The Role of Serine-Glyoxylate Aminotransferase and Malyl-CoA Lyase in the Metabolism of Methylococcus capsulatus Bath.

The genome of aerobic methanotroph Methylococcus capsulatus Bath possesses genes of three biochemical pathways of C1-carbon assimilation: the ribulose monophosphate cycle, the Calvin-Benson-Bassham cycle, and the partial serine cycle. Numerous studies have demonstrated that during methanotrophic growth cells of Methylococcus capsulatus Bath express key enzymes of these routes. In this study, the role of the serine cycle key enzymes, serine-glyoxylate aminotransferase (Sga) and malyl-CoA lyase (Mcl) in metabolism of Methylococcus capsulatus Bath was investigated by gene inactivation. The Δsga mutant obtained by double homologous recombination showed a prolonged lag phase, and after the lag period, the growth rate became similar to that of the wild type strain. The elevated intracellular levels of glutamate, serine, glycine, alanine, methionine, leucine, and succinate suggested significant metabolic changes in the mutant cells. Deletion of the mcl gene resulted in very poor growth and glycine only partially improved growth of the mutant strain. Cells of Δmcl mutant possess lower content of histidine, but enhanced level of alanine, leucine, and lysine than those of the wild type strain. Our data imply the importance of the serine cycle enzymes in metabolism of the methanotroph as well as relationships of the three C1 assimilation pathways in the gammaproteobacterial methanotrophs.

SARS-CoV-2 herd immunity of the Kyrgyz population in 2021.

In the fight against coronavirus infection, control of the immune response is of decisive importance, an important component of which is the seroprevalence of antibodies to SARS-CoV-2. Immunity to SARS-CoV-2 is formed either naturally or artificially through vaccination. The purpose of this study was to assess the seroprevalence of antibodies to SARS-CoV-2 in the population of Kyrgyzstan. A cross-sectional randomized study of seroprevalence was carried out according to a program developed by Rospotrebnadzor and the St. Petersburg Pasteur Institute, taking into account WHO recommendations. The ethics committees of the Association of Preventive Medicine (Kyrgyzstan) and the St. Petersburg Pasteur Institute (Russia) approved the study. Volunteers (9471) were recruited, representing 0.15% (95% CI 0.14-0.15) of the total population, randomized by age and region. Plasma antibodies (Abs) to the nucleocapsid antigen (Nag) were determined. In vaccinated individuals, Abs to the SARS-CoV-2 receptor-binding domain antigen (RBDag) were determined. Differences were considered statistically significant at p < 0.05. The SARS-CoV-2 Nag Ab seroprevalence was 48.7% (95% CI 47.7-49.7), with a maximum in the 60-69 age group [59.2% (95% CI 56.6-61.7)] and a minimum in group 1-17 years old [32.7% (95 CI: 29.4-36.1)]. The highest proportion of seropositive individuals was in the Naryn region [53.3% (95% CI 49.8-56.8)]. The lowest share was in Osh City [38.1% (95% CI 32.6-43.9)]. The maximum SARS-CoV-2 Nag seropositivity was found in the health-care sector [57.1% (95% CI 55.4-58.8)]; the minimum was seen among artists [38.6% (95% CI 26.0-52.4)]. Asymptomatic SARS-CoV-2 Nag seropositivity was 77.1% (95% CI 75.6-78.5). Vaccination with Sputnik V or Sinopharm produced comparable Ab seroprevalence. SARS-CoV-2 Nag seropositivity in the Kyrgyz population was 48.75% (95% CI 47.7-49.7), with the mass vaccination campaign undoubtedly benefitting the overall situation.

Mitochondria-targeted mesoporous silica nanoparticles noncovalently modified with triphenylphosphonium cation: Physicochemical characteristics, cytotoxicity and intracellular uptake.

Novel nanocomposite system based on mesoporous silica nanoparticles (MSNs) noncovalently modified with hexadecyltriphenylphosphonium bromide (HTPPB) has been prepared, thoroughly characterized and used for encapsulation of model cargo Rhodamine B (RhB). The high encapsulation efficacy of this dye by HTPPB-modified mesoporous particles was demonstrated by spectrophotometry and thermography techniques. The bioavailability of MSN@HTPPB was testified. Cytotoxicity assay revealed that a marked suppression of M-HeLa cancer cells (epithelioid carcinoma of the cervix) occurs at concentration of 0.06 µg/mL, while the higher viability of Chang liver normal cell line was preserved in the concentration range of 0.98-0.06 µg/mL. Hemolysis assay demonstrated that only 2% of red blood cells are destructed at ~ 30 µg/mL concentration. This allows us to select the most harmless compositions based on MSN@HTPPB with minimal side effects toward normal cells and recommend them for the development of antitumor formulations. Fluorescence microscopy technique testified satisfactory penetration of HTPPB-modified carriers into M-HeLa cells. Importantly, modification of the MSN with HTPPB is shown to promote efficient delivery to mitochondria. To the best of our knowledge, it is one of the first successful examples of noncovalent surface modification of the MSNs with lipophilic phosphonium cation that improves targeted delivery of loads to mitochondria.

Phenotypic and Molecular Epidemiology of ESBL-, AmpC-, and Carbapenemase-Producing Escherichia coli in Northern and Eastern Europe.

Extended-spectrum beta-lactamases (ESBL) and AmpC producing-Escherichia coli have spread worldwide, but data about ESBL-producing-E. coli in the Northern and Eastern regions of Europe is scant. The aim of this study has been to describe the phenotypical and molecular epidemiology of different ESBL/AmpC/Carbapenemases genes in E. coli strains isolated from the Baltic States (Estonia, Latvia, and Lithuania), Norway and St. Petersburg (Russia), and to determine the predominant multilocus sequence type and single nucleotide polymorphisms diversity of E. coli isolates deduced by whole genome sequencing (WGS). A total of 10,780 clinical E. coli strains were screened for reduced sensitivity to third-generation cephalosporins. They were collected from 21 hospitals located in Estonia, Latvia, Lithuania, Norway and St. Petersburg during a 5 month period in 2012. The overall prevalence of ESBL/AmpC strains was 4.7% by phenotypical test and 3.9% by sequencing. We found more strains with the ESBL/AmpC phenotype and genotype in St. Petersburg and Latvia than other countries. Of phenotypic E. coli strains, 85% contained confirmed ESBL genes (including bla CTX-M, bla TEM- 29, bla TEM- 71), AmpC genes (bla CMY- 59, bla ACT- 12 / - 15 / - 20, bla ESC- 6, bla FEC- 1, bla DHA- 1), or carbapenemase genes (bla NDM- 1). bla CTX-M- 1, bla CTX-M - 14 and bla CTX-M- 15 were found in all countries, but bla CTX-M- 15 prevalence was higher in Latvia than in St. Petersburg (Russia), Estonia, Norway and Lithuania. The dominating AmpC genes were bla CMY- 59 in the Baltic States and Norway, and bla DHA- 1 in St. Petersburg. E. coli strains belonged to 83 different sequence types, of which the most prevalent was ST131 (40%). In conclusion, we generally found low ESBL/AmpC/Carbapenemase prevalence in E. coli strains isolated in Northern/Eastern Europe. However, several inter-country differences in distribution of particular genes and multilocus sequence types were found.

Application of Molecular Methods for Carbapenemase Detection.

This study has evaluated the correlation between different carbapenemases detection methods on carbapenem non-susceptible Klebsiella pneumoniae strains from Northern and Eastern Europe; 31 institutions in 9 countries participated in the research project, namely Finland, Estonia, Latvia, Lithuania, Russia, St. Petersburg, Poland, Belarus, Ukraine, and Georgia. During the research program, a total of 5,001 clinical K. pneumoniae isolates were screened for any carbapenem non-susceptibility by the disk diffusion method, Vitek 2 or Phoenix system following the EUCAST guideline on detection of resistance mechanisms, version 1.0. Strains isolated from outpatients and hospitalized patients from April 2015 to June 2015 were included. All types of samples (blood, pus, urine, etc.) excluding fecal screening or fecal colonization samples have been represented. In total, 171 carbapenemase screening-positive K. pneumoniae isolates (3.42%) were found and characterized. Several methods were used for detection of carbapenemases production, including Luminex assay (PCR and hybridization), whole genome sequencing, MALDI-TOF based Imipenem degradation assay, and immunochromatography testing. Minimal inhibitory concentration determination for Meropenem by agar-based gradient method was also used. Finally, 83 K. pneumoniae strains were carbapenemase negative by all confirmation methods (49.4% of all screening-positive ones), 74 - positive by three methods (44.0%), 8 - positive by two methods (4.8%) and 3 - positive by only one method (1.8%). The sensitivity of the tests was 96.3% for Whole genome sequencing and MALDI-TOF assay (both three undetected cases), and 95.1% for Luminex-Carba (4 undetected cases). The most commonly detected carbapenemases were NDM (n = 54) and OXA-48 (n = 26), followed by KPC-2, VIM-5, and OXA-72 (one case of each). Our results showed that different types of carbapenemases can be detected in the countries involved in the project. The sensitivity of our methods for carbapenemase detection (including screening as a first step and further confirmation tests) was >95%, but we would recommend using different methods to increase the sensitivity of detection and make it more precise.

Automated plaque analysis for the prognostication of major adverse cardiac events.

OBJECTIVE: The purpose of this study is to assess the value of an automated model-based plaque characterization tool for the prediction of major adverse cardiac events (MACE). METHODS: We retrospectively included 45 patients with suspected coronary artery disease of which 16 (33%) experienced MACE within 12 months. Commercially available plaque quantification software was used to automatically extract quantitative plaque morphology: lumen area, wall area, stenosis percentage, wall thickness, plaque burden, remodeling ratio, calcified area, lipid rich necrotic core (LRNC) area and matrix area. The measurements were performed at all cross sections, spaced at 0.5 mm, based on fully 3D segmentations of lumen, wall, and each tissue type. Discriminatory power of these markers and traditional risk factors for predicting MACE were assessed. RESULTS: Regression analysis using clinical risk factors only resulted in a prognostic accuracy of 63% with a corresponding area under the curve (AUC) of 0.587. Based on our plaque morphology analysis, minimal cap thickness, lesion length, LRNC volume, maximal wall area/thickness, the remodeling ratio, and the calcium volume were included into our prognostic model as parameters. The use of morphologic features alone resulted in an increased accuracy of 77% with an AUC of 0.94. Combining both clinical risk factors and morphological features in a multivariate logistic regression analysis increased the accuracy to 87% with a similar AUC of 0.924. CONCLUSION: An automated model based algorithm to evaluate CCTA-derived plaque features and quantify morphological features of atherosclerotic plaque increases the ability for MACE prognostication significantly compared to the use of clinical risk factors alone.

The genes and enzymes of sucrose metabolism in moderately thermophilic methanotroph Methylocaldum szegediense O12.

Four enzymes involved in sucrose metabolism: sucrose phosphate synthase (Sps), sucrose phosphate phosphatase (Spp), sucrose synthase (Sus) and fructokinase (FruK), were obtained as his-tagged proteins from the moderately thermophilic methanotroph Methylocaldum szegediense O12. Sps, Spp, FruK and Sus demonstrated biochemical properties similar to those of other bacterial counterparts, but the translated amino acid sequences of Sps and Spp displayed high divergence from the respective microbial enzymes. The Sus of M. szegediense O12 catalyzed the reversible reaction of sucrose cleavage in the presence of ADP or UDP and preferred ADP as a substrate, thus implying a connection between sucrose and glycogen metabolism. Sus-like genes were found only in a few methanotrophs, whereas amylosucrase was generally used in sucrose cleavage in this group of bacteria. Like other microbial fructokinases, FruK of M. szegediense O12 showed a high specificity to fructose.

Evaluating the Association between Enlarged Perivascular Spaces and Disease Worsening in Multiple Sclerosis.

BACKGROUND AND PURPOSE: Enlarged perivascular spaces (EPVSs) have been associated with relapses and brain atrophy in multiple sclerosis (MS). We investigated the association of EPVS with clinical and MRI features of disease worsening in a well-characterized cohort of relapsing-remitting MS patients prospectively followed for up to 10 years. METHODS: Baseline EPVSs were scored on 1.5T MRI in 30 converters to moderate-severe disability, and 30 nonconverters matched for baseline characteristics. RESULTS: EPVS scores were not significantly different between converters and nonconverters, nor associated with accrual of lesions or brain atrophy. CONCLUSIONS: Our preliminary findings from a relatively small study sample argue against a potential use of EPVS as early indicator of risk for disease worsening in relapsing-remitting MS patients in a clinical setting. Although the small sample size and clinical 1.5T MRI may have limited our ability to detect a significant effect, we provided estimates of the association of EPVS with clinical and MRI indicators of disease worsening in a well-characterized cohort of MS patients.

The effect of alcohol and red wine consumption on clinical and MRI outcomes in multiple sclerosis.

BACKGROUND: Alcohol and in particular red wine have both immunomodulatory and neuroprotective properties, and may exert an effect on the disease course of multiple sclerosis (MS). OBJECTIVE: To assess the association between alcohol and red wine consumption and MS course. METHODS: MS patients enrolled in the Comprehensive Longitudinal Investigation of Multiple Sclerosis at the Brigham and Women's Hospital (CLIMB) who completed a self-administered questionnaire about their past year drinking habits at a single time point were included in the study. Alcohol and red wine consumption were measured as servings/week. The primary outcome was the Expanded Disability Status Scale (EDSS) at the time of the questionnaire. Secondary clinical outcomes were the Multiple Sclerosis Severity Score (MSSS) and number of relapses in the year before the questionnaire. Secondary MRI outcomes included brain parenchymal fraction and T2 hyperintense lesion volume (T2LV). Appropriate regression models were used to test the association of alcohol and red wine intake on clinical and MRI outcomes. All analyses were controlled for sex, age, body mass index, disease phenotype (relapsing vs. progressive), the proportion of time on disease modifying therapy during the previous year, smoking exposure, and disease duration. In the models for the MRI outcomes, analyses were also adjusted for acquisition protocol. RESULTS: 923 patients (74% females, mean age 47 ± 11 years, mean disease duration 14 ± 9 years) were included in the analysis. Compared to abstainers, patients drinking more than 4 drinks per week had a higher likelihood of a lower EDSS score (OR, 0.41; p = 0.0001) and lower MSSS (mean difference, - 1.753; p = 0.002) at the time of the questionnaire. Similarly, patients drinking more than 3 glasses of red wine per week had greater odds of a lower EDSS (OR, 0.49; p = 0.0005) and lower MSSS (mean difference, - 0.705; p = 0.0007) compared to nondrinkers. However, a faster increase in T2LV was observed in patients consuming 1-3 glasses of red wine per week compared to nondrinkers. CONCLUSIONS: Higher total alcohol and red wine intake were associated with a lower cross-sectional level of neurologic disability in MS patients but increased T2LV accumulation. Further studies should explore a potential cause-effect neuroprotective relationship, as well as the underlying biological mechanisms.

Characterizing Clinical and MRI Dissociation in Patients with Multiple Sclerosis.

BACKGROUND AND PURPOSE: Two common approaches for measuring disease severity in multiple sclerosis (MS) are the clinical exam and brain magnetic resonance imaging (MRI) scan. Although most patients show similar disease severity on both measures, some patients have clinical/MRI dissociation. METHODS: Subjects from a comprehensive care MS center who had a concurrent brain MRI, spinal cord MRI, clinical examination, and patient reported outcomes were classified into three groups based on the Expanded Disability Status Scale (EDSS) and cerebral T2 hyperintense lesion volume (T2LV). The first group was the low lesion load/high disability group (LL/HD) with T2LV < 2 ml and EDSS ≥ 3. The second group was the high lesion load/low disability group (HL/LD) with T2LV > 6 ml and EDSS ≤ 1.5. All remaining subjects were classified as not dissociated. The three groups were compared using regression techniques for unadjusted analyses and to adjust for age, disease duration, and gender. RESULTS: Twenty-two subjects were classified as LL/HD (4.1%; 95% CI: 2.6%, 6.2%), and 50 subjects were classified as HL/LD (9.4%; 95% CI: 7.0%, 12.2%). Subjects in the LL/HD group were more likely to have a progressive form of MS and had significantly lower physical quality of life in adjusted and unadjusted analysis. Subjects in HL/LD had significantly more gadolinium-enhancing lesions, and subjects in the LL/HD group had significantly more cervical spinal cord lesions. CONCLUSIONS: Our results indicate that dissociation may occur between physical disability and cerebral lesion volume in either direction in patients with MS. Type of MS, brain atrophy, and spinal cord lesions may help to bridge this dissociation.

Global phylogeography and evolutionary history of Shigella dysenteriae type 1.

Together with plague, smallpox and typhus, epidemics of dysentery have been a major scourge of human populations for centuries(1). A previous genomic study concluded that Shigella dysenteriae type 1 (Sd1), the epidemic dysentery bacillus, emerged and spread worldwide after the First World War, with no clear pattern of transmission(2). This is not consistent with the massive cyclic dysentery epidemics reported in Europe during the eighteenth and nineteenth centuries(1,3,4) and the first isolation of Sd1 in Japan in 1897(5). Here, we report a whole-genome analysis of 331 Sd1 isolates from around the world, collected between 1915 and 2011, providing us with unprecedented insight into the historical spread of this pathogen. We show here that Sd1 has existed since at least the eighteenth century and that it swept the globe at the end of the nineteenth century, diversifying into distinct lineages associated with the First World War, Second World War and various conflicts or natural disasters across Africa, Asia and Central America. We also provide a unique historical perspective on the evolution of antibiotic resistance over a 100-year period, beginning decades before the antibiotic era, and identify a prevalent multiple antibiotic-resistant lineage in South Asia that was transmitted in several waves to Africa, where it caused severe outbreaks of disease.

Fatigue predicts disease worsening in relapsing-remitting multiple sclerosis patients.

BACKGROUND: It is unclear whether fatigue is a consequence or a predictive trait of disease worsening. OBJECTIVE: To investigate the predictive value of fatigue toward conversion to confirmed moderate-severe disability in patients with relapsing-remitting multiple sclerosis (RRMS). METHODS: We retrospectively selected from the Comprehensive Longitudinal Investigations in MS at the Brigham and Women's Hospital (CLIMB) study cohort RRMS patients who converted to confirmed (⩾2 years) Expanded Disability Status Scale (EDSS) score ⩾3 within a follow-up period ⩾3 years. We contrasted the Modified Fatigue Impact Scale (MFIS) score of 33 converters, obtained at least 1 year before conversion to EDSS ⩾3, with that of 33 non-converter RRMS patients matched for baseline characteristics. RESULTS: Total MFIS score was higher in converter versus non-converter MS patients (median 37 vs 13; p < 0.0001). EDSS and Center for Epidemiological Studies Depression scale (CES-D) scores were also higher in the converters (median EDSS 1.5 vs 0, p < 0.0001; median CES-D 30 vs 24, p < 0.0001) and were both associated with MFIS score (EDSS: rho = 0.42, p = 0.0005; CES-D: rho = 0.72, p < 0.0001). After adjusting for EDSS and CES-D in multivariate analysis, MFIS remained a significant predictor of subsequent conversion to confirmed EDSS ⩾3. CONCLUSION: Fatigue is a promising indicator of risk for conversion to confirmed moderate-severe disability in RRMS patients.

Handling changes in MRI acquisition parameters in modeling whole brain lesion volume and atrophy data in multiple sclerosis subjects: Comparison of linear mixed-effect models.

Magnetic resonance imaging (MRI) of the brain provides important outcome measures in the longitudinal evaluation of disease activity and progression in MS subjects. Two common measures derived from brain MRI scans are the brain parenchymal fraction (BPF) and T2 hyperintense lesion volume (T2LV), and these measures are routinely assessed longitudinally in clinical trials and observational studies. When measuring each outcome longitudinally, observed changes may be potentially confounded by variability in MRI acquisition parameters between scans. In order to accurately model longitudinal change, the acquisition parameters should thus be considered in statistical models. In this paper, several models for including protocol as well as individual MRI acquisition parameters in linear mixed models were compared using a large dataset of 3453 longitudinal MRI scans from 1341 subjects enrolled in the CLIMB study, and model fit indices were compared across the models. The model that best explained the variance in BPF data was a random intercept and random slope with protocol specific residual variance along with the following fixed-effects: baseline age, baseline disease duration, protocol and study time. The model that best explained the variance in T2LV was a random intercept and random slope along with the following fixed-effects: baseline age, baseline disease duration, protocol and study time. In light of these findings, future studies pertaining to BPF and T2LV outcomes should carefully account for the protocol factors within longitudinal models to ensure that the disease trajectory of MS subjects can be assessed more accurately.

Using multiple imputation to efficiently correct cerebral MRI whole brain lesion and atrophy data in patients with multiple sclerosis.

Automated segmentation of brain MRI scans into tissue classes is commonly used for the assessment of multiple sclerosis (MS). However, manual correction of the resulting brain tissue label maps by an expert reader remains necessary in many cases. Since automated segmentation data awaiting manual correction are "missing", we proposed to use multiple imputation (MI) to fill-in the missing manually-corrected MRI data for measures of normalized whole brain volume (brain parenchymal fraction-BPF) and T2 hyperintense lesion volume (T2LV). Automated and manually corrected MRI measures from 1300 patients enrolled in the Comprehensive Longitudinal Investigation of Multiple Sclerosis at the Brigham and Women's Hospital (CLIMB) were identified. Simulation studies were conducted to assess the performance of MI with missing data both missing completely at random and missing at random. An imputation model including the concurrent automated data as well as clinical and demographic variables explained a high proportion of the variance in the manually corrected BPF (R(2)=0.97) and T2LV (R(2)=0.89), demonstrating the potential to accurately impute the missing data. Further, our results demonstrate that MI allows for the accurate estimation of group differences with little to no bias and with similar precision compared to an analysis with no missing data. We believe that our findings provide important insights for efficient correction of automated MRI measures to obviate the need to perform manual correction on all cases.

Detection of carbapenemase-producing enterobacteriaceae in the baltic countries and st. Petersburg area.

The spread of carbapenemase-producing Enterobacteriaceae is a global problem; however, no exact data on the epidemiology of carbapenemase in the Baltic countries and St. Petersburg area is available. We aimed to evaluate the epidemiology of carbapenemase-producing Escherichia coli and Klebsiella pneumoniae in the Baltic States and St. Petersburg, Russia, and to compare the different methods for carbapenemase detection. From January to May 2012, all K. pneumoniae (n = 1983) and E. coli (n = 7774) clinical isolates from 20 institutions in Estonia, Latvia, Lithuania, and St. Petersburg, Russia were screened for carbapenem susceptibility. The IMP, VIM, GIM, NDM, KPC, and OXA-48 genes were detected using real-time PCR and the ability to hydrolyze ertapenem was determined using MALDI-TOF MS. Seventy-seven strains were found to be carbapenem nonsusceptible. From these, 15 K. pneumoniae strains hydrolyzed ertapenem and carried the bla NDM gene. All of these strains carried integron 1 and most carried integron 3 as well as genes of the CTX-M-1 group. No carbapenemase-producing E. coli or K. pneumoniae strains were found in Estonia, Latvia, or Lithuania; however, NDM-positive K. pneumoniae was present in the hospital in St. Petersburg, Russia. A MALDI-TOF MS-based assay is a suitable and cost-effective method for the initial confirmation of carbapenemase production.

Evaluation of circulating osteopontin levels in an unselected cohort of patients with multiple sclerosis: relevance for biomarker development.

BACKGROUND: Osteopontin (OPN) is a pleiotropic protein with important roles in inflammation and immunity that has been suggested as a candidate biomarker for disease activity in multiple sclerosis (MS). OBJECTIVE: We evaluated plasma levels of OPN in an unselected cohort of MS patients, to determine its potential as a biomarker for disease subtype and/or disease activity in a regular clinical setting. METHODS: We analyzed OPN plasma levels in 492 consecutive MS patients, using a commercial enzyme-linked immunosorbent assay (ELISA). RESULTS: OPN levels were higher in relapsing-remitting and secondary progressive MS, compared to healthy controls. Treatment with natalizumab or glatiramer acetate was associated with lower OPN levels. There was no significant association between the OPN levels and disease activity, as measured by clinical or radiological criteria. One-third of patients with high OPN levels had concurrent disorders that may also be associated with increased OPN expression, and which may mask a modest effect of MS disease activity on OPN levels. CONCLUSION: Our data do not support a role for circulating OPN levels as a biomarker for disease activity in a heterogeneous clinical setting, but does not rule out a potential role in the cerebrospinal fluid, in a controlled setting such as a clinical trial, or in concert with other biomarkers.

Circulating microRNAs as biomarkers for disease staging in multiple sclerosis.

OBJECTIVE: MicroRNAs (miRNAs) are single-stranded, small noncoding RNAs that regulate gene expression. Because they are stable in serum, they are being developed as biomarkers for cancer and other diseases. In multiple sclerosis (MS), miRNAs have been studied in cell populations but not in the circulation. In MS, a major challenge is to develop immune biomarkers to monitor disease. We asked whether circulating miRNAs could be identified in MS and whether they are linked to disease stage and/or disability. METHODS: A total of 368 miRNAs were measured in ethylenediaminetetraacetic acid plasma in 10 relapsing-remitting MS (RRMS) patients, 9 secondary progressive MS (SPMS) patients, and 9 healthy controls (HCs) using miRCURY LNA™ Universal RT microRNA polymerase chain reaction panels. Nineteen miRNAs from this discovery set were validated using qPCR on an independent set of 50 RRMS patients, 51 SPMS patients, and 32 HCs. RESULTS: We found that circulating miRNAs are differentially expressed in RRMS and SPMS versus HCs and in RRMS versus SPMS. We also found miRNAs to be linked to Expanded Disability Status Scale (EDSS). hsa-miR-92a-1* was identified in the largest number of comparisons. It was different in RRMS versus SPMS, and RRMS versus HCs, and showed an association with EDSS and disease duration. miR-92 has target genes involved in cell cycle regulation and cell signaling. The let-7 family of miRNAs differentiated SPMS from HCs and RRMS from SPMS. let-7 miRNAs regulate stem cell differentiation and T cell activation, activate Toll-like receptor 7, and are linked to neurodegeneration. hsa-miR-454 differentiated RRMS from SPMS, and hsa-miR-145 differentiated RRMS from HCs and RRMS from SPMS. Interestingly, the same circulating miRNAs (let-7 and miR-92) that were differentially expressed in RRMS versus SPMS also differentiated amyotrophic lateral sclerosis (ALS) from RRMS subjects, but were not different between SPMS and ALS, suggesting that similar processes may occur in SPMS and ALS. INTERPRETATION: Our results establish circulating miRNAs as a readily accessible blood biomarker to monitor disease in MS.

Neurovascular coupling is impaired in slow walkers: the MOBILIZE Boston Study.

OBJECTIVE: Neurovascular coupling may be involved in compensatory mechanisms responsible for preservation of gait speed in elderly people with cerebrovascular disease. Our study examines the association between neurovascular coupling in the middle cerebral artery and gait speed in elderly individuals with impaired cerebral vasoreactivity. METHODS: Twenty-two fast and 20 slow walkers in the lowest quartile of cerebral vasoreactivity were recruited from the MOBILIZE Boston Study. Neurovascular coupling was assessed in bilateral middle cerebral arteries by measuring cerebral blood flow during the N-Back task. Cerebral white matter hyperintensities were measured for each group using magnetic resonance imaging. RESULTS: Neurovascular coupling was attenuated in slow compared to fast walkers (2.8%; 95% confidence interval [CI], -0.9 to 6.6 vs 8.2%; 95% CI, 4.7-11.8; p = 0.02). The odds ratio of being a slow walker was 6.4 (95% CI, 1.7-24.9; p = 0.007) if there was a high burden of white matter hyperintensity; however, this risk increased to 14.5 (95% CI, 2.3-91.1; p = 0.004) if neurovascular coupling was also attenuated. INTERPRETATION: Our results suggest that intact neurovascular coupling may help preserve mobility in elderly people with cerebral microvascular disease.

A randomized controlled double-masked trial of albuterol add-on therapy in patients with multiple sclerosis.

BACKGROUND: Interleukin 12 (IL-12), a cytokine that promotes generation of helper T cells subtype 1, is increased in multiple sclerosis. Albuterol sulfate, a ß2-adrenergic agonist, reduces IL-12 expression, so we tested the effect of albuterol as an add-on treatment to glatiramer acetate therapy. OBJECTIVES: To investigate the clinical and immunologic effects of albuterol treatment as an add-on therapy in patients starting glatiramer acetate treatment. DESIGN: Single-center double-masked clinical trial. SETTING: Academic research. Patients Subjects with relapsing-remitting multiple sclerosis. MAIN OUTCOME MEASURES: In this single-center double-masked clinical trial, subjects with relapsing-remitting multiple sclerosis were randomized to receive a subcutaneous injection of glatiramer acetate (20 mg) plus an oral dose of placebo daily for 2 years or a subcutaneous injection of glatiramer acetate (20 mg) plus an oral dose of albuterol daily for 2 years. The primary clinical efficacy measurement was the change in Multiple Sclerosis Functional Composite at 2 years, and the primary immunologic end point was the change in expression of IL-13 and interferon γ at each study time point. The classification level of evidence from this trial is C for each question, as this is the first class II clinical trial addressing the efficacy of glatiramer acetate plus albuterol. RESULTS: Forty-four subjects were randomized to receive glatiramer acetate plus albuterol or glatiramer acetate plus placebo, and 39 subjects contributed to the analysis. Improvement in the Multiple Sclerosis Functional Composite was observed in the glatiramer acetate plus albuterol group at the 6-month (P = .005) and 12-month (P = .04) time points but not at the 24-month time point. A delay in the time to first relapse was also observed in the glatiramer acetate plus albuterol group (P = .03). Immunologically, IL-13 and interferon-γ production decreased in both treatment groups, and a treatment effect on IL-13 production was observed at the 12-month time point (P < .05). Adverse events were generally mild, and only 3 moderate or severe events were considered related to the treatment. CONCLUSION: Treatment with glatiramer acetate plus albuterol is well tolerated and improves clinical outcomes in patients with multiple sclerosis. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT00039988.