Alterations in the innate and adaptive immune system in a real-world cohort of multiple sclerosis patients treated with ocrelizumab.

B cell depletion by the anti-CD20 antibody ocrelizumab is effective in relapsing-remitting (RR) and primary progressive (PP) multiple sclerosis (MS). We investigated immunological changes in peripheral blood of a real-world MS cohort after 6 and 12 months of ocrelizumab. All RRMS and most PPMS patients (15/20) showed treatment response. Ocrelizumab not only reduced CD20+ B cells, but also numbers of CD20+ T cells. Absolute numbers of monocytes, dendritic cells and CD8+ T cells were increased, while CD56hi natural killer cells were reduced after ocrelizumab. The residual B cell population shifted towards transitional and activated, IgA+ switched memory B cells, double negative B cells, and antibody-secreting cells. Delaying the treatment interval by 2-3 months increased mean B cell frequencies and enhanced naive B cell repopulation. Ocrelizumab reduced plasma levels of interleukin(IL)-12p70 and interferon(IFN)-α2. These findings will contribute to understanding ineffective treatment responses, dealing with life-threatening infections and further unravelling MS pathogenesis.

Dimethyl fumarate induces a persistent change in the composition of the innate and adaptive immune system in multiple sclerosis patients.

The effects of dimethyl fumarate (DMF) on the immune system in multiple sclerosis (MS) are not completely elucidated. In this study, an extensive immunophenotypic analysis of innate and adaptive immune cells of DMF-treated MS patients was performed. Peripheral blood immune cell phenotypes were determined using flow cytometry in a follow-up study of 12 MS patients before, after 3 and 12 months of DMF treatment and a cross-sectional study of 25 untreated and 64 DMF-treated MS patients. Direct effects of DMF on B cells were analyzed in vitro. After 12 months of DMF treatment, percentages of monocytes, natural killer cells, naive T and B cells and transitional B cells increased. Percentages of (effector) memory T cells, (non) class-switched memory B cells and double negative B cells decreased together with CD4+ T cells expressing interferon-γ (IFN-γ), granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin-17 (IL-17). DMF treatment was fully effective as of 6 months and directly induced apoptosis and decreased expression of costimulatory CD40, antigen presentation molecule MHCII and B cell activating factor receptor (BAFFR) on B cells. DMF induced a persistent change of the immune system of MS patients, directly induced apoptosis and reduced expression of functional markers on B cells.

Differential effects of leptin on adiponectin expression with weight gain versus obesity.

BACKGROUND/OBJECTIVE: Adiponectin exerts beneficial effects by reducing inflammation and improving lipid metabolism and insulin sensitivity. Although the adiponectin level is lower in obese individuals, whether weight gain reduces adiponectin expression in humans is controversial. We sought to investigate the role of weight gain, and consequent changes in leptin, on altering adiponectin expression in humans. METHODS/RESULTS: Forty-four normal-weight healthy subjects were recruited (mean age 29 years; 14 women) and randomized to either gain 5% of body weight by 8 weeks of overfeeding (n=34) or maintain weight (n=10). Modest weight gain of 3.8±1.2 kg resulted in increased adiponectin level (P=0.03), whereas weight maintenance resulted in no changes in adiponectin. Further, changes in adiponectin correlated positively with changes in leptin (P=0.0085). In-vitro experiments using differentiated human white preadipocytes showed that leptin increased adiponectin mRNA and protein expression, whereas a leptin antagonist had opposite effects. To understand the role of leptin in established obesity, we compared adipose tissue samples obtained from normal-weight versus obese subjects. We noted, first, that leptin activated cellular signaling pathways and increased adiponectin mRNA in the adipose tissue from normal-weight participants, but did not do so in the adipose tissue from obese participants. Second, we noted that obese subjects had increased caveolin-1 expression, which attenuates leptin-dependent increases in adiponectin. CONCLUSIONS: Modest weight gain in healthy individuals is associated with increases in adiponectin levels, which correlate positively with changes in leptin. In vitro, leptin induces adiponectin expression, which is attenuated by increased caveolin-1 expression. In addition, the adipose tissue from obese subjects shows increased caveolin-1 expression and impaired leptin signaling. This leptin signal impairment may prevent concordant increases in adiponectin levels in obese subjects despite their high levels of leptin. Therefore, impaired leptin signaling may contribute to low adiponectin expression in obesity and may provide a target for increasing adiponectin expression, hence improving insulin sensitivity and cardio-metabolic profile in obesity.

Anti-SPAG16 antibodies in primary progressive multiple sclerosis are associated with an elevated progression index.

BACKGROUND AND PURPOSE: Sperm-associated antigen 16 (SPAG16), a sperm protein which is upregulated in reactive astrocytes in multiple sclerosis (MS) lesions, has recently been identified as a novel autoantibody target in MS. The aim of this study was to investigate whether anti-SPAG16 antibody levels differ between MS subtypes (relapsing-remitting, RR; primary or secondary progressive, PP, SP) and whether antibody positivity is associated with clinical characteristics. METHODS: Plasma anti-SPAG16 antibody levels were determined by recombinant protein enzyme-linked immunosorbent assay (ELISA) in 374 MS patients (274 RRMS, 39 SPMS and 61 PPMS) and 106 healthy controls. RESULTS: Significantly elevated anti-SPAG16 antibodies were found in 22% of MS patients with 93% specificity. Anti-SPAG16 seropositivity was associated with an increased Expanded Disability Status Scale (EDSS) in overall MS. A higher proportion of PPMS patients showed anti-SPAG16 antibody reactivity (34%) compared to RRMS (19%) and SPMS (26%), and presented with higher anti-SPAG16 antibody levels. Seropositive PPMS patients had a significantly increased progression index compared to seronegative patients. CONCLUSIONS: Anti-SPAG16 antibodies are associated with an increased EDSS in overall MS, indicating that they are linked to a worse MS disease outcome. Moreover, the presence of anti-SPAG16 antibodies may be a biomarker for a more severe disease in PPMS patients, as indicated by an increased progression index.

Diagnostic accuracy of body mass index to identify obesity in older adults: NHANES 1999-2004.

BACKGROUND: Body composition changes with aging lead to increased adiposity and decreased muscle mass, making the diagnosis of obesity challenging. Conventional anthropometry, including body mass index (BMI), while easy to use clinically may misrepresent adiposity. We determined the diagnostic accuracy of BMI using dual-energy X-ray absorptiometry (DEXA) in assessing the degree of obesity in older adults. METHODS: The National Health and Nutrition Examination Surveys 1999-2004 were used to identify adults aged ⩾60 years with DEXA measures. They were categorized (yes/no) as having elevated body fat by gender (men: ⩾25%; women ⩾35%) and by BMI ⩾25 and ⩾30 kg m(-)(2). The diagnostic performance of BMI was assessed. Metabolic characteristics were compared in discordant cases of BMI/body fat. Weighting and analyses were performed per NHANES (National Health and Nutrition Examination Survey) guidelines. RESULTS: We identified 4984 subjects (men: 2453; women: 2531). Mean BMI and % body fat was 28.0 kg m(-2) and 30.8% in men, and 28.5 kg m(-)(2) and 42.1% in women. A BMI ⩾30 kg m(-)(2) had a low sensitivity and moderately high specificity (men: 32.9 and 80.8%, concordance index 0.66; women: 38.5 and 78.5%, concordance 0.69) correctly classifying 41.0 and 45.1% of obese subjects. A BMI ⩾25 kg m(-2) had a moderately high sensitivity and specificity (men: 80.7 and 99.6%, concordance 0.81; women: 76.9 and 98.8%, concordance 0.84) correctly classifying 80.8 and 78.5% of obese subjects. In subjects with BMI <30 kg m(-)(2), body fat was considered elevated in 67.1% and 61.5% of men and women, respectively. For a BMI ⩾30 kg m(-)(2), sensitivity drops from 40.3% to 14.5% and 44.5% to 23.4%, whereas specificity remains elevated (>98%), in men and women, respectively, in those 60-69.9 years to subjects aged ⩾80 years. Correct classification of obesity using a cutoff of 30 kg m(-)(2) drops from 48.1 to 23.9% and 49.0 to 19.6%, in men and women in these two age groups. CONCLUSIONS: Traditional measures poorly identify obesity in the elderly. In older adults, BMI may be a suboptimal marker for adiposity.

Diagnostic performance of body mass index to identify obesity as defined by body adiposity in children and adolescents: a systematic review and meta-analysis.

BACKGROUND: The ideal means of identifying obesity in children and adolescents has not been determined although body mass index (BMI) is the most widely used screening tool. OBJECTIVE: We performed a systematic review and meta-analysis of studies assessing the diagnostic performance of BMI to detect adiposity in children up to 18 years. METHODS: Data sources were EMBASE, MEDLINE, Cochrane, Database of Systematic Reviews Cochrane CENTRAL, Web of Science and SCOPUS up to March 2013. Studies providing measures of diagnostic performance of BMI and using body composition technique for body fat percentage measurement were included. RESULTS: Thirty-seven eligible studies that evaluated 53 521 patients, with mean age ranging from 4 to 18 years were included in the meta-analysis. Commonly used BMI cut-offs for obesity showed pooled sensitivity to detect high adiposity of 0.73 (confidence interval [CI] 0.67-0.79), specificity of 0.93 (CI 0.88-0.96) and diagnostic odds ratio of 36.93 (CI 20.75-65.71). Males had lower sensitivity. Moderate heterogeneity was observed (I(2) = 48%) explained in meta-regression by differences across studies in race, BMI cut-off, BMI reference criteria (Center for Disease Control vs. International Obesity Task Force) and reference standard method assessing adiposity. CONCLUSION: BMI has high specificity but low sensitivity to detect excess adiposity and fails to identify over a quarter of children with excess body fat percentage.

Clinical characteristics of patients with resistant hypertension: the RESIST-POL study.

Recent studies indicate that resistant hypertension (RHTN) is present in about 12% of the treated hypertensive population. However, patients with true RHTN (confirmed out of the office) have not been widely studied. We prospectively studied 204 patients (123 male, 81 female, mean age 48.4 years, range 19-65 years) with truly RHTN (ambulatory daytime mean blood pressure >135/85 mm Hg). We evaluated the frequency of obstructive sleep apnea (OSA), renal artery stenosis (RAS), primary aldosteronism (PA) and other secondary forms of hypertension (HTN) and conditions. Mild, moderate and severe OSA were present in 55 (27.0%), 38 (18.6%) and 54 (26.5%) patients, respectively. Secondary forms of HTN were diagnosed in 49 patients (24.0%), the most frequent being PA (15.7%) and RAS (5.4%). Metabolic syndrome (MS) was present in 65.7% of patients. Excessive sodium excretion was evident in 33.3% of patients and depression in 36.8% patients. In patients with RHTN, OSA and MS were the most frequent conditions, frequently overlapping with each other and also with PA. Our data indicate that in the vast majority of patients with truly RHTN, at least one of three co-morbidities-OSA, MS and PA-is present. Other conditions, even though less frequent, should also be taken into the consideration.

A novel method for making human monoclonal antibodies.

We have developed a B cell immortalization method for low B cell numbers per well using simultaneous B cell stimulation by CpG2006 and B cell infection by Epstein-Barr virus (EBV), followed by an additional CpG2006 and interleukin-2 (IL-2) stimulus. Using this method, immunoglobulin G (IgG)-producing immortalized B cell lines were generated from peripheral blood IgG(+)CD22(+) B cells with an efficiency of up to 83%. Antibody can already be obtained from the culture supernatant after 3-4 weeks. Moreover, clonality analysis demonstrated monoclonality in 87% of the resulting immortalized B cell lines. Given the high immortalization efficiency and monoclonality rate, evidence is provided that no further subcloning is necessary. An important application of this B cell immortalization method is the characterization of (autoreactive) antibodies from patients with autoimmune disease. This could eventually lead to the identification of new autoantigens, disease markers or targets for therapy.

Diagnostic performance of body mass index to identify obesity as defined by body adiposity: a systematic review and meta-analysis.

OBJECTIVE: We performed a systematic review and meta-analysis of studies that assessed the performance of body mass index (BMI) to detect body adiposity. DESIGN: Data sources were MEDLINE, EMBASE, Cochrane, Database of Systematic Reviews, Cochrane CENTRAL, Web of Science, and SCOPUS. To be included, studies must have assessed the performance of BMI to measure body adiposity, provided standard values of diagnostic performance, and used a body composition technique as the reference standard for body fat percent (BF%) measurement. We obtained pooled summary statistics for sensitivity, specificity, positive and negative likelihood ratios (LRs), and diagnostic odds ratio (DOR). The inconsistency statistic (I2) assessed potential heterogeneity. RESULTS: The search strategy yielded 3341 potentially relevant abstracts, and 25 articles met our predefined inclusion criteria. These studies evaluated 32 different samples totaling 31 968 patients. Commonly used BMI cutoffs to diagnose obesity showed a pooled sensitivity to detect high adiposity of 0.50 (95% confidence interval (CI): 0.43-0.57) and a pooled specificity of 0.90 (CI: 0.86-0.94). Positive LR was 5.88 (CI: 4.24-8.15), I (2)=97.8%; the negative LR was 0.43 (CI: 0.37-0.50), I (2)=98.5%; and the DOR was 17.91 (CI: 12.56-25.53), I (2)=91.7%. Analysis of studies that used BMI cutoffs >or=30 had a pooled sensitivity of 0.42 (CI: 0.31-0.43) and a pooled specificity of 0.97 (CI: 0.96-0.97). Cutoff values and regional origin of the studies can only partially explain the heterogeneity seen in pooled DOR estimates. CONCLUSION: Commonly used BMI cutoff values to diagnose obesity have high specificity, but low sensitivity to identify adiposity, as they fail to identify half of the people with excess BF%.

Analysis of antibody reactivity in paired cerebrospinal fluid and serum of a relapsing remitting multiple sclerosis patient.

Increasing evidence indicates an involvement of B cells in multiple sclerosis (MS). However, little is known about antigenic targets recognized by antibodies present in blood and cerebrospinal fluid (CSF) of MS patients. This study was therefore aimed at identifying the antigen reactivity of antibodies present in CSF and compares the identified antibody profile with that of the serum of the same patient using cDNA phage display. Selection rounds on paired CSF and serum of this patient identified 13 antigenic targets of which 5 were enriched by serum antibodies and 2 were identified by CSF antibodies. Interestingly, the six remaining antigenic targets were shown to be recognized by both CSF and serum antibodies. These findings point towards both common as well as distinct antibody profiles in CSF and serum of MS patients.

B cell characterization and reactivity analysis in multiple sclerosis.

B cells are one of the key players in the pathogenesis of multiple sclerosis (MS). The peripheral B cell distributions are similar in healthy persons and MS patients. In healthy controls, B cells are rarely present in the cerebrospinal fluid (CSF) while in MS patients, a clonally expanded B cell population is detected. This consists of memory B cells, centroblasts and antibody-secreting plasma blasts and plasma cells that are responsible for intrathecal immunoglobulin G production and oligoclonal band formation in more than 90% of MS patients. Unfortunately, the targets of the autoreactive B cells and antibodies remain largely unknown. Various candidate antigens have been identified but often their involvement in the disease process is still unclear. Most studies characterizing these target antigens examined autoantibodies by analyzing sera or CSF of MS patients. An alternative approach is focusing on the clonally expanded B cells. In this way B cells directed against myelin, astroglia and axons have been denoted in MS patients. B cell immortalization, that is based on the antibody-producing potential of Epstein-Barr virus (EBV) transformed B cells, can be used to expand B cells from MS patients for the production of antibodies, that ultimately can be analysed for target identification.

Accuracy of body mass index in diagnosing obesity in the adult general population.

BACKGROUND: Body mass index (BMI) is the most widely used measure to diagnose obesity. However, the accuracy of BMI in detecting excess body adiposity in the adult general population is largely unknown. METHODS: A cross-sectional design of 13 601 subjects (age 20-79.9 years; 49% men) from the Third National Health and Nutrition Examination Survey. Bioelectrical impedance analysis was used to estimate body fat percent (BF%). We assessed the diagnostic performance of BMI using the World Health Organization reference standard for obesity of BF%>25% in men and>35% in women. We tested the correlation between BMI and both BF% and lean mass by sex and age groups adjusted for race. RESULTS: BMI-defined obesity (> or =30 kg m(-2)) was present in 19.1% of men and 24.7% of women, while BF%-defined obesity was present in 43.9% of men and 52.3% of women. A BMI> or =30 had a high specificity (men=95%, 95% confidence interval (CI), 94-96 and women=99%, 95% CI, 98-100), but a poor sensitivity (men=36%, 95% CI, 35-37 and women=49%, 95% CI, 48-50) to detect BF%-defined obesity. The diagnostic performance of BMI diminished as age increased. In men, BMI had a better correlation with lean mass than with BF%, while in women BMI correlated better with BF% than with lean mass. However, in the intermediate range of BMI (25-29.9 kg m(-2)), BMI failed to discriminate between BF% and lean mass in both sexes. CONCLUSIONS: The accuracy of BMI in diagnosing obesity is limited, particularly for individuals in the intermediate BMI ranges, in men and in the elderly. A BMI cutoff of> or =30 kg m(-2) has good specificity but misses more than half of people with excess fat. These results may help to explain the unexpected better survival in overweight/mild obese patients.

Dynamic QT/RR coupling in patients with pacemakers.

The dynamic coupling between heart rate intervals (RR) and ventricular repolarization (QT) is analyzed. The analysis is based on measurements of 11 patients with pacemaker. In each measurement, there are at least 4 abrupt changes of RR preset by the pacemaker. With such a protocol, RR changes are important and well defined while disturbing factors and noise sources (such as those related with motion of patient) are minimized. The QT/RR coupling was described by 3 parameters (a1, b2, b3) transfer function (TRF) selected on the basis of a statistical analysis of performances of different TRF models. We found that our model is by far the best in its class: with more parameters (higher order models) the residuals remain almost the same while the extra parameters display variability much larger than that of our parameters. For all measurements, our TRF model describes more than 70% of QT variability. Within the patient set, we found interesting differences concerning dynamic non-linearity (response times longer with decreasing RR intervals than with increasing RR).

Exploring cDNA phage display for autoantibody profiling in the serum of multiple sclerosis patients: optimization of the selection procedure.

We applied a cDNA phage display method called serological antigen selection (SAS) to identify immunogenic targets that evoke an autoantibody response in the serum of multiple sclerosis (MS) patients. This method involves the display of a cDNA expression library, in this study a MS brain library, on filamentous phage and subsequent selection using patient immunoglobulin G (IgG). To apply the SAS technology for autoantibodies in the serum of MS patients, an optimization was necessary to deplete cDNA products that encode IgG fragments derived from B cells present in the MS brain plaques. We describe a differential screening procedure in which positive selection rounds on MS serum and negative selection rounds on healthy control serum were alternated to optimize the selection procedure. As a result, a substantial decrease of IgG-displaying phage clones was observed after each negative selection round, thereby preventing an overgrowth of IgG-displaying phage clones. Our depletion strategy was therefore successful in preventing the enrichment of IgG-displaying phage clones. This approach will facilitate the identification of possible MS-related antigens.

Obesity-related cardiovascular disease: implications of obstructive sleep apnea.

Obesity and obstructive sleep apnea (OSA) often coexist. OSA has been linked to cardiovascular disease. Thus, OSA may contribute to the cardiovascular consequences of obesity. In this review, we explore clinical and pathophysiological interactions between obesity, cardiovascular disease and OSA. We discuss the mechanisms whereby OSA may contribute to hypertension, atherosclerosis, insulin resistance and atrial fibrillation associated with obesity, and emphasize the potential implications for understanding why only a subgroup of obese patients develop cardiovascular disease. Identification of the OSA-dependent and OSA-independent pathways in the cardiovascular pathophysiology of obesity may hold clinical and therapeutic promise.

Noninvasive detection of vessel stiffness from continuous blood pressure recordings in hypertensive subjects.

This paper presents results of blood pressure dynamicity analysis aimed at vessel stiffness detection and subsequent cardiac risk stratification. We analyzed ECG and BP parameters from 12 normotensive young healthy volunteers, 10 old healthy volunteers, and two groups of hypertensive patients -- 12 young non-medicated hypertensive subjects with no other known complications and 16 hypertensive non-medicated subjects with confirmed obesity (according to waist circumference), hyperlipidemia or diabetes mellitus. The dynamic parameters obtained from a derivative continuous blood pressure signal provide additional information about vessel compliance. They can differentiate hypertensive subjects according to the level of cardiovascular risk.

Profiling the autoantibody repertoire by serological antigen selection.

The identification of disease related autoantigens targeted by pathogenic T- and B-cell responses is crucial for the development of improved therapies for autoimmune diseases. To identify immunogenic targets recognized by the humoral immune response, we have recently applied a novel and powerful molecular approach, named 'serological antigen selection' (SAS). This method involves the display of a cDNA expression library on filamentous phage and subsequent selection on patient immunoglobulin G (IgG). In the present study, we have cloned a cDNA repertoire from a multiple sclerosis (MS) patient in pVI phage display vectors and performed selections on pooled MS cerebrospinal fluid (CSF) samples immobilized with anti-human IgG. To further streamline this procedure, we report an optimized SAS procedure in which we have successfully established methods for enrichment of MS-specific candidate antigens. In conclusion, the broad applicability of the SAS method makes it a highly promising method for investigating the autoimmune repertoire.

Recognition, diagnosis and management of obesity after myocardial infarction.

OBJECTIVE: We investigated the documentation of obesity as a medical problem, and subsequent management recommendations, in patients after myocardial infarction (MI). DESIGN: We performed a cross-sectional analysis of a randomly selected sample of 627 patients discharged after an MI, from five US teaching hospitals between 1/1/01 and 12/31/02. Information was extracted from clinical notes using standardized definitions. RESULTS: Mean body mass index (BMI) was 31+/-13 kg/m2, which was documented in only 14% of patients and had to be calculated post hoc in the rest. Waist circumference and waist/hip ratio were not documented at all; 83% of patients were overweight, 55% obese, and 8% morbidly obese. In only 20% of patients with BMI> or =30 kg/m2 was the diagnosis of obesity documented either as a current medical problem, as part of past medical history or as a final diagnosis. A dietary counseling was carried out in 61% of patients with BMI> or =25 kg/m2 and in 61% of patients with BMI<25 kg/m2, P=0.96. Weight loss was described as part of the goals/plan at discharge in 7% of overweight and 9% of obese patients. There was no change in either the level of recognition of obesity (22 vs 19%, P=0.3) or in the proportion of obese patients for whom weight loss was described as part of the goals/plan at discharge (8 vs 10%, P=0.7) before (n=301) compared to after (n=326) the Call to Action in Obesity by the Surgeon General in December 2001. CONCLUSION: Obesity is underecognized, underdiagnosed and undertreated in persons with acute MI.

Sleep apnea and hypertension.

Obstructive sleep apnea (OSA) is the most common form of sleep-disordered breathing and frequently coexists with obesity. Almost 15 million Americans are affected by this disorder. This prevalence is likely increasing, given the current epidemic of obesity. Recent data confirm an association between sleep apnea and several cardiovascular disease conditions, suggesting that OSA may be a new risk factor for coronary artery disease, heart failure, heart rhythm disturbances and hypertension, independent of body mass index. In this review, the authors focus on the nature of the association between OSA and hypertension, the evidence suggesting a causal interaction, and discuss the potential pathophysiologic mechanisms responsible. These mechanisms include activation of the sympathetic and renin-angiotensin-aldosterone systems (RAAS), oxidative stress, and systemic and vascular inflammation, all of which could link OSA to a sustained increase in blood pressure. The authors also review potential therapeutic strategies for the hypertensive patient with OSA.