Functional characterization of BRCA1 gene variants by mini-gene splicing assay

Mutational screening of the breast cancer susceptibility gene BRCA1 leads to the identification of numerous pathogenic variants such as frameshift and nonsense variants, as well as large genomic rearrangements. The screening moreover identifies a large number of variants, for example, missense, silent, and intron variants, which are classified as variants of unknown clinical significance owing to the lack of causal evidence. Variants of unknown clinical significance can potentially have an impact on splicing and therefore functional examinations are warranted to classify whether these variants are pathogenic or benign. Here we validate a mini-gene splicing assay by comparing the results of 24 variants with previously published data from RT-PCR analysis on RNA from blood samples/lymphoblastoid cell lines. The analysis showed an overall concordance of 100%. In addition, we investigated 13 BRCA1 variants of unknown clinical significance or putative variants affecting splicing by in silico analysis and mini-gene splicing assay. Both the in silico analysis and mini-gene splicing assay classified six BRCA1 variants as pathogenic (c.80+1G>A, c.132C>T (p.=), c.213−1G>A, c.670+1delG, c.4185+1G>A, and c.5075−1G>C), whereas six BRCA1 variants were classified as neutral (c.-19-22_-19-21dupAT, c.302−15C>G, c.547+14delG, c.4676−20A>G, c.4987−21G>T, and c.5278−14C>G) and one BRCA1 variant remained unclassified (c.670+16G>A). In conclusion, our study emphasizes that in silico analysis and mini-gene splicing assays are important for the classification of variants, especially if no RNA is available from the patient. This knowledge is crucial for proper genetic counseling of patients and their family members.     

Exposure to second-hand smoke and reproductive outcomes depending on maternal asthma

Tobacco consumption and exposure to second-hand smoke (SHS) are associated with reduced birth weight. One issue that has not been clarified previously is that of the potential higher risk of this outcome in mothers with asthma. We assessed the role of prenatal maternal tobacco use and SHS on reproductive outcomes and assessed the interaction with maternal history of asthma. Data was collected from the INMA study, a maternal birth cohort selected from the general population established in Spain in 2002. We measured cotinine at the 32nd week of pregnancy in 2,219 females. Diagnosed maternal asthma was self-reported during pregnancy. 35% of mothers reported not being exposed to smoking or SHS during pregnancy. Active smoking (i.e. self-reported or cotinine >50 ng·mL(-1)) was related to a 134 g decrease in birth weight and a relative risk of 1.8 for small for gestational age and fetal growth restriction. These results were not modified by maternal asthma. Maternal asthma had a similar frequency in all exposure groups. Non SHS-exposed females had the lowest prevalence of asthma. SHS (i.e. cotinine 20-50 ng·mL(-1)) decreased birth weight by 32 g among those without maternal asthma, but these differences were not statistically significant (95% CI -88.76-24.76). Maternal asthma did not promote these effects. Maternal history of asthma did not modify the effects of smoking on reproductive outcomes in a cohort sampled from the general population.     

Inverse Association Between Birth Weight,Birth Length and Serum Total Cholesterol in Adulthood

Objectives - To investigate whether impaired fetal growth, measured by low birth weight and short birth length, is linked with raised levels of serum lipids and increased risk and mortality of coronary heart disease. Design - The association between birth length, birth weight, Ponderal Index and total serum cholesterol was examined in 545 Danish men and women aged 31 to 51 years who participated in the Ebeltoft Health Promotion Project in Denmark. Results - No associations were found in women. For men, a negative association was found between birth weight and serum total cholesterol, with a fall in mean serum total cholesterol from 6.03 mmol/l at birth weight below 3300 g to 5.64 mmol/l at birth weight above 4000. A similar association was found between birth length and serum cholesterol, with a mean value of 6.23 mmol/l at birth length below 51 cm and a mean value of 5.56 mmol/l at birth length above 54 cm. No associations were found for Ponderal Index. Between 3% and 8% of the variance in serum total cholesterol could be explained by the statistical models used in this study. Conclusion - Our findings support the hypothesis of a negative association between birth weight, birth length and elevated serum cholesterol in adult life, but only in men.     

Exercise training in older patients with systolic heart failure: Adherence,exercise capacity,inflammation and glycemic control

Objectives. Training improves exercise capacity in patients with heart failure (CHF) but most evidence is on selected younger patients with systolic CHF. Design. All patients diagnosed with CHF over 3 years were screened for inclusion and exclusion criteria. Fifty two patients with systolic CHF (LVEF<45, NYHA II-III) received supervised exercise training twice weekly for 8 weeks. Results. Mean age was 68.2 (+/?SD 11.3) years. Despite marked improvements in physical fitness (workload, 6 minute walk test, incremental shuttle walk test and sit to stand test), there were no changes in serological markers of glycemic control (glucose, insulin, glycerol, free fatty acids, HbA1c), inflammation and endothelial function (hsCRP, orosomucoid, interleukin 6, TNF-alpha, urine-orosomucoid and -albumin/creatinin), lipid metabolism, NT-proBNP or other regulatory hormones (cortisol, epinephrine and IGF-1). There were no changes in quality of life. Conclusions. The effect of exercise training in these older CHF-patients was not as impressive as reported in younger and more selected patients. More studies on the efficiency of exercise training that reflect the age- and co-morbidity of the majority of CHF-patients are needed.     

CSF α-synuclein improves diagnostic and prognostic performance of CSF tau and Aβ in Alzheimer’s disease

Alzheimer’s disease (AD) and Lewy body diseases (LBD), e.g., Parkinson’s disease (PD) dementia and dementia with Lewy bodies (DLB), are common causes of geriatric cognitive impairments. In addition, AD and LBD are often found in the same patients at autopsy; therefore, biomarkers that can detect the presence of both pathologies in living subjects are needed. In this investigation, we report the assessment of α-synuclein (α-syn) in cerebrospinal fluid (CSF) and its association with CSF total tau (t-tau), phosphorylated tau₁₈₁ (p-tau₁₈₁), and amyloid beta_1-42 (Aβ_1-42) in subjects of the Alzheimer’s Disease Neuroimaging Initiative (ADNI; n = 389), with longitudinal clinical assessments. A strong correlation was noted between α-syn and t-tau in controls, as well as in patients with AD and mild cognitive impairment (MCI). However, the correlation is not specific to subjects in the ADNI cohort, as it was also seen in PD patients and controls enrolled in the Parkinson’s Progression Markers Initiative (PPMI; n = 102). A bimodal distribution of CSF α-syn levels was observed in the ADNI cohort, with high levels of α-syn in the subjects with abnormally increased t-tau values. Although a correlation was also noted between α-syn and p-tau₁₈₁, there was a mismatch (α-syn–p-tau₁₈₁-Mis), i.e., higher p-tau₁₈₁ levels accompanied by lower α-syn levels in a subset of ADNI patients. We hypothesize that this α-syn–p-tau₁₈₁-Mis is a CSF signature of concomitant LBD pathology in AD patients. Hence, we suggest that inclusion of measures of CSF α-syn and calculation of α-syn–p-tau₁₈₁-Mis improves the diagnostic sensitivity/specificity of classic CSF AD biomarkers and better predicts longitudinal cognitive changes.     

Longitudinal associations of lifetime adiposity with leukocyte telomere length and mitochondrial DNA copy number

Adiposity may cause adverse health outcomes by increasing oxidative stress and systemic inflammation, which can be reflected by altered telomere length (TL) and mitochondrial DNA copy number (mtCN) in peripheral blood leukocytes. However, little is known about the influence of lifetime adiposity on TL and mtCN in later life. This study was performed to investigate the associations of lifetime adiposity with leukocyte TL and mtCN in 9613 participants from the Nurses’ Health Study. A group-based trajectory modelling approach was used to create trajectories of body shape from age 5 through 60 years, and a genetic risk score (GRS) was created based on 97 known adiposity susceptibility variants. Associations of body shape trajectories and GRS with dichotomized TL and mtCN were assessed by logistic regression models. After adjustment for lifestyle and dietary factors, compared with the lean-stable group, the lean-marked increase group had higher odds of having below-median TL (OR?=?1.18, 95% CI 1.04, 1.35; P?=?0.01), and the medium-marked increase group had higher odds of having below-median mtCN (OR?=?1.28, 95% CI 1.00, 1.64; P?=?0.047). There was a suggestive trend toward lower mtCN across the GRS quartiles (P for trend?=?0.07). In conclusion, telomere attrition may be accelerated by marked weight gain in middle life, whereas mtCN is likely to be reduced persistently by adiposity over the life course. The findings indicate the importance of lifetime weight management to preserve functional telomeres and mitochondria.     

Subclinical affective and cognitive fluctuations in Parkinson's disease: a randomized double-blind double-dummy study of Oral vs. Intrajejunal Levodopa

Journal of Neurology - Chronic levodopa treatment in Parkinson’s disease (PD) may promote undesirable motor and non-motor fluctuations. Compared to chronic oral levodopa treatment, continuous...