, including neutrophil inflammation, lymphocyte activation, signal transduction, and metabolism. The DEGs were enriched in differentiation, activation of immune cells, and cytokine signaling. Examination of the clinical serum samples also demonstrated significantly increased expressions of LTB4R mRNA (P=0.044) and protein (P=0.008) in AML patients, and LTB4R mRNA expression was positively correlated with the expression of the immune checkpoint HAVCR2 (r= 0.466, P=0.040).@*CONCLUSION@#LTB4R can serve as a novel biomarker and independent prognostic indicator of AML and its expression patterns provide insights into the crosstalk of leukemogenesis signaling pathways involving tumor immunity and metabolism.

Root canal contamination or exposure to lipopolysaccharide differentially modulate prostaglandin E 2 and leukotriene B 4 signaling in apical periodontitis

Abstract Purpose To evaluate the kinetics of apical periodontitis development in vivo , induced either by contamination of the root canals by microorganisms from the oral cavity or by inoculation of bacterial lipopolysaccharide (LPS) and the regulation of major enzymes and receptors involved in the arachidonic acid metabolism. Methodology Apical periodontitis was induced in C57BL6 mice (n=96), by root canal exposure to oral cavity (n=48 teeth) or inoculation of LPS (10 µL of a suspension of 0.1 µg/µL) from E. coli into the root canals (n= 48 teeth). Healthy teeth were used as control (n=48 teeth). After 7, 14, 21 and 28 days the animals were euthanized and tissues removed for histopathological and qRT-PCR analyses. Histological analysis data were analyzed using two-way ANOVA followed by Sidak's test, and qRT-PCR data using two-way ANOVA followed by Tukey's test (α=0.05). Results Contamination by microorganisms led to the development of apical periodontitis, characterized by the recruitment of inflammatory cells and bone tissue resorption, whereas inoculation of LPS induced inflammatory cells recruitment without bone resorption. Both stimuli induced mRNA expression for cyclooxygenase-2 and 5-lipoxygenase enzymes. Expression of prostaglandin E 2 and leukotriene B 4 cell surface receptors were more stimulated by LPS. Regarding nuclear peroxisome proliferator-activated receptors (PPAR), oral contamination induced the synthesis of mRNA for PPARδ, differently from inoculation of LPS, that induced PPARα and PPARγ expression. Conclusions Contamination of the root canals by microorganisms from oral cavity induced the development of apical periodontitis differently than by inoculation with LPS, characterized by less bone loss than the first model. Regardless of the model used, it was found a local increase in the synthesis of mRNA for the enzymes 5-lipoxygenase and cyclooxygenase-2 of the arachidonic acid metabolism, as well as in the surface and nuclear receptors for the lipid mediators prostaglandin E2 and leukotriene B4.

Pathogenic role leukotriene B4 in lung injury induced by lung-protective mechanical ventilation in rabbits / 南方医科大学学报

OBJECTIVE@#To elucidate the pathogenic role of leukotriene B4 (LTB4) in pulmonary hyper-permeability and inflammation induced by lung-protective mechanical ventilation (LPMV) in rabbits.@*METHODS@#Thirty-two healthy Japanese white rabbits were randomized into 4 groups for treatment with vehicle or bestatin (a leukotriene A4 hydrolase inhibitor that inhibits LTB4 production) administered intragastrically at the daily dose of 8 mg/kg for 5 days, followed by sham operation (group S and group BS, respectively, in which the rabbits were anesthetized only) or LPMV (group PM and group BPM, respectively, in which the rabbits received ventilation with 50% oxygen at a tidal volume of 8 mL/kg for 5 h). The concentrations of LTB4 and cyclic adenosine monophosphate (cAMP) in the lung tissues were analyzed by ELISA. cAMP content, protein kinase A (PKA) protein expression and the Rap1-GTP protein to total Rap1 protein ratio were determined to assess the activities of cAMP/PKA and Rap1 signaling pathways. The lung injury was evaluated by assessing lung permeability index, lung wet/dry weight ratio, polymorphonuclear leukocyte (PMN) count in bronchoalveolar lavage fluid (BALF), pulmonary myeloperoxidase (MPO) activity and lung histological scores.@*RESULTS@#None of the examined parameters differed significantly between group S and group BS. All the parameters with the exception of lung histological score increased significantly in group PM and group BPM as compared to those in group S (@*CONCLUSIONS@#LPMV can induce LTB4 overproduction to down-regulate cAMP/PKA and Rap1 signaling pathways in the lungs of rabbits, which results in lung hyper-permeability and inflammation. Bestatin can inhibit LTB4 production in the lungs to protect against LPMV-induced lung hyper-permeability and inflammation.

Leukotriene B4 in equine asthma syndrome: what do we know so far? / Leucotrieno B4 na síndrome asmática equina: o que sabemos até o momento?

The term "equine asthma syndrome" (EAS) was recently proposed due to the resemblance of the equine disease to human asthma. Leukotrienes cause constriction of the bronchi, especially in the lower airways and increase mucus secretion in the respiratory system. Leukotriene B4 (LTB4) has been discovered as a strong chemotactic factor, which plays a role in neutrophil migration. The immunologic background of EAS remains not fully elucidated despite many studies on the pathogenesis. This study aimed to evaluate the LTB4 concentration in the bronchoalveolar lavage fluid (BALF) of horses with and without pulmonary inflammatory disease. Thirty-five mixed breed horses were studied and LTB4 was determined by using specific ELISA Kit. The horses were grouped by 2 different criteria for statistical analysis of data: 1) according to the values for BALF citology and 2) according to the detection of LTB4 in BALF. There was significant difference of effect of age on the LTB4 detection in equine BALF. Younger animals were the majority where it was possible to detect LTB4 values in LBA. In conclusion, there was an effect of age on the detection of LTB4 in equine BALF, where LTB4 levels were more easily detected in younger animals than older animals and the results of this study raise the possibility of considering future studies with the objective of establishing the real role and the best moment to detect LTB4 in BALF of the equine asthma syndrome.(AU)

Recentemente, o termo "síndrome da asma equina" (SAE) foi proposto devido à semelhança da doença equina à asma humana. Os leucotrienos causam constrição dos brônquios, especialmente nas vias aéreas posteriores e aumentam a secreção de muco no sistema respiratório. O leucotrieno B4 (LTB4) foi descoberto como um forte fator quimiotático, que desempenha um papel na migração de neutrófilos. O fundo imunológico do SAE permanece não completamente elucidado apesar de muitos estudos sobre a patogênese. Este estudo teve como objetivo avaliar a concentração de LTB4 no lavado broncoalveolar (LBA) de equinos com e sem doença inflamatória pulmonar. Trinta e cinco cavalos de raças mistas foram estudados e o LTB4 foi determinado usando o kit ELISA específico. Os animais foram agrupados por dois critérios diferentes para análise estatística dos dados: 1) de acordo com os valores para citologia do LBA e 2) de acordo com a detecção do LTB4 no LBA. Houve diferença significativa do efeito da idade na detecção do LTB4 no LBA equino. Os animais mais jovens foram a maioria onde foi possível detectar os valores de LTB4 no LBA. Em conclusão, houve um efeito da idade na detecção de LTB4 em LBA equino, onde os níveis de LTB4 foram mais facilmente detectados em animais jovens do que em animais mais velhos e foi possível detectar a concentração de LTB4 no LBA equino e os resultados deste estudo levantam a possibilidade de considerar futuros estudos com o objetivo de estabelecer o real papel e o melhor momento para detectar LTB4 no LBA da síndrome asmática equina.(AU)

Comparison of effect of desloratadine citrate disodium tablets and loratadine dispersible tablets in treatment of patients with allergic rhinitis / 中国基层医药

Objective To compare the efficacy and adverse reactions of desloratadine citrate disodium tablets and loratadine dispersible tablets in treatment of patients with allergic rhinitis,and their influence on leukotriene B4(LTB4),interleukin-4 (IL-4),interleukin-10 (IL-10) and interferon gamma (INF-γ).Methods From June 2015 to June 2016,a total of 110 patients with allergic rhinitis in the Second People's Hospital of Cangnan County were selected and randomly divided into control group and observation group according to the digital table,with 55 cases in each group.The control group was given loratadine dispersible tablets,while the observation group was given desloratadine citrate disodium tablets.Both two groups were treated for 14 days.The clinical efficacy,adverse reactions and changes of LTB4,IL-4,IL-10,INF-γ levels before and after treatment were compared between the two groups.Results The total effective rate of the observation group was 94.5 ％,which was significantly higher than 80.0％ of the control group(x2 =6.310,P ＜ 0.05).The incidence rate of adverse reactions in the observation group was 16.4％,which was similar to 20.0％ in the control group.Before treatment,there were no statistically significant differences in LTB4,IL-4,IL-10,INF-γ levels between the two groups (all P ＞ 0.05).After 2 weeks of treatment,the levels of LTB4,IL-4,IL-10 and INF-γ in the observation group were (67.74 ±10.15) ng/L,(52.37 ± 5.12) μg/L,(81.26 ± 11.78) μg/L,(94.47 ± 7.87) μg/L,respectively,which in the control group were (80.32 ± 9.97) ng/L,(62.95 ± 5.45) μg/L,(96.32 ± 11.57) μg/L,(86.74 ± 7.63) μg/L,respectively,there were statistically significant differences between the two groups (t =7.124,5.262,4.654,3.718,all P ＜ 0.05).Conclusion Both desloratadine citrate disodium tablets and loratadine dispersible tablets can effectively treat patients with allergic rhinitis,improve the symptoms and physical signs,reduce the levels of LTB4,IL-4 and IL-10,increase the level of INF-γ,and the adverse reactions are less and slight,but the efficacy of desloratadine citrate disodium tablets is better than loratadine dispersible tablets.

Drug repurposing of histone deacetylase inhibitors that alleviate neutrophilic inflammation in acute lung injury and idiopathic pulmonary fibrosis via inhibiting leukotriene a4 hydrolase and blocking LTB4 biosynthesis / 中国药理学与毒理学杂志

OBJECTIVE Leukotriene B4 (LTB4) biosynthesis and subsequently neutrophilic inflam?mation may provide a potential strategy for the treatment of acute lung injury (ALI) or idiopathic pulmonary fibrosis (IPF). To provide a potential strategy for the treatment of ALI or IPF, we identified potent inhibi?tors of Leukotriene A4 hydrolase (LTA4H), a key enzyme in the biosynthesis of LTB4. METHODS In this study, we identified two known histone deacetylase (HDAC) inhibitors, suberanilohydroxamic acid (SAHA) and its analogue 4-(dimethylamino)-N-〔7-(hydroxyamino)-7-oxoheptyl〕benzamide (M344), as effective inhibitors of LTA4H using enzymatic assay, thermofluor assay, and X- ray crystallographic investigation. We next tested the effect of SAHA and M344 on endogenous LTB4 biosynthesis in neutrophils by ELISA and neutrophil migration by transwell migration assay. A murine experimental model of ALI was induced by lipopolysaccharide(LPS) inhalation. Histopathological analysis of lung tissue using H&E staining revealed the serious pulmonary damage caused by LPS treatment and the effect of the SAHA. We next examined mRNA and protein levels of pro-inflammatory cytokines in lung tissue and bronchoalveolar lavage fluid using qRT- PCR and ELISA to further investigate the underlying mechanisms of anti-inflammatory activities by SAHA. We also investigated the effects of SAHA and M344 on a murine experimental model of bleomycin (BLM)-induced IPF model. RESULTS The results of enzymatic assay and X-ray crystallography showed that both SAHA and M344 bind to LTA4H, signif?icantly decrease LTB4 levels in neutrophil, and markedly diminish early neutrophilic inflammation in mouse models of ALI and IPF under a clinical safety dose. CONCLUSION Collectively, SAHA and M344 would provide promising agents with well-known clinical safety for potential treatment in patients with ALI and IPF via pharmacologically inhibiting LAT4H and blocking LTB4 biosynthesis.

A rare case of Sjogren-Larsson syndrome with recurrent pneumonia and asthma / 소아과

Sjogren-Larsson syndrome (SLS) is a rare autosomal recessive neurocutaneous disorder with worldwide incidence of 0.4 per 100,000 people. It is characterized by the triad of congenital ichthyosis, spastic diplegia or quadriplegia, and mental retardation. Herein we report a 2-year-old male child with SLS, asthma, and recurrent pneumonia. SLS was confirmed by a molecular genetics study that revealed a deletion mutation in the ALDH3A2 gene. An ALDH3A2 gene mutation results in dysfunction of the microsomal enzyme fatty aldehyde dehydrogenase and impaired metabolism and accumulation of leukotriene B4, which is a key molecule and a pro-inflammatory mediator in developing allergic diseases, especially asthma. An increased level of leukotriene B4 has been reported in SLS patients. As far as we are aware, this is the first report of SLS associated with asthma and recurrent pneumonia. In conclusion, pediatricians should be aware of and evaluate patients with SLS for possible associated asthma and allergic disorders.

Antagonist of leukotriene B4 receptor 1 attenuates cisplatin induced acute kidney injury in mice and its associated mechanism / 中华肾脏病杂志

Objective To investigate the effect of pretreatment with U75302,antagonist of leukotriene B4 receptor 1 (BLT1),on cisplatin induced acute kidney injury in mice and its immunoregulatory mechanism.Methods Healthy C57BL/6 mice were randomized into four subgroups:1.healthy control group;2.cisplatin group;3.U75302 control group;4.cisplatin + U75302 group,n=6.Group 2 and 4 received intraperitoneal injection of cisplatin (20 mg/kg) on day 0,group 3 and 4received intraperitoneal injection of U75302 (5 μg/mouse) on day 0 and day 2.Mice were sacrificed on the 3rd day and blood and kidney were collected.Renal function and histological changes were estimated,the infiltration of immune cells were determined by flow cytometry,the level of peroxidase (MPO) in kidney were determined by colorimetry,relative expression of TNF-α,IL-1β,CXCL1,CXCL2 were detected by Real-time PCR.Results Compared with healthy control group,levels of BUN,Scr were higher in cisplatin group with serious tubular structural damage.There were more neutrophils,macrophages,CD4+ T lymphocytes,CD8+ T lymphocytes in kidneys of cisplatin group,the level of MPO and relative expression of TNF-α,IL-1β,CXCL1,CXCL2 were also higher in cisplatin group.Compared with cisplatin group,lower BUN [(17.75±1.80) mmol/L vs (42.6±6.66) mmol/L,P ＜0.05],Scr were found in cisplatin+ U75302 group with less tubular structural damage.Meanwhile,U75302 reduced infiltration of neutrophils [(146±13)×103/g vs (296±66) ×103/g,P ＜ 0.05],macrophages [(245± 13)× 103/g vs (420±78)× 103/g,P ＜ 0.05] in the kidney.Levels of MPO [(1.756±0.283) U/g vs (3.308±0.577) U/g,P＜0.05] and relative expression of TNF-α,IL-1β,CXCL1,CXCL2 were also lower.Conclusions BLT1 antagonist U75302 protects mice against AKI induced by cisplatin,and the mechanism is associated with reduced infiltration of inflammatory cells in kidney and the inhibition of kidney inflammation.

Effects of U75302, blocker of leukotriene B4 receptor 1, on cellular immunity and inflammation in septic mice / 第二军医大学学报

Objective To observe the influence of U75302, blocker of leukotriene B4 receptor 1, on immune function in septic mice, so as to explore the implication of blocking leukotriene B4 receptor 1 for treatment of sepsis. Methods Experimental sepsis model was induced by cecal ligation and puncture (CLP). A total of 18 male C57BL/6 mice were randomly divided into sham group (n=6), CLP group (n=6) and CLP + U75302 intraperitoneal injection group (n = 6). The peripheral blood cytokine tumor necrosis factor- α (TNF-α) and interleukin-10 (IL-10) levels, peritoneal lavage fluid Gr-1+ cell count and thymus T lymphocytes apoptosiswere detected in mice of three groups 24 h after surgery. Results Compared with the CLP group, CLP + U75302 intraperitoneal injection group had significantly reduced blood TNF-α level (by 43%, P< 0. 05), significantly increased IL-10 levll (by 88%, P<0. 05), signficantly decreased Gr-1+ cell counts in peritoneal lavage fluid (P<0. 05), and significantly decreased apoptosis of thymus T lymphocytes (P<0. 01). Conclusion Blocking leukotriene B4 receptor 1 with U75302 may decrease peripheral TNF-α level, increase IL-10 level, and improve cellular immunity, which may be involved in inflammation of sepsis.

mRNA and protein expression of leukotriene B4 receptor 2 during transition from colitis to colon cancer / 国际药学研究杂志

Objective: To observe mRNA and protein expression of leukotriene B4 (LTB4) receptor 2 (BLT2)in mice during the course of development from colitis to colitis-associated cancer. Methods: ICR mice were used to establish the animal model of colitis-associated colon cancer and randomly divided into control group and experimental group. We began to administer inducer on d0. Mice were sacrificed at 2, 3, 5, 7, 9, 13, and 18w, respectively. Histopathological changes in colons were examined. The protein and mRNA expression of BLT2 in colons were measured by immunohistochemical assay and real-time quantitative PCR, respectively. Results: Histological study showed that with the extension of time, the lesions of mice colons aggravated, first a mild inflammation, then atypical hyperplasia, and finally to colon cancer. Immunohistochemical staining showed that the expression of BLT2 protein was low in normal colon, but high in inflammatory lesions, especially in inflammatory cells. There was no BLT2 expression in atypical hyperplasia and cancer cells, while BLT2 was highly expressed in the stroma and lumans of cancer tissue. Compared with the control group, mRNA expression of BLT2 in the experimental group was significantly higher at 2, 13 and 18w (P<0.05); and very significantly higher at 3, 5, 7 and 9w(P<0.01). Conclusion: The mouse model of colitis-associated colon cancer was developed in this study. The expression of BLT2 changed in the course of colitis development, indicating that BLT2 may play an important role in the transition process from colitis to colon cancer.

mRNA and protein expression of leukotriene B4 receptor 2 during transition from colitis to colon cancer / 国际药学研究杂志

Objective To observe mRNA and protein expression of leukotriene B4(LTB4)receptor 2(BLT2)in mice during the course of development from colitis to colitis-associated cancer. Methods ICR mice were used to establish the animal model of colitis-associated colon cancer and randomly divided into control group and experimental group. We began to administer inducer on d0. Mice were sacrificed at 2,3,5,7,9,13,and 18w,respectively. Histopathological changes in colons were examined. The protein and mRNA expression of BLT2 in colons were measured by immunohistochemical assay and real-time quantitative PCR, respectively. Results Histological study showed that with the extension of time,the lesions of mice colons aggravated,first a mild inflammation,then atypical hyperplasia,and finally to colon cancer. Immunohistochemical staining showed that the expression of BLT2 protein was low in normal colon,but high in inflammatory lesions,especially in inflammatory cells. There was no BLT2 expression in atypical hyperplasia and cancer cells,while BLT2 was highly expressed in the stroma and lumans of cancer tissue. Compared with the control group,mRNA expression of BLT2 in the experimental group was significantly higher at 2,13 and 18w(P<0.05);and very significantly higher at 3,5,7 and 9w (P<0.01). Conclusion The mouse model of colitis-associated colon cancer was developed in this study. The expression of BLT2 changed in the course of colitis development,indicating that BLT2 may play an important role in the transition process from colitis to colon cancer.

Increased inflammatory mediator in exhaled breath condensate from asthmatic children

PURPOSE: There has recently been increasing interest in the use of exhaled breath condensate (EBC) as a simple noninvasive means for understanding the physiology of asthma. The aim of this study was to evaluate the levels of leukotriene B4 (LTB4) and eosinophil cationic protein (ECP) in the EBC of asthmatic children. METHODS: We measured LTB4 and ECP levels in EBC from children aged 6-14 years, including healthy children (n=25) and asthmatic children (n=25). We also measured serum LTB4 and serum ECP. Pulmonary function tests and methacholine challenge tests were performed on all subjects. RESULTS: Exhaled LTB4 levels were increased significantly in patients with asthma compared to normal subjects (7.1+/-3.7 pg/mL vs. 2.2+/-1.7 pg/mL, P<0.05). Serum LTB4 levels were not significantly different in patients with asthma compared to normal subjects (674.7+/-484.1 pg/mL vs. 487.1+/-272.0 pg/mL, P=0.156,) and no significant correlations were found between exhaled and serum LTB4 concentrations in children with asthma (r=0.052, P=0.758). Exhaled ECP levels were not significantly different in patients with asthma compared to normal subjects (P=0.419). Serum ECP levels were significantly increased in patients with asthma compared to normal subjects (44.37+/-32.14 microg/L vs. 16.40+/-13.23 microg/L, P=0.001). CONCLUSION: We found significantly elevated LTB4 levels in the EBC of asthmatic children. Our results suggest that EBC may be one of the supportive tools to measure airway inflammation in children with asthma.

The expression and clinical significance of the serum IL-17/LTB4/IgE in children with asthma / 国际儿科学杂志

Objective To explore the impact of levels of serum IL-17,Leukotriene B4 and IgE on pathogenesis of childhood asthma.Methods Totally 60 children with asthma acute exacerbation ( 29 children with mild asthma,31 children with moderate-severe asthma) were selected as study group,24 healthy children were selected as control group.Serum IL-17 and LTB4 were measured with euryzemLinked immunosorbent assay,serum IgE was determined with enzyme-linked fluoroimmuneassay by pharmacia CAP Sytem,PMN was determined with automatic blood analyser,pulmonary function was measured in the study group.Results ( 1 ) The level of serum IL-17 ( 1.15 ± 0.10 μg/L,2.80 ± 2.30 μg/L,0.83 ± 0.10 μg/L),LTB4 (2.22 ± 1.01 μg/L,8.79 ± 9.36 μg/L,1.94 ± 1.13 μg/L) and IgE( 123.70 ±86.94 μg/L,322.27 ±332.28 μg/L,24.27 ±7.64 μg/L) were significantly different among mild asthma group,moderate-severe asthma group and control group( P ＜ 0.001 ).( 2 )The N％ of mild asthma group,moderate-severe asthma group and study group were( 55.06 ± 1 1.15 ) ％,( 64.44± 11.87)％,(47.96 ± 13.52)％,L％ were(42.20 ± 11.04)％,(33.93 ± 10.02)％,(49.65 ± 13.02)％,and there were significant differences in N％ and L％ between study group and control group( P ＜ 0.05 ).( 3 ) There were significant positive correlations between the serum IL-17 levels and IgE,LTB4 and IgE,IL-17 and LTB4 in asthmatic children( P ＜0.05).(4) There were significant negative correlcations between the level of serum IL-17,LTB4 and FEVI,PEF( P ＜0.001 ).There were significant positive correlations between serum IL-17,LTB4 and N％ (P ＜0.001 ).(5)There were not correlations between the level of serum IgE and FEV1,PEF and N％in asthmatic children( P ＞0.05 ).Conclusion The levels of serum IL-17,LTB4 and IgE participated in pathogenesis on asthmatic children patients.

Evidence for eosinophil recruitment, leukotriene B4 production and mast cell hyperplasia following Toxocara canis infection in rats

It is well known that eosinophilia is a key pathogenetic component of toxocariasis. The objective of the present study was to determine if there is an association between peritoneal and blood eosinophil influx, mast cell hyperplasia and leukotriene B4 (LTB4) production after Toxocara canis infection. Oral inoculation of 56-day-old Wistar rats (N = 5-7 per group) with 1000 embryonated eggs containing third-stage (L3) T. canis larvae led to a robust accumulation of total leukocytes in blood beginning on day 3 and peaking on day 18, mainly characterized by eosinophils and accompanied by higher serum LTB4 levels. At that time, we also noted increased eosinophil numbers in the peritoneal cavity. In addition, we observed increased peritoneal mast cell number in the peritoneal cavity, which correlated with the time course of eosinophilia during toxocariasis. We also demonstrated that mast cell hyperplasia in the intestines and lungs began soon after the T. canis larvae migrated to these compartments, reaching maximal levels on day 24, which correlated with the complete elimination of the parasite. Therefore, mast cells appear to be involved in peritoneal and blood eosinophil infiltration through an LTB4-dependent mechanism following T. canis infection in rats. Our data also demonstrate a tight association between larval migratory stages and intestinal and pulmonary mast cell hyperplasia in the toxocariasis model.

Low-dose UVB irradiation stimulates matrix metalloproteinase-1 expression via a BLT2-linked pathway in HaCaT cells

Skin exposure to low-dose ultraviolet B (UVB) light up-regulates the expression of matrix metalloproteinase-1 (MMP-1), thus contributing to premature skin aging (photo-aging). Although cyclooxygenase-2 (COX-2) and its product, prostaglandin E2 (PGE2), have been associated with UVB-induced signaling to MMP expression, very little are known about the roles of lipoxygenases and their products, especially leukotriene B4 (LTB4) and 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE), in MMP-1 expression in skin keratinocytes. In the present study, we demonstrate that BLT2, a cell surface receptor for LTB4 and 12(S)-HETE, plays a critical role in UVB-mediated MMP-1 upregulation in human HaCaT keratinocytes. Moreover, our results demonstrated that BLT2-mediated MMP-1 upregulation occurs through a signaling pathway dependent on reactive oxygen species (ROS) production and the subsequent stimulation of ERK. Blockage of BLT2 via siRNA knockdown or with the BLT2-antagonist LY255283 completely abolished the up-regulated expression of MMP-1 induced by low-dose UVB irradiation. Finally, when HaCaT cells were transiently transfected with a BLT2 expression plasmid, MMP-1 expression was significantly enhanced, along with ERK phosphorylation, suggesting that BLT2 overexpression alone is sufficient for MMP-1 up-regulation. Together, our results suggest that the BLT2-ROS-ERK-linked cascade is a novel signaling mechanism for MMP-1 upregulation in low-dose UVB-irradiated keratinocytes and thus potentially contributes to photo-aging.

Detection of plasma RANTES, eotaxin, TNF-α and LTB4 in patients with chronic urticaria / 中华皮肤科杂志

Objective To investigate the plasma levels of regulated upon activation normal T cell expressed and secreted (RANTES), eotaxin, tumor necrosis factor (TNF)-α and leukotriene B4 (LTB4) in patients with chronic urticaria and their roles in the pathogenesis of chronic urticaria. Methods Forty-one patients with chronic urticaria were included into this study along with 20 normal human controls. Patients were graded into three groups, I.e. Mild group (n = 11), moderate group (n = 21) and severe group (n = 9), according to their symptom score. All patients were treated with mizolastine 10 mg per day for 4 weeks. ELISA was used to study the plasma levels of RANTES, eotaxin, TNF-α and LTB4 in normal controls and patients before and after treatment. Results The plasma levels of RANTES, eotaxin, TNF-α and LTB4 were (52.5 ± 10.2) g/L, (58.4 ± 16.1) g/L, (35.1 ± 9.6) ng/L and (109.4 ± 21.7) ng/L, respectively, in untreatedpatients with chronic urticaria, compared to (33.7 ± 9.4) g/L, (48.3 ± 13.6) g/L, (21.3 ± 8.9) ng/L and(77.8 ± 11.6) ng/L, respectively, in normal controls(P < 0.01, 0.05, 0.01, 0.01, respectively). Increased plasmalevels of RANTES, eotaxin, TNF-α and LTB4 were observed in patients with moderate or severe chronic urticaria compared with those with mild chronic urticaria (both P < 0.05), whereas there was no significant difference between patients with severe and mild chronic urticaria (P < 0.05). After treatment with mizolas-fine the plasma levels ofRANTES, eotaxin, TNF-α and LTB4 were (36.3 ± 8.9) g/L, (46.3 ± 10.2) g/L, (23.2 ± 7.5) ng/L and (83.1 ± 14.2) ng/L respectively, significantly lower than those in patients before treatment (all P < 0.01), but showed no difference from those in normal controls (all P > 0.05). Conclu-sions The plasma levels of RANTES, eotaxin, TNF-α and LTB4 are elevated in patients with chronic urticaria, and they exhibits a positive correlation tendency with disease activity. After treatment with mizolastine, a significant decrease is observed in the plasma levels of RANTES, eotaxin, TNF-α and LTB4, which hints that RANTES, eotaxin, TNF-α and LTB4 may play a certain role in the pathogenesis of chronic urticaria.

The effect of juvenile rheumatoid arthritis serum on the LTB4-BLT2 signal pathway of mice dendritic cells / 中华风湿病学杂志

Objective To investigate the effect of iuvenile rheumatoid arthritis(JRA) serum (leukotriene B4,LTB4) LTB4-BLT2 on mice DCs.Methods Bone marrow(BM)-derived DCs from healthy mouse were purified.and induced by cytokine IL-4 and GM-CSF to immature DCs and then differentiated to mature DCs under the stimulation of LPS in vitro.DCs were evaluated by light microscope and flow cvtometry.The concentrations of LTB4 in DCs supernatant,normal serum,active JRA,and that of the co-cuItured with BLT2 antagonist LY255283 were detected by ELISA.The expression of BLT2 protein and mRNA in DCs was examined by immunocytochemistry,immunofluorescence and RT-PCR.Meanwhile,the expression of BLT2 in DCs after 18 h co-cultured with normal serum,in serum of active JRA and that of BLT2 antagonist (LY255283)group was assayed by flow cytometry respectively.Results LTB4-BLT2 was expressed by DCs.Not only BLT2 mRNA but also its protein was expressed in DCs.The concentration of LTB4 Was(17±3)pg/ml,(82±20)pg/ml,(82±20)pg/ml and(24±6)pg/ml,(115±20)pg/ml,(91±11)pg/ml in normal serum group,active JRA group and LY255283 group before and after 1 8 h,respectively.The expression Was higher in serum of active JRA group than that of normal sertlm group(P<0.01)and there was a tendency to be higher when compared with LY255283 group(P<0.05).The DC BLT2 expression was 27.7±2.9,46.3±8.7 and 30.3±5.5 in normal serum group,serum of active JRA group and LY255283 grbup after 1 8h respectively.The expression was stronger in active JRA group than other groups(P<0.05).Conclusion DC can develop a LTB4-BLT2 signal pathway by BLT2 with autocrine and/or extrinsic LTB4.The overexpression of this pathway may be involved in the initiating and activation of JRA.

Change of lipxin A4, leukotriene B4 and 15-1ipoxygenase in chiMren with acute poststreptococcal glomerulonephritis / 中华肾脏病杂志

ObjectiveTo investigate the change of lipoxin A4 (LXA4), leuotriene B4(LTB4) in blood and urine and leukocyte 15-lipoxygenase (15-LO) of the children with acute poststreptococcal glomendonephritis (APSGN) and to evaluate its significance. MethodsBlood and urinary levels of LXA4 and LTB4 were measured with ELISA within 3 days (acute phase), 10 to 14 days (early resolution phase) and 6 to 8 weeks (late resolution phase) respectively after onset of APSGN in 22 patients. In 8 children with APSGN, expression level of leukocyte 15-LO mRNA was examined with RT-PCR. Leukocyte LTB4 synthesis was assessed with ELISA. Chemotactic effect of LTB4, LXA4 and 15-S-hydroxyeicosatetraenoic acid (15-S-HETE) on neutrophils was determined by in vitro chemotaxis assay. Twenty-two healthy children were served as control. ResultsBlood and urinary levels of LXA4 and leukocyte 15-LO mRNA were up-regnlated in acute phase, further increased in early resolution phase, and decreased in late resolution phase of APSGN, which were stir higher than those in the controls (P＜0.01). Blood and urinary levels of LTB4 were increased in acute phase (P＜0.01) and then were decreased in early resolution phase and hte resolution phase of APSGN, which were still higher than those in the controls (P＜0.01). Administration of 15-S-HETE or LXA4 in vitro inhibited LTB4-induced chemotactic effect on neutrophils of the patients,and inhibited the production of leukocyte LTB4. ConclusionsChanges of blood and urinary levels of LXA4 and LTB4 in early resolution phase of APSGN are contrary. 15-S-HETE and LXA4may play a role in anti-inflammation and resolution of APSGN via inhibiting LTB4.