Influence of different spectra of NOTCH3 variants on the clinical phenotype of CADASIL - experience from Slovakia.

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is the most common hereditary vascular disorder causing ischaemic attacks and strokes in middle-aged adults. Though the clinical spectrum includes some typical symptoms, recognition of the disease, especially at an earlier stage, is very difficult because of the highly variable manifestation and incomplete clinical picture. Characteristic brain MRI findings and the presence of pathogenic variants in the NOTCH3 gene are fundamental for CADASIL diagnosis. In this paper, we provide the first comprehensive report on CADASIL patients from Slovakia. Altogether, we identified 23 different pathogenic variants in 35 unrelated families. In our cohort of patients with clinical suspicion of CADASIL, we found a causal genetic defect and confirmed the diagnosis in 10.2% of cases. We present the case reports with up-to-date unpublished NOTCH3 variants and describe their phenotype-genotype correlation: p.(Cys65Phe), p.(Pro86Leu/Ser502Phe), p.(Arg156*), p.(Cys408Arg), p.(Tyr423Cys), p.(Asp1720His), and p.(Asp1893Thrfs*13). The most frequently described location for pathogenic variants was in exon 4, whereas the most common single variant was p.Arg1076Cys in exon 20. Based on the results of our study, we propose a re-evaluation of the criteria for the selection of patients suitable for NOTCH3 gene analysis. We hereby state that the currently used protocol of a high score requirement is not ideal for assessing molecular analysis, and it will be desirable to be less strict in criteria for genetic testing.

Distribution of the most common polymorphisms in TYMS gene in Slavic population of central Europe.

Thymidylate synthetase (TS) plays a critical role in the de novo synthesis of dTMP inside the cell. Therefore, TS is a suitable target for cytotoxic drugs such as fluoropyrimidines. Drug efficacy and toxicity depend on the intracellular level of TS, which is significantly influenced by the polymorphisms in the 5'UTR (TSER - rs45445694, TSER*3G>C - rs2853542) and 3'UTR (1494del TTAAAG - rs151264360) of TYMS gene. Polymorphic variants of TYMS gene affect TS activity via gene expression and transcript stability. Patients who undergo fluoropyrimidine therapy may benefit from genetic testing prior to the administration of chemotherapy. At the 5' terminus of TYMS, there is a polymorphic region represented by a variable number of 28bp long tandem repeats (2-9 tandems) with the G or C nucleotide variant (SNP G>C). The 3'end of TYMS gene may decrease the stability of mRNA in the case of 6 base deletion (1494del6, D). In our study, we have focused on testing of TYMS gene polymorphisms, determination of TYMS variant frequencies in Western Slavic population and comparison of Slovak population with other populations.We performed identification of 5'UTR (rs45445694 - TSER*2 or TSER*3; rs2853542 - TSER*3G>C; TSER*3+ins6) and 3'UTR (rs151264360/1494del6/D) polymorphic regions of TYMS gene among 96 volunteers by PCR-RFLP and fragment analysis. Slovak frequencies of selected polymorphisms were established as follows: the frequency of TSER*2, TSER*3, TSER*3G>C, 1494del6/D and I to be 41%, 59%, 34%, 37.5% and 62.5% respectively. The high resolution of the capillary electrophoresis technique allowed among TSER*3 group identification of a subgroup of four individuals with rare 6bp insertion in 3R allele, id est 2.1% TSER*3+ins6 allele frequency. In our study, we have revealed individuals with rare G>C substitution in the first 28bp tandem repeat of TSER*2 promoter enhancer region (rs183205964) as well, the overall frequency of this polymorphic allele in Slovak population was 2.1%. Our results proved that Slovak population is in Hardy-Weinberg equilibrium and proportion of TYMS polymorphisms is in accordance with other published data.

ERVW-1 gene polymorphisms related to preeclampsia.

OBJECTIVES: Identification of genetic association between the gene ERVW-1 and preeclampsia. BACKGROUND: Preeclampsia is a multifactorial disease affecting women during pregnancy and it is one of the main causes of perinatal and maternal morbidity and mortality. The pathophysiology of preeclampsia is very complex and several aspects of the disease have not been elucidated yet. Abnormal placentation frequently occurs during severe preeclampsia. Protein syncytin 1, a product of the ERVW-1 gene, plays a crucial role in the syncytiotrophoblast differentiation and optimal placentation. The syncytin 1 expression is disturbed during preeclampsia. The main focus of this study was the analysis of the ERVW-1 regulatory regions and identification of DNA polymorphisms associated with preeclamptic cases in Slovak population. METHODS: Regulatory region of gene ERVW-1 was analyzed by sequencing to identify genetic variants. RESULTS: We identified four DNA variants, namely rs4727276, rs148592540, rs569899772 and rs555416193, in samples of Slovak population. CONCLUSION: No relation between polymorphisms and preeclampsia was observed, indicating that further investigations with a larger sampling are still required. However, our work represents new original approach in genetic differential diagnosis of preeclampsia with possible useful findings in the future (Tab. 3, Fig. 1, Ref. 34).

GAI - distinct genotype and phenotype characteristics in reported Slovak patients.

OBJECTIVES: The clinical, biochemical and genetic findings in two Slovak patients with glutaric aciduria type I (GAI) are presented. BACKGROUND: GAI is a rare autosomal recessive neuro-metabolic disorder caused by deficiency of glutaryl-CoA dehydrogenase, which is involved in the catabolic pathways of lysine, hydroxylysine and tryptophan. This enzymatic defect gives rise to elevated levels of glutaric acid (GA), 3-hydroxyglutaric acid (3-OH-GA) and glutarylcarnitine (C5DC) in body fluids. METHODS: Biochemical and molecular-genetic tests were performed. Urinary organic acids were analysed by Gas Chromatography/Mass Spectrometry (GC/MS) and the entire coding region of the GCDH gene, including flanking parts, was sequenced. RESULTS: We found the presence of typical metabolic profile and novel causal pathogenic variants in both GAI patients. CONCLUSION: We present the first report of two Slovak patients with GAI, which differed in the clinical and biochemical phenotype significantly. They were diagnosed by two distinct approaches - selective and newborn screening. Their diagnosis was complexly confirmed by biochemical and later on molecular-genetic examinations. Though we agreed with a thesis that early diagnostics might positively influenced patient's health outcome, contradictory facts should be considered. Supposed extremely low prevalence of GAI patients in the general population and/or the existence of asymptomatic individuals with a questionable benefit of the applied therapeutic intervention for them lead to doubts whether the inclusion of disease into the newborn screening programme is justified well enough (Tab. 1, Fig. 3, Ref. 41).

Peroxisomal D-bifunctional protein deficiency: First case reports from Slovakia.

D-bifunctional protein deficiency (#OMIM 261515) is a rare autosomal recessive hereditary metabolic disorder causing severe clinical and biochemical abnormalities that are usually fatal in the course of the first years of life. This disease is classified as single enzyme peroxisomal disorder affecting the ß-oxidation pathway in this compartment. In this paper we present a full overview of the clinical presentation, magnetic resonance imaging, biochemical and molecular data of two Slovak D-bifunctional protein deficient patients. In the clinical presentation of both patients severe generalized hypotonia, depression of neonatal reflexes, craniofacial dysmorphism and seizures dominated starting from the second day of life. In both patients, who died up to two years of life, we found elevated plasma levels of very long chain fatty acids and we identified the presence of causative mutations in the HSD17B4 gene. In the first case, we found the homozygous mutation c.46G>A, which is responsible for a defect in the dehydrogenase domain. In the second patient, the heterozygous mutations c.1369A>G and c.1516C>T were present and functionally they are related to the hydratase domain of the protein. This combination of mutations in the second patient is very rare and has not been reported until now. The presence of mutations was examined in all family members, and the resulting data were successfully utilized for prenatal diagnosis.

Frequencies of polymorphisms in CYP2C9 and VKORC1 genes influencing warfarin metabolism in Slovak population: implication for clinical practice.

OBJECTIVES: The study was aimed at establishing an effective molecular-genetic method for detecting polymorphisms in genes CYP2C9 and VKORC1, which affect the pharmacogenetics of warfarin, and at determining their prevalence in Slovak population. BACKGROUND: Warfarin, derivative of coumarin, belongs to the most commonly prescribed oral anticoagulants with narrow therapeutic index. An insufficient dose of warfarin can result in failure to produce the antithrombotic effect, whereas an overdose increases the risk of bleeding. It was proven that genetic variability in two genes, CYP2C9 a VKORC1, has a significant influence on the individual's response to the dosage of warfarin. METHODS: In a control group of 112 randomly selected individuals, we tested the frequency of selected single nucleotide polymorphisms including CYP2C9*2 (430C>T), CYP2C9*3 (1075A>C), VKORC1*2 (1173C>T) by allele-specific Real-Time PCR and VKORC1*2 (-1639G>A) by using PCR-RFLP. RESULTS: Due to the combination of frequent alleles CYP2C9*2, CYP2C9*3 and VKORC1*2 in Slovak population we determine that 25% of population need a standard 5-mg daily dose of warfarin, while 44%, 23%, and 8% need 4 mg, 3 mg and 2 mg of warfarin per day. CONCLUSION: Slovak population is in Hardy-Weinberg equilibrium and frequencies of SNPs were in accordance with other published results in European populations (Tab. 5. Fig. 3, Ref. 51).

The influence of silicon substitution on the properties of spherical- and whisker-like biphasic α-calcium-phosphate/hydroxyapatite particles.

In this work, the influence of the morphology of hydroxyapatite particles on silicon substitution through hydrothermal synthesis performed under the same conditions was investigated. Spherical- and whisker-like hydroxyapatite particles were obtained starting from calcium-nitrate, sodium dihydrogen phosphate, disodium-ethylenediaminetetraacetic acid and urea (used only for the synthesis of whisker-like particles) dissolved in aqueous solutions. Silicon was introduced into the solution using tetraethylorthosilicate. X-ray diffraction, infrared spectroscopy, scanning electron microscopy, energy-dispersive X-ray spectroscopy and transmission electron microscopy indicate that silicon doping induce different phase compositions and bioactivity of spherical- and whisker-like hydroxyapatite particles obtained under the same hydrothermal conditions. Silicon-substituted, spherical hydroxyapatites particles showed greater phase transformation to silicon-substituted α- calcium-phosphate compared with whiskers-like hydroxyapatite particles synthesized with the same amount of added silicon. Metabolic activity assay performed with SaOs2 osteosarcoma cells showed better biocompatibility of annealed biphasic spherical-like particles compared with annealed whiskerlike particles while dried spherical-like particles induce high cytotoxicity effect.

Association between time-to-presentation and clinical outcome in patients with subarachnoid hemorrhage: an observational study.

BACKGROUND: Headache is the most common presenting symptom of subarachnoid hemorrhage (SAH), ranging from mild headache to the "worst headache of my life". As headache is often non-specific, patients may not seek immediate medical attention, though prompt medical and surgical management is expected to improve clinical outcomes. In this study, we explore the independent association between duration from onset of symptoms to presentation at an emergency department (ED) and clinical outcomes after SAH. METHODS: Participants with a primary diagnosis of nontraumatic SAH were identified from consecutive patients at 11 regional stroke centres participating in the Registry of the Canadian Stroke Network (RCSN, 2003-2005). Hunt and Hess score (H+H), and modified Rankin Scale (mRS) at discharge were collected on SAH cases by trained nurse-abstractors. For analysis, patients were categorized into patients with mild-moderate dependency (mRS 0-3) and those with severe dependence or death (mRS 4-6) at hospital discharge. Multivariable regression analyses were used to determine the association between 'time to presentation' and clinical outcomes, independent of comorbidities. RESULTS: Of 721 SAH patients included in the RCSN, 642 (89.0%) had the interval between 'time last seen normal' and time of ED presentation recorded. Mean duration from symptom onset to ED arrival was 27.04 hours (+/- 2.02). One hundred and sixty-six patients (25.9%) presented to the ED more than 24 hours after onset of symptoms. On multivariable analysis, there was no association between time to presentation and severe disability or death at hospital discharge (OR 1.0 [95% CI 0.95-1.01]); 30-day mortality (OR 1.0 [95% CI 0.91-1.02]; or six-month mortality (OR 1.0 [95% CI 1.0-1.02]). Increasing H+H score and age were significantly associated with increased odds of death and severe dependence at hospital discharge. CONCLUSIONS: In this observational study, duration from symptom onset to hospital presentation was not independently associated with death or severe disability at hospital discharge following SAH. Age and H+H score were independent predictors of clinical outcome after non-traumatic SAH.

The antihyperalgesic effect of levetiracetam in an inflammatory model of pain in rats: mechanism of action.

BACKGROUND AND PURPOSE: Levetiracetam, a novel antiepileptic drug, has recently been shown to have antinociceptive effects in various animal models of pain. The purpose of this study was to investigate the antihyperalgesic effect of levetiracetam and its mechanism of action, by examining the involvement of GABAergic, opioidergic, 5-hydroxytryptaminergic (5-HTergic) and adrenergic systems in its effect, in a rat model of inflammatory pain. EXPERIMENTAL APPROACH: Rats were intraplantarly injected with the pro-inflammatory compound carrageenan. A paw pressure test was used to determine: (i) the effect of levetiracetam on carrageenan-induced hyperalgesia; and (ii) the effects of bicuculline (selective GABA(A) receptor antagonist), naloxone (non-selective opioid receptor antagonist), methysergide (non-selective 5-HT receptor antagonist) and yohimbine (selective alpha(2)-adrenoceptor antagonist) on the antihyperalgesic action of levetiracetam. RESULTS: Levetiracetam (10-200 mg.kg(-1); p.o.) significantly reduced, in a dose-dependent manner, the inflammatory hyperalgesia induced by carrageenan. The antihyperalgesic effect of levetiracetam was significantly decreased after administration of bicuculline (0.5-2 mg.kg(-1); i.p.), naloxone (1-3 mg.kg(-1); i.p.), methysergide (0.25-1 mg.kg(-1); i.p.) and yohimbine (1-3 mg.kg(-1); i.p.). CONCLUSIONS AND IMPLICATIONS: These results show that levetiracetam produced antihyperalgesia which is at least in part mediated by GABA(A), opioid, 5-HT and alpha(2)-adrenergic receptors, in an inflammatory model of pain. The efficacy of levetiracetam in this animal model of inflammatory pain suggests that it could be a potentially important agent for treating inflammatory pain conditions in humans.

Major salivary gland sonography in Sjögren's syndrome: diagnostic value of a novel ultrasonography score (0-12) for parenchymal inhomogeneity.

OBJECTIVE: To validate ultrasonographic criteria for examination of the major salivary glands in the diagnosis of primary Sjögren's syndrome (SS). METHOD: A total of 209 consecutive patients with rheumatic diseases were selected according to the American-European Consensus Group (AECG) classification criteria for SS. One hundred and fifteen patients had primary SS, 44 had secondary SS, and 50 had sicca symptoms, and 36 subjects served as asymptomatic controls. This cohort was analysed for size, echogenicity, parenchymal inhomogeneity, focal changes, and posterior borders of the major salivary glands by ultrasonography (US). A novel US score for parenchymal inhomogeneity (0-12) was assigned and its diagnostic accuracy evaluated. RESULTS: Ultrasonographic abnormalities of salivary glands were detected in 107/115 (93.0%) patients with primary SS, in 12/44 (27.3%) with secondary SS, in 25/50 (50.0%) with sicca symptoms, and in 4/36 (11.1%) asymptomatic controls. Area under the receiver operating characteristic curve (AUC-ROC) for US inhomogeneity score was highly significant [0.96 +/- 0.01; 95% confidence interval (CI) 0.94-0.99, p < 0.000] for primary SS, with a sensitivity to specificity ratio of 91/83 for parotid and 93/90 for submandibular glands. Setting the cut-off US inhomogeneity score at 6 resulted in the best ratio of specificity (90.0%) to sensitivity (95.1%), with a positive predictive value of 72% and a negative predictive value of 96%. A US inhomogeneity score >or= 6 was closely correlated with positive biopsy (p < 0.000) and scintigraphy findings (p < 0.000). CONCLUSIONS: We demonstrate the high diagnostic value of a novel US score for parenchymal inhomogeneity (0-12) that could serve as a useful single US criterion in the evaluation of salivary gland involvement in primary SS.

[The role of cholesterol in embryogenesis and the Smith-Lemli-Opitzov syndrom]. / Uloha cholesterolu v embryogenéze a Smith-Lemli-Opitzov syndróm.

The role of cholesterol in cell biology has been known for years. The sight of cholesteol biological function has changed after the discovery that the genetic disorder Smith-Lemli-Opitz syndrome is caused by a defect in cholesterol biosynthetic pathway. Cholesterol has an important role in regulation and modification of Hedgehog proteins, what links cholesterol to early embryonic development. Hedgehog proteins comprise a family of secreted signaling molecules that are essential for embryonic patterning and morphogenesis. The deficit of cholesterol during embryogenesis causes severe abnormalities in SLOS because of disrupt autoprocessing of hedgehog proteins. SLOS is an autosomal recessive disorder of sterol metabolism. The underlying pathogenetic basis for SLOS has been shown to be a deficiency of 7-dehydrocholesterol reductase, which catalyzes the last step in cholesterol biosynthesis. Reduced enzyme activity leads to a deficit of cholesterol and accumulation of precursor sterols. The human 7-dehydrocholesterol reductase gene (DHCR7) is localized on chromosome 11q 12-13.

Analyzing Thorough QT Study 1 & 2 in the Telemetric and Holter ECG Warehouse (THEW) using Hannover ECG System HES : A validation study.

Following the ICH E14 clinical evaluation guideline [1], the measurement of QT/QTc interval prolongation has become the standard surrogate biomarker for cardiac drug safety assessment and the faith of a drug development. In Thorough QT (TQT) study, a so-called positive control is employed to assess the ability of this study to detect the endpoint of interest, i.e. the QT prolongation by about five milliseconds. In other words the lower bound of the one-sided 95% confidence interval (CI) must be above 0 [ms]. Fully automated detection of ECG fiducial points and measurement of the corresponding intervals including QT intervals and RR intervals vary between different computerized algorithms. In this work we demonstrate the ability and reliability of Hannover ECG System (HES(®)) to assess drug effects by detecting QT/QTc prolongation effects that meet the threshold of regulatory concern as mentioned by using THEW database studies namely TQT studies one and two.

The 10-year follow-up data of the Euro-Lupus Nephritis Trial comparing low-dose and high-dose intravenous cyclophosphamide.

OBJECTIVE: To update the follow-up of the Euro-Lupus Nephritis Trial (ELNT), a randomised prospective trial comparing low-dose (LD) and high-dose (HD) intravenous (IV) cyclophosphamide (CY) followed by azathioprine (AZA) as treatment for proliferative lupus nephritis. PATIENTS AND METHODS: Data for survival and kidney function were prospectively collected during a 10-year period for the 90 patients randomised in the ELNT, except in 6 lost to follow-up. RESULTS: Death, sustained doubling of serum creatinine and end-stage renal disease rates did not differ between the LD and HD group (5/44 (11%) vs 2/46 (4%), 6/44 (14%) vs 5/46 (11%) and 2/44 (5%) vs 4/46 (9%), respectively) nor did mean serum creatinine, 24 h proteinuria and damage score at last follow-up. Most patients in both groups were still treated with glucocorticoids, other immunosuppressant agents and blood pressure lowering drugs. After 10 years of follow-up, the positive predictive value for a good outcome of an early drop in proteinuria in response to initial immunosuppressive therapy was confirmed. CONCLUSION: The data confirm that a LD IVCY regimen followed by AZA-the "Euro-Lupus regimen"-achieves good clinical results in the very long term.

Surface roughness of ultra-thin silver films sputter deposited on a glass.

Silver was sputter deposited on a glass with a thin film thickness ranging from 10 to 80 nm. Scanning tunnelling microscopy was used to study the morphology of the obtained Ag-glass surfaces and to estimate the surface roughness. An equation for the surface roughness of the thin film was evaluated using parameters related to the thin film features: the surface roughness of the substrate, the compressibility of the thin film and the film thickness. The experimental results were fitted using the evaluated equation, and the conditions favouring lower or higher surface roughness were analyzed.

Clinical and serological follow-up of 71 patients with anti-mitochondrial type 5 antibodies.

Antimitochondrial M5 type antibodies (AMA M5) have been described in patients with antiphospholipid syndrome (APS), thrombocytopenia and autoimmune hemolytic anemia. The aim of this study was to describe the clinical and immunological characteristics of a series of patients with AMA M5. We analyzed 71 patients with AMA M5 seen consecutively at our centres during the last 8 years. The clinical and immunological characteristics of diseases expression were compared with 70 consecutive disease control patients without AMA M5. Compared with the control group, AMA M5 positive patients presented higher prevalence of false positive VDRL test (P < 0.001) and thrombocytopenia (P = 0.002) with lower levels of anti-beta2 glycoprotein I antibodies. In AMA M5 patients (56 female, 15 male) a heterogeneous group of disorders were found. Twenty-seven (38%) patients fulfilled the Sapporo criteria for the classification of the APS. Laboratory criteria were met in 55 (77.5%), and clinical criteria in 31 (43.7%) patients. Six patients initially presented with non-criteria features of APS during follow-up period developed APS. Younger patients with AMA M5 should be carefully observed for the development of APS, even in the absence of serological criteria, while elderly must be screened for monoclonal gammopathy and hematological disorders.

Role of alpha2-adrenoceptors in the local peripheral antinociception by carbamazepine in a rat model of inflammatory mechanical hyperalgesia.

The anticonvulsant carbamazepine was recently shown to possess local peripheral antinociceptive properties. In this study, we investigated whether alpha2-adrenergic receptors are involved in the local peripheral antihyperalgesic effects of carbamazepine and determined the type of interaction between carbamazepine and clonidine, an alpha2-adrenoceptor agonist. Intraplantar (i.pl.) coadministration of either carbamazepine (100-1000 nmol/paw) or clonidine (1.9-3.7 nmol/paw) with the proinflammatory compound concanavalin A (Con A; 0.8 mg/paw) caused a significant dose- and time-dependent reduction of the difference between the forces exerted by a rat's hind paws in a modified paw-pressure test. The coadministration of 260 and 520 nmol/paw (i.pl.) yohimbine, an alpha2-adrenoceptor antagonist, with carbamazepine, significantly depressed the local antihyperalgesic effect in a dose- and time-dependent manner whereas yohimbine by itself did not have any effect. The administration of a mixture of carbamazepine and clonidine at fixed dose fractions (1/4, 1/2 and 3/4) of ED50 caused a significant and dose-dependent reduction of Con A-induced hyperalgesia. Isobolographic analysis revealed an additive interaction. These results suggest that alpha2-adrenoceptors play a role in the local peripheral antihyperalgesic effects of carbamazepine and that local peripheral coadministration of carbamazepine with clonidine results in an additive antihyperalgesic effect.

Adrenoleukodystrophy--a new mutation identified.

INTRODUCTION: X-linked adrenoleukodystrophy from the group of peroxisomal disorders presents with an extensive spectrum of phenotypes. The mutation affects the ABCD1 gene encoding a peroxisomal membrane protein. So far, its detailed function has not been clarified. However, it plays an essential role in the ethiopathogenesis of X-linked adrenoleukodystrophy. Its defect causes accumulation of the very long chain fatty acids in the tissues of the central and peripheral nervous system, adrenal glands and in the body fluids. PURPOSE: To review the clinical presentations and diagnostic issues in X-adrenoleukodystrophy diagnosed in the one affected family. METHODS: A case report. Measurement of very long chain fatty acids. Molecular analysis of the adrenoleukodystrophy gene. RESULTS: A new "unique" mutation in the initiation codon in the first'exon of ABCD1 gene was identified. We present a phenotype description of a patient with this mutation. CONCLUSIONS: X-linked adrenoleukodystrophy is a disease with the incidence rate approximately 1:16,800. Detection of new mutations contributes to better understanding of this rare disease and makes the diagnostic more available and precise. The importance of an adequate diagnosis is justified not only by a different therapeutic approach, but also by the need of prenatal diagnostics and the need of genetic counselling in the affected families. As demonstrated in our case, it is necessary to consider this diagnosis also in the adult age, e.g. within the differential diagnosis of spastic paraparesis (Tab. 1, Fig. 4, Ref 23). Full Text (Free, PDF) www.bmrj.sk.

Characterization of antigen-presenting cells in human apical periodontitis lesions by flow cytometry and immunocytochemistry.

AIM: To analyse phenotypic characteristics of antigen-presenting cells (APC), isolated from human periapical lesions by flow cytometry and immunocytochemistry. METHODOLOGY: Sixteen periapical lesions were digested for 15 min with 0.05% collagenase. Mononuclear cells, separated from other inflammatory cells by density centrifugation, were processed for flow cytometry and/or immunocytochemistry. Single and double immunostainings were performed using monoclonal antibodies specific for human CD45, CD3, CD19, CD14, HLA-DR, CD1a, CD83 and CD123. RESULTS: Antigen-presenting cells (HLA-DR(+) cells) represented 32.9 +/- 17.8% of total mononuclear cells. Amongst them, B cells (HLA-DR(+) CD19(+)) were the predominant APC population, followed by activated macrophages (HLA-DR(+) CD14(+)), dendritic cells (DC) (HLA-DR(+) CD14(-) CD19(-) CD3(-)) and activated T cells (HLA-DR(+) CD3(+)). Based on the predominance of T cells (CD3(+)) or B cells and plasma cells (CD19(+) and CD19(lo), respectively) amongst mononuclear cell infiltrates, lesions were divided into T- and B-types. The percentage of DC in T-type lesions (27.1 +/- 6.8% of total HLA-DR(+) cells) was higher, compared with B-type lesions (10.3 +/- 5.2%) (P < 0.01). Within the DC population, the percentages of CD1a (Langerhans cell type) and CD123 (probably plasmacytoid DC type) did not differ significantly between the groups (P > 0.05). However, the percentage of mature DC (CD83(+)) was significantly higher in T-type periapical lesions (P < 0.05). CONCLUSIONS: Flow cytometry and immunocytochemistry are suitable methods for phenotypic analysis of APC after their isolation from human periapical lesions. APC, that were phenotypically heterogeneous, constituted a significant component of infiltrating cells. Lesions with the predominance of T cells were characterized by a higher proportion of mature DC (HLA-DR(+)CD83(+) cells) than lesions with predominance of B cells/plasma cells.

Peripheral anti-hyperalgesia by oxcarbazepine: involvement of adenosine A1 receptors.

In this study we determined whether oxcarbazepine (OXC) could produce local peripheral antinociceptive effects in a rat model of inflammatory hyperalgesia, and whether adenosine receptors were involved. When coadministered with the pro-inflammatory compound concanavalin A, OXC (1000-3000 nmol/paw) caused a significant dose- and time-dependent anti-hyperalgesia. Caffeine (1000-1500 nmol/paw), a nonselective adenosine receptor antagonist, as well as 1,3-dipropyl-8-cyclopentylxanthine (DPCPX) (10-30 nmol/paw), a selective A1 receptor antagonist, coadministered with OXC, significantly depressed its anti-hyperalgesic effect. Drugs injected into the contralateral hind paw did not produce significant effects. These results indicate that OXC produces local peripheral anti-hyperalgesic effects, which is mediated via peripheral A1 receptors.