LACC1 polymorphisms in inflammatory bowel disease and juvenile idiopathic arthritis.

The function of the Laccase domain-containing 1 (LACC1) gene is unknown, but genetic variation at this locus has been reported to consistently affect the risk of Crohn's disease (CD) and leprosy. Recently, a LACC1 missense mutation was found in patients suffering from monogenic forms of CD, but also systemic juvenile idiopathic arthritis. We tested the hypothesis that LACC1 single nucleotide polymorphisms (SNPs), in addition to CD, are associated with juvenile idiopathic arthritis (JIA, non-systemic), and another major form of inflammatory bowel disease, ulcerative colitis (UC). We selected 11 LACC1 tagging SNPs, and tested their effect on disease risk in 3855 Swedish individuals from three case-control cohorts of CD, UC and JIA. We detected false discovery rate corrected significant associations with individual markers in all three cohorts, thereby expanding previous results for CD also to UC and JIA. LACC1's link to several inflammatory diseases suggests a key role in the human immune system and justifies further characterization of its function(s).

Long-term outcome of resective epilepsy surgery in Norwegian children.

AIM: The aim of this study was to evaluate the long-term results of resective surgery on children with difficult-to-treat epilepsy in Norway. METHODS: In the period 1995-2004, 64 surgical procedures (54 resections and 10 functional hemispherotomies) were performed in 54 children. The children's medical records were retrospectively reviewed at a minimum of 2 years after surgery. We sent a questionnaire regarding their epilepsy (seizures, usage of antiepileptic drugs) and general functioning (social situation, motor, language, cognition, behavioural or emotional problems, any remedial action) to the children/parents after a mean follow-up period of 7 years. RESULTS: 55.5% of the children were seizure-free. The success rate varied according to the type of surgery. Best results were found after functional hemispherotomies and temporal lobe resections, as nine of 10 (90%) and 10 of 19 (53%) of these patients, respectively, became seizure-free. In addition to a better seizure control, 71% of the children/parents reported of a better cognitive and psychosocial functioning. CONCLUSION: The results of epilepsy surgery in this paediatric cohort are very edifying, and it is our impression that this treatment option is underused in Norway.

Parental SCN1A mutation mosaicism in familial Dravet syndrome.

Different SCN1A mutations are known to cause a variety of phenotypes, such as generalized epilepsy with febrile seizures plus (GEFS+), Dravet syndrome and familial hemiplegic migraine (FHM). In Dravet syndrome, most mutations are de novo and familial cases are rare. In this study, Dravet syndrome is observed in two maternal half sisters. They have healthy fathers and their common mother has never experienced seizures, but has a lifelong history of migraine. Direct sequencing of DNA extracted from blood revealed a heterozygous SCN1A nonsense mutation c.3985C>T in the sisters, but not in the mother. The mutation induces a premature stop codon and probably leads to a non-functional protein. Further examination of the mother's DNA showed that she has a mosaicism of the mutation. This report of parental SCN1A nonsense mutation mosaicism in familial Dravet syndrome suggests that mosaicism might be more common than previously suspected and emphasizes the importance of taking mosaicism into account in genetic counselling of Dravet syndrome and SCN1A mutations. Furthermore, whether the migraine of the mother could be influenced by her SCN1A mutation mosaicism is not known, but increased awareness of migraine in future studies of SCN1A related epilepsies could clarify this intriguing link between migraine and epilepsy.

Elevation of cytokines in peritoneal fluid and blood in patients with liver cirrhosis.

BACKGROUND/AIMS: Liver cirrhosis, described as the endstage of a necroinflammatory process, is often accompanied by ascites formation. The rationale for this study was the hypothesis that patients with liver cirrhosis have a low-grade chronic inflammatory response, which leads to an increased amount of proinflammatory cytokines accumulated in ascites. Twenty-five patients with liver cirrhosis complicated by ascites and twelve healthy volunteers were prospectively included in the study. METHODOLOGY: Ascites and blood samples from the patients were obtained for analysis of inflammatory cytokines using enzyme-linked immunosorbent assay methodology. Blood samples were taken from the healthy volunteers to obtain reference values. RESULTS: Plasma and ascites concentrations of interleukin-1alpha, interleukin-6, and tumor necrosis factor-alpha were significantly elevated in the patients compared with plasma levels in the group of healthy controls. Significant elevation of interleukin-10 concentrations was found in ascites but not in plasma in the patients. There was no significant difference in interleukin-10 levels between patient and control plasma. CONCLUSIONS: The findings suggest that elevated cytokine concentrations in ascites and serum could perpetuate an inflammatory reaction that may be a source of preservation of an ongoing systemic inflammatory reaction. This may contribute to the maintenance, and even progress, of the liver dysfunction, leading to exaggerated ascites development.

Effect of age and body weight on neurohumoral variables in healthy Cavalier King Charles spaniels.

OBJECTIVE: To evaluate the effect of age and body weight on several neurohumoral variables that are commonly altered in heart failure in Cavalier King Charles Spaniels. ANIMALS: 17 healthy privately owned Cavalier King Charles Spaniels, 10 males and 7 females, ranging in age from 0.4 to 9.7 years, and ranging in body weight from 6.6 to 12.2 kg. PROCEDURE: The clinical condition of the dogs was evaluated by physical examination, thoracic radiography, and echocardiography. Plasma nitrate and nitrite (P-NN), N-terminal atrial natriuretic and brain natriuretic peptides (NT-ANP and BNP, respectively), endothelin (ET-1), urine cyclic guanosine monophosphate (U-cGMP), and urine nitrate and nitrite (U-NN) concentrations were analyzed. RESULTS: Plasma concentrations of NT-ANP and P-NN increased significantly with age, but plasma NT-ANP and P-NN also correlated significantly, irrespective of age. A modest increase of left atrial size did not explain the increase of NT-ANP and P-NN with age. Concentration of ET-1 correlated positively with heart rate; heart rate did not change with age. Weight had a negative impact on NT-ANP, P-NN, and U-cGMP concentrations and left atrial relative size. CONCLUSIONS AND CLINICAL RELEVANCE: Age-matched controls are essential for evaluation of NT-ANP and P-NN concentrations and left atrial size. Weight may alter reference values of plasma NT-ANP, P-NN, and urine cGMP concentrations. Natriuretic peptides can be used as further evidence that heart failure exists. The increased plasma concentrations of NT-ANP (but not BNP) and P-NN with aging reflect neurohumoral physiologic changes that must be distinguished from pathologic changes in patients with heart failure.

The effect of lamotrigine on epileptiform discharges in young patients with drug-resistant epilepsy.

PURPOSE: In a double-blind crossover study with lamotrigine (LTG), we investigated a possible relationship between the clinical responses and changes of the amount of epileptiform activity in EEG. METHODS: Twelve patients, aged 4-21 years, with generalized drug-resistant epilepsy who had responded to LTG, completed the study. Twenty-four-hour video-EEGs were taken during control, placebo, and drug phases. The amounts of epileptiform discharges were estimated and compared with the clinical effects. RESULTS: The duration of periods of repeated epileptiform discharges was significantly reduced during the LTG phase compared with the placebo phase (n = 12, p = 0.04). Ten patients showed a reduction of the amount of epileptiform discharges in the LTG phase by a mean of 81% (range, 17-100%). Periods of repeated epileptiform discharges with duration longer than 30 s were significantly reduced in length (p = 0.03) and number (p = 0.04) during the LTG phase compared with the placebo phase. Shorter periods of epileptiform discharges and the numbers of single epileptiform discharges were not affected. In five patients there was a seizure reduction (>/=50%) concomitant with the reduction of epileptiform discharges in the EEG. The behavior improved during LTG treatment in all patients. The patients became more alert, and their concentration and performance improved, according to their parents and the medical personnel. CONCLUSIONS: LTG in dosages of 1-8 mg/kg body weight was found to depress relatively long episodes of interictal, repetitive, epileptiform activity in young patients with drug-resistant epilepsy, whereas short episodes were not affected. It depressed seizures in about half of the patients studied but gave improvements of behavior in all patients.

Analysis of CSF amino acids in young patients with generalised refractory epilepsy during an add-on study with lamotrigine.

The effect of add-on administration of lamotrigine (1-12 mg/kg per day, 2-12 months) on the levels of neurotransmission related amino acids including gamma-aminobutyric acid (GABA), glutamate, aspartate, glycine and antiepileptic drugs (AEDs) in lumbar cerebrospinal fluid (CSF) was studied in 22 children and young adults with generalised therapy resistant epilepsy. Two lumbar punctures were performed, one prior to, and one following a mean of 5 months (2-12 months) of lamotrigine treatment. Lamotrigine decreased seizure incidence and severity in 12 of the 22 patients without influencing CSF GABA, glutamate, aspartate or glycine levels. Lamotrigine did not alter the concentrations of AEDs in CSF or plasma. However, CSF GABA levels were 86% higher in those patients also treated with gamma-vinyl-GABA (vigabatrin, GVG) compared with patients treated with other combinations and this was not altered by co-medication with lamotrigine. The proposed mechanism of action of lamotrigine, namely that it may inhibit glutamate release in the CNS, is not reflected by changes in CSF glutamate levels. The present findings indicate that CSF GABA, glutamate, aspartate and glycine levels may not be useful as in vivo neurochemical markers in young patients responding to the therapeutic dose of lamotrigine used in this study.

The genome sequence of Rickettsia prowazekii and the origin of mitochondria.

We describe here the complete genome sequence (1,111,523 base pairs) of the obligate intracellular parasite Rickettsia prowazekii, the causative agent of epidemic typhus. This genome contains 834 protein-coding genes. The functional profiles of these genes show similarities to those of mitochondrial genes: no genes required for anaerobic glycolysis are found in either R. prowazekii or mitochondrial genomes, but a complete set of genes encoding components of the tricarboxylic acid cycle and the respiratory-chain complex is found in R. prowazekii. In effect, ATP production in Rickettsia is the same as that in mitochondria. Many genes involved in the biosynthesis and regulation of biosynthesis of amino acids and nucleosides in free-living bacteria are absent from R. prowazekii and mitochondria. Such genes seem to have been replaced by homologues in the nuclear (host) genome. The R. prowazekii genome contains the highest proportion of non-coding DNA (24%) detected so far in a microbial genome. Such non-coding sequences may be degraded remnants of 'neutralized' genes that await elimination from the genome. Phylogenetic analyses indicate that R. prowazekii is more closely related to mitochondria than is any other microbe studied so far.

The efficacy of lamotrigine in children and adolescents with refractory generalized epilepsy: a randomized, double-blind, crossover study.

PURPOSE: We report a double-blind, placebo-controlled crossover study of lamotrigine (LTG) as add-on treatment in therapy-resistant, generalized epilepsy in children and adolescents (n = 30). METHODS: Twenty patients had Lennox-Gastaut syndrome. Each patient acted as his or her own control. LTG and placebo were randomly added to existing antiepileptic medication (AEDs). The LTG dosage was individualized in an open phase preceding the placebo/treatment phase. Patients who responded to LTG in the open phase went on to the double-blind phase. "Responders " were defined as patients with a >50% seizure reduction or less severe seizures or both, or improved behavior or improved motor skills or both. "Nonresponders" were defined as children who showed no positive effects of LTG with plasma levels of < or = 10 microg/ml or children who had adverse events during the open phase. RESULTS: There was a clear statistically significant reduction of seizure frequency in LTG compared with placebo treatment. None of the children studied showed abnormal biochemical or hematologic findings, or changes in plasma levels of concomitantly administered AEDs. CONCLUSIONS: LTG is a well-tolerated and effective treatment in children with intractable generalized epilepsies, including those with Lennox-Gastaut syndrome. The study design allowed a double-blind placebo-controlled assessment of LTG although the participating children used 19 different AED combinations at entry.

No increase in carbamazepine-10,11-epoxide during addition of lamotrigine treatment in children.

It has been suggested that lamotrigine (LTG) may enhance the toxicity of carbamazepine (CBZ) by increasing the concentration of the active metabolite carbamazepine-10,11-epoxide (CBZ-E) in adult patients. The authors investigated this hypothesis in an add-on study in 11 children and 3 adolescents, aged 6-22 years, who had been treated for more than 1 year with CBZ in monotherapy or with CBZ in combination with one or two other antiepileptic drugs. The LTG dosage was increased step by step until clinical response or side effects were observed. The plasma concentrations of LTG, CBZ, and CBZ-E were monitored during steady state conditions before and after the addition of LTG. It was found that LTG had no effect on mean CBZ concentrations and that it decreased rather than increased the mean plasma concentration of CBZ-E from 6.4 +/- 2.6 to 4.9 +/- 2.4 mumol/l (mean +/- SD, n = 14, P = 0.019). Observed side effects were diplopia in two children, agitation in two, and increased number of seizures in one. None of these five patients had unusually high CBZ-E levels when the side effect developed. It is concluded that addition of lamotrigine in children treated with carbamazepine children does not result in a pharmacokinetic interaction with a toxic accumulation of carbamazepine-10,11-epoxide.

Leukocyte hydrogen peroxide production in a surgical wound in mice. The effects of an amide local anaesthetic.

The oxygen metabolism of polymorphonuclear leukocytes (PMN) is of importance in local tissue repair processes. Amide local anaesthetics are commonly used to relieve surgical wound pain. The cellular effects of local anaesthetics in vivo is poorly described in the literature. However, interactions between amide local anaesthetics and the oxygen metabolism of leukocytes have been reported. To extend that knowledge, this paper investigates the influence of lidocaine treatment on the production of hydrogen peroxide (H2O2) by leukocyte oxygen metabolism. A soft tissue chamber model in the mouse was used, allowing measurements of the H2O2 production spontaneously and after phorbol myristate acetate (PMA) addition, from two different leukocyte pools. Exudate leukocytes were generally more reactive to PMA stimulation in comparison to tissue chamber adherent leukocytes. Topically administered lidocaine significantly influenced the number of leukocytes in the wound exudate at 24 h postoperatively. Exudate leukocytes, topically exposed to lidocaine, showed an enhanced H2O2 production in comparison to leukocytes receiving lidocaine systemically. At 6 days, the viability and the H2O2 production differed significantly between the group receiving topically applied lidocaine in comparison to placebo. We conclude that the wound healing process may be effected by topically applied lidocaine, administered in clinical doses, at least via interference with leukocyte oxygen metabolism.

Pharmacokinetic interactions between lamotrigine and other antiepileptic drugs in children with intractable epilepsy.

PURPOSE: We wished to determine the oral pharmacokinetics of lamotrigine LTG and to assess possible interactions with other AEDs in an unselected population of children. Concentration data in plasma and in CSF for lamotrigine as well as for the other AEDs are presented. METHODS: Thirty-one children, children and young adults aged > 2 years with intractable generalized epilepsy despite adequate duration and dose of at least three conventional AEDs were studied. RESULTS: There was a linear relation between the dose administered and the maximal plasma concentration, indicating that saturation of absorption or elimination mechanisms did not occur in the dose range studied. The median elimination half-life (t1/2) in patients receiving concomitant valproate (VPA) was 43.3 h; in patients receiving carbamazepine (CBZ) and/or phenobarbital (PB), it was 14.1 h; and in patients receiving both VPA and CBZ/ PB or other antiepileptic drugs (AEDs), it was 28.9 h. No clinically important changes in the plasma levels of CBZ, VPA, valproate, ethosuximide, or PB were observed in the follow-up period (2-12 months). No dose adjustments of concomitant AEDs were necessary. The plasma concentration of clonazepam (CZP) was reduced when LTG was introduced. CONCLUSIONS: The complex interaction between LTG and other AEDs in children with intractable epilepsy makes therapeutic drug monitoring (TDM) desirable.

Monocyte activation on titanium-sputtered polystyrene surfaces in vitro: the effect of culture conditions on interleukin-1 release.

The release of interleukin-1 alpha (IL-1 alpha) by human peripheral blood monocytes cultured for 24 and 48 h on polystyrene (PS) and titanium-sputtered polystyrene (Ti) was evaluated. Magnetron sputtering of the PS surfaces resulted in a formation of a 50-nm-thick coat, consisting of an outer layer of TiO2. Monocytes released IL-1 alpha without the addition of exogenous stimuli. A doubling of the culture time from 24 to 48 h did not have a major effect on the amount of IL-1 alpha released. The IL-1 alpha levels were increased by addition of lipopolysaccharide (LPS). High concentrations of PS particles (1 and 3 microns diameter) were equally effective stimuli for IL-1 alpha release as LPS. Preadsorption of fibronectin to culture plates augmented LPS-stimulated IL-1 alpha secretion, whereas preadsorbed fibrinogen had an inhibitory effect. Our observation indicate a direct activation of monocytes by PS and Ti, resulting in IL-1 alpha secretion, which is modified by protein adsorption and exogenous stimuli.

The Rickettsia prowazekii genome: a random sequence analysis.

We describe here the first general survey of the genomic content and the coding capacity of the 1.1 Mb genome of Rickettsia prowazekii based on an analysis of a total of 200 kb of unique sequence data collected in a random manner. Based on nucleotide distribution profiles, we estimate that the R. prowazekii genome may have a coding density of 60%-70% and that it may contain a total of circa 800 genes. Here, we have tentatively identified and classified 173 of these genes. Our analysis suggests that the R. prowazekii genome is a highly derived, reduced genome that has lost many genes involved in amino acid biosynthetic pathways and regulatory functions. Furthermore, the R. prowazekii genome seems to lack glycolytic genes, but it does contain genes encoding components of the tricarboxylic acid cycle as well as of the electron transport system. We have also encountered a family of homologous genes coding for ATP/ADP translocases, as observed in several mitochondrial genomes. We relate these findings to previous phylogenetic studies that suggest that Rickettsia and mitochondria share a common ancestor.

Ex vivo analysis of leukocyte hydrogen peroxide production using a bi-plate model in mice.

Implantation of artificial materials is followed by inflammation and wound healing, where phagocytic cells play an important role. The mechanisms whereby the implant surface may elicit and modulate leukocyte functions in vivo are not understood, partly due to the technical difficulties of examining the local inflammatory events in vicinity of the material-tissue interface with conventional biochemical and immunological techniques. In the present study a newly developed biplate implant was inserted subcutaneously in the mouse. Leukocytes from the local inflammatory exudate and leukocytes associated to the surface of the implants were retrieved after 1 and 6 days and separately assayed with respect to hydrogen peroxide (H2O2) production ex vivo. Implantation caused a local accumulation of predominantly mononuclear cells in the surrounding subcutaneous tissue. The H2O2 production was found to be low in both the subcutaneous exudate and the implant-associated leukocytes, irrespective of implant material and implantation times. However, ex vivo-stimulation with phorbol myristate acetate (PMA) caused an enhanced H2O2 production. These observations show that biplate implants do not maximally activate the oxidative metabolism of the recruited leukocytes. The exudate leukocytes were more responsive to PMA stimulation in comparison with implant-associated leukocytes, indicating that properties of the implant surface and possibly surface-associated proteins could modify the responsiveness of the phagocytic cells at the implant site. Our results suggest that the present biplate model may be suitable for further studies on local production of oxygen metabolites and function of leukocytes at implanted biomaterials.

Leukocyte recruitment and turnover in an experimental inflammatory exudate.

The turn-over of leukocytes at sites of inflammation in vivo is to a large extent uninvestigated, mainly due to the technical difficulties associated with sampling and analysis of the inflammatory exudate. This paper investigates the immigration of fluorescently labeled granulocytes into exudate chambers at 8 h and at 1, 3, and 6 days after implantation into abdominal muscle of rat. In each experiment, the circulating granulocytes were labeled by intravenous administration of the DNA-labeling fluorochrome Hoechst 33342 and allowed to migrate into the chamber during 6 h before harvesting the chamber exudate. The rate of granulocyte immigration into the chamber varied considerably over time, showing a minimum at 3 days after implantation. The resulting kinetic pattern of granulocyte numbers in the exudate showed a two-step appearance, different from that of earlier determinations in soft tissue. A comparison between the calculated rates of granulocyte immigration and the total number of granulocytes present in the exudate at different times indicated that all immigrated cells survived in the chamber for the entire observation period of 6 days.

Flunarizine of limited value in children with intractable epilepsy.

Fourteen ambulatory children and adolescents with intractable epilepsy were studied in an open phase II study to investigate the pharmacokinetics and pharmacodynamics of flunarizine as an add-on treatment. Flunarizine was given in increasing doses starting with 0.1-0.3 mg/kg/day until effect was observed or a steady-state plasma concentration of 50-60 ng/ml was reached. Treatment was continued for 3 months at steady state. Pharmacokinetics were determined during the immediate posttreatment period. Positive antiepileptic effect (> or = 50% reduction in seizure frequency) was observed in 4 of 14 patients (29%; 95% CI: 52-5). Independently of antiepileptic effect, 10 of 14 parents (71.4%; 95% CI: 95-48) observed positive cognitive effects. In all patients treatment was withdrawn due to either lack of effect or weight gain. Flunarizine was rapidly absorbed; mean time of peak concentration (Tmax) was 2.7 hours (range: 1-8). The mean terminal half-life was 23.2 days (range: 7-48), the total plasma clearance of flunarizine per fraction of the dose absorbed (CLp/F) was 0.28 ml/min/kg (range: 0.07-042), and the volume of distribution of flunarizine per fraction of the dose absorbed (Vd/F) was 187 L/kg (range: 99-348). We conclude that flunarizine (0.1-0.3 mg/kg/day) seems to be of limited antiepileptic value in children with intractable epilepsy. The pharmacokinetic profile of flunarizine complicates its clinical use.

Leucocyte accumulation and leukotriene B4 release in response to polyglactin 910 and expanded polytetrafluoroethylene in hollow chambers in the rat.

The acute inflammatory reaction elicited after implantation of polymer membranes used clinically to promote bone healing and augmentation was studied in a soft tissue titanium chamber model. The two materials compared were non-degradable expanded polytetrafluoroethylene (ePTFE) and degradable polyglactin 910, a copolymer of 90% polyglycolic acid and 10% polylactic acid. The membranes were implanted in the titanium chamber for 24 h and 6 d. The number of leucocytes increased for both materials, whereas the leukotriene B4 (LTB4) content decreased over time. In both groups polymorphonuclear granulocytes predominated. The number of leucocytes was significantly higher in chambers with polyglactin 910 than ePTFE. In contrast, the LTB4 content was higher in chambers with ePTFE than polyglactin 910. No differences in cell viability were observed between the materials tested. This study shows that both degradable and non-degradable polymers elicit a marked influx and activation of inflammatory cells during early healing in soft tissues.

Inhibitory effects of amide local anaesthetics on stimulus-induced human leukocyte metabolic activation, LTB4 release and IL-1 secretion in vitro.

The anti-inflammatory effects of the amide local anaesthetics lidocaine and bupivacaine were evaluated in vitro by examination of the metabolic activation and secretory responses of human polymorphonuclear granulocytes (PMNGs) and mononuclear cells. Pretreatment with lidocaine or bupivacaine had a dose-dependent inhibitory effect on PMNG luminol-amplified chemiluminescence stimulated by bovine serum albumin (BSA)/anti-BSA immune complexes (IC) or by serum-opsonized zymosan (SOZ) particles. Both lidocaine and bupivacaine inhibited the release of the inflammatory mediators leukotriene B4 (LTB4) and interleukin-1 (IL-1) evaluated by radioimmunoassay (RIA). Pretreatment of suspended PMNGs and monocytes with the anaesthetics caused a marked inhibition of LTB4 release when the cells were stimulated with SOZ. In short-term (24 h) cultures of mononuclear cells the addition of lidocaine or bupivacaine reduced, in a dose-dependent manner, the level of IL-1 detected after stimulation with lipopolysaccharide (LPS). In all three assays (chemiluminescence, LTB4 and IL-1 RIA) bupivacaine was found to be more potent than lidocaine. The present results show that amide local anaesthetics have marked suppressive effects on the metabolic activation and secretory functions of leukocytes stimulated by different agonists. Although the detailed mechanisms for these effects are not known, they may explain part of the potent anti-inflammatory actions of local anaesthetics previously described in vivo.