Organelle tethering, pore formation and SNARE compensation in the late endocytic pathway.

To provide insights into the kiss-and-run and full fusion events resulting in endocytic delivery to lysosomes, we investigated conditions causing increased tethering and pore formation between late endocytic organelles in HeLa cells. Knockout of the soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) VAMP7 and VAMP8 showed, by electron microscopy, the accumulation of tethered lysosome-associated membrane protein (LAMP)-carrier vesicles around multivesicular bodies, as well as the appearance of 'hourglass' profiles of late endocytic organelles attached by filamentous tethers, but did not prevent endocytic delivery to lysosomal hydrolases. Subsequent depletion of the SNARE YKT6 reduced this delivery, consistent with it compensating for the absence of VAMP7 and VAMP8. We also investigated filamentous tethering between multivesicular bodies and enlarged endolysosomes following depletion of charged multi-vesicular body protein 6 (CHMP6), and provide the first evidence that pore formation commences at the edge of tether arrays, with pore expansion required for full membrane fusion.

Altered pituitary morphology as a sign of benign hereditary chorea caused by TITF1/NKX2.1 mutations.

Benign hereditary chorea (BHC) is a rare genetically heterogeneous movement disorder, in which conventional neuroimaging has been reported as normal in most cases. Cystic pituitary abnormalities and features of empty sella have been described in only 7 patients with BHC to date. We present 4 patients from 2 families with a BHC phenotype, 3 of whom underwent targeted pituitary MR imaging and genetic testing. All four patients in the two families displayed a classic BHC phenotype. The targeted pituitary MR imaging demonstrated abnormal pituitary sella morphology. Genetic testing was performed in three patients, and showed mutations causing BHC in three of the patients, as well as identifying a novel nonsense mutation of the TITF1/NKX2-1 gene in one of the patients. The presence of the abnormal pituitary sella in two affected members of the same family supports the hypothesis that this sign is a distinct feature of the BHC phenotype spectrum due to mutations in the TITF1 gene. Interestingly, these abnormalities seem to develop in adult life and are progressive. They occur in at least 26% of patients affected with Brain-lung-thyroid syndrome. As a part of the management of these patients we recommend to perform follow-up MRI brain with dedicated pituitary imaging also in adult life as the abnormality can occur years after the onset of chorea.

Retinal Architecture in Autosomal Recessive Spastic Ataxia of Charlevoix-Saguenay (ARSACS): Insights into Disease Pathogenesis and Biomarkers.

BACKGROUND: Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) causes unique retinal abnormalities, which have not been systematically investigated. OBJECTIVE: To deeply phenotype the retina in ARSACS in order to better understand its pathogenesis and identify potential biomarkers. METHODS: We evaluated 29 patients with ARSACS, 66 with spinocerebellar ataxia (SCA), 38 with autosomal recessive cerebellar ataxia (ATX), 22 with hereditary spastic paraplegia (SPG), 21 cases of papilledema, and 20 healthy controls (total n = 196 subjects). Participants underwent visual acuity assessment, intraocular pressure measurement, fundoscopy, and macular and peripapillary optical coherence tomography (OCT). Macular layers thicknesses in ARSACS were compared with those of age-matched healthy controls. Ophthalmologists analyzed the scans for abnormal signs in the different patient groups. Linear regression analysis was conducted to look for associations between retinal changes and age, age at onset, disease duration, and Scale for the Assessment and Rating of Ataxia (SARA) scores in ARSACS. RESULTS: Only patients with ARSACS exhibited peripapillary retinal striations (82%) on fundoscopy, and their OCT scans revealed foveal hypoplasia (100%), sawtooth appearance (89%), papillomacular fold (86%), and macular microcysts (18%). Average peripapillary retinal nerve fiber layer (pRNFL) was thicker in ARSACS than in SCA, ATX, SPG, and controls; a cut-off of 121 µm was 100% accurate in diagnosing ARSACS. All macular layers were thicker in ARSACS when compared to healthy controls. RNFL thickness in the inferior sector of the macula positively correlated with SARA scores. CONCLUSIONS: Retinal abnormalities are highly specific for ARSACS, and suggest retinal hyperplasia due to abnormal retinal development. OCT may provide potential biomarkers for future clinical trials. © 2021 International Parkinson and Movement Disorder Society.

Prediction of Survival With Long-Term Disease Progression in Most Common Spinocerebellar Ataxia.

BACKGROUND: Spinocerebellar ataxias are rare dominantly inherited neurodegenerative diseases that lead to severe disability and premature death. OBJECTIVE: To quantify the impact of disease progression measured by the Scale for the Assessment and Rating of Ataxia on survival, and to identify different profiles of disease progression and survival. METHODS: Four hundred sixty-two spinocerebellar ataxia patients from the EUROSCA prospective cohort study, suffering from spinocerebellar ataxia type 1, spinocerebellar ataxia type 2, spinocerebellar ataxia type 3, and spinocerebellar ataxia type 6, and who had at least two measurements of Scale for the Assessment and Rating of Ataxia score, were analyzed. Outcomes were change over time in Scale for the Assessment and Rating of Ataxia score and time to death. Joint model was used to analyze disease progression and survival. RESULTS: Disease progression was the strongest predictor for death in all genotypes: An increase of 1 standard deviation in total Scale for the Assessment and Rating of Ataxia score increased the risk of death by 1.28 times (95% confidence interval: 1.18-1.38) for patients with spinocerebellar ataxia type 1; 1.19 times (1.12-1.26) for spinocerebellar ataxia type 2; 1.30 times (1.19-1.42) for spinocerebellar ataxia type 3; and 1.26 times (1.11-1.43) for spinocerebellar ataxia type 6. Three subgroups of disease progression and survival were identified for patients with spinocerebellar ataxia type 1: "severe" (n = 13; 12%), "intermediate" (n = 31; 29%), and "moderate" (n = 62; 58%). Patients in the severe group were more severely affected at baseline with higher Scale for the Assessment and Rating of Ataxia scores and frequency of nonataxia signs compared to those in the other groups. CONCLUSION: Rapid ataxia progression is associated with poor survival of the most common spinocerebellar ataxia. Theses current results have implications for the design of future interventional studies of spinocerebellar ataxia. © 2019 International Parkinson and Movement Disorder Society.

Optical coherence tomography in autosomal recessive spastic ataxia of Charlevoix-Saguenay.

Autosomal recessive spastic ataxia of Charlevoix-Saguenay is a rare neurodegenerative disorder caused by mutations in the SACS gene. Thickened retinal nerve fibres visible on fundoscopy have previously been described in these patients; however, thickening of the retinal nerve fibre layer as demonstrated by optical coherence tomography appears to be a more sensitive and specific feature. To test this observation, we assessed 292 individuals (191 patients with ataxia and 101 control subjects) by peripapillary time-domain optical coherence tomography. The patients included 146 with a genetic diagnosis of ataxia (17 autosomal spastic ataxia of Charlevoix-Saguenay, 59 Friedreich's ataxia, 53 spinocerebellar ataxias, 17 other genetically confirmed ataxias) and 45 with cerebellar ataxia of unknown cause. The controls included 13 asymptomatic heterozygotes for SACS mutations and 88 unaffected controls. The cases with autosomal recessive spastic ataxia of Charlevoix-Saguenay included 11 previously unpublished SACS mutations, of which seven were nonsense and four missense mutations. Most patients were visually asymptomatic and had no previous history of ophthalmic complaints and normal or near normal visual test results. None had visual symptoms directly attributable to the retinal changes. Twelve of the 17 cases (70.6%) had thickened retinal nerve fibres visible on fundoscopy. All patients with autosomal recessive spastic ataxia of Charlevoix-Saguenay had thickening of the peripapillary retinal nerve fibre layer on optical coherence tomography, whereas all the remaining cases and controls except one showed normal or reduced average peripapillary retinal nerve fibre layer thickness on optical coherence tomography. We propose a cut-off value of 119 µm in average peripapillary retinal nerve fibre layer thickness, which provides a sensitivity of 100% and specificity of 99.4% amongst patients affected with ataxia. This is the largest cohort of patients with this condition to undergo systematic evaluation by optical coherence tomography. This is a useful tool in identifying cases of autosomal recessive spastic ataxia of Charlevoix-Saguenay from other causes of ataxia. Visualization of thickened retinal fibres by direct fundoscopy is less sensitive. We therefore advocate the use of this technique in the assessment of possible cases of this condition.

The impact of genetic counseling on patient engagement in a specialty cancer clinic.

The identification of patient outcomes unique to the field of genetic counseling has become a recent priority of the profession. Current health-care efforts have targeted patient engagement as an outcome capable of improving population health and reducing health-care costs. This study analyzed patient engagement levels among 182 participants who underwent genetic counseling for gastrointestinal (GI) cancer risk assessment in an outpatient specialty clinic. Patients seen at the UPMC Hereditary GI Tumor Program completed a validated patient engagement measure, the Altarum Consumer Engagement (ACE), prior to undergoing genetic counseling and again three months after enrollment. Paired t test analysis was conducted to assess the changes in Total ACE scores, and within the following three domains: Navigation, Informed Choice, and Commitment. In the sample of 182 participants, Total ACE scores increased after genetic counseling (by 5.7%; p < .0001), as did all three domains (Commitment p = .0008; Navigation p = .0008; and Informed Choice p = .0016). This study is the first known report of patient engagement levels in individuals undergoing genetic counseling in a specialty cancer clinic and suggests that genetic counseling improves patient engagement levels.

The Healthy Health Care Workplace: a Competitive Advantage.

PURPOSE OF REVIEW: This review provides an evidence-based approach to improve health, reduce excessive medical costs, and optimize productivity for health care employees and their families. What opportunities and challenges are unique or specific to health care organizations particularly as aligned with value and competitive advantage in achieving the Quadruple Aim? RECENT FINDINGS: An integrated approach to improving health and performance is essential. The strategy includes the employer "environment" (broadly defined), health and medical care behaviors, care delivery modalities, benefits alignment, and a supportive, total health and productivity integrated data analytic capability. Epigenetics and lifestyle medicine represent a promising direction in accelerating the prevention, treatment, and reversal of common chronic disease. An integrated health and productivity approach, emerging science, and practices can accelerate health care systems' goal to improve employee health and organizational competitiveness. Additional published examples of health care specific employer experience will further advance refinement of existing models and tailoring to the medical care setting.

Compound heterozygous FXN mutations and clinical outcome in friedreich ataxia.

OBJECTIVE: Friedreich ataxia (FRDA) is an inherited neurodegenerative disease characterized by ataxia and cardiomyopathy. Homozygous GAA trinucleotide repeat expansions in the first intron of FXN occur in 96% of affected individuals and reduce frataxin expression. Remaining individuals are compound heterozygous for a GAA expansion and a FXN point/insertion/deletion mutation. We examined disease-causing mutations and the impact on frataxin structure/function and clinical outcome in FRDA. METHODS: We compared clinical information from 111 compound heterozygotes and 131 individuals with homozygous expansions. Frataxin mutations were examined using structural modeling, stability analyses and systematic literature review, and categorized into four groups: (1) homozygous expansions, and three compound heterozygote groups; (2) null (no frataxin produced); (3) moderate/strong impact; and (4) minimal impact. Mean age of onset and the presence of cardiomyopathy and diabetes mellitus were compared using regression analyses. RESULTS: Mutations in the hydrophobic core of frataxin affected stability whereas surface residue mutations affected interactions with iron sulfur cluster assembly and heme biosynthetic proteins. The null group of compound heterozygotes had significantly earlier age of onset and increased diabetes mellitus, compared to the homozygous expansion group. There were no significant differences in mean age of onset between homozygotes and the minimal and moderate/strong impact groups. INTERPRETATION: In compound heterozygotes, expression of partially functional mutant frataxin delays age of onset and reduces diabetes mellitus, compared to those with no frataxin expression from the non-expanded allele. This integrated analysis of categorized frataxin mutations and their correlation with clinical outcome provide a definitive resource for investigating disease pathogenesis in FRDA.

When the heart rules the head: ischaemic stroke and intracerebral haemorrhage complicating infective endocarditis.

Sir William Osler meticulously described the clinical manifestations of infective endocarditis in 1885, concluding that: 'few diseases present greater difficulties in the way of diagnosis … which in many cases are practically insurmountable'. Even with modern investigation techniques, diagnosing infective endocarditis can be hugely challenging, yet is critically important in patients presenting with stroke (both cerebral infarction and intracranial haemorrhage), its commonest neurological complication. In ischaemic stroke, intravenous thrombolysis carries an unacceptably high risk of intracranial haemorrhage, while in intracerebral haemorrhage, mycotic aneurysms require urgent treatment to avoid rebleeding, and in all cases, prompt treatment with antibiotics and valve surgery may be life-saving. Here, we describe typical presentations of ischaemic stroke and intracerebral haemorrhage caused by infective endocarditis. We review the diagnostic challenges, the importance of rapid diagnosis, treatment options and controversies.

A non-canonical ESCRT pathway, including histidine domain phosphotyrosine phosphatase (HD-PTP), is used for down-regulation of virally ubiquitinated MHC class I.

The Kaposi's sarcoma-associated herpes virus (KSHV) K3 viral gene product effectively down-regulates cell surface MHC class I. K3 is an E3 ubiquitin ligase that promotes Lys(63)-linked polyubiquitination of MHC class I, providing the signal for clathrin-mediated endocytosis. Endocytosis is followed by sorting into the intralumenal vesicles (ILVs) of multivesicular bodies (MVBs) and eventual delivery to lysosomes. The sorting of MHC class I into MVBs requires many individual proteins of the four endosomal sorting complexes required for transport (ESCRTs). In HeLa cells expressing the KSHV K3 ubiquitin ligase, the effect of RNAi-mediated depletion of individual proteins of the ESCRT-0 and ESCRT-I complexes and three ESCRT-III proteins showed that these are required to down-regulate MHC class I. However, depletion of proteins of the ESCRT-II complex or of the ESCRT-III protein, VPS20 (vacuolar protein sorting 20)/CHMP6 (charged MVB protein 6), failed to prevent the loss of MHC class I from the cell surface. Depletion of histidine domain phosphotyrosine phosphatase (HD-PTP) resulted in an increase in the cell surface concentration of MHC class I in HeLa cells expressing the KSHV K3 ubiquitin ligase. Rescue experiments with wild-type (WT) and mutant HD-PTP supported the conclusion that HD-PTP acts as an alternative to ESCRT-II and VPS20/CHMP6 as a link between the ESCRT-I and those ESCRT-III protein(s) necessary for ILV formation. Thus, the down-regulation of cell surface MHC class I, polyubiquitinated by the KSHV K3 ubiquitin ligase, does not employ the canonical ESCRT pathway, but instead utilizes an alternative pathway in which HD-PTP replaces ESCRT-II and VPS20/CHMP6.

Rabies virus envelope glycoprotein targets lentiviral vectors to the axonal retrograde pathway in motor neurons.

Rabies pseudotyped lentiviral vectors have great potential in gene therapy, not least because of their ability to transduce neurons following their distal axonal application. However, very little is known about the molecular processes that underlie their retrograde transport and cell transduction. Using multiple labeling techniques and confocal microscopy, we demonstrated that pseudotyping with rabies virus envelope glycoprotein (RV-G) enabled the axonal retrograde transport of two distinct subtypes of lentiviral vector in motor neuron cultures. Analysis of this process revealed that these vectors trafficked through Rab5-positive endosomes and accumulated within a non-acidic Rab7 compartment. RV-G pseudotyped vectors were co-transported with both the tetanus neurotoxin-binding fragment and the membrane proteins thought to mediate rabies virus endocytosis (neural cell adhesion molecule, nicotinic acetylcholine receptor, and p75 neurotrophin receptor), thus demonstrating that pseudotyping with RV-G targets lentiviral vectors for transport along the same pathway exploited by several toxins and viruses. Using motor neurons cultured in compartmentalized chambers, we demonstrated that axonal retrograde transport of these vectors was rapid and efficient; however, it was not able to transduce the targeted neurons efficiently, suggesting that impairment in processes occurring after arrival of the viral vector in the soma is responsible for the low transduction efficiency seen in vivo, which suggests a novel area for improvement of gene therapy vectors.

Epigenetic and neurological effects and safety of high-dose nicotinamide in patients with Friedreich's ataxia: an exploratory, open-label, dose-escalation study.

BACKGROUND: Friedreich's ataxia is a progressive degenerative disorder caused by deficiency of the frataxin protein. Expanded GAA repeats within intron 1 of the frataxin (FXN) gene lead to its heterochromatinisation and transcriptional silencing. Preclinical studies have shown that the histone deacetylase inhibitor nicotinamide (vitamin B3) can remodel the pathological heterochromatin and upregulate expression of FXN. We aimed to assess the epigenetic and neurological effects and safety of high-dose nicotinamide in patients with Friedreich's ataxia. METHODS: In this exploratory, open-label, dose-escalation study in the UK, male and female patients (aged 18 years or older) with Friedreich's ataxia were given single doses (phase 1) and repeated daily doses of 2-8 g oral nicotinamide for 5 days (phase 2) and 8 weeks (phase 3). Doses were gradually escalated during phases 1 and 2, with individual maximum tolerated doses used in phase 3. The primary outcome was the upregulation of frataxin expression. We also assessed the safety and tolerability of nicotinamide, used chromatin immunoprecipitation to investigate changes in chromatin structure at the FXN gene locus, and assessed the effect of nicotinamide treatment on clinical scales for ataxia. This study is registered with ClinicalTrials.gov, number NCT01589809. FINDINGS: Nicotinamide was generally well tolerated; the main adverse event was nausea, which in most cases was mild, dose-related, and resolved spontaneously or after dose reduction, use of antinausea drugs, or both. Phase 1 showed a dose-response relation for proportional change in frataxin protein concentration from baseline to 8 h post-dose, which increased with increasing dose (p=0·0004). Bayesian analysis predicted that 3·8 g would result in a 1·5-times increase and 7·5 g in a doubling of frataxin protein concentration. Phases 2 and 3 showed that daily dosing at 3·5-6 g resulted in a sustained and significant (p<0·0001) upregulation of frataxin expression, which was accompanied by a reduction in heterochromatin modifications at the FXN locus. Clinical measures showed no significant changes. INTERPRETATION: Nicotinamide was associated with a sustained improvement in frataxin concentrations towards those seen in asymptomatic carriers during 8 weeks of daily dosing. Further investigation of the long-term clinical benefits of nicotinamide and its ability to ameliorate frataxin deficiency in Friedreich's ataxia is warranted. FUNDING: Ataxia UK, Ataxia Ireland, Association Suisse de l'Ataxie de Friedreich, Associazione Italiana per le Sindromi Atassiche, UK National Institute for Health Research, European Friedreich's Ataxia Consortium for Translational Studies, and Imperial Biomedical Research Centre.

A novel de novo mutation of the TITF1/NKX2-1 gene causing ataxia, benign hereditary chorea, hypothyroidism and a pituitary mass in a UK family and review of the literature.

Benign hereditary chorea (BHC) is a rare autosomal dominant condition characterized by early onset, non-progressive chorea, usually caused by mutations in the thyroid transcription factor-1 gene (TITF1). We describe a novel mutation arising de novo in a proband presenting in infancy with delayed walking and ataxia. She later developed chorea, then hypothyroidism and a large cystic pituitary mass. Her daughter presented in infancy with delayed walking and ataxia and went on to develop non-progressive chorea and a hormonally inactive cystic pituitary mass. Mutational analysis of the whole coding region of the TITF1 gene was undertaken and compared with a population study of 160 control subjects. This showed that both affected subjects have a heterozygous A > T substitution at nucleotide 727 of the TITF1 gene changing lysine to a stop codon at residue 211. Genetic analysis of parents and siblings of the proband confirmed that the mutation arose de novo in the proband. The mutated lysine is an evolutionarily highly conserved amino acid in the protein homoeodomain (HD) where most point mutations associated with BHC are located. The range of mutations in BHC is reviewed with particular emphasis on pituitary abnormalities. Cystic pituitary masses and abnormalities of the sella turcica are reported in just 6.4 % of published cases. This is a new nonsense mutation associated with ataxia, benign chorea and pituitary abnormalities which further extends the phenotype of this condition. Mutational screening of TITF1 is important in cases of sporadic or dominant juvenile-onset ataxia, with mild chorea where no other cause is found, particularly if pituitary abnormalities are seen on imaging.

Accurate whole human genome sequencing using reversible terminator chemistry.

DNA sequence information underpins genetic research, enabling discoveries of important biological or medical benefit. Sequencing projects have traditionally used long (400-800 base pair) reads, but the existence of reference sequences for the human and many other genomes makes it possible to develop new, fast approaches to re-sequencing, whereby shorter reads are compared to a reference to identify intraspecies genetic variation. Here we report an approach that generates several billion bases of accurate nucleotide sequence per experiment at low cost. Single molecules of DNA are attached to a flat surface, amplified in situ and used as templates for synthetic sequencing with fluorescent reversible terminator deoxyribonucleotides. Images of the surface are analysed to generate high-quality sequence. We demonstrate application of this approach to human genome sequencing on flow-sorted X chromosomes and then scale the approach to determine the genome sequence of a male Yoruba from Ibadan, Nigeria. We build an accurate consensus sequence from >30x average depth of paired 35-base reads. We characterize four million single-nucleotide polymorphisms and four hundred thousand structural variants, many of which were previously unknown. Our approach is effective for accurate, rapid and economical whole-genome re-sequencing and many other biomedical applications.

Comparison of traditional copromicroscopy with image analysis devices for detection of gastrointestinal nematode infection in sheep.

Sustainable parasite control practices are necessary to combat the negative effects of gastrointestinal nematodes on animal health and production while reducing the selection pressure for anthelmintic resistance. Parasite diagnostic tests can inform treatment decisions, the timing and effectiveness of treatment and enable livestock breeding programmes. In recent years new diagnostic methods have been developed, some incorporating machine learning (ML), to facilitate the detection and enumeration of parasite eggs. It is important to understand the technical characteristics and performance of such new methods compared to long standing and commonly utilised methods before they are widely implemented. The aim of the present study was to trial three new diagnostic tools relying on image analysis (FECPAKG2, Micron and OvaCyte) and to compare them to traditional manual devices (McMaster and Mini-FLOTAC). Faecal samples were obtained from 41 lambs naturally infected with gastrointestinal nematodes. Samples were mixed and separated into 2 aliquots for examination by each of the 5 methods: McMaster, Mini-FLOTAC, FECPAKG2, Micron and OvaCyte. The techniques were performed according to their respective standard protocols and results were collected by trained staff (McMaster and Mini-FLOTAC) or by the device (FECPAKG2, Micron and OvaCyte). Regarding strongyle worm egg count, McMaster values varied from 0 to 9,000 eggs per gram (EPG). When comparing replicate aliquots, both the Mini-FLOTAC and Micron methods displayed similar repeatability to McMaster. However, we found FECPAKG2 and OvaCyte significantly less precise than McMaster. When comparing parasite egg enumeration, significant positive linear correlations were established between McMaster and all other methods. No difference was observed in EPG between McMaster and Mini-FLOTAC or FECPAKG2; however, Micron and OvaCyte returned significantly higher and lower EPG, respectively, compared to McMaster. The number of eggs ascribed to other parasite species was not sufficient for performing a robust statistical comparison between all methods. However, it was noted that FECPAKG2 generally did not detect Strongyloides papillosus eggs, despite these being detected by other methods. In addition, Moniezia spp and Trichuris spp eggs were detected by OvaCyte and Mini-FLOTAC, respectively, but not by other methods. The observed variation between traditional and new methods for parasite diagnostics highlights the need for continued training and enhancing of ML models used and the importance of developing clear guidelines for validation of newly developed methods.

American College of Lifestyle Medicine Expert Consensus Statement: Lifestyle Medicine for Optimal Outcomes in Primary Care.

OBJECTIVE: Identify areas of consensus on integrating lifestyle medicine (LM) into primary care to achieve optimal outcomes. METHODS: Experts in both LM and primary care followed an a priori protocol for developing consensus statements. Using an iterative, online process, panel members expressed levels of agreement with statements, resulting in classification as consensus, near consensus, or no consensus. RESULTS: The panel identified 124 candidate statements addressing: (1) Integration into Primary Care, (2) Delivery Models, (3) Provider Education, (4) Evidence-base for LM, (5) Vital Signs, (6) Treatment, (7) Resource Referral and Reimbursement, (8) Patient, Family, and Community Involvement; Shared Decision-Making, (9) Social Determinants of Health and Health Equity, and (10) Barriers to LM. After three iterations of an online Delphi survey, statement revisions, and removal of duplicative statements, 65 statements met criteria for consensus, 24 for near consensus, and 35 for no consensus. Consensus was reached on key topics that included LM being recognized as an essential component of primary care in patients of all ages, including LM as a foundational element of health professional education. CONCLUSION: The practice of LM in primary care can be strengthened by applying these statements to improve quality of care, inform policy, and identify areas for future research.

Co-enzyme Q10 and idebenone use in Friedreich's ataxia.

Friedreich's ataxia is a debilitating progressive neurodegenerative disease associated with cardiomyopathy and other features. The underlying cause is a deficiency of the mitochondrial protein frataxin which causes mitochondrial iron deposition, increased oxidative stress and impaired adenosine triphosphate production. Over the last 15 years, multiple clinical trials have assessed the efficacy of antioxidant agents in this disease. This article reviews trials of the two most important agents, namely co-enzyme Q10 and idebenone.

Clinical features of Friedreich's ataxia: classical and atypical phenotypes.

One hundred and fifty years since Nikolaus Friedreich's first description of the degenerative ataxic syndrome which bears his name, his description remains at the core of the classical clinical phenotype of gait and limb ataxia, poor balance and coordination, leg weakness, sensory loss, areflexia, impaired walking, dysarthria, dysphagia, eye movement abnormalities, scoliosis, foot deformities, cardiomyopathy and diabetes. Onset is typically around puberty with slow progression and shortened life-span often related to cardiac complications. Inheritance is autosomal recessive with the vast majority of cases showing an unstable intronic GAA expansion in both alleles of the frataxin gene on chromosome 9q13. A small number of cases are caused by a compound heterozygous expansion with a point mutation or deletion. Understanding of the underlying molecular biology has enabled identification of atypical phenotypes with late onset, or atypical features such as retained reflexes. Late-onset cases tend to have slower progression and are associated with smaller GAA expansions. Early-onset cases tend to have more rapid progression and a higher frequency of non-neurological features such as diabetes, cardiomyopathy, scoliosis and pes cavus. Compound heterozygotes, including those with large deletions, often have atypical features. In this paper, we review the classical and atypical clinical phenotypes of Friedreich's ataxia.

Plasma cytokines, chemokines and cellular immune responses in pre-school Nigerian children infected with Plasmodium falciparum.

BACKGROUND: Malaria is a major cause of morbidity and mortality worldwide with over one million deaths annually, particularly in children under five years. This study was the first to examine plasma cytokines, chemokines and cellular immune responses in pre-school Nigerian children infected with Plasmodium falciparum from four semi-urban villages near Ile-Ife, Osun State, Nigeria. METHODS: Blood was obtained from 231 children (aged 39-73 months) who were classified according to mean P. falciparum density per µl of blood (uninfected (n = 89), low density (<1,000, n = 51), medium density (1,000-10,000, n = 65) and high density (>10,000, n = 22)). IL-12p70, IL-10, Nitric oxide, IFN-γ, TNF, IL-17, IL-4 and TGF-ß, C-C chemokine RANTES, MMP-8 and TIMP-1 were measured in plasma. Peripheral blood mononuclear cells were obtained and examined markers of innate immune cells (CD14, CD36, CD56, CD54, CD11c AND HLA-DR). T-cell sub-populations (CD4, CD3 and γδTCR) were intracellularly stained for IL-10, IFN-γ and TNF following polyclonal stimulation or stimulated with malaria parasites. Ascaris lumbricoides was endemic in these villages and all data were analysed taking into account the potential impact of bystander helminth infection. All data were analysed using SPSS 15 for windows and in all tests, p <0.05 was deemed significant. RESULTS: The level of P. falciparum parasitaemia was positively associated with plasma IL-10 and negatively associated with IL-12p70. The percentage of monocytes was significantly decreased in malaria-infected individuals while malaria parasitaemia was positively associated with increasing percentages of CD54+, CD11c+ and CD56+ cell populations. No association was observed in cytokine expression in mitogen-activated T-cell populations between groups and no malaria specific immune responses were detected. Although A. lumbricoides is endemic in these villages, an analysis of the data showed no impact of this helminth infection on P. falciparum parasitaemia or on immune responses associated with P. falciparum infection. CONCLUSIONS: These findings indicate that Nigerian children infected with P. falciparum exhibit immune responses associated with active malaria infection and these responses were positively associated with increased P. falciparum parasitaemia.