Pathology-supported genetic testing as a method for disability prevention in multiple sclerosis (MS). Part I. Targeting a metabolic model rather than autoimmunity.

In this Review (Part I), we investigate the scientific evidence that multiple sclerosis (MS) is caused by the death of oligodendrocytes, the cells that synthesize myelin, due to a lack of biochemical and nutritional factors involved in mitochondrial energy production in these cells. In MS, damage to the myelin sheaths surrounding nerve axons causes disruption of signal transmission from the brain to peripheral organs, which may lead to disability. However, the extent of disability is not deterred by the use of MS medication, which is based on the autoimmune hypothesis of MS. Rather, disability is associated with the loss of brain volume, which is related to the loss of grey and white matter. A pathology-supported genetic testing (PSGT) method, developed for personalized assessment and treatment to prevent brain volume loss and disability progression in MS is discussed. This involves identification of MS-related pathogenic pathways underpinned by genetic variation and lifestyle risk factors that may converge into biochemical abnormalities associated with adverse expanded disability status scale (EDSS) outcomes and magnetic resonance imaging (MRI) findings during patient follow-up. A Metabolic Model is presented which hypothesizes that disability may be prevented or reversed when oligodendrocytes are protected by nutritional reserve. Evidence for the validity of the Metabolic Model may be evaluated in consecutive test cases following the PSGT method. In Part II of this Review, two cases are presented that describe the PSGT procedures and the clinical outcomes of these individuals diagnosed with MS.

Pathology-supported genetic testing as a method for disability prevention in multiple sclerosis (MS). Part II. Insights from two MS cases.

In Part I of this Review we evaluated the scientific evidence for a Metabolic Model of multiple sclerosis (MS). Part II outlines the implementation of an adaptive pathology-supported genetic testing (PSGT) algorithm aimed at preventing/reversing disability in two illustrative MS cases, starting with a questionnaire-based risk assessment, including family history and lifestyle factors. Measurement of iron, vitamin B12, vitamin D, cholesterol and homocysteine levels identified biochemical deficits in both cases. Case 1, after following the PSGT program for 15 years, had an expanded disability status scale (EDSS) of 2.0 (no neurological sequelae) together with preserved brain volume on magnetic resonance imaging (MRI). A novel form of iron deficiency was identified in Case 1, as biochemical testing at each hospital submission due to MS symptoms showed low serum iron, ferritin and transferrin saturation, while hematological status and erythrocyte sedimentation rate measurement of systemic inflammation remained normal. Case 2 was unable to walk unaided until her EDSS improved from 6.5 to 4.0 over 12 months after implementation of the PSGT program, with amelioration of her suboptimal biochemical markers and changes to her diet and lifestyle, allowing her to regain independence. Genotype-phenotype correlation using a pathway panel of functional single nucleotide variants (SNVs) to facilitate clinical interpretation of whole exome sequencing (WES), elucidated the underlying metabolic pathways related to the biochemical deficits. A cure for MS will remain an elusive goal if separated from nutritional support required for production and maintenance of myelin, which can only be achieved by a lifelong investment in wellness.

Fractional anisotropy of white matter, disability and blood iron parameters in multiple sclerosis.

Multiple sclerosis (MS) is a disorder related to myelin damage, which can be investigated by neuroimaging techniques such as fractional anisotropy (FA), a measure of microstructural white matter properties. The objectives of this study were to investigate (1) the relationship between FA and disability using an extremes of outcome approach, and (2) whether blood iron parameters were associated with FA and/or disability. Patients diagnosed with MS (n = 107; 14 males and 93 females) had iron parameter tests and disability determinations using the Expanded Disability Status Scale (EDSS). FA was recorded in 48 white matter tracts in 11 of the female patients with MS and 12 female controls. RESULTS: In patients with high disability scores the mean FA was significantly lower (0.34 ± 0.067) than in the control group (0.45 ± 0.036; p = 0.04), while patients with low disability had mean FA values (0.44 ± 0.014) similar to controls (p = 0.5). Positive associations were found between FA and the iron parameters serum iron, ferritin and percentage transferrin saturation (%Tfsat) in all the white matter tracts. For % Tfsat, the associations were highly significant in 14 tracts (p < 0.01; r-values 0.74-0.84) and p < 0.001 (r = 0.83) in the superior fronto occipital fasciculus (LH). In the whole patient group a trend was found towards an inverse association between the EDSS and the %Tfsat (r = -0.26, p = 0.05) after excluding male gender and smoking as confounders, suggesting reduced disability in the presence of higher blood iron parameters. Additionally, significant inverse associations between disease duration and haemoglobin (p = 0.04) as well as %Tfsat (p = 0.02) suggested that patients with MS may experience a decrease in blood iron concentrations over time.

Exome Sequencing in a Family with Luminal-Type Breast Cancer Underpinned by Variation in the Methylation Pathway.

Panel-based next generation sequencing (NGS) is currently preferred over whole exome sequencing (WES) for diagnosis of familial breast cancer, due to interpretation challenges caused by variants of uncertain clinical significance (VUS). There is also no consensus on the selection criteria for WES. In this study, a pathology-supported genetic testing (PSGT) approach was used to select two BRCA1/2 mutation-negative breast cancer patients from the same family for WES. Homozygosity for the MTHFR 677 C>T mutation detected during this PSGT pre-screen step was considered insufficient to cause bilateral breast cancer in the index case and her daughter diagnosed with early-onset breast cancer (<30 years). Extended genetic testing using WES identified the RAD50 R385C missense mutation in both cases. This rare variant with a minor allele frequency (MAF) of <0.001 was classified as a VUS after exclusion in an affected cousin and extended genotyping in 164 unrelated breast cancer patients and 160 controls. Detection of functional polymorphisms (MAF > 5%) in the folate pathway in all three affected family members is consistent with inheritance of the luminal-type breast cancer in the family. PSGT assisted with the decision to pursue extended genetic testing and facilitated clinical interpretation of WES aimed at reduction of recurrence risk.

Multiple sclerosis-like diagnosis as a complication of previously treated malaria in an iron and vitamin D deficient Nigerian patient.

In contrast to malaria, multiple sclerosis (MS) is infrequently found in Black Africans. We describe a 29 year old Nigerian female who developed an MS-like condition with symptoms similar to relapsing-remitting MS following malaria infection, leading to a diagnosis of MS. However, absence of hyperintense lesions in the brain and spinal cord presented a conundrum since not all the diagnostic criteria for MS were met. Pathology supported genetic testing (PSGT) was applied to combine family and personal medical history, lifestyle factors, and biochemical test results for interpretation of genetic findings. This approach provides a means of identifying risk factors for different subtypes of demyelinating disease. The patient was subsequently treated according to an individualised intervention program including nutritional supplementation as well as a change in diet and lifestyle. Deficiencies of vitamin B12, iron and vitamin D were addressed. Genetic analysis revealed absence of the HLA DRB1*1501 allele, considered to be the most prominent genetic risk factor for MS. Extended mutation analysis identified variations in three genes in the folate-vitamin B12 metabolic pathway, which could have increased the patient's sensitivity to the antifolate drugs used to treat the malaria. A glutathione-S-transferase GSTM1 null allele, previously associated with neurological complications of malaria, was also detected. Furthermore, a heterozygous variation in the iron-related transmembrane protease serine 6 (TMPRSS6) gene, rs855791 was found, which could have impacted the patient's iron status following two successive blood donations and exposure to malaria preceding the MS diagnosis. PSGT identifies relevant risk factors for demyelinating disorders resembling MS and uses the data for individualised treatment programs, and to systematically build a database that can provide evidence in large patient cohorts. Follow-up investigations may be suggested, such as whole exome sequencing in selected cases, to ensure that remyelination and restoration of function are achieved.

Genomic medicine and risk prediction across the disease spectrum.

Genomic medicine is based on the knowledge that virtually every medical condition, disease susceptibility or response to treatment is caused, regulated or influenced by genes. Genetic testing may therefore add value across the disease spectrum, ranging from single-gene disorders with a Mendelian inheritance pattern to complex multi-factorial diseases. The critical factors for genomic risk prediction are to determine: (1) where the genomic footprint of a particular susceptibility or dysfunction resides within this continuum, and (2) to what extent the genetic determinants are modified by environmental exposures. Regarding the small subset of highly penetrant monogenic disorders, a positive family history and early disease onset are mostly sufficient to determine the appropriateness of genetic testing in the index case and to inform pre-symptomatic diagnosis in at-risk family members. In more prevalent polygenic non-communicable diseases (NCDs), the use of appropriate eligibility criteria is required to ensure a balance between benefit and risk. An additional screening step may therefore be necessary to identify individuals most likely to benefit from genetic testing. This need provided the stimulus for the development of a pathology-supported genetic testing (PSGT) service as a new model for the translational implementation of genomic medicine in clinical practice. PSGT is linked to the establishment of a research database proven to be an invaluable resource for the validation of novel and previously described gene-disease associations replicated in the South African population for a broad range of NCDs associated with increased cardio-metabolic risk. The clinical importance of inquiry concerning family history in determining eligibility for personalized genotyping was supported beyond its current limited role in diagnosing or screening for monogenic subtypes of NCDs. With the recent introduction of advanced microarray-based breast cancer subtyping, genetic testing has extended beyond the genome of the host to also include tumor gene expression profiling for chemotherapy selection. The decreasing cost of next generation sequencing over recent years, together with improvement of both laboratory and computational protocols, enables the mapping of rare genetic disorders and discovery of shared genetic risk factors as novel therapeutic targets across diagnostic boundaries. This article reviews the challenges, successes, increasing inter-disciplinary integration and evolving strategies for extending PSGT towards exome and whole genome sequencing (WGS) within a dynamic framework. Specific points of overlap are highlighted between the application of PSGT and exome or WGS, as the next logical step in genetically uncharacterized patients for whom a particular disease pattern and/or therapeutic failure are not adequately accounted for during the PSGT pre-screen. Discrepancies between different next generation sequencing platforms and low concordance among variant-calling pipelines caution against offering exome or WGS as a stand-alone diagnostic approach. The public reference human genome sequence (hg19) contains minor alleles at more than 1 million loci and variant calling using an advanced major allele reference genome sequence is crucial to ensure data integrity. Understanding that genomic risk prediction is not deterministic but rather probabilistic provides the opportunity for disease prevention and targeted treatment in a way that is unique to each individual patient.

The fat mass and obesity-associated FTO rs9939609 polymorphism is associated with elevated homocysteine levels in patients with multiple sclerosis screened for vascular risk factors.

UNLABELLED: The previously reported link between homocysteine and obesity, both identified as established risk factors for multiple sclerosis (MS), has not previously been studied in relation to the fat mass and obesity-associated (FTO) gene. AIM: To investigate the mechanism underlying homocysteine accumulation in MS patients. A total of 114 patients and 195 population-matched controls were analysed for the FTO rs9939609 polymorphism. Homocysteine levels were measured in a subgroup of 60 patients and 87 controls screened for multiple vascular risk factors. After adjustment for potential confounders, the risk-associated FTO rs9939609 A-allele was associated with raised homocysteine levels (p = 0.003) in patients diagnosed with MS, but not in controls. Homocysteine levels correlated positively with body mass index (BMI) (p = 0.045) and total cholesterol levels (p = 0.048). Both homocysteine (p = 0.011) and BMI (p = 0.017) were significantly reduced with higher intake of folate in the diet. Higher BMI also correlated with increased intake of saturated/trans fat (p < 0.01) and low physical activity (p < 0.006). Daily intake of at least five fruits and vegetables had a favourable lowering effect on the Expanded Disability Status Scale (EDSS) (p = 0.035), while smoking increased MS disability (p < 0.001). This study has shown for the first time that having a diagnosis of MS moderates the effect of the FTO rs9939609 polymorphism on homocysteine levels. This is consistent with the role of FTO in demethylation and epigenetic changes. Identification of FTO rs9939609 reinforces the importance of adequate fruit, vegetable and folate and restriction of saturated/trans fat intake in the diet.

Disability in multiple sclerosis is associated with vascular factors: An ultrasound study.

BACKGROUND: Although multiple sclerosis (MS) is an immune-related disorder, pharmaceutical interventions targeting the immune system do not stop or reverse disability progression; the major challenge for this condition. Studies show that disability progression in MS is associated with vascular comorbidity and brain volume loss, indicating that a multi-targeted approach is required to prevent debilitation. The aim of the present study was to examine the associations between vascular ultrasound, disability, biochemistry and lifestyle data in people with MS (pwMS). METHODS: Extracranial vascular ultrasound was performed on 51 pwMS and 25 age-matched controls. Sonographic interrogation determined carotid intima-media thickness (cIMT) and abnormal blood flow patterns. Disability was assessed using the Expanded Disability Status Scale (EDSS). Biochemical and lifestyle data were obtained for all participants. RESULTS: The EDSS had a highly significant positive association with the cIMT of the right (r = 0.63; p = 0.001) and left (r = 0.49; p = 0.001) common carotid arteries and negative associations with the peak systolic blood flow velocity of the right vertebral artery (r = -0.42; p = 0.01) as well as end-diastolic velocity of the left internal carotid artery (r = -0.47; p = 0.01). These associations were significantly influenced by biochemical and lifestyle factors. Both cIMT and age showed significant associations with the EDSS. When cIMT was adjusted for age in a regression analysis, the association between the EDSS and the cIMT remained significant (p < 0.01), while the age association was reduced to being significant only at 10% (p = 0.06). There was no association between the use of MS medication and the EDSS (p = 0.56). CONCLUSION: PwMS who had increased cIMT, a surrogate marker for atherosclerosis, and reduced carotid artery blood flow velocities were at risk for greater disability over and above the effect of aging. These findings provide important information for disease management and disability prevention in pwMS. Modification of diet and lifestyle may promote the unhindered flow of essential nutritional factors into the brain in pwMS.

Pathology-supported genetic testing presents opportunities for improved disability outcomes in multiple sclerosis.

Background: Lipid metabolism may impact disability in people with multiple sclerosis (pwMS). Methods: Fifty-one pwMS entered an ultrasound and MRI study, of whom 19 had followed a pathology-supported genetic testing program for more than 10 years (pwMS-ON). Genetic variation, blood biochemistry, vascular blood flow velocities, diet and exercise were investigated. Results: pwMS-ON had significantly lower (p < 0.01) disability (Expanded Disability Status Scale) than pwMS not on the program (1.91 ± 0.75 vs 3.87 ± 2.32). A genetic variant in the lipid transporter FABP2 gene (rs1799883; 2445G>A, A54T) was significantly associated (p < 0.01) with disability in pwMS not on the program, but not in pwMS-ON (p = 0.88). Vascular blood flow velocities were lower in the presence of the A-allele. Conclusion: Pathology-supported genetic testing may provide guidance for lifestyle interventions with a significant impact on improved disability in pwMS.

This study investigated the role of a genetic variant that increases saturated fat absorption and may make people with multiple sclerosis (MS) more susceptible to disability progression. Of 51 people with MS, 19 had followed a program which includes normalization of blood test results and daily intake of unsaturated fatty acids for more than 10 years, while the others had not. The latter group had significantly greater disability than the people who had followed the program, suggesting that the unsaturated fatty acids modulated the effect of the genetic variant. Six MS cases are presented as examples, including a marathon athlete (Case 1) and a patient who showed a dramatic decrease in disability from being wheelchair-bound for 15 years to walking freely (Case 2).

Detecting the most critical clinical variables of COVID-19 breakthrough infection in vaccinated persons using machine learning.

Background: COVID-19 vaccines offer different levels of immune protection but do not provide 100% protection. Vaccinated persons with pre-existing comorbidities may be at an increased risk of SARS-CoV-2 breakthrough infection or reinfection. The aim of this study is to identify the critical variables associated with a higher probability of SARS-CoV-2 breakthrough infection using machine learning. Methods: A dataset comprising symptoms and feedback from 257 persons, of whom 203 were vaccinated and 54 unvaccinated, was used for the investigation. Three machine learning algorithms - Deep Multilayer Perceptron (Deep MLP), XGBoost, and Logistic Regression - were trained with the original (imbalanced) dataset and the balanced dataset created by using the Random Oversampling Technique (ROT), and the Synthetic Minority Oversampling Technique (SMOTE). We compared the performance of the classification algorithms when the features highly correlated with breakthrough infection were used and when all features in the dataset were used. Result: The results show that when highly correlated features were considered as predictors, with Random Oversampling to address data imbalance, the XGBoost classifier has the best performance (F1 = 0.96; accuracy = 0.96; AUC = 0.98; G-Mean = 0.98; MCC = 0.88). The Deep MLP had the second best performance (F1 = 0.94; accuracy = 0.94; AUC = 0.92; G-Mean = 0.70; MCC = 0.42), while Logistic Regression had less accurate performance (F1 = 0.89; accuracy = 0.88; AUC = 0.89; G-Mean = 0.89; MCC = 0.68). We also used Shapley Additive Explanations (SHAP) to investigate the interpretability of the models. We found that body temperature, total cholesterol, glucose level, blood pressure, waist circumference, body weight, body mass index (BMI), haemoglobin level, and physical activity per week are the most critical variables indicating a higher risk of breakthrough infection. Conclusion: These results, evident from our unique data source derived from apparently healthy volunteers with cardiovascular risk factors, follow the expected pattern of positive or negative correlations previously reported in the literature. This information strengthens the body of knowledge currently applied in public health guidelines and may also be used by medical practitioners in the future to reduce the risk of SARS-CoV-2 breakthrough infection.

Pathology supported genetic testing and treatment of cardiovascular disease in middle age for prevention of Alzheimer's disease.

Chronic, multi-factorial conditions caused by a complex interaction between genetic and environmental risk factors frequently share common disease mechanisms, as evidenced by an overlap between genetic risk factors for cardiovascular disease (CVD) and Alzheimer's disease (AD). Single nucleotide polymorphisms (SNPs) in several genes including ApoE, MTHFR, HFE and FTO are known to increase the risk of both conditions. The E4 allele of the ApoE polymorphism is the most extensively studied risk factor for AD and increases the risk of coronary heart disease by approximately 40%. It furthermore displays differential therapeutic responses with use of cholesterol-lowering statins and acetylcholinesterase inhibitors, which may also be due to variation in the CYP2D6 gene in some patients. Disease expression may be triggered by gene-environment interaction causing conversion of minor metabolic abnormalities into major brain disease due to cumulative risk. A growing body of evidence supports the assessment and treatment of CVD risk factors in midlife as a preventable cause of cognitive decline, morbidity and mortality in old age. In this review, the concept of pathology supported genetic testing (PSGT) for CVD is described in this context. PSGT combines DNA testing with biochemical measurements to determine gene expression and to monitor response to treatment. The aim is to diagnose treatable disease subtypes of complex disorders, facilitate prevention of cumulative risk and formulate intervention strategies guided from the genetic background. CVD provides a model to address the lifestyle link in most chronic diseases with a genetic component. Similar preventative measures would apply for optimisation of heart and brain health.

The conundrum of iron in multiple sclerosis--time for an individualised approach.

Although the involvement of immune mechanisms in multiple sclerosis (MS) is undisputed, some argue that there is insufficient evidence to support the hypothesis that MS is an autoimmune disease, and that the difference between immune- and autoimmune disease mechanisms has yet to be clearly delineated. Uncertainties surrounding MS disease pathogenesis and the modest efficacy of currently used disease modifying treatments (DMTs) in the prevention of disability, warrant the need to explore other possibilities. It is evident from the literature that people diagnosed with MS differ widely in symptoms and clinical outcome--some patients have a benign disease course over many years without requiring any DMTs. Attempting to include all patients into a single entity is an oversimplification and may obscure important observations with therapeutic consequences. In this review we advocate an individualised approach named Pathology Supported Genetic Testing (PSGT), in which genetic tests are combined with biochemical measurements in order to identify subgroups of patients requiring different treatments. Iron dysregulation in MS is used as an example of how this approach may benefit patients. The theory that iron deposition in the brain contributes to MS pathogenesis has caused uncertainty among patients as to whether they should avoid iron. However, the fact that a subgroup of people diagnosed with MS show clinical improvement when they are on iron supplementation emphasises the importance of individualised therapy, based on genetic and biochemical determinations.

Assessment of Epstein-Barr virus in blood from patients with multiple sclerosis.

Viruses such as Epstein-Barr virus (EBV) which can establish latent infections in the central nervous system or the immune system have been associated with chronic neurological disorders, including multiple sclerosis. Results vary, therefore the aim of this study was to investigate the presence of EBV using both viral DNA and antibody screening techniques, using PCR and ELISA assays respectively, to evaluate viral presence in blood from control subjects and patients with multiple sclerosis. Viral gene sequences for latent proteins EBNA-1 and LMP-1 and lytic gene BamH1-W were present equally in both patients and controls (<7%). Anti-EBV-VCA IgG positive cases were present in >99% of all study subjects, and anti-EBV-VCA IgG immune status ratio showed a near-significant positive correlation with the EDSS in patients with multiple sclerosis. In contrast, Anti-EBV-VCA IgM positive cases were significantly increased in patients (controls: 23.3%; patients; 41.9%; P = 0.046). The IgM to IgG immune status ratio was near-significantly higher in patients with relapse episodes in the year preceding blood sampling (P = 0.058). Results from this and previous studies have shown higher prevalence rates for EBV evaluating anti-EBV IgM positive cases against viral DNA positive cases. Also, IgM, an innate immune response, showed an association with relapse episodes, suggesting viral re-activation as a contributing factor to these relapses.

Red blood cell membrane fluidity in the etiology of multiple sclerosis.

Organisms adjust the order, or fluidity, of their cellular membranes in response to changes in their physiochemical environment by adjusting the lipid composition of their membranes. We investigated membrane fluidity using the phospholipid, fatty acid and cholesterol content of red blood cells (RBCs) from multiple sclerosis (MS) patients and correlated this with C-reactive protein (CRP) as well as with the severity of neurological outcome as measured by the Kurtzke Expanded Disability Status Scale (EDSS) and its Functional System Scores. The study group consisted of 31 patients with MS and 30 healthy control subjects. Phospholipids were determined using a colorimetric assay, fatty acids by gas chromatography, cholesterol by an enzymatic assay and CRP by a Beckman nephelometer. Cell membrane fluidity was calculated according to previously established formulae. RBC membrane fluidity as measured by the saturated to polyunsaturated fatty acid ratio was higher in patients than in controls (P = 0.04). The phosphatidylethanolamine saturated to polyunsaturated fatty acid ratio showed highly significant positive correlations with the EDSS and CRP < 5 microg/ml. CRP showed significant inverse correlations with the saturated nature but positive correlations with the ordered-crystalline-phase to liquid-crystalline-phase lipid ratio. In this study we show that membrane fluidity as measured by the relationship between membrane fatty acids, phospholipids and cholesterol is closely interrelated with inflammation and disease outcome in patients with MS. In conclusion, our findings suggest that the membrane lipid composition of patients with MS and, consequently, membrane fluidity are altered, which seems to be influenced by the inflammatory status.

Changes in erythrocyte membrane fatty acids during a clinical trial of eicosapentaenoic acid (EPA) supplementation in schizophrenia.

In a previously reported double-blind, placebo-controlled trial of eicosapentaenoic acid (EPA) as supplemental treatment in 40 patients with schizophrenia, we found significant improvement in symptoms as measured by the Positive and Negative Syndrome Scale (PANSS) compared to placebo (Emsley et al. 2002). Here we report changes in fatty acid composition of erythrocyte membranes in the same sample (n = 16 in each group). After 12 weeks of receiving EPA, levels of several saturated and mono-unsaturated fatty acids decreased significantly while levels of n-3 fatty acids increased significantly compared to the placebo group. Increases of n-3 and n-6 fatty acids in the erythrocyte membranes were greater in subjects who improved more than 20% on overall symptoms. Changes in fatty acids correlated significantly with improvement in PANSS sub-scale scores, more so in females than in males. Docosahexaenoic acid (DHA) (22:6n-3) levels increased less than expected, suggesting a possible defect in synthesis or incorporation of DHA into membranes in schizophrenia. Improvement in dyskinesia correlated significantly with an increase in alpha-linolenic acid (18:3n-3; p = 0.03), and a decrease in 20:1n-9 (p = 0.005).

Identification of an iron-responsive subtype in two children diagnosed with relapsing-remitting multiple sclerosis using whole exome sequencing.

BACKGROUND: Multiple sclerosis is a disorder related to demyelination of axons. Iron is an essential cofactor in myelin synthesis. Previously, we described two children (males of mixed ancestry) with relapsing-remitting multiple sclerosis (RRMS) where long-term remission was achieved by regular iron supplementation. A genetic defect in iron metabolism was postulated, suggesting that more advanced genetic studies could shed new light on disease pathophysiology related to iron. METHODS: Whole exome sequencing (WES) was performed to identify causal pathways. Blood tests were performed over a 10 year period to monitor the long-term effect of a supplementation regimen. Clinical wellbeing was assessed quarterly by a pediatric neurologist and regular feedback was obtained from the schoolteachers. RESULTS: WES revealed gene variants involved in iron absorption and transport, in the transmembrane protease, serine 6 (TMPRSS6) and transferrin (TF) genes; multiple genetic variants in CUBN, which encodes cubilin (a receptor involved in the absorption of vitamin B12 as well as the reabsorption of transferrin-bound iron and vitamin D in the kidneys); SLC25A37 (involved in iron transport into mitochondria) and CD163 (a scavenger receptor involved in hemorrhage resolution). Variants were also found in COQ3, involved with synthesis of Coenzyme Q10 in mitochondria. Neither of the children had the HLA-DRB1*1501 allele associated with increased genetic risk for MS, suggesting that the genetic contribution of iron-related genetic variants may be instrumental in childhood MS. In both children the RRMS has remained stable without activity over the last 10 years since initiation of nutritional supplementation and maintenance of normal iron levels, confirming the role of iron deficiency in disease pathogenesis in these patients. CONCLUSION: Our findings highlight the potential value of WES to identify heritable risk factors that could affect the reabsorption of transferrin-bound iron in the kidneys causing sustained iron loss, together with inhibition of vitamin B12 absorption and vitamin D reabsorption (CUBN) and iron transport into mitochondria (SLC25A37) as the sole site of heme synthesis. This supports a model for RRMS in children with an apparent iron-deficient biochemical subtype of MS, with oligodendrocyte cell death and impaired myelination possibly caused by deficits of energy- and antioxidant capacity in mitochondria.

Maternal separation of rat pups increases the risk of developing depressive-like behavior after subsequent chronic stress by altering corticosterone and neurotrophin levels in the hippocampus.

Children that are abused have an increased risk for developing psychiatric disorders later in life, because of the negative effects of stress on the developing brain. We used a maternal separation model in rats to see how neurotrophins, stress hormones, behavior and the anti-oxidant potential of serum are affected. Rat pups were separated from their mothers for 3h/day on days 2-14. Maternal separation caused changes in levels of NGF and NT-3 in the dorsal and ventral hippocampus, increased basal corticosterone levels and decreased ACTH levels after acute restraint stress. The anti-oxidant potential of the rat serum was significantly lower in the maternal separation group. Depressive-like behavior, measured during a forced swim test, was seen in maternally separated rats after additional chronic stress during adulthood. Maternal separation caused downregulation of neurotrophins in the ventral hippocampus, possibly as an effect of high corticosterone levels, but compensatory mechanisms against cell death may be involved as neurotrophin levels increased in the dorsal hippocampus. Decreased anti-oxidant potential of serum could have been an effect of downregulated neurotrophin levels.

Safety of the omega-3 fatty acid, eicosapentaenoic acid (EPA) in psychiatric patients: results from a randomized, placebo-controlled trial.

Omega-3 fatty acids, particularly eicosapentaenoic acid (EPA), are increasingly being used by psychiatric patients. Most studies have concentrated on efficacy aspects, while little is known about their safety and tolerability in psychiatric populations. This study aimed to assess the effects of EPA treatment on body mass, glucose metabolism, lipid profiles, prolactin secretion, bleeding time, haematology and liver functions. Eighty-four subjects with schizophrenia were treated with either EPA 2 g/day or placebo in addition to their antipsychotic medication for 12 weeks, in a randomized, controlled trial. Forty-seven entered a 40-week open-label extension phase of EPA 2 g/day. Seventy-four patients were included in the analysis. Six patients discontinued from the EPA group and 14 in the placebo group. Adverse event reporting was similar for the two groups. While there were no significant between-group differences, in the blinded phase the EPA group showed a significant increase in body mass index (BMI) and bleeding time. In the open-label extension, there was again a modest increase in BMI. Total cholesterol and HDL levels were significantly decreased. EPA 2 g/day is generally well tolerated. Clinicians should be aware of possible increases in bleeding time, as well as changes in weight and lipid metabolism.