Prognostic value of power doppler ultrasonography for equine superficial digital flexor tendon injury in thoroughbred racehorses.

The potential value of hypervascularity detected with power Doppler ultrasonography (PDU) within equine superficial digital flexor tendon (SDFT) as a prognostic factor of SDFT injury is not clear. The purpose of this study was to test the hypothesis that hypervascularity within SDFT is one of the risk factors for subsequent severe SDFT injury and to evaluate the prognostic value. A prospective cohort study of 97 Thoroughbred racehorses without any clinical signs of SDFT injury was conducted. Six variables of age, body weight, sex, the cross-sectional area of SDFT, PDU signal within SDFT and experience of steeplechase were assessed for the possibility of risk factors of subsequent SDFT injury in follow-up period of 1 year. Multivariable logistic regression analyses were used for assessment of the odds ratios (ORs) and 95 % confidence intervals (CIs) of SDFT injury. Multivariable logistic regression analysis revealed that the PDU signal within SDFT was a risk factor for the development of SDFT injury in follow-up period (P = 0.017). The adjusted OR of SDFT injury was significantly higher in PDU positive group than in PDU negative group (OR 3.17, 95 % CIs 1.20-8.35). Although further studies are required, these results would be useful for early detection and/or prevention of development for clinical severe SDFT injury.

Prognostic nutritional index and early mortality with percutaneous endoscopic gastrostomy.

BACKGROUND: Although percutaneous endoscopic gastrostomy (PEG) is a well-accepted and less invasive method of feeding tube placement in patients with swallowing difficulties, complications and early death after PEG have been reported. AIM: This study aimed to evaluate predictive factors associated with 30-day mortality after PEG, and to assess the utility of nutritional supporting period before PEG in reducing early mortality following PEG. DESIGN: An observational study. METHODS: We retrospectively analyzed 268 patients who underwent PEG at Sapporo Shirakaba-dai Hospital from 2006 to 2010, using clinical and laboratory data to analyze predictive factors associated with early death after PEG. Then, we prospectively assessed 152 consecutive patients assessed for eligibility for PEG from 2011 to 2014. We assessed the patients' nutritional condition using Onodera's prognostic nutritional index (PNI), and supported nutrition for more than 10 days before PEG in patients with a poor nutritional index (PNI < 37). RESULTS: In both univariate and multivariate analyses in the retrospective study, Onodera's PNI of less than 37 was the only predictive factor for early mortality. In the second study, among the 115 patients who finally underwent PEG, early mortality rates improved to 1.7% from 5.2% in the first study. Conversely, 32% of patients with malnutrition who did not undergo PEG died within 30 days. CONCLUSION: Nutritional status might be a predictive factor for early mortality after PEG. In patients with poor nutritional status, nutritional supporting period before PEG might improve the outcomes and reduce unnecessary PEG.

A retrospective cohort study investigating risk factors for the failure of Thoroughbred racehorses to return to racing after superficial digital flexor tendon injury.

A retrospective cohort study was conducted to investigate risk factors for the failure of Thoroughbred racehorses to return to racing after an injury of the superficial digital flexor tendon (SDFT). Successful return was defined as the completion of five or more races after SDFT injury. The official Japan Racing Association (JRA) medical records of racehorses with a core-type SDFT injury were reviewed for clinical variables related to the characteristics of the horse and the severity of SDFT injuries at the time of diagnosis. Data on racing outcomes were obtained from the official JRA racing database. Risk factors were screened using univariable logistic regression and subsequent multivariable model building. Forty-nine of 346 (14.2%) horses successfully returned to racing after SDFT injuries. Multivariable model building revealed that an increase in the total number of injured zones (defined as the total number of zones in which the injured hypoechoic area was observed at the time of ultrasonographic diagnosis of SDFT injury) was associated with an increased risk of failure to return to racing after SDFT injury. Horse characteristics, such as age, body mass and sex, were not associated with a successful return to racing. In the rehabilitation of cases with larger (longer) lesions, more effective and careful medical management may be needed for an improvement in the athletic outcomes.

Urinary concentrations of 15-epimer of lipoxin A(4) are lower in patients with aspirin-intolerant compared with aspirin-tolerant asthma.

BACKGROUND: Although an abnormality in arachidonic acid metabolism may be responsible for aspirin-intolerant asthma (AIA), there is little knowledge about the concentrations of urinary lipoxin A(4) (LXA(4)) and the 15-epimer of LXA(4) (15-epi-LXA(4)) in relation to asthma severity in AIA subjects. OBJECTIVE: The purpose of this study is to estimate urinary LXA(4) and the 15-epimer concentrations to investigate lipoxins in AIA. METHODS: In this study, we examined AIA, aspirin-tolerant asthma (ATA) and healthy control groups. The AIA and ATA groups were subdivided into the severe asthma and non-severe asthma subgroups. Urinary LXA(4), 15-epi-LXA(4) and leukotriene E(4) (LTE(4) ) were quantified using enzyme immunoassay after separating these compounds using high-performance liquid chromatography. RESULTS: The urinary LXA(4) concentration was significantly lower than the 15-epi-LXA(4) concentration in the asthmatic subjects. The AIA group showed significantly lower urinary 15-epi-LXA(4) (P < 0.01) and higher urinary LTE(4) concentrations (P < 0.05) than the ATA group. Comparison of 15-epi-LXA(4) concentrations between the severe asthmatic and non-severe asthmatic subjects in the AIA and ATA groups revealed that the decreased 15-epi-LXA(4) concentration may be related to aspirin intolerance, but not asthma severity. Receiver operator characteristic curves demonstrated that the concentration ratio of LTE(4) to 15-epi-LXA(4) was superior to 15-epi-LXA(4) concentration and LTE(4) concentration as a predictive factor for aspirin intolerance. CONCLUSIONS AND CLINICAL RELEVANCE: We have demonstrated for the first time that urinary 15-epi-LXA(4) concentration is significantly higher than LXA(4) concentration in both the AIA and ATA groups. 15-Epi-LXA(4) concentration was significantly lower in the AIA group with an increased urinary LTE(4) concentration than in the ATA group. An imbalance between proinflammatory cysteinyl-leukotrienes and anti-inflammatory 15-epi-LXA(4) may be involved in AIA pathogenesis.

Concentration of 14,15-leukotriene C4 (eoxin C4) in bronchoalveolar lavage fluid.

BACKGROUND: There has been no information about the concentration of 14,15-leukotriene C4, which is generated by 15- and 12-lipoxygenase and has been recently named eoxin C4, in biological fluids. OBJECTIVE: To determine the clinical concentrations of eoxin C4 in various respiratory inflammatory diseases, we quantified eoxin C4 in relation to the concentrations of cysteinyl-leukotrienes (CysLTs) and 15-hydroxyeicosatetraenoic acid (15-HETE) in bronchoalveolar lavage fluid (BALF). METHODS: BALF fluid was obtained from patients with a number of inflammatory lung diseases. Eoxin C4 and CysLTs were quantified by enzyme immunoassay in combination with high-performance liquid chromatography. Eoxin C4 immunoassay does not detect eoxin D4 or eoxin E4. 15-HETE was quantified by gas chromatography-mass spectrometry using (18)O-labeled compounds as an internal standard. RESULTS: The concentration of eoxin C4 (median 1.4, range <1.12-6.7 pg/mL) was significantly lower than that of eoxin C4 or CysLTs (P<0.0001). The concentration of 15-HETE significantly correlated with those of LTC4 and CysLTs or the number and the percentage of eosinophils in BALF. On the other hand, eoxin C4 concentration did not correlate with eosinophil number or CysLTs concentration in BALF. CONCLUSIONS: This is the first study demonstrating the presence of eoxin C4 in human biological fluids. Further studies are necessary to elucidate the pathophysiological role of eoxin C4 in some respiratory inflammatory diseases.

CHFR, a potential tumor suppressor, downregulates interleukin-8 through the inhibition of NF-kappaB.

The mitotic checkpoint gene CHFR (checkpoint with forkhead and ring finger domains) is silenced in various human cancers by promoter hypermethylation, suggesting that CHFR is a tumor suppressor. Here, we show that CHFR functions as a negative regulator of the nuclear factor-kappaB (NF-kappaB) pathway. Expression of CHFR inhibited NF-kappaB reporter activity, whereas knockdown of CHFR activated reporter activity. These activities are independent of its RING finger domain. Furthermore, we found that CHFR physically interacts with p65 in cells. Electrophoretic mobility shift assays (EMSAs) and ELISA-based NF-kappaB-binding assays showed that CHFR negatively regulated transcriptional activity of p65. In addition, our data show that interleukin (IL)-8 is significantly downregulated by CHFR, and that the migration of human endothelial cells is suppressed in culture medium conditioned from CHFR-expressing cancer cells. Using a xenograft model, we show that neovascularization is suppressed by adenovirus-mediated transfer of CHFR. These results indicate that expression of CHFR markedly reduces the expression of IL-8 through the inhibition of NF-kappaB. As the NF-kappaB signaling pathway plays a critical role in the development and progression of cancer, our findings show the functional relationship between epigenetic alteration and inflammation/angiogenesis in human cancer cells, thereby showing several potential targets for therapeutic intervention.

Increased production of cysteinyl leukotrienes and prostaglandin D2 during human anaphylaxis.

BACKGROUND: Anaphylaxis is a life-threatening syndrome resulting from the sudden release of mast cell- and basophil-derived mediators into the circulation. However, pathological evidence of the association between inflammatory mediators and human anaphylaxis is insufficient. OBJECTIVE: The aim of this study was to better understand the relationship between in vivo production of inflammatory mediators and the pathogenesis of anaphylaxis. We also sought to evaluate mast cell activation in anaphylaxis. METHODS: We measured the concentrations of various inflammatory mediators in urine samples, which were collected from 32 anaphylactic patients during the onset of anaphylaxis and during clinical remission, 21 patients with asthma on acute exacerbation and 15 healthy control subjects. Blood and urine specimens were collected from the patients after provocation test. Urinary leukotriene E4 (LTE4), 9alpha, 11beta-prostaglandin F2 (9alpha, 11beta-PGF2), eosinophil-derived neurotoxin (EDN) and leukotriene B4 glucuronide (LTBG) concentrations were determined by enzyme immunoassay, and the activity of plasma platelet-activating factor acetylhydrolase and serum tryptase concentration were measured using commercially available kits. RESULTS: Significantly higher concentrations of urinary LTE4 and 9alpha, 11beta-PGF2, which immediately decreased during clinical remission, were observed in the anaphylactic patients than in asthmatic patients on acute exacerbation and healthy control subjects. Concentrations of EDN and LTBG were not significantly different among the anaphylactic patients, asthmatic patients on acute exacerbation and healthy subjects. There was a significant correlation between urinary LTE4 and 9alpha, 11beta-PGF2 concentrations in the anaphylactic patients (r=0.672, P=0.005, n=32). In addition, LTE4 concentration in patients with anaphylactic shock is significantly elevated compared with that in patients without anaphylactic shock. CONCLUSIONS: This is a report on the significant increase in urinary LTE4 and 9alpha, 11beta-PGF2 concentrations during anaphylaxis. Urinary LTE4 and 9alpha, 11beta-PGF2 concentrations may be a reliable marker of endogenous production of inflammatory mediators associated with anaphylaxis.

Comparison of cysteinyl leukotriene concentrations between exhaled breath condensate and bronchoalveolar lavage fluid.

BACKGROUND: Collection of exhaled breath condensate (EBC) is a simple, non-invasive method of obtaining samples from the airways and it can be repeated in short intervals without side effects; therefore, it provides an opportunity to monitor the changes in concentration of inflammatory mediators in the airways. However, EBC analysis still has several unresolved issues. OBJECTIVE: To better understand the characteristics of EBC, we compared cysteinyl leukotriene (CysLT) concentrations between bronchoalveolar lavage fluid (BALF) and EBC. We also attempted to correct CysLT concentrations in BALF and EBC diluted with saline and water vapour using biological markers. METHODS: EBC was collected from 14 patients with idiopathic pulmonary fibrosis before bronchoscopy. We measured CysLT concentrations and also quantified tyrosine, urea and total protein as possible biomarkers for correcting dilution. RESULTS: (1) We have validated the quantification of CysLTs in EBC. (2) Although a significant correlation was observed among tyrosine and urea concentrations in BALF, urea and total protein concentrations were below the detection limit in EBC. (3) CysLT concentrations were higher in BALF than in EBC (median, 15.96 pg/mL vs. 5.5 pg/mL; P=0.001) and there was no correlation of CysLT concentrations in BALF with those in EBC. A significant correlation of the ratio of total CysLT concentration to tyrosine concentration (CysLT/Y) in EBC with that in BALF was observed (r=0.547, P=0.043). (4) CysLT/Y in EBC correlated with serum KL-6 concentration and total cell count in BALF, and CysLT/Y in BALF also correlated with exhaled NO concentration and %VC. CONCLUSIONS: CysLT/Y in EBC significantly correlated with that in BALF and some clinical parameters correlated with CysLT/Y. Tyrosine concentration may be used to correct the dilution error for CysLT concentrations, and CysLT/Y in EBC can be a surrogate marker for CysLT concentrations in BALF.

Increased urinary leukotriene E4 concentration in patients with eosinophilic pneumonia.

Although eosinophils produce cysteinyl leukotrienes (CysLTs) in large quantities, information on the relationship between CysLTs and eosinophilic pneumonia (EP) is lacking. Inflammatory mediator concentrations in urine were quantified to clarify the relationship between CysLT concentrations and EP severity. Leukotriene (LT)E(4), eosinophil-derived neurotoxin (EDN), 9alpha,11beta-prostaglandin F2 and LTB(4) glucuronide concentrations were quantified in the urine of: EP patients during acute exacerbation and clinical remission; asthmatic patients during acute exacerbation and under stable conditions; and healthy control subjects. The urinary LTE(4) and EDN concentrations of EP patients during acute exacerbation were significantly higher than those of asthmatic patients and healthy subjects, and decreased immediately during clinical remission. The urinary LTE(4) concentration was associated with the urinary EDN concentration of EP patients during acute exacerbation. The urinary LTE(4) concentration significantly correlated with the diffusing capacity of the lung for carbon monoxide in EP patients during acute exacerbation. The increased urinary concentrations of leukotriene and eosinophil-derived neurotoxin were associated with acute exacerbation in eosinophilic pneumonia patients. The increased leukotriene concentration significantly correlated with diffusing capacity of the lung for carbon monoxide, suggesting that the monitoring of leukotriene concentration may aid in the management of eosinophilic pneumonia patients.

Frequent epigenetic inactivation of SFRP genes and constitutive activation of Wnt signaling in gastric cancer.

Activation of Wnt signaling has been implicated in gastric tumorigenesis, although mutations in APC (adenomatous polyposis coli), CTNNB1 (beta-catenin) and AXIN are seen much less frequently in gastric cancer (GC) than in colorectal cancer. In the present study, we investigated the relationship between activation of Wnt signaling and changes in the expression of secreted frizzled-related protein (SFRP) family genes in GC. We frequently observed nuclear beta-catenin accumulation (13/15; 87%) and detected the active form of beta-catenin in most (12/16; 75%) GC cell lines. CpG methylation-dependent silencing of SFRP1, SFRP2 and SFRP5 was frequently seen among GC cell lines (SFRP1, 16/16, 100%; SFRP2, 16/16, 100%; SFRP5, 13/16, 81%) and primary GC specimens (SFRP1, 42/46, 91%; SFRP2, 44/46, 96%; SFRP5, 30/46, 65%), and treatment with the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine rapidly restored SFRP expression. Ectopic expression of SFRPs downregulated T-cell factor/lymphocyte enhancer factor transcriptional activity, suppressed cell growth and induced apoptosis in GC cells. Analysis of global expression revealed that overexpression of SFRP2 repressed Wnt target genes and induced changes in the expression of numerous genes related to proliferation, growth and apoptosis in GC cells. It thus appears that aberrant SFRP methylation is one of the major mechanisms by which Wnt signaling is activated in GC.

Involvement of eicosanoids and surfactant protein D in extrinsic allergic alveolitis.

The pathophysiology of extrinsic allergic alveolitis (EAA) involves oxidative lung damage as well as interstitial and alveolar inflammation. Macrophages and mast cells are inflammatory components of EAA that produce both leukotrienes (LTs) and prostaglandin D2 (PGD2). In addition, PGD2 is also produced by the free-radical-catalysed peroxidation of arachidonic acid during oxidative stress. Urinary 8-iso prostaglandin F2alpha (8-isoPGF2alpha) and serum surfactant protein D (SP-D) are considered appropriate biomarkers of oxidative stress and interstitial lung disease activity, respectively. The present study aimed to assess the association of these biomarkers with the pathophysiology of EAA. Two cases of acute EAA caused by the inhalation of fungi spores were reported. Eight asthmatic patients and six healthy control subjects were also enrolled in the current study. The serum SP-D and urinary eicosanoid (LTE4, PGD2 metabolite (9alpha,11betaPGF2), 8-isoPGF2alpha) concentrations markedly increased during the acute exacerbation phase. These concentrations decreased following corticosteroid therapy in the EAA patients. There was a significant correlation between serum SP-D and urinary 9alpha,11betaPGF2 concentrations in the EAA patients. In conclusion, although the present study proposes that serum surfactant protein-D and urinary eicosanoids are new biomarkers involved in the various immunological responses in extrinsic allergic alveolitis, further large-scale studies are needed to investigate the role of these compounds, not just as biomarkers, but also as biological potentiators of extrinsic allergic alveolitis.

Aberrant DNA methylation associated with silencing BNIP3 gene expression in haematopoietic tumours.

Hypoxia is a key factor contributing to the progression of human neoplasias and to the development of resistance to chemotherapy. BNIP3 is a proapoptotic member of the Bcl-2 protein family involved in hypoxia-induced cell death. We evaluated the expression and methylation status of BNIP3 gene to better understand the role of epigenetic alteration of its expression in haematopoietic tumours. Methylation of the region around the BNIP3 transcription start site was detected in four acute lymphocytic leukaemia, one multiple myeloma and one Burkitt lymphoma cell lines, and was closely associated with silencing the gene. That expression of BNIP3 was restored by treatment with 5-aza2'-deoxycytidine (5-aza-dC), a methyltransferase inhibitor, which confirmed the gene to be epigenetically inactivated by methylation. Notably, re-expression of BNIP3 using 5-aza2-dC also restored hypoxia-mediated cell death in methylated cell lines. Acetylation of histone H3 in the 5' region of the gene, which was assessed using chromatin immunoprecipitation assays, correlated directly with gene expression and inversely with DNA methylation. Among primary tumours, methylation of BNIP3 was detected in five of 34 (15%) acute lymphocytic leukaemias, six of 35 (17%) acute myelogenous leukaemias and three of 14 (21%) multiple myelomas. These results suggest that aberrant DNA methylation of the 5' CpG island and histone deacetylation play key roles in silencing BNIP3 expression in haematopoietic tumours.

Increase in urinary leukotriene B4 glucuronide concentration in patients with aspirin-intolerant asthma after intravenous aspirin challenge.

BACKGROUND: Aspirin challenge of aspirin-intolerant asthma (AIA) patients causes a significant increase in leukotriene E4 (LTE4) concentration in urine. However, knowledge on leukotriene B4 (LTB4) generation in patients with AIA is insufficient. Recent research has demonstrated that exogenously administered LTB4 is excreted as glucuronide into the urine in human healthy subjects. OBJECTIVE: The purpose of this study is to estimate urinary LTB4 glucuronide (LTBG) concentration in the clinically stable condition in healthy subjects and asthmatic patients and to investigate changes in urinary LTBG concentration in patients with AIA after aspirin challenge. METHODS: A provocation test was performed by intravenous aspirin challenge. After urine was hydrolysed by beta-glucuronidase, the fraction containing LTB4 was purified by high-performance liquid chromatography and LTB4 concentration was quantified by enzyme immunoassay. Urinary LTBG concentration was calculated as the difference between the concentration obtained with hydrolysis and that without hydrolysis. RESULTS: (1) After hydrolysis, the presence of urinary LTB4 was verified by gas chromatography-mass spectrometry-selected ion monitoring. (2) The urinary LTBG concentration was significantly higher in the asthmatic patients than in the healthy subjects (median, 5.37 pg/mg creatinine [range 1.2-13] vs. 3.32 pg/mg creatinine [range, 0.14-10.5], P = 0.0159). (3) The patients with AIA (n = 7), but not those with aspirin-tolerant asthma (n = 6), showed significant increases in LTBG and LTE4 excretions after aspirin challenge. (4) When the concentrations after aspirin challenge were analysed simultaneously, a significant linear correlation was observed between urinary LTBG concentration and urinary LTE4 concentration in patients with AIA (Spearman's rank correlation test, r = 0.817, P = 0.0003). CONCLUSION: LTBG is present in human urine, albeit at a concentration lower than urinary LTE4. In addition to a marked increase in cysteinyl-leukotriene production, aspirin challenge induced LTB4 production in AIA patients.

Urinary 3-bromotyrosine and 3-chlorotyrosine concentrations in asthmatic patients: lack of increase in 3-bromotyrosine concentration in urine and plasma proteins in aspirin-induced asthma after intravenous aspirin challenge.

BACKGROUND: Eosinophil peroxidase and myeloperoxidase halogenate tyrosine residues in plasma proteins and generate 3-bromotyrosine (BY) and 3-chlorotyrosine (CY), respectively. OBJECTIVES: (1) To estimate urinary concentrations of BY and CY in asthmatic patients. (2) To investigate BY concentration in relation to urinary leukotriene E4 (LTE4) concentration in order to evaluate the activation of eosinophils in patients with aspirin-induced asthma (AIA). METHODS: BY and CY were quantified with a gas chromatograph-mass spectrometer using (13)C-labelled compounds as internal standards. RESULTS: (1) Activation of eosinophils and neutrophils by immobilized IgG1 induced preferential formation of BY and CY, respectively. (2) A significantly higher concentration of BY was observed in the urine from asthmatic patients than in that from healthy control subjects (45+/-21.7 vs. 22.6+/-10.8 ng/mg-creatinine, P<0.01). CY concentration was also elevated in the urine from asthmatic patients (4.4+/-3.2 vs. 1.5+/-1.0 ng/mg-creatinine, P<0.01). (3) After intravenous aspirin challenge of aspirin-induced asthmatic patients, the concentration of BY in urine did not significantly change. No significant change was also observed in the ratio of BY concentration to total tyrosine concentration in plasma proteins. In contrast, the concentration of urinary LTE4 significantly increased after the intravenous aspirin challenge. CONCLUSION: Determination of BY and CY concentrations may be useful for monitoring the activation of eosinophils and neutrophils in asthmatic patients, respectively. After aspirin challenge of AIA patients, the increased concentration of urinary LTE4 did not accompany changes in BY concentration in both urine and plasma proteins. These results may preclude the activation of eosinophils after aspirin challenge in patients with AIA.

Inactivation of class II transactivator by DNA methylation and histone deacetylation associated with absence of HLA-DR induction by interferon-gamma in haematopoietic tumour cells.

By presenting immunogenic peptides at the cell surface, major histocompatibility complex (MHC) class II molecules play a key role in the control of adaptive immune responses. Whether expressed constitutively or induced by interferon-gamma, expression of MHC class II molecules is regulated via coactivator class II transactivator (CIITA); moreover, suppression of their expression is one mechanism by which cancer cells escape host immunity. In this study, we surveyed the relationship between the expression of one MHC class II antigen, HLA-DR, and its coactivators in a group of haematopoietic cell lines, and explored the role of the aberrant DNA methylation in silencing HLA-DR expression. Among 26 cell lines studied, HLA-DR expression was lost from eight T-cell and two myeloid leukaemia cell lines, and this loss was closely associated with suppression of CIITA-PIV expression. Notably, nine of the 10 cell lines that lost CIITA-PIV expression showed methylation of the gene's 5' CpG island. Thus, DNA methylation is believed to inhibit the expression of MHC class II molecules in haematopoietic tumour cells by silencing its coactivator, CIITA-PIV. Furthermore, methylation of CIITA-PIV was detected in seven of 32 primary acute myeloid leukaemia specimens, indicating that epigenetic alteration is not a cell line-specific phenomenon. Collectively, these data suggest that, by suppressing expression of MHC class II molecules, epigenetic inactivation of CIITA provides a survival advantage to a subset of haematopoietic tumours.

Nasal blockage and urinary leukotriene E4 concentration in patients with seasonal allergic rhinitis.

BACKGROUND: Cysteinyl-leukotrienes have been reported to have a primary role in the induction of nasal blockage of allergic rhinitis. However, there has been little experimental evidence that substantiates the relationship between nasal blockage severity and urinary leukotriene E4 (U-LTE4) concentration in patients with seasonal allergic rhinitis (SAR). METHODS: The concentrations of urinary mediators in 20 SAR patients were measured using an enzyme immunoassay to determine the relationship between nasal blockage severity and U-LTE4 concentration in patients with SAR. RESULTS: The basal U-LTE4 concentration was significantly higher in SAR patients with severe nasal blockage than in those with mild nasal blockage and in healthy control subjects. Although U-LTE4 concentrationwas significantly higher in patients with both asthma and SAR than in SAR patients with mild nasal blockage, no significant difference in the U-LTE4 concentration between patients with both asthma and SAR and SAR patients with severe nasal blockage was found. There was a significant correlation between U-LTE4 and urinary 9alpha11beta-prostoglandin F2 (9alpha11betaPGF2) concentrations (rs = 0.51, P = 0.02) in SAR patients. CONCLUSIONS: Although specific sites and cells of cysteinyl-leukotriene biosynthesis could not be determined in this study, severe nasal blockage is associated with the increased excretion level of U-LTE4.

A comparative study of eicosanoid concentrations in sputum and urine in patients with aspirin-intolerant asthma.

BACKGROUND: Although many studies have assumed that the overproduction of cysteinyl- leukotrienes (cys-LTs) and an imbalance of arachidonic acid metabolism may be plausible causes for the pathogenesis of aspirin-intolerant asthma (AIA), there has been little experimental evidence to substantiate this notion in lower airways of patients with AIA. OBJECTIVES: The purpose of this study was to compare the eicosanoid concentrations in sputum and urine from patients with AIA. METHODS: The concentrations of sputum cys-LTs, prostaglandin E2 (PGE2), PGF2alpha, PGD2 and thromboxane B2 were measured to assess local concentrations of eicosanoids in patients with AIA and in those with aspirin-tolerant asthma (ATA). The concentrations of two urinary metabolites, leukotriene E4 (LTE4) and 9alpha11betaPGF2, were also measured to corroborate the relationship between the eicosanoid biosynthesis in the whole body and that in lower airways. RESULTS: The concentration of PGD2 in sputum was significantly higher in patients with AIA than in those with ATA (median, 5.3 pg/mL vs. 3.1 pg/mL, P < 0.05), but there was no significant difference in the concentration of the corresponding metabolite, 9alpha11betaPGF2, between the two groups. No differences were noted in the concentrations of other prostanoids in sputum between the two groups. The sputum cys-LT concentrations showed no differences between the two groups, in spite of the observation that the concentration of urinary LTE4 was significantly higher in patients with AIA than in those with ATA (median, 195.2 pg/mg-cre vs. 122.1 pg/mg-cre, P < 0.05). There was a significant correlation among the concentration of cys-LTs, the number of eosinophils and the concentration of eosinophil-derived neurotoxin (EDN) in sputum. CONCLUSION: The urinary concentration of LTE4 does not necessary reflect cys-LT biosynthesis in lower airways. A significantly higher concentration of PGD2 in sputum from patients with AIA suggests the possible ongoing mast cell activation in lower airways.

Levels of cysteinyl leukotriene receptor mRNA in human peripheral leucocytes: significantly higher expression of cysteinyl leukotriene receptor 2 mRNA in eosinophils.

BACKGROUND: Cysteinyl leukotrienes (CysLTs) have been implicated as important contributors in the pathophysiology of asthma and their biological effects are mediated by at least two distinct G-protein-coupled receptors. cDNA sequences of cysteinyl leukotriene receptor 1 (CysLTR1) and cysteinyl leukotriene receptor 2 (CysLTR2) have recently been elucidated. OBJECTIVES: Our aim is to explore gene expression and the comparative expression of CysLTR1 mRNA and CysLTR2 mRNA in human peripheral blood leucocytes. METHODS: Gene expression of CysLTR1 and CysLTR2 mRNAs in human peripheral blood eosinophils, neutrophils, monocytes and T lymphocytes has been measured by competitive reverse transcription-polymerase chain reactions using RNA or DNA competitors. RESULTS: (a) When cellular levels of CysLTR1 mRNA were normalized to those of G3PDH mRNA, the relative concentration of CysLTR1 mRNA in eosinophils (43.8 +/- 37.2, n = 29) was significantly higher than that in neutrophils (18.7 +/- 23.3, n = 11), monocytes (0.93 +/- 1.1, n = 10) and T lymphocytes (3.4 +/- 2.4, n = 11). (b) When measured using each DNA competitor, mRNAs for both types of CysLTR coexisted in each type of leucocyte. The ratio of CysLTR1 mRNA to CysLTR2 mRNA was significantly lower in eosinophils (0.65 +/- 0.42, n = 12) than in neutrophils (6.9 +/- 4.9, n = 12), monocytes (1.8 +/- 0.9, n = 10) and T lymphocytes (4.5 +/- 5.7, n = 10). (c) Human umbilical vein endothelial cells expressed CysLTR2 mRNA, but not CysLTR1 mRNA. CONCLUSION: These studies reveal that CysLTR1 mRNA and, in particular, CysLTR2 mRNA are abundantly expressed at high levels in eosinophils, raising the possibility that CysLTR2 may have an important physiological role in eosinophils and a CysLTR2 antagonist may be a good target for preventing signal transduction by CysLTs in eosinophils.