Serum C-reactive protein and white blood cell level as markers of successful percutaneous drainage of acute sterile peripancreatic fluid collection.

BACKGROUND: Percutaneous drainage is not a widely used therapeutic method recently for evacuating peripancreatic sterile fluid collections in patients with severe acute pancreatitis.However, many clinical studies have proved its positive effects. AIM: We tested the changes in serum laboratory parameters:C-reactive protein (CRP), complement factor 3-4 (C 3-4),tumor necrosis factor a (TNF-a), amylase, lipase and white blood cell (WBC) count in patients treated by percutaneous drainage. PATIENTS AND METHODS: 10 patients with severe acute pancreatitis with peripancreatic fluid collection were monitored.Laboratory parameters and the amount of drained fluid were measured on the 1st, 5th and 10th day. Statistical analysis was performed by using Statistica for Windows (Version 7.0)software. P values less than 0.05 were considered statistically significant. RESULTS: We found significant positive correlation between the CRP and WBC serum level and volumes of the drained fluid. We used these parameters as markers of successful percutaneous drainage in case of patients with severe acute pancreatitis complicated with sterile peripancreatic fluid.There was no significant change in the levels of C 3-4,tumor necrosis factor-Î+-, amylase and lipase. CONCLUSIONS: Monitoring of serum CRP and WBC levels maybe recommended for follow up after percutaneous drainage of peripancreatic fluid. ABBREVIATIONS: CRP: C-reactive Protein TNFÎ+-: Tumour Necrosis Factor a, C3-4: Complement 3-4 WBC: White Blood Cell CT: Computed Tomography.

The effects of various doses of bacterial lipopolysaccharide on the expression of CD63 and the release of histamine by basophils of atopic and non-atopic patients.

OBJECTIVE: We tested the effect of various doses of bacterial lipopolysaccharide (LPS, endotoxin) on the expression of CD63 and the in vitro release of histamine by basophils stimulated with ragweed allergen in patients with or without ragweed and mite allergies. METHODS: The peripheral blood of 11 patients with ragweed allergy, 10 patients with mite allergy and 14 control patients was incubated with ragweed allergen extract following pretreatment with varying doses of LPS. The expression of CD63 in basophils was measured by flow cytometry, and the release of histamine was determined by ELISA. RESULTS: In the samples of patients with ragweed allergy that were exposed to specific allergen, only high doses of LPS significantly elevated the expression of CD63 (200 ng/ml; 1,000 EU/ml) and the release of histamine (2,000 ng/ml; 10,000 EU/ml). There was no effect of LPS in any other cases. CONCLUSIONS: Bacterial LPS (endotoxin) concentrations higher than 200 ng/ml (1,000 EU/ml), which rarely occurs in nature, could only activate the basophils from atopic patients whilst in the presence of the specific allergen. Thus, the restoration of the urban, "microbe-poor" milieu with endotoxin (as LPS) can be a promising and harmless approach for allergy prevention.

Analysis of serum cytokine levels in children with chronic cough associated with Toxocara canis infection.

Toxocara infection is associated with an increased prevalence of airway symptoms and may be a possible aetiologic agent of chronic cough. The occurrence of toxocariasis in Hungary is mild and/or sporadic. The purpose of this study was to investigate the levels of serum cytokines (IL-1, IL-2, IL-4, IL-5, IL-6, IL-10, IL-13, IFN-gamma and TNF-alpha) and total IgE, the blood eosinophil count, the results of skin prick and non-specific bronchus provocation tests in Toxocara-seropositive children with chronic cough relative to those in healthy controls. The patients exhibited moderate eosinophilia, significantly elevated levels of serum total IgE, IL-6, IL-10, IL-13 and IFN-gamma, and higher skin reactivity to common allergens, whereas the bronchial hyperreactivity was similar in the two groups. The protective proinflammatory cytokines (IL-6, IFN-gamma and IL-13) in association with the anti-inflammatory cytokine (IL-10) were simultaneously increased in Toxocara-infected children with chronic cough. During infections, the activation and suppression of immune processes occur simultaneously and cytokines of Th1/Th2 and regulatory T cells contribute to the regulation of the immune response evoked by helminth infections (depending on the parasite load, the timing and duration of the infection and the status of the host immune system).

Serum and urinary cytokine levels of SLE patients.

Systemic lupus erythematosus (SLE) is a chronic, relapsing, polysystemic autoimmune disease with various clinical signs. The prognosis of SLE patients is influenced by neuropsychiatric and renal involvement. Lupus nephritis (LN) is present in 40-60% of patients. Classical laboratory parameters are not sensitive and specific in prediction renal flares, over the last few years there has been a growing interest in searching novel lupus biomarkers predicting future flares. Our goal was to detect serum and urinary level of cytokines in 36 patients with lupus nephritis (34 female and 2 male, mean age: 43.36 +/- 11.53 years), 23 patients with SLE without renal involvement (19 women and 4 men, mean age: 54 +/- 8.71) (both groups followed by the 3rd Department of Internal Medicine, Division of Clinical Immunology, University of Debrecen) and 30 healthy controls (23 female and 7 male, mean age: 45.5 +/- 12.4). Serum IL-1 (interleukin), IL-2 (both p < 0.05), IL-6, IL-13 and IFN-gamma (p < 0.001) levels were significantly higher in lupus nephritis patients, as compared to patients with SLE without renal involvement and healthy controls. Urinary level of IL-1 and TNF-alpha were significantly higher in SLE patients without renal disease (p = 0.012 and p < 0.001), while urinary IFN-gamma was significantly higher in LN patients (p = 0.002). Measurement of IL-6 level in SLE patients could help to predict future renal involvement of SLE patients.

Different effects of bortezomib on the expressions of various protein kinase C isoenzymes in T cells of patients with systemic lupus erythematosus and in Jurkat cells.

The effects of proteosome inhibitor Bortezomib (BZ) were studied in vitro for 24 h on the protein kinase C (PKC) profiles, rates of proliferation and apoptosis in Jurkat cells and lymphocytes of 10 patients with systemic lupus erythematosus (SLE) and nine healthy subjects. The expressions of PKC proteins, the rates of proliferation and apoptosis were determined. The effects of BZ were different in the Jurkat and lupus T cells. Whereas BZ elevated the expression of PKC θ, δ and ξ isoenzymes in the Jurkat cells, it was unable to do that in the lupus T cells. BZ induced a dose-dependent increase in the apoptosis of Jurkat cells, while decreased the proliferation. The same effect of BZ was observed on the apoptosis of lymphocytes both in SLE and healthy subjects at concentrations higher than the therapeutic dose. We conclude that BZ treatment in vitro was not able to restore the SLE-specific defect (decrease) in the expression of PKC isoenzymes in the T cells as it was expected. This can be a limiting factor in the positive clinical effects of BZ in lupus.

Anti-TNF-alpha antibody (infliximab) therapy supports the recovery of eNOS and VEGFR2 protein expression in endothelial cells.

The aim of this study is to investigate the effect of sera obtained from patients of Crohn's disease treated by anti-TNF-alpha antibody (Infliximab) on the expression of endothelial nitric oxide synthase (eNOS) and vascular endothelial growth factor receptor-2 (VEGFR2) protein in human umbilical vein endothelial cells (HUVEC) cultured in vitro. HUVEC was cultured in the presence of sera derived from patients before and after treatment, or from healthy individuals. Effects of sera on the expression of eNOS and VEGFR2 were monitored by determination of mRNA and protein levels using real time quantitative PCR and Western blot analysis, respectively. The serum of Crohn's patients contained elevated levels of TNF-alpha (34±1.80 pg/mL), which resulted in a decrease in the protein level of eNOS in HUVEC with a simultaneous induction of VEGFR2. Infliximab treatment normalized the expression level of these proteins by decreasing TNF-alpha level, particularly in those cases when clinical healing was also recorded, and it also conferred restitution of the level of angiogenic cytokines. Results suggest that altered angiogenesis possibly contributes to the initiation and perpetuation of inflammatory processes in inflammatory bowel disease (IBD). Endothelial dysfunction, a selective feature of Crohn's disease is beneficially affected by intravascular TNF-alpha neutralization.

Assessment of IgG antibodies to oxidized LDL in patients with acute coronary syndrome.

OBJECTIVES: Circulating IgG antibodies to oxidized low-density lipoprotein (anti-oxLDL) have been implicated in the development of atherosclerotic plaques. In this study, we investigated the prognostic value of IgG anti-oxLDL antibodies in patients with acute coronary syndrome (ACS). METHODS: In total 54 patients with ACS and 41 matched healthy controls were involved in this prospective study. Serum IgG anti-oxLDL levels were assessed by ELISA. RESULTS: Higher IgG anti-oxLDL levels were found in patients with ACS versus controls (22.8 ± 23.3 vs. 7.5 ± 5.27 EU/ml, p < 0.0001). IgG anti-oxLDL concentrations were significantly higher in ACS patients with unstable clinical complications (circulatory insufficiency, malignant arrhythmias, recurring ischaemic pain, positive stress-test, need for urgent coronary intervention or sudden cardiac death) versus those without such complications (30.0 vs. 11.7 EU/ml, p < 0.001). Twelve patients (22%) were taking statins. Patients on statins had a significant reduction in clinical complications (33%) versus patients not receiving statin therapy (61%). IgG anti-oxLDL levels were also different in these two groups (11.4 vs. 25.8 EU/ml, respectively; p = 0.03). Serum IgG anti-oxLDL levels correlated with the subsequent development of unstable coronary events. Levels of anti-oxLDL significantly decreased in response to statin therapy, independently of its lipid-lowering effect. CONCLUSIONS: Anti-oxLDL antibodies are involved in ACS. The association of anti-oxLDL with unstable clinical complications may indicate the role of this antibody in plaque destabilization.

Altered T-cell and regulatory cell repertoire in patients with diffuse cutaneous systemic sclerosis.

OBJECTIVES: The aim of this study was to evaluate a wide spectrum of peripheral immune-competent cell types, reflecting overall disturbances in immune homeostasis, characteristic of systemic sclerosis (SSc). We also assessed visceral organ involvement and evaluated the relationship between cell proportions and clinical symptoms of the disease. METHODS: Twenty-one patients with diffuse cutaneous SSc (dcSSc) and 15 healthy individuals participated in the study. Peripheral blood lymphocyte subgroups were quantified by flow cytometry, soluble cytokines were assessed by enzyme-linked immunosorbent assay (ELISA), serum complement levels were measured by nephelometry, and autoantibodies were determined by indirect immunofluorescence staining and ELISA technique. Functional tests of regulatory T (Treg) cells were also carried out. RESULTS: Patients with SSc had higher percentages of activated CD3+/HLA-DR+ T cells. Comparing naive vs. memory subsets of CD4+ and CD8+ T cells, a shift towards central memory phenotype was observed in SSc. Natural killer (NK) and T-helper (Th)17 cell percentages were increased, while NKT, Th1, Treg type 1 (Tr1), and CD4+CD25+ Treg cell percentages were decreased in patients. Moreover, the suppressor activity of CD4+CD25+ Treg cells was lower in SSc. Negative correlations occurred between modified Rodnan skin score (MRSS) and Tr1 cell percentages and between complement levels and CD4+CD25+ Treg cells. We also found decreased interleukin (IL)-10 levels in SSc. CONCLUSIONS: Our data suggest that the increased Th17/CD4+CD25+ Treg ratio and the altered regulatory function of CD4+CD25+ Treg cells play an important role in the development of SSc. Moreover, our study reveals the potential role of the decreased profile of IL-10-producing Tr1 cells in the progression of disproportionate immune responses in SSc.

Clinical and immunoserological characteristics of the transition from primary to overlap antiphospholipid syndrome.

Antiphospholipid syndrome (APS) is a distinct clinical entity characterized by arterial and venous thromboembolic events, recurrent fetal loss and the presence of antiphospholipid antibodies in the patients' sera. In primary APS, there is no detectable underlying disease, while overlap APS is associated with clinical syndromes including systemic autoimmune diseases, infections, or malignancies. We carried out a retrospective analysis of serological and clinical manifestations as well as assessed outcome-measures in 165 patients with primary APS. Thrombotic manifestations and possible signs of autoimmune diseases were determined at the time of the diagnosis, followed by the analysis of recurrent thrombotic events and effects of therapy during the follow-up period. Among the 165 patients with primary APS at onset, 105 patients (63%) remained primary APS after a mean 5.2 years of follow-up. In 14% of the patients, subsequently APS became associated with various characteristics of undifferentiated connective tissue disease. Finally 23% of patients evolved into a definitive systemic autoimmune disease during a mean 9.75 years of follow-up. Recurrent thrombotic events were registered in 24% of patients. Our results suggest that primary APS may be considered as a potential early phase of a dynamic transition towards a well-defined systemic autoimmune disease.

Cells with regulatory function of the innate and adaptive immune system in primary Sjögren's syndrome.

The aim of the present study was to describe subsets of cells with regulatory properties in primary Sjögren's syndrome (pSS), and to correlate these cell populations with clinical symptoms. Among the 32 investigated patients, 23 had extraglandular manifestations (EGMs), while nine had only glandular symptoms. Twenty healthy individuals served as controls. The percentages of natural killer (NK), natural killer T cells (NK T), interleukin (IL)-10 producing T regulatory type 1 (Tr1) cells and CD4(+)CD25(+) regulatory T cells (T(reg)) cells were determined by flow cytometry and serum cytokine levels of IL-4, IL-6, IL-10, tumour necrosis factor (TNF)-alpha and interferon (IFN)-gamma were evaluated by enzyme-linked immunosorbent assay (ELISA). Functional tests were carried out to assess the suppressor properties of T(reg) cells in patients and controls. Peripheral NK, NK T and Tr1 cell percentages were elevated in pSS, while CD4(+)CD25(+) T(reg) cells showed reduced frequencies in patients compared to controls. In pSS, elevated percentages of NK T, Tr1 and CD4(+)CD25(+) T(reg) cells were observed in patients with EGMs, when compared to patients with sicca symptoms only. CD4(+)CD25(+) T(reg) cell percentages showed a negative correlation with sialometry values. The in vitro functional assay demonstrated lower suppression activity of CD4(+)CD25(+) T(reg) cells in patients compared to controls. Serum IL-6 and TNF-alpha levels were elevated, while IL-10 was decreased in patients compared to controls. Negative correlation was found between IL-10 levels and the percentages of Tr1 cells. Changes in the investigated subsets of regulatory cells in pSS may contribute to the development and progression of the disease.

Regulatory T cells in atopic dermatitis: epidermal dendritic cell clusters may contribute to their local expansion.

BACKGROUND: Regulatory T cells (Tregs) have an essential role in tolerance and immune regulation. However, few and controversial data have been published to date on the role and number of these cells in atopic dermatitis (AD). OBJECTIVES: To investigate the number of CD4+CD25+FOXP3+ Tregs and interleukin 10-producing T regulatory type 1 (Tr1) cells in patients with AD. METHODS: Peripheral blood and skin biopsy samples from atopy patch test (APT)-positive patients with acute- and chronic-phase AD were investigated. Immunohistochemistry was applied to identify CD4+CD25+FOXP3+ Tregs in the skin, while flow cytometry was used to detect CD4+CD25highFOXP3+ Tregs and Tr1 cells in the peripheral blood. RESULTS: In the peripheral blood samples of patients with AD significantly elevated numbers of Tr1 cells were found. Although neither the absolute number nor the percentage of CD4+CD25highFOXP3+ Tregs showed significant alteration in the peripheral blood of patients, increased numbers of FOXP3+ Tregs were detected in skin biopsy specimens. All of the APT-positive skin samples showed epidermal dendritic cell aggregates, morphologically consistent with so-called Langerhans cell microgranulomas, which also contained intermingled FOXP3+ Tregs. CONCLUSIONS: Tr1 cell numbers were elevated in the peripheral blood and increased numbers of CD4+CD25highFOXP3+ Tregs were detected in the skin of patients with AD. The epidermal dendritic cell clusters in APT-positive lesional skin showed a close connection to the FOXP3+ Tregs.

Immunological features of primary anti-phospholipid syndrome in connection with endothelial dysfunction.

OBJECTIVES: To describe how certain peripheral immune parameters reflect the inflammatory alterations in patients with primary APS. METHODS: Twenty-eight patients with newly diagnosed primary APS were studied. The control group included 26 patients with stable coronary disease and 38 healthy individuals. Peripheral blood lymphocyte subgroups were quantified, intracellular cytokines were measured by flow cytometry, soluble cytokines and auto-antibodies were assessed using ELISA. Endothelial dysfunction was evaluated by measuring endothelium-dependent (flow-mediated; FMD) vasodilation. Carotid duplex ultrasound was performed to quantify the carotid artery intima-media thickness (IMT). Stiffness parameters, augmentation index (AIx) and pulse wave velocity (PWV) were assessed by TensioClinic technology. RESULTS: Serum IL-4 and IL-6 levels were significantly higher in APS. CD4+IL10+ and CD8+IL10+ cell percentages in APS were significantly increased compared with controls. Th 0 and T cytotoxic 0 cell percentages were significantly decreased in patients compared with controls. FMD in APS was significantly lower, while IMT was higher than that of controls. FMD showed strong association with stiffness parameters, AIx and PWV. A significant negative linear correlation was detected between PWV and CD8+IL10+ cell percentages and significant positive linear correlation was found between PWV and CD8+IL10- cell percentage. CONCLUSION: In APS, the orchestrated pro-inflammatory cascade can eventually result in endothelial dysfunction, leading to the characteristic vascular abnormalities of the disease.

Frequencies of genetic polymorphisms of TLR4 and CD14 and of HLA-DQ genotypes in children with celiac disease, type 1 diabetes mellitus, or both.

OBJECTIVES: Besides the central role of the adaptive immune system, a disturbance of innate immunity is also involved in the pathogenesis of celiac disease (CD). Inasmuch as CD and type 1 diabetes mellitus (T1DM) frequently coexist because of a common genetic predisposition, our aim was to study the frequency of CD14 C-260T and TLR4 A+896G single nucleotide polymorphisms (SNPs) and the distribution of HLA-DQ genotypes in children affected by CD, T1DM, or both. PATIENTS AND METHODS: TLR4 and CD14 SNPs were tested by polymerase chain reaction, followed by restriction fragment length polymorphism analysis in 80 children with T1DM, 100 children with CD, and 47 children with both CD and T1DM. Determination of HLA-DQ alleles was done by sequence-specific polymerase chain reaction. Frequencies were compared with those of healthy control children. RESULTS: The prevalence of the homozygous CD14 C-260TT genotype was significantly (P = 0.0081) lower in children with T1DM but not in those with CD and T1DM, compared with control children. No difference was found in the genotype and allele frequencies of TLR4 between the studied groups. In patients with T1DM, the frequency of the homozygous HLA-DQ8 genotype was significantly higher than in CD, whereas the frequency of homozygous or heterozygous HLA-DQ2 genotypes did not differ from that in control children. In patients with CD, both homozygous and heterozygous HLA-DQ2 genotypes were significantly more frequent than in the control and T1DM groups, and no elevation in the frequency of the HLA-DQ8 genotypes was observed. In patients with T1DM and those with CD and T1DM, the occurrence of HLA-DQ2/8 heterozygosity was significantly higher than in children with CD only and in control children. CONCLUSIONS: Our results suggest that in patients with T1DM, the CD14 C-260TT homozygous genotype increases the risk for the development of CD. The distribution of HLA-DQ genotype is different in children with CD and T1DM than in children with CD or T1DM only. Determination of the HLA-DQ genotype in children with T1DM may help in estimating the risk for the development of CD.

Use of three lumen catheter facilitates bowel movement after pancreato-duodenectomy.

BACKGROUND/AIMS: The advantages of jejunal nutrition in postoperative bowel paralysis following pancreato-duodenectomy were analyzed. METHODOLOGY: Patients resected for pancreatic cancer received 25 kcal/kg/day and were followed up for 10 days postoperatively. Nasojejunal tube ensured enteral feeding in 16 patients (Gr. I), 6 patients (Gr. II) were nourished parenterally. Laboratory parameters, outcome were compared. Bowel movements were registered. Patients of Gr.1 received 25 kcal/kg parenterally. Jejunal nutriment (1.5 cal/mL) followed gradually up to 1500mL. Parenteral nutriment decreased reflecting enteral intake. Patients of Gr. II were nourished parenterally only for 8 days. Laboratory data were measured preoperatively, on the 1st, 4th, 10th days. RESULTS: The first stool appeared on the 4th day in Gr. I In Gr. II the bowel movement was delayed by 8 days. Laboratory data from the 1st, and 10th days were compared. In Gr. I serum total protein increased from 48.06 to 58.7g/L (p<0.001), serum albumin from 27.5 to 32.2g/L (p<0.02), CRP decreased from 117.8 to 74.1mg/L (p<0.035). No changes were significant in Gr. II. Length of hospitalization, weight loss did not differ between the 2 groups. CONCLUSIONS: Immediately postoperative use of a three-luminal tube ensured early enteral nutrition, improved serum total protein, albumin values and facilitated bowel movements.

The severity of systemic lupus erythematosus negatively correlates with the increasing number of CD4+CD25(high)FoxP3+ regulatory T cells during repeated plasmapheresis treatments of patients.

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease characterized by increased pathologic autoantibody production. A decrease in the number of CD4+CD25(high)FoxP3+ regulatory T cells can play a key role in the loss of tolerance to self antigens. Our aim was to determine the absolute number of peripheral CD4+CD25(high)FoxP3+ T cells in 44 patients with SLE, furthermore, to measure the changes in the number of CD+CD25(high)FoxP3+ T cells in 5 patients with severe SLE treated with repeated plasmapheresis for 4-6 days in comparison to the changes in the activity of disease (SLEDAI). Percent of CD4+CD25(high)FoxP3+ T cells were measured by flow cytometry. The absolute number of peripheral CD4+CD25(high)FoxP3+ T cells was significantly decreased in the 44 patients with SLE compared to the healthy controls n = 32 (0.012 +/- 0.006 vs. 0.038 +/- 0.017 G/L, p < 0.05). In the 5 patients with severe SLE the repeated plasmapheresis treatments increased the peripheral number of CD4+CD25(high)FoxP3+ T cells. As the number of CD4+CD25(high)FoxP3+ T cells increased during the treatment, the activity of disease (the value of SLE activity index) decreased. In the peripheral blood of SLE patients not only the ratio was decreased (as it was published earlier) but also the absolute number of these regulatory T cells. The repeated plasmapheresis treatments of SLE patients induced a significant increase in the number of peripheral CD4+CD25(high)FoxP3+ T cells in parallel to the decrease in the values of SLEDAI (the activity of disease). This phenomenon is, among others, possibly due to the elimination of interpheron-alpha and lymphocytotoxic antibodies during plasmapheresis.

Measurement of natural (CD4+CD25high) and inducible (CD4+IL-10+) regulatory T cells in patients with systemic lupus erythematosus.

Abnormalities of regulatory T cells may play an important role in the loss of self-tolerance, which is a major characteristic of lupus. The objective of this study was to determine the ratio and the number of natural CD4+CD25highFoxp3+ and inducible CD4+IL-10+ regulatory T cells in lupus patients and to search correlation with disease activity. Seventy-two Hungarian lupus patients were enrolled in the study. Fourty-one age- and sex matched healthy donors served as controls. Flow cytometry was used for the quantification of CD4+CD25high Foxp3+ (nTreg) and CD4+IL-10+ (iTreg) cells. The ratio (3.06 +/- 1.45%) and the number (0.019 +/- 0.012 x 10(9)/L) of nTreg cells decreased in lupus significantly (P < 0.001 in both) as compared to normal controls (4.26 +/- 1.01% and 0.039 +/- 0.017 x 10(9)/L). The ratio of iTreg cells were significantly higher in patients than in controls (20.92 +/- 14.02% versus 15.49 +/- 11.65%, P < 0.03), but the number of these cell type did not differ in significant manner (0.314 +/- 0.236 x 10(9)/L versus 0.259 +/- 0.183 x 10(9)/L). The 19 active patients were characterised by significantly higher disease activity index (SLEDAI 8.63 +/- 2.95 versus 1.74 +/- 1.68, P < 0.001) and anti-DNA concentration (117.85 +/- 145.89 versus 37.36 +/- 68.85 IU/mL, P = 0.001) as compered to the 52 inactive patients. Furthermore, active patients required higher dose of methylprednisolon than inactive ones (14.8 +/- 10.6 versus 4.8 +/- 3.4 mg/day, P < 0.001). However, we did not find statistical significant difference in the number and ratio of the examined cell populations regarding to disease activity. Altered ratio and number of both natural and inducible regulatory T cells may play a role in the pathogenesis of lupus. There are small but appreciable difference in the number of regulatory T cells between inactive patients and healthy controls. It suggests that immunoregulatory deficiencies are present in the inactive stage of the disease also.

Adenosine inhibits the release of arachidonic acid and its metabolites (AAM) in activated human peripheral mononuclear cells.

OBJECTIVE: The effects of adenosine (Ado) and subtype-specific activators of adenosine receptors (A(1), A(2A), A(2B) and A(3)) were studied on the release of arachidonic acid (AA) and its metabolites (AAM) from human peripheral mononuclear cells (monocytes). MATERIALS AND METHOD: Adenosine and the selective agonists and antagonists of adenosine receptors were used. (3)H-AA and its metabolites released into the medium were determined by measurement of the total (3)H radioactivity released without separating the AAM. RESULTS: In the cells activated by protein kinase C specific phorbol ester (phorbol 12-myristate 13-acetate) and Ca(2+) ionophore (A23187), adenosine and two subtype-specific receptor agonists, CPA(A(1)) and CGS 21680 (A(2A)) induced concentration-dependent inhibition of the release of AAM, whereas stimulation of A(2B) or A(3) receptors was ineffective. The rank order of potency for the inhibition of AAM release was as follows: CGS 21680 = CPA > adenosine > NECA (in the presence of ZM 24185 and DPCPX as A(2A) and A(1) adenosine receptor antagonists, respectively) = IB-MECA. Adenosine inhibited the release of AAM only at and above the concentration of 100 muM, whereas the inhibitory effect of A(1) and A(2A) receptor specific agonists appeared at a concentration of 10(-7) M. CONCLUSIONS: It can be concluded that adenosine physiologically may not have a significant effect on the AAM release of circulating monocytes, but in pathological conditions, where the local Ado concentrations increases, this nucleoside, through activation of A(2A) and A(1) receptors can exert, at least in part, an antiinflammatory action by decreasing proinflammatory AAM production.

Diagnostic significance of HLA-DQ typing in patients with previous coeliac disease diagnosis based on histology alone.

BACKGROUND: Coeliac disease is strongly associated with human leukocyte antigen (HLA)-DQ2 or DQ8 genotypes. The diagnosis is based on demonstrating crypt-hyperplastic villous atrophy, endomysial or transglutaminase antibodies and correlation of disease activity with gluten intake. AIM: To evaluate the clinical utility of HLA-DQ typing, when coeliac disease diagnosis had previously been established solely by histology. METHODS: HLA-DQ alleles, endomysial and transglutaminase antibodies were investigated and histology slides reviewed in 70 patients diagnosed 2-25 years earlier by small-intestinal biopsy but without measuring endomysial or transglutaminase antibodies. Patients without DQ2 or DQ8 or without unequivocal villous atrophy were followed-up on free diet by using serology and biopsies. RESULTS: All 40 endomysial/transglutaminase antibodies positive patients carried DQ2 or DQ8, and 39 of them had severe villous atrophy. Only 56% of patients without endomysial or transglutaminase antibodies positivity had DQ2 or DQ8 (P < 0.001). Seropositivity and relapse developed in 4 of 11 DQ2 positive but in none of 15 DQ2 and DQ8 negative patients on long-term gluten exposure. CONCLUSIONS: Coeliac disease diagnosis based solely on histology is not always reliable. HLA-DQ typing is important in identifying DQ2 and DQ8 negative subjects who need revision of their diagnosis, but it does not have additive diagnostic value if endomysial positivity is already known.

Regulatory T cells in peripheral blood of patients with mixed connective tissue disease.

OBJECTIVE: To determine the percentage and absolute number of CD4+ regulatory T-cells (Treg) in 48 patients with mixed connective tissue disease (MCTD). METHODS: Data were classified on the basis of the stage of the disease: 17 patients were in the active stage and 31 in the inactive stage. The absolute number of CD4+/intracellular interleukin-10+ (IL-10+) and CD4+CD25+high Treg cells was determined by flow cytometry. The percentage of CD4+CD25+high suppressor T-cells was determined on the basis of Foxp3 expression. RESULTS: The percentage and the absolute number of CD4+CD25+high Treg cells were lower in patients than in healthy controls (p<0.04), and were further decreased in patients with active MCTD and were lower than in the inactive stage (p<0.01). There was an increase in the percentage and absolute number of CD4+IL-10+ Treg cells in patients with MCTD compared to the healthy controls (p<0.02). The percentage of CD4+IL-10+ Treg cells was higher in the active stage of MCTD than in the inactive stage of the disease (p<0.005). However, we did not find any significant difference in the absolute number of CD4+IL-10+ Treg cells between the patients in the active and inactive stages of MCTD. CONCLUSION: Our results suggest that the decrease in the number of CD4+CD25+high Treg cells in an important factor in the immunoregulatory disturbance in patients with MCTD. We suggest that the increase in the number of CD4+IL-10+ Treg cells is a compensatory mechanism aiming to restore the balance between type 1 and type 2 cytokines in MCTD.