HLA-DPB1 and HLA class I confer risk of and protection from narcolepsy.

Type 1 narcolepsy, a disorder caused by a lack of hypocretin (orexin), is so strongly associated with human leukocyte antigen (HLA) class II HLA-DQA1(∗)01:02-DQB1(∗)06:02 (DQ0602) that very few non-DQ0602 cases have been reported. A known triggering factor for narcolepsy is pandemic 2009 influenza H1N1, suggesting autoimmunity triggered by upper-airway infections. Additional effects of other HLA-DQ alleles have been reported consistently across multiple ethnic groups. Using over 3,000 case and 10,000 control individuals of European and Chinese background, we examined the effects of other HLA loci. After careful matching of HLA-DR and HLA-DQ in case and control individuals, we found strong protective effects of HLA-DPA1(∗)01:03-DPB1(∗)04:02 (DP0402; odds ratio [OR] = 0.51 [0.38-0.67], p = 1.01 × 10(-6)) and HLA-DPA1(∗)01:03-DPB1(∗)04:01 (DP0401; OR = 0.61 [0.47-0.80], p = 2.07 × 10(-4)) and predisposing effects of HLA-DPB1(∗)05:01 in Asians (OR = 1.76 [1.34-2.31], p = 4.71 × 10(-05)). Similar effects were found by conditional analysis controlling for HLA-DR and HLA-DQ with DP0402 (OR = 0.45 [0.38-0.55] p = 8.99 × 10(-17)) and DP0501 (OR = 1.38 [1.18-1.61], p = 7.11 × 10(-5)). HLA-class-II-independent associations with HLA-A(∗)11:01 (OR = 1.32 [1.13-1.54], p = 4.92 × 10(-4)), HLA-B(∗)35:03 (OR = 1.96 [1.41-2.70], p = 5.14 × 10(-5)), and HLA-B(∗)51:01 (OR = 1.49 [1.25-1.78], p = 1.09 × 10(-5)) were also seen across ethnic groups in the HLA class I region. These effects might reflect modulation of autoimmunity or indirect effects of HLA class I and HLA-DP alleles on response to viral infections such as that of influenza.

Cerebrospinal fluid cytokine levels in type 1 narcolepsy patients very close to onset.

Type 1 narcolepsy is caused by a loss of hypocretin (orexin) signaling in the brain. Genetic data suggests the disorder is caused by an autoimmune attack on hypocretin producing neurons in hypothalamus. This hypothesis has however not yet been confirmed by consistent findings of autoreactive antibodies or T-cells in patient samples. One explanation for these negative results may be that the autoimmune process is no longer active when patients present to the clinic. With increasing awareness in recent years, more and more patients have been diagnosed closer and closer to disease onset. In this study, we tested whether an active immune process in the brain could be detected in these patients, as reflected by increased cytokine levels in the cerebrospinal fluid (CSF). Using multiplex analysis, we measured the levels of 51 cytokines and chemokines in the CSF of 40 type 1 narcolepsy patients having varying disease duration. For comparison, we used samples from 9 healthy controls and 9 patients with other central hypersomnia. Cytokine levels did not differ significantly between controls and patients, even in 5 patients with disease onset less than a month prior to CSF sampling.

Reduced CSF hypocretin-1 levels are associated with cluster headache.

BACKGROUND: Cluster headache (CH) is a debilitating disorder characterized by unilateral, severe pain attacks with accompanying autonomic symptoms, often waking the patient from sleep. As it exhibits strong chronobiological traits and genetic studies have suggested a link with the hypocretin (HCRT) system, the objective of this study was to investigate HCRT-1 in CH patients. METHODS: Cerebrospinal fluid HCRT-1 concentration was measured in 12 chronic and 14 episodic CH patients during an active bout, and in 27 healthy controls. The patients were well characterized and clinical features compared to the HCRT concentration. RESULTS: We found significantly lower HCRT levels both in chronic (p = 0.0221) and episodic CH (p = 0.0005) patients compared with controls. No significant relationship was found with other clinical features. CONCLUSIONS: This is the first report of significantly reduced HCRT concentrations in CH patients. We speculate that decreased HCRT may reflect insufficient antinociceptive activity of the hypothalamus. The mechanism of the antinociceptive effect of HCRT is not known and requires further investigation. This study supports the hypothesis of a connection between arousal regulation and CH.

EGFR signaling patterns are regulated by its different ligands.

EGF receptor (EGFR) and its signaling have been investigated for many years, but how its different ligands regulate signaling has not been thoroughly explored. When investigating EGFR activation and downstream signaling in HeLa cells using a panel of ligands, we found a ligand-dependent differential activation of EGFR and the signaling pathways Akt, PLCγ and STAT with HB-EGF and BTC being the most potent ligands. All the tested ligands induced full activation of Erk signaling at 1 nM, whereas only HB-EGF and partly BTC and EGF induced strong activation of Akt, STAT3 and PLCγ at this concentration. Interestingly, we also found that the high activation potencies of HB-EGF and BTC could only partially be explained by their binding affinities, and are therefore likely to be regulated by other mechanisms. We thus suggest that the signaling pathways initiated from the EGFR vary depending on the ligands bound in a cell specific manner.

Clinical, polysomnographic and genome-wide association analyses of narcolepsy with cataplexy: a European Narcolepsy Network study.

The aim of this study was to describe the clinical and PSG characteristics of narcolepsy with cataplexy and their genetic predisposition by using the retrospective patient database of the European Narcolepsy Network (EU-NN). We have analysed retrospective data of 1099 patients with narcolepsy diagnosed according to International Classification of Sleep Disorders-2. Demographic and clinical characteristics, polysomnography and multiple sleep latency test data, hypocretin-1 levels, and genome-wide genotypes were available. We found a significantly lower age at sleepiness onset (men versus women: 23.74 ± 12.43 versus 21.49 ± 11.83, P = 0.003) and longer diagnostic delay in women (men versus women: 13.82 ± 13.79 versus 15.62 ± 14.94, P = 0.044). The mean diagnostic delay was 14.63 ± 14.31 years, and longer delay was associated with higher body mass index. The best predictors of short diagnostic delay were young age at diagnosis, cataplexy as the first symptom and higher frequency of cataplexy attacks. The mean multiple sleep latency negatively correlated with Epworth Sleepiness Scale (ESS) and with the number of sleep-onset rapid eye movement periods (SOREMPs), but none of the polysomnographic variables was associated with subjective or objective measures of sleepiness. Variant rs2859998 in UBXN2B gene showed a strong association (P = 1.28E-07) with the age at onset of excessive daytime sleepiness, and rs12425451 near the transcription factor TEAD4 (P = 1.97E-07) with the age at onset of cataplexy. Altogether, our results indicate that the diagnostic delay remains extremely long, age and gender substantially affect symptoms, and that a genetic predisposition affects the age at onset of symptoms.

Differential effects of EGFR ligands on endocytic sorting of the receptor.

Endocytic downregulation is a pivotal mechanism turning off signalling from the EGF receptor (EGFR). It is well established that whereas EGF binding leads to lysosomal degradation of EGFR, transforming growth factor (TGF)-alpha causes receptor recycling. TGF-alpha therefore leads to continuous signalling and is a more potent mitogen than EGF. In addition to EGF and TGF-alpha, five EGFR ligands have been identified. Although many of these ligands are upregulated in cancers, very little is known about their effect on EGFR trafficking. We have compared the effect of six different ligands on endocytic trafficking of EGFR. We find that, whereas they all stimulate receptor internalization, they have very diverse effects on endocytic sorting. Heparin-binding EGF-like growth factor and Betacellulin target all EGFRs for lysosomal degradation. In contrast, TGF-alpha and epiregulin lead to complete receptor recycling. EGF leads to lysosomal degradation of the majority but not all EGFRs. Amphiregulin does not target EGFR for lysosomal degradation but causes fast as well as slow EGFR recycling. The Cbl ubiquitin ligases, especially c-Cbl, are responsible for EGFR ubiquitination after stimulation with all ligands, and persistent EGFR phosphorylation and ubiquitination largely correlate with receptor degradation.

Rapid eye movement sleep behaviour disorder in patients with narcolepsy is associated with hypocretin-1 deficiency.

Rapid eye movement sleep behaviour disorder is characterized by dream-enacting behaviour and impaired motor inhibition during rapid eye movement sleep. Rapid eye movement sleep behaviour disorder is commonly associated with neurodegenerative disorders, but also reported in narcolepsy with cataplexy. Most narcolepsy with cataplexy patients lack the sleep-wake, and rapid eye movement sleep, motor-regulating hypocretin neurons in the lateral hypothalamus. In contrast, rapid eye movement sleep behaviour disorder and hypocretin deficiency are rare in narcolepsy without cataplexy. We hypothesized that rapid eye movement sleep behaviour disorder coexists with cataplexy in narcolepsy due to hypocretin deficiency. In our study, rapid eye movement sleep behaviour disorder was diagnosed by the International Classification of Sleep Disorders (2nd edition) criteria in 63 narcolepsy patients with or without cataplexy. Main outcome measures were: rapid eye movement sleep behaviour disorder symptoms; short and long muscle activations per hour rapid eye movement and non-rapid eye movement sleep; and periodic and non-periodic limb movements per hour rapid eye movement and non-rapid eye movement sleep. Outcome variables were analysed in relation to cataplexy and hypocretin deficiency with uni- and multivariate logistic/linear regression models, controlling for possible rapid eye movement sleep behaviour disorder biasing factors (age, gender, disease duration, previous anti-cataplexy medication). Only hypocretin deficiency independently predicted rapid eye movement sleep behaviour disorder symptoms (relative risk = 3.69, P = 0.03), long muscle activations per hour rapid eye movement sleep (ln-coefficient = 0.81, P < 0.01), and short muscle activations per hour rapid eye movement sleep (ln-coefficient = 1.01, P < 0.01). Likewise, periodic limb movements per hour rapid eye movement and non-rapid eye movement sleep were only associated with hypocretin deficiency (P < 0.01). A significant association between hypocretin deficiency and cataplexy was confirmed (P < 0.01). In a sub-analysis, hypocretin deficiency suggested the association of periodic limb movements and rapid eye movement sleep behaviour disorder outcomes (symptoms, non-periodic short and long muscle activity) in rapid eye movement sleep. Our results support the hypothesis that hypocretin deficiency is independently associated with rapid eye movement sleep behaviour disorder in narcolepsy. Thus, hypocretin deficiency is linked to the two major disturbances of rapid eye movement sleep motor regulation in narcolepsy: rapid eye movement sleep behaviour disorder and cataplexy. Hypocretin deficiency is also significantly associated with periodic limb movements in rapid eye movement and non-rapid eye movement sleep, and provides a possible pathophysiological link between rapid eye movement sleep behaviour disorder and periodic limb movements in narcolepsy. The study supports the hypothesis that an impaired hypocretin system causes a general instability of motor regulation during wakefulness, rapid eye movement and non-rapid eye movement sleep in human narcolepsy.

Validation of the ICSD-2 criteria for CSF hypocretin-1 measurements in the diagnosis of narcolepsy in the Danish population.

STUDY OBJECTIVES: The International Classification of Sleep Disorders (ICSD-2) criteria for low CSF hypocretin-1 levels (CSF hcrt-1) still need validation as a diagnostic tool for narcolepsy in different populations because inter-assay variability and different definitions of hypocretin deficiency complicate direct comparisons of study results. DESIGN AND PARTICIPANTS: Interviews, polysomnography, multiple sleep latency test, HLA-typing, and CSF hcrt-1 measurements in Danish patients with narcolepsy with cataplexy (NC) and narcolepsy without cataplexy (NwC), CSF hcrt-1 measurements in other hypersomnias, neurological and normal controls. Comparisons of hypocretin deficiency and frequency of HLA-DQB1*0602-positivity in the Danish and eligible NC and NwC populations (included via MEDLINE search), by (re)calculation of study results using the ICSD-2 criterion for low CSF hcrt-1 (< 30% of normal mean). MEASUREMENTS AND RESULTS: In Danes, low CSF hcrt-1 was present in 40/46 NC, 3/14 NwC and 0/106 controls (P < 0.0001). Thirty-nine of 41 NC and 4/13 NwC patients were HLA-DQB1*0602-positive (P < 0.01). Hypocretin-deficient NC patients had higher frequency of cataplexy, shorter mean sleep latency, more sleep onset REM periods (P < 0.05) and more awakenings (NS) than did NC patients with normal CSF hcrt-1. Across populations, low CSF hcrt-1 and HLA-DQB1*0602-positivity characterized the majority of NC (80% to 100%, P = 0.53; 80% to 100%, P = 0.11) but a minority of NwC patients (11% to 29%, P = 0.75; 29% to 89%, P = 0.043). CONCLUSION: The study provides evidence that hypocretin deficiency causes a more severe NC phenotype. The ICSD-2 criterion for low CSF hcrt-1 (< 30% of normal mean) is valid for diagnosing NC, but not NwC. HLA-typing should precede CSF hcrt-1 measurements because hypocretin deficiency is rare in HLA-DQB1*0602-negative patients.

Anti-Tribbles homolog 2 (TRIB2) autoantibodies in narcolepsy are associated with recent onset of cataplexy.

STUDY OBJECTIVE: Recent studies have found increased autoantibodies against Tribbles homolog 2 (anti-TRIB2) and anti-streptolysin O (ASO) in narcolepsy. In this study, we replicated this finding with a primary focus on recent onset cases. PARTICIPANTS AND METHODS: Participants included (1) 90 cases with cataplexy, (2) 57 cases without cataplexy, and (3) 156 age-sex matched controls, including 73 human leukocyte antigen (HLA)-DQB1*0602 allele carriers. A radioligand binding assay was used to detect anti-TRIB2 antibodies. RESULTS: Anti-TRIB2 antibodies were prevalent in HLA-DQB1*0602 positive cases with cataplexy (25.0% of 76) and rare in cases without cataplexy (3.5% of 57, OR = 9.2, 95% CI = 2.5 - 33.5, P = 6.0 x 10(-4)) or controls (4.5% of 156, OR = 7.1, 95% CI = 3.1 - 16.2, P = 9.3 x 10(-6)). Anti-TRIB2 positivity in controls was not associated with DQB1*0602. In DQB1*0602 narcolepsy-cataplexy cases, the presence of anti-TRIB2 was associated with short disease duration (2.3 years from cataplexy onset), with 41.0% positive in this group (OR = 7.4 versus cases with onset > 2.3 years, 95% CI = 1.9- 28.5, P = 9.0 x 10(-4)). Anti-TRIB2 positivity in 39 DQB1*0602 positive recent onset cases was associated with increased ASO antibody (> 200 IU) (OR = 6.2, 95% CI = 1.6 - 24.6, P = 0.01), but did not correlate with age, gender, or body mass index. CONCLUSION: Anti-TRIB2 autoantibodies are strongly associated with narcolepsy close to cataplexy onset (< or = 2.3 years). Anti-TRIB2 was rarely found in cases without cataplexy or with distant onset.

Emergency drug kits at the Danish hospital pharmacies: varying management and challenges.

OBJECTIVES: Access to emergency drug kits (EDK) during medical emergencies can be life-saving; however, recent doubts about the quality of the kits have been expressed. Procurements of pharmaceuticals to the five regional authorities in Denmark are serviced by Amgros, a public sector organisation owned by the regions and established to create economies of scale and achieve administrative savings by centralisation. This means that Amgros calls for tenders for the supply of pharmaceuticals to the hospital pharmacies. The Hospital Pharmacy in the North Denmark Region does not currently have an effective method to manage Amgros procurements in relation to EDKs. Thus, the objectives were to explore how quality in the management and packing of EDKs is assured and maintained at different hospital pharmacies in Denmark and how this is affected by Amgros procurements. METHODS: The hospital pharmacies in Denmark were enrolled in a cross-sectional study. Information about the management and challenges of the EDKs was inquired by means of a questionnaire. Responses were analysed by simple statistics. RESULTS: All eight hospital pharmacies in Denmark completed the questionnaire, and the distribution between single-use and reusable packaging was nearly equal. The hospital pharmacies comply with a variation of regulations of which good distribution practice is the most common. Six hospital pharmacies experience challenges with drug replacements in the EDKs and only one hospital pharmacy complies completely with the Amgros procurement. The majority of the hospital pharmacies use parameters such as price of the new drug and potential expense for new packaging in their decision of whether to comply with the Amgros procurement. CONCLUSION: The management of the EDKs varies greatly among the hospital pharmacies in Denmark, and national requirements are therefore encouraged to ensure the quality. The challenges experienced with drug replacements reflect that complying with the Amgros procurement can be troublesome.

Effect of glycosylation on the extracellular domain of the Ag43 bacterial autotransporter: enhanced stability and reduced cellular aggregation.

Autotransporters constitute the biggest group of secreted proteins in Gram-negative bacteria and contain a membrane-bound beta-domain and a passenger domain secreted to the extracellular environment via an unusually long N-terminal sequence. Several passenger domains are known to be glycosylated by cytosolic glycosyl transferases, promoting bacterial attachment to mammalian cells. In the present study we describe the effect of glycosylation on the extracellular passenger domain of the Escherichia coli autotransporter Ag43alpha, which induces frizzy colony morphology and cell settling. We identify 16 glycosylation sites and suggest two possible glycosylation motifs for serine and threonine residues. Glycosylation stabilizes against thermal and chemical denaturation and increases refolding kinetics. Unexpectedly, glycosylation also reduces the stabilizing effect of Ca(2+) ions, removes the ability of Ca(2+) to promote cell adhesion, reduces the ability of Ag43alpha-containing cells to form bacterial amyloid and increases the susceptibility of the resulting amyloid to proteolysis. In addition, our results indicate that Ag43alpha folds without a stable intermediate, unlike pertactin, indicating that autotransporters may arrive at the native state by a variety of different mechanisms despite a common overall structure. A small but significant fraction of Ag43alpha can survive intact in the periplasm if expressed without the beta-domain, suggesting that it is able to adopt a protease-resistant structure prior to translocation across the membrane. The present study demonstrates that glycosylation may play significant roles in structural and functional properties of bacterial autotransporters at many different levels.

Hypocretin-deficient narcolepsy patients have abnormal brain activation during humor processing.

STUDY OBJECTIVES: To assess brain activation patterns in response to fun-rated and neutral-rated movies we performed functional magnetic resonance imaging (fMRI) during a humor-paradigm in narcolepsy type 1 (NT1) patients with cataplexy (muscle atonia triggered by emotions) and controls. METHODS: The fMRI-humor-paradigm consisted of short movies (25/30 with a humorous punchline; 5/30 without a humorous punchline [but with similar build-up/anticipation]) rated by participants based on their humor experience. We included 41 NT1 patients and 44 controls. Group-level inferences were made using permutation testing. RESULTS: Permutation testing revealed no group differences in average movie ratings. fMRI analysis found no group differences in brain activations to fun-rated movies. Patients showed significantly higher activations compared to controls during neutral-rated movies; including bilaterally in the thalamus, pallidum, putamen, amygdala, hippocampus, middle temporal gyrus, cerebellum, brainstem and in the left precuneus, supramarginal gyrus, and caudate. We found no brain overactivation for patients during movies without a humorous punchline (89.0% neutral-rated). Group analyses revealed significantly stronger differentiation between fun-rated and neutral-rated movies in controls compared with patients (patients showed no significant differentiation), including bilaterally in the inferior frontal gyrus, thalamus, putamen, precentral gyrus, lingual gyrus, supramarginal gyrus, occipital areas, temporal areas, cerebellum and in the right hippocampus, postcentral gyrus, pallidum, and insula. CONCLUSION: Patients showed significantly higher activations in several cortical and subcortical regions during neutral-rated movies, with no differentiation from activations during fun-rated movies. This lower threshold for activating the humor response (even during neutral-rated movies), might represent insight into the mechanisms associated with cataplexy.

Psychiatric symptoms in patients with post-H1N1 narcolepsy type 1 in Norway.

STUDY OBJECTIVES: Several studies have reported psychiatric comorbidity in patients with narcolepsy type 1 (NC1). The primary aim of this study was to explore the extent of psychiatric symptoms in a cohort of Norwegian NC1 patients, most of whom were H1N1-vaccinated. We also wanted to explore possible causes of the psychiatric symptoms seen in NC1. METHODS: Cross-sectional study. Psychiatric symptoms were assessed by the Achenbach System of Empirically Based Assessment (ASEBA) Child Behavior Check List (CBCL) in children and by Adult Self Report (ASR) in adults. RESULTS: The mean (SD) total T-scores were 58.6 (9.2) for children and 57.0 (9.8) for adults, these being mainly driven by internalizing problems. Internalizing symptom T-scores showed that 37.5% of the children and 33.3% of the adults were in the clinical range of concern. T-scores were lower when the questionnaire's sleep-related items were excluded. However, 27.5% of children and 22.2% of adults still remained within the total psychiatric symptoms clinical range. Psychiatric symptoms and excessive daytime sleepiness were not associated. However, in children fragmented sleep, measured by sleep-stage shift index was significantly negatively associated with all the psychiatric summary scores (all p ≤ 0.020), and awakening index was negatively associated with externalizing (p = 0.042) and total summary scores (p = 0.042). In adults, awakening index, but not sleep-stage shift index, was positively associated with internalizing score (p = 0.015). Hypocretin-1 levels showed no association with psychiatric symptoms. CONCLUSIONS: We found a high prevalence of psychiatric symptoms in NC1 patients. Fragmented sleep was significantly associated with psychiatric symptoms.

CD8+ T cells from patients with narcolepsy and healthy controls recognize hypocretin neuron-specific antigens.

Narcolepsy Type 1 (NT1) is a neurological sleep disorder, characterized by the loss of hypocretin/orexin signaling in the brain. Genetic, epidemiological and experimental data support the hypothesis that NT1 is a T-cell-mediated autoimmune disease targeting the hypocretin producing neurons. While autoreactive CD4+ T cells have been detected in patients, CD8+ T cells have only been examined to a minor extent. Here we detect CD8+ T cells specific toward narcolepsy-relevant peptides presented primarily by NT1-associated HLA types in the blood of 20 patients with NT1 as well as in 52 healthy controls, using peptide-MHC-I multimers labeled with DNA barcodes. In healthy controls carrying the disease-predisposing HLA-DQB1*06:02 allele, the frequency of autoreactive CD8+ T cells was lower as compared with both NT1 patients and HLA-DQB1*06:02-negative healthy individuals. These findings suggest that a certain level of CD8+ T-cell reactivity combined with HLA-DQB1*06:02 expression is important for NT1 development.

Changes in quality of life in individuals with narcolepsy type 1 after the H1N1-influenza epidemic and vaccination campaign in Norway: a two-year prospective cohort study.

OBJECTIVE: Cross-sectional studies show a lower health-related quality of life (HRQoL) in individuals with narcolepsy. We aimed to describe changes in HRQoL after two years of multidisciplinary follow-up in a cohort of mainly post-H1N1 vaccination narcolepsy type-1 (NT1) patients in Norway. METHODS: Prospective-cohort study. Narcolepsy diagnosis was based on the International Classification of Sleep Disorders (third edition). Psychiatric comorbidity was assessed using the Achenbach System of Empirically Based Assessment (ASEBA). HRQoL was evaluated with the Pediatric Quality of Life Inventory (PedsQL™ Generic Core Scales 4.0) at baseline and follow-up. Mean follow-up time was 20.7 (2.7) months. RESULTS: Thirty one patients (18 females) with NT1, mean age 14.6 (SD = 4.8) years answered questionnaires at baseline and follow-up. On a group level, the PedsQL Total Health Summary score significantly improved by a mean of 5.9 (95%CI = 0.4, 11.9), p = 0.038; this was mainly driven by improvements in the Physical Health Summary score by 9.8 (3.0, 16.5) points, p = 0.006 and the School Functioning Scale score by 7.5 (1.0, 13.9) points p = 0.025. The Total ASEBA score was correlated with PedsQL Total Health Summary score at baseline, but not with changes in HRQoL. Sodium oxybate (Xyrem®) treatment at follow up was positively associated with changes in PedsQL Total Health Summary score, after adjusting for age and gender, p = 0.027. CONCLUSION: HRQoL in NT1 patients improved after two years of follow-up. The use of sodium oxybate (Xyrem®) at follow-up was associated with increases in HRQoL. Psychiatric comorbidity was correlated with HRQoL at baseline but did not predict changes in HRQoL at follow-up.

Comparison of subjective and objective measures of constipation - Employing a new method for categorizing gastrointestinal symptoms.

INTRODUCTION: Correlations between subjective and objective measures of constipation have seldom been demonstrated. This could be due to multiple confounding factors in clinical studies and the broad span of symptoms represented in questionnaires used to assess constipation. We developed a new method for categorizing gastrointestinal (GI) symptoms into relevant symptom groups, and used this in a controlled experimental study aimed to investigate whether GI transit times and colonic volumes were correlated to self-reported GI symptoms. METHODS: Twenty-five healthy male participants were enrolled in a randomized, double-blinded, placebo-controlled, five-day crossover study with the treatments oxycodone and placebo. Objective measures of GI transit times and colonic volumes were obtained by the means of the 3D-Transit System and magnetic resonance colonography, whereas subjective GI symptoms were measures via three validated questionnaires. The symptoms were then categorized into five groups; "abdominal symptoms", "defecation difficulties", "incomplete bowel evacuation", "reduced bowel movement frequency", and "stool symptoms". Spearman's rank order correlation was used to determine correlations between the five groups of symptoms and the objective measures. RESULTS: No correlations between the GI symptoms and transit times or colonic volumes were found (all P > 0.05). DISCUSSION: GI transit times and colonic volumes were not correlated to self-reported GI symptoms even in a controlled experimental study and when symptoms were categorized into relevant symptom groups. Thus, both subjective and objective measures must be considered relevant when assessing constipation in clinical and research settings, ensuring that both physiological aspects as well as the severity and impact of symptoms experienced by patients can be assessed.

Widespread white matter changes in post-H1N1 patients with narcolepsy type 1 and first-degree relatives.

Study Objectives: To assess white matter involvement in H1N1-vaccinated hypocretin deficient patients with narcolepsy type 1 (NT1) compared with first-degree relatives (a potential risk group) and healthy controls. Methods: We compared four diffusion tensor imaging-based microstructural indices (fractional anisotropy [FA], mean diffusivity [MD], radial diffusivity [RD], and axial diffusivity [AD]) in 57 patients with NT1 (39 females, mean age 21.8 years, 51/57 H1N1-vaccinated, 57/57 HLA-DQB1*06:02-positive, 54/54 hypocretin-deficient), 54 first-degree relatives (29 females, mean age 19.1 years, 37/54 H1N1-vaccinated, 32/54 HLA-DQB1*06:02-positive), and 55 healthy controls (38 females, mean age 22.3 years). We tested for differences between these groups, for parametric effects (controls > first-degree relatives > patients) and associations in patients (cerebrospinal fluid [CSF] hypocretin-1 and disease duration) and first-degree relatives (HLA-DQB1*06:02 and H1N1-vaccination). We employed tract-based spatial statistics and used permutation testing and threshold-free cluster enhancement for inference. Results: Patients with NT1 had a widespread, bilateral pattern of significantly lower FA compared with first-degree relatives and healthy controls. Additionally, patients with NT1 also exhibited significantly higher RD and lower AD in several focal white matter clusters. The parametric model showed that first-degree relatives had intermediate values. Full sample of patients with NT1 showed no significant associations with disease duration or CSF hypocretin-1. Conclusions: Our study suggests widespread abnormal white matter involvement far beyond the already known focal hypothalamic pathology in NT1, possibly reflecting the combined effects of the loss of the widely projecting hypothalamic hypocretin neurons, and/or secondary effects of wake/sleep dysregulation. These findings demonstrate the importance of white matter pathology in NT1.

Antibodies in narcolepsy-cataplexy patient serum bind to rat hypocretin neurons.

Autoimmunity is considered the most likely cause of human narcolepsy-cataplexy, but no specific autoantibodies or antigen(s) have yet been identified. By means of indirect avidin-biotin immunohistochemical method, we searched for antibodies in serum from narcolepsy-cataplexy patients and controls that bind to rat hypocretin neurons. No staining was found in eight out of nine narcolepsy-cataplexy patients or controls. The serum from one narcolepsy-cataplexy patient, however, strongly produced staining of the membrane and superficial cytoplasm of neurons in the lateral hypothalamus. Dual staining revealed that the vast majority of the hypocretin-positive neurons were positive, but nonhypocretin neurons in the same area were binding antibodies from the patient's serum. These results show that antibodies bind to specific hypocretin- and nonhypocretin-containing neurons in the hypothalamus and indicate the presence of autoantibodies in narcolepsy patients.