Clinical characteristics and proteome modifications in two Charcot-Marie-Tooth families with the AARS1 Arg326Trp mutation.

BACKGROUND: Aminoacyl tRNA-synthetases are ubiquitously-expressed enzymes that attach amino acids to their cognate tRNA molecules. Mutations in several genes encoding aminoacyl tRNA-synthetases, have been associated with peripheral neuropathy, i.e. AARS1, GARS1, HARS1, YARS1 and WARS1. The pathogenic mechanism underlying AARS1-related neuropathy is not known. METHODS: From 2012 onward, all probands presenting at Telemark Hospital (Skien, Norway) with peripheral neuropathy were screened for variants in AARS1 using an "in-house" next-generation sequencing panel. DNA from patient's family members was examined by Sanger sequencing. Blood from affected family members and healthy controls were used for quantification of AARS1 mRNA and alanine. Proteomic analyses were conducted in peripheral blood mononuclear cells (PBMC) from four affected family members and five healthy controls. RESULTS: Seventeen individuals in two Norwegian families affected by Charcot-Marie-Tooth disease (CMT) were characterized in this study. The heterozygous NM_001605.2:c.976C > T p.(Arg326Trp) AARS1 mutation was identified in ten affected family members. All living carriers had a mild to severe length-dependent sensorimotor neuropathy. Three deceased obligate carriers aged 74-98 were reported to be unaffected, but were not examined in the clinic. Proteomic studies in PBMC from four affected individuals suggest an effect on the immune system mediated by components of a systemic response to chronic injury and inflammation. Furthermore, altered expression of proteins linked to mitochondrial function/dysfunction was observed. Proteomic data are available via ProteomeXchange using identifier PXD023842. CONCLUSION: This study describes clinical and neurophysiological features linked to the p.(Arg326Trp) variant of AARS1 in CMT-affected members of two Norwegian families. Proteomic analyses based on of PBMC from four CMT-affected individuals suggest that involvement of inflammation and mitochondrial dysfunction might contribute to AARS1 variant-associated peripheral neuropathy.

A clinical interview versus prospective headache diaries in the diagnosis of menstrual migraine without aura.

OBJECTIVES: The objective of this article is to compare the diagnosis of menstrual migraine without aura (MM) from a clinical interview with prospective headache diaries in a population-based study. MATERIAL AND METHODS: A total of 237 women with self-reported migraine in at least half of menstruations were interviewed by a neurologist about headache and diagnosed according to the International Classification of Headache Disorders II (ICHD II). Additionally, the MM criteria were expanded to include other types of migraine related to menstruation. Subsequently, all women were asked to complete three month prospective headache diaries. RESULTS: A total of 123 (52%) women completed both clinical interview and diaries. Thirty-eight women were excluded from the analyses: Two had incomplete diaries and 36 women recorded ≤1 menstruation, leaving 85 diaries eligible for analysis. Sensitivity, specificity, positive and negative predictive value and Kappa for the diagnosis of MM in clinical interview vs. headache diary were 82%, 83%, 90%, 71% and 0.62 (95% CI 0.45-0.79). Using a broader definition of MM, Kappa was 0.64 (95% CI 0.47-0.83). CONCLUSION: A thorough clinical interview is valid for the diagnosis of MM. When this is undertaken, prospective headache diaries should not be mandatory to diagnose MM but may be necessary to exclude a chance association.

Hereditary peripheral neuropathies diagnosed by next-generation sequencing.

BACKGROUND: Next-generation sequencing (NGS) is a genetic technique used to determine the order of nucleotides in DNA. The technique has proved to be more efficient than the traditional method, Sanger sequencing, for sequencing multiple genes. NGS is now being used to diagnose disorders in which multiple genes are involved. This study has examined whether next-generation sequencing produces a greater number of positive diagnoses than its traditional counterpart in patients with suspected hereditary peripheral neuropathy. MATERIAL AND METHOD: This study is a retrospective review of samples from 103 patients investigated for hereditary peripheral neuropathy, received by Telemark Hospital in the period 2012-14. After exclusion of duplication/deletion of PMP22, 96 samples were analysed by NGS with physical enrichment of 52 hereditary peripheral neuropathy genes. RESULTS: A genetic cause was identified in 35 patients (34%) with peripheral neuropathy, of which 28 (27%) were point mutations identified by NGS. INTERPRETATION: Of the pathogenic point mutations identified in this study, 12 were in genes that would previously have been analysed by Sanger sequencing in our department, whereas 16 were in genes that would not previously have been tested.

Headache in sleep apnea syndrome: epidemiology and pathophysiology.

This review investigates the relation between obstructive sleep apnea and sleep apnea headache, migraine and tension-type headache. Focus is made on studies from the general population with interviews conducted by a physician and obstructive sleep apnea confirmed by polysomnography. Obstructive sleep apnea syndrome is observed in 3% of the middle-aged population. The prevalence of sleep apnea headache in this population is 12%-18%, while morning headache with similar symptomatology as sleep apnea headache occur in 5%-8% of the general population. People with sleep apnea headache did have significantly more minutes below 90% oxygen saturation (23.1 min vs. 1.9 min, p = 0.002), higher level of average oxygen desaturation (5.9% vs. 4.5%, p < 0.001) and lower average of the lowest oxygen saturation (80.9% vs. 88.5%, p < 0.001) than people with morning headache. A comparison of those with obstructive sleep apnea with or without sleep apnea headache showed no significant differences. Thus, oxygen desaturation alone cannot explain the pathophysiology of sleep apnea headache. Obstructive sleep apnea and migraine, and obstructive sleep apnea and tension-type headache are not related in the general population. The cause of sleep apnea headache remains to be elucidated.

Prevalence of menstrual migraine: a population-based study.

AIM: To present data from a population-based epidemiological study on menstrual migraine. MATERIAL AND METHODS: Altogether, 5000 women aged 30-34 years were screened for menstrual migraine. Women with self-reported menstrual migraine in at least half of their menstrual cycles were invited to an interview and examination. We expanded the International Classification of Headache Disorders III beta appendix criteria on menstrual migraine to include both migraine without aura and migraine with aura, as well as probable menstrual migraine with aura and migraine without aura. RESULTS: A total of 237 women were included in the study. The prevalence among all women was as follows: any type of menstrual migraine 7.6%; menstrual migraine without aura 6.1%; menstrual migraine with aura 0.6%; probable menstrual migraine without aura 0.6%; probable menstrual migraine with aura 0.3%. The corresponding figures among female migraineurs were: any type of menstrual migraine 22.0%, menstrual migraine without aura 17.6%, menstrual migraine with aura 1.7%, probable menstrual migraine without aura 1.6% and probable menstrual migraine with aura 1.0%. CONCLUSION: More than one of every five female migraineurs aged 30-34 years have migraine in ≥50% of menstruations. The majority has menstrual migraine without aura and one of eight women had migraine with aura in relation to their menstruation. Our results indicate that the ICHD III beta appendix criteria of menstrual migraine are not exhaustive.

A Multicenter Study of Left Ventricular Assist Device-Related Gastrointestinal Bleeding.

INTRODUCTION: Continuous left ventricular assist devices (LVADs) offer hemodynamic support in advanced and decompensated heart failure but are often complicated by gastrointestinal bleeding (GIB) in medically fragile patients. METHODS: We performed a retrospective analysis of 475 consecutive patients who underwent LVAD implantation at the Massachusetts General Hospital and Tufts Medical Center from 2008 to 2019 and identified 128 patients with clinically significant GIB. Clinical characteristics of each bleeding event, including procedures and interventions, were recorded. We examined LVAD patients with overt and occult presentations to determine diagnostic endoscopic yield and analyzed predictors of recurrent GIB. RESULTS: We identified 128 unique patients with LVAD implantation complicated by GIB. No significant difference was observed based on study center, underlying cardiomyopathy, race/ethnicity, serum indices, and medications used. Overt bleeders presented more commonly during LVAD implantation admission ( P = 0.001) than occult bleeders. Occult bleed presentations had only 1 lower and no middle GI bleed source identified, despite similar workups to overt bleeds. Destination therapy (e.g., among nontransplant candidates) LVAD implantation (odds ratio 2.38, 95% confidence interval 1.05-5.58) and a history of GIB (odds ratio 3.85, 95% confidence interval 1.29-12.7) were independently associated with an increased risk of recurrent GIB-related hospitalization. DISCUSSION: Our findings confirm a high rate of GIB, especially in destination LVAD patients, and show a low diagnostic yield for colonoscopy and middle GI bleed assessments in LVAD patients with occult bleeds. Overt bleeding was more common and associated with vascular malformations. Although endoscopic interventions stopped active hemorrhage, GIB often recurred.

A Norwegian population-based study on the risk and prevalence of obstructive sleep apnea. The Akershus Sleep Apnea Project (ASAP).

The Berlin Questionnaire (BQ) is a widely used screening tool for obstructive sleep apnea (OSA), but its performance in the general population setting is unknown. The prevalence of OSA in middle-aged adults is not known in Norway. Accordingly, the aims of the current study were to evaluate the utility of the BQ for OSA screening in the general population and to estimate the prevalence of OSA in Norway. The study population consisted of 29,258 subjects (aged 30-65 years, 50% female) who received the BQ by mail. Of these, 16,302 (55.7%) responded. Five-hundred and eighteen subjects were included in the clinical sample and underwent in-hospital polysomnography. Screening properties and prevalence were estimated by a statistical model that adjusted for bias in the sampling procedure. Among the 16,302 respondents, 24.3% (95% confidence interval (CI)=23.6-25.0%) were classified by the BQ to be at high-risk of having OSA. Defining OSA as an apnea-hypopnea index (AHI) ≥5, the positive predictive value of the BQ was estimated to be 61.3%, the negative predictive value 66.2%, the sensitivity 37.2% and the specificity 84.0%. Estimated prevalences of OSA were 16% for AHI≥5 and 8% for AHI≥15. In conclusion, the BQ classified one out of four middle-aged Norwegians to be at high-risk of having OSA, but the screening properties of the BQ were suboptimal. The estimated prevalence of OSA was comparable to previous estimates from general populations in the USA, Australia and Europe.

A Rare Case of Waldenstrom Macroglobulinemia of the Rectosigmoid Colon.

Waldenstrom macroglobulinemia is an uncommon mature B-cell lymphoma characterized by monoclonal immunoglobulin M protein in peripheral blood and lymphoplasmacytic cells in bone marrow and/or extramedullary sites. The gastrointestinal tract is a rare site of involvement. The diagnosis is based on clinicopathologic findings, although somatic mutations, such as MYD88, can aid in the diagnosis. We present a patient with irregular stools diagnosed with Waldenstrom macroglobulinemia involving the rectosigmoid colon by histopathology and immunohistochemistry on colonic biopsies, immunoglobulin M protein in serum, clonal plasma cells in bone marrow, and MYD88 mutation in colonic and bone marrow specimens.

MFN2 point mutations occur in 3.4% of Charcot-Marie-Tooth families. An investigation of 232 Norwegian CMT families.

BACKGROUND: Point mutations in the mitofusin 2 (MFN2) gene has been identified exclusively in Charcot-Marie-Tooth type 2 (CMT2), and in a single family with intermediate CMT. MFN2 point mutations are probably the most common cause of CMT2. METHODS: Two-hundred and thirty-two consecutive unselected and unrelated CMT families with available DNA from all regions in Norway were included. We screened for point mutations in the MFN2 gene. RESULTS: We identified four known and three novel point mutations in 8 unrelated CMT families. The novel point mutations were not found in 100 healthy controls. This corresponds to 3.4% (8/232) of CMT families have point mutations in the MFN2 gene. The phenotypes were compatible with CMT1 in two families, CMT2 in four families, intermediate CMT in one family and distal Hereditary Motor Neuropathy (dHMN) in one family. This corresponds to 2.3% of CMT1, 5.5% of CMT2, 12.5% of intermediate CMT and 6.7% of dHMN families have a point mutation in the MFN2 gene. Point mutations in the MFN2 gene is likely to be the fourth most common cause to CMT after duplication of the peripheral myelin protein 22 (PMP22) gene, and point mutations in the Connexin32 (Cx32) and myelin protein zero (MPZ) genes. CONCLUSIONS: The identified known and novel point mutations in the MFN2 gene expand the clinical spectrum from CMT2 and intermediate CMT to also include possibly CMT1 and the dHMN phenotypes. Thus, genetic analyses of the MFN2 gene should not be restricted to persons with CMT2.

An Unusual Cause of Lower Gastrointestinal Bleeding: Cecal Dieulafoy's Lesion.

Colonic Dieulafoy's lesions are an exceptionally rare cause of lower gastrointestinal (GI) bleeding. These lesions are almost exclusively found in the upper GI tract based on previous reviews. We present a case of an 81-year-old man who presented with melena progressing to hematochezia and was found to have a cecal Dieulafoy's lesion on colonoscopy. Hemostasis with clipping was achieved and allowed for the resumption of anticoagulation. This case demonstrates the importance of considering this diagnosis in lower GI bleeding when evidence of more common causes may not be present, especially considering these lesions amenability to endoscopic therapy.

Dubin-Johnson Syndrome Presenting During Cardiac Transplantation Evaluation.

Dubin-Johnson syndrome is a rare, benign disorder that results in conjugated hyperbilirubinemia. The disease manifests as intermittent jaundice without long-term hepatic or other clinical complications. This article reports a case of Dubin-Johnson syndrome, which was identified during cardiac transplant evaluation for cardiomyopathy secondary to a polyglycogen storage disease. The patient successfully underwent an orthotopic heart transplant. Postoperatively, her conjugated hyperbilirubinemia increased as compared to her baseline but resolved after several weeks. This report briefly reviews the hepatic manifestations in patients with Dubin-Johnson syndrome undergoing major surgery and highlights urinary coproporphyrin as a useful diagnostic test for Dubin-Johnson syndrome.

Sp1 regulates osteopontin expression in SW480 human colon adenocarcinoma cells.

BACKGROUND: Osteopontin (OPN) mediates cancer metastasis. Mechanisms regulating OPN expression in human colorectal cancer are unknown. Using SW480 colon adenocarcinoma cells, we hypothesized that transcription determines OPN expression. METHODS: SW480 constitutively express OPN. Transient transfection and deletion analysis of human OPN promoter (full-length 2.1 kb)-luciferase constructs identified cis-regulatory regions. Gelshift and chromatin immunoprecipitation (ChIP) assays identified the trans-regulatory nuclear protein. Using in vitro adhesion, migration, and invasion studies, siRNA was used to determine the functional effect of decreased nuclear protein expression. RESULTS: A cis-regulatory promoter region, nt-80 to nt-108, upregulated OPN transcription. Gelshift assays demonstrated specific binding of nuclear proteins. Competition with unlabeled mutant oligonucleotides indicated that the region, nt-94 to nt-104 (TGGGCTGGGC), was essential for protein binding in gelshift assays. Confirmatory ChIP assays showed the corresponding nuclear protein to be Sp1. Sp1 expression was ablated with siRNA (si-Sp1), resulting in decreased OPN-dependent adhesion, migration, and invasion by 50%, 70%, and 65%, respectively. Exogenous addition of OPN to si-Sp1 cells restored adhesion, migration, and invasion indices. CONCLUSIONS: In SW480 human colon cancer cells, we conclude that Sp1 mediated expression of the tumor metastasis protein, OPN, regulates in vitro functional correlates of tumor metastasis.

Two novel connexin32 mutations cause early onset X-linked Charcot-Marie-Tooth disease.

BACKGROUND: X-linked Charcot-Marie Tooth (CMT) is caused by mutations in the connexin32 gene that encodes a polypeptide which is arranged in hexameric array and form gap junctions. METHODS: We describe two novel mutations in the connexin32 gene in two Norwegian families. RESULTS: Family 1 had a c.225delG (R75fsX83) which causes a frameshift and premature stop codon at position 247. This probably results in a shorter non-functional protein structure. Affected individuals had an early age at onset usually in the first decade. The symptoms were more severe in men than women. All had severe muscle weakness in the legs. Several abortions were observed in this family. Family 2 had a c.536 G>A (C179Y) transition which causes a change of the highly conserved cysteine residue, i.e. disruption of at least one of three disulfide bridges. The mean age at onset was in the first decade. Muscle wasting was severe and correlated with muscle weakness in legs. The men and one woman also had symptom from their hands. The neuropathy is demyelinating and the nerve conduction velocities were in the intermediate range (25-49 m/s). Affected individuals had symmetrical clinical findings, while the neurophysiology revealed minor asymmetrical findings in nerve conduction velocity in 6 of 10 affected individuals. CONCLUSION: The two novel mutations in the connexin32 gene are more severe than the majority of previously described mutations possibly due to the severe structural change of the gap junction they encode.

Copy number variations in a population-based study of Charcot-Marie-Tooth disease.

Copy number variations (CNVs) are important in relation to diversity and evolution but can sometimes cause disease. The most common genetic cause of the inherited peripheral neuropathy Charcot-Marie-Tooth disease is the PMP22 duplication; otherwise, CNVs have been considered rare. We investigated CNVs in a population-based sample of Charcot-Marie-Tooth (CMT) families. The 81 CMT families had previously been screened for the PMP22 duplication and point mutations in 51 peripheral neuropathy genes, and a genetic cause was identified in 37 CMT families (46%). Index patients from the 44 CMT families with an unknown genetic diagnosis were analysed by whole-genome array comparative genomic hybridization to investigate the entire genome for larger CNVs and multiplex ligation-dependent probe amplification to detect smaller intragenomic CNVs in MFN2 and MPZ. One patient had the pathogenic PMP22 duplication not detected by previous methods. Three patients had potentially pathogenic CNVs in the CNTNAP2, LAMA2, or SEMA5A, that is, genes related to neuromuscular or neurodevelopmental disease. Genotype and phenotype correlation indicated likely pathogenicity for the LAMA2 CNV, whereas the CNTNAP2 and SEMA5A CNVs remained potentially pathogenic. Except the PMP22 duplication, disease causing CNVs are rare but may cause CMT in about 1% (95% CI 0-7%) of the Norwegian CMT families.

Genetic diagnosis of Charcot-Marie-Tooth disease in a population by next-generation sequencing.

Charcot-Marie-Tooth (CMT) disease is the most prevalent inherited neuropathy. Today more than 40 CMT genes have been identified. Diagnosing heterogeneous diseases by conventional Sanger sequencing is time consuming and expensive. Thus, more efficient and less costly methods are needed in clinical diagnostics. We included a population based sample of 81 CMT families. Gene mutations had previously been identified in 22 families; the remaining 59 families were analysed by next-generation sequencing. Thirty-two CMT genes and 19 genes causing other inherited neuropathies were included in a custom panel. Variants were classified into five pathogenicity classes by genotype-phenotype correlations and bioinformatics tools. Gene mutations, classified certainly or likely pathogenic, were identified in 37 (46%) of the 81 families. Point mutations in known CMT genes were identified in 21 families (26%), whereas four families (5%) had point mutations in other neuropathy genes, ARHGEF10, POLG, SETX, and SOD1. Eleven families (14%) carried the PMP22 duplication and one family carried a MPZ duplication (1%). Most mutations were identified not only in known CMT genes but also in other neuropathy genes, emphasising that genetic analysis should not be restricted to CMT genes only. Next-generation sequencing is a cost-effective tool in diagnosis of CMT improving diagnostic precision and time efficiency.

Sleep apnea headache: a growing concern in an increasingly obese population?

Sleep apnea headache is a recurrent universal pressing headache without accompanying symptoms at awakening that resolves within 4 h. The diagnosis requires polysomnography-verified apnea hypopnea index≥5, that is, obstructive sleep apnea (OSA). Morning headache has similar symptomatology without OSA. The prevalence of sleep apnea headache is 10-15% in people with OSA, whereas morning headache occurs in 5%. The severity of OSA only slightly affects the prevalence of sleep apnea headache. The pathophysiology of sleep apnea headache remains an enigma, since average oxygen desaturation and lowest oxygen saturation are similar in OSA people without sleep apnea headache. Migraine and tension-type headache are unrelated to OSA. Thus, growing concern of sleep apnea headache in an increasingly obese population is unfounded with our current knowledge.

Charcot-Marie-Tooth caused by a copy number variation in myelin protein zero.

INTRODUCTION: Charcot-Marie-Tooth disease (CMT) is the most common inherited disorder of the peripheral nervous system. The majority has a duplication of the peripheral myelin protein 22. CMT is otherwise caused by point mutations or small insertions/deletions in one of the 44 known CMT genes. METHODS AND RESULTS: Conventional sequencing of six CMT genes were followed by Multiplex Ligation-dependent Probe Amplification (MLPA), array Comparative Genomic Hybridization (aCGH) and breakpoint analysis in a large Norwegian CMT pedigree. Affected had an extra copy of the myelin protein zero (MPZ) gene. CONCLUSION: To our knowledge this is the first non-peripheral myelin protein 22 copy number variation to cause Charcot-Marie-Tooth disease.

Two novel missense mutations in the myelin protein zero gene causes Charcot-Marie-Tooth type 2 and Déjérine-Sottas syndrome.

BACKGROUND: The Charcot-Marie-Tooth (CMT) phenotype caused by mutation in the myelin protein zero (MPZ) gene varies considerably, from early onset and severe forms to late onset and milder forms. The mechanism is not well understood. The myelin protein zero (P0) mediates adhesion in the spiral wraps of the Schwann cell's myelin sheath. The crystalline structure of the extracellular domain of the myelin protein zero (P0ex) is known, while the transmembrane and intracellular structure is unknown. FINDINGS: One novel missense mutation caused a milder late onset CMT type 2, while the second missense mutation caused a severe early onset phenotype compatible with Déjérine-Sottas syndrome. CONCLUSIONS: The phenotypic variation caused by different missense mutations in the MPZ gene is likely caused by different conformational changes of the MPZ protein which affects the functional tetramers. Severe changes of the MPZ protein cause dysfunctional tetramers and predominantly uncompacted myelin, i.e. the severe phenotypes congenital hypomyelinating neuropathy and Déjérine-Sottas syndrome, while milder changes cause the phenotypes CMT type 1 and 2.