Neurofilament levels are associated with blood-brain barrier integrity, lymphocyte extravasation, and risk factors following the first demyelinating event in multiple sclerosis.

BACKGROUND: Increased blood brain barrier (BBB) permeability, CNS inflammation and neuroaxonal damage are pathological hallmarks in early multiple sclerosis (MS). OBJECTIVE: To investigate the associations of neurofilament light chain (NfL) levels with measures of BBB integrity and central nervous system (CNS) inflammation in MS during the first demyelinating event. METHODS: Blood and cerebrospinal fluid (CSF) were obtained from 142 MS (McDonald 2017) treatment-naive patients from the SET study (63% female; age: 29.7 ± 7.9 years) following the disease onset. NfL, albumin, immunoglobulin G (IgG), and immunoglobulin M (IgM) levels were measured in CSF and blood samples. Albumin quotient was computed as a marker of BBB integrity. Immune cell subset counts in CSF were measured using flow cytometry. MS risk factors, such as Human leukocyte antigen DRB1 locus gene (HLA DRB1)*1501, anti-Epstein-Barr virus (EBV) antibodies, and 25-hydroxy vitamin D3, were also measured. RESULTS: Higher serum NfL (sNfL) levels were associated with higher albumin quotient (p < 0.001), CSF CD80+ (p = 0.012), and CD80+ CD19+ (p = 0.015) cell frequency. sNfL levels were also associated with contrast-enhancing and T2 lesions on brain magnetic resonance imaging (MRI; all p ⩽ 0.001). Albumin quotient was not associated with any of the MS risk factors assessed. sNfL levels were associated with anti-EBV viral capsid antigen (VCA) IgG levels (p = 0.0026). CONCLUSION: sNfL levels during the first demyelinating event of MS are associated with greater impairment of BBB integrity, immune cell extravasation, and brain lesion activity on MRI.

Peripheral blood lymphocytes immunophenotyping predicts disease activity in clinically isolated syndrome patients.

BACKGROUND: Clinically isolated syndrome (CIS) represents first neurological symptoms suggestive of demyelinating lesion in the central nervous system (CNS). Currently, there are no sufficient immunological or genetic markers predicting relapse and disability progression, nor there is evidence of the efficacy of registered disease modifying treatments (DMTs), such as intramuscular interferon beta1a. The aim of the study is to evaluate immunological predictors of a relapse or disability progression. METHODS: One hundred and eighty one patients with CIS were treated with interferon beta1a and followed over the period of 4 years. Lymphocyte subsets were analyzed by flow cytometry. A Kaplan-Meier estimator of survival probability was used to analyze prognosis. For statistical assessment only individual differences between baseline values and values at the time of relapse or confirmed disability progression were analysed. RESULTS: Higher levels of B lymphocytes predicted relapse-free status. On the other hand, a decrease of the naïve subset of cells (CD45RA+ in CD4+) after 12, 24, and 36 months of follow-up were associated with an increased risk of confirmed disability progression. CONCLUSION: Our data suggest that the quantification of lymphocyte subsets in patients after the first demyelinating event suggestive of MS may be an important biomarker.

Protective associations of HDL with blood-brain barrier injury in multiple sclerosis patients.

The purpose of this work was to investigate the associations of serum cholesterol and apolipoproteins with measures of blood-brain barrier (BBB) permeability and CNS inflammation following the first clinical demyelinating event. This study included 154 patients [67% female; age, 29.5 ± 8.2 years (mean ± SD)] enrolled in a multi-center study of interferon ß1-a treatment following the first demyelinating event. Blood and cerebrospinal fluid (CSF) were obtained at screening prior to treatment. A comprehensive serum lipid profile and multiple surrogate markers of BBB breakdown and CNS immune activity were obtained. Higher levels of serum HDL cholesterol (HDL-C) and ApoA-I were associated with lower CSF total protein level, CSF albumin level, albumin quotient, and CSF IgG level (all P ≤ 0.001 for HDL-C and all P < 0.01 for ApoA-I). HDL-C was also associated with CSF CD80+ (P < 0.001) and with CSF CD80+CD19+ (P = 0.007) cell frequencies. Higher serum HDL is associated with lower levels of BBB injury and decreased CD80+ and CD80+CD19+ cell extravasation into the CSF. HDL may potentially inhibit the initiation and/or maintenance of pathogenic BBB injury following the first demyelinating event.

Complement System Deficiencies in Elite Athletes.

BACKGROUND: Although regular physical activity improves immune competency and reduces the prevalence of inflammatory diseases, strenuous training in elite athletes is associated with an increased susceptibility to infectious complications. Therefore, the objective of our study was to assess the routinely examined parameters of the complement system in elite athletes. The study was carried out in a cohort of elite athletes (n = 134) and healthy control subjects (n = 110). In all subjects, besides a routine laboratory check-up, serum concentrations of the C3 and C4 complement components, mannose-binding lectin (MBL), as well as activation of all three complement pathways were determined. RESULTS: Compared to healthy controls, lower C3 and C4 complement component concentrations were observed in elite athletes (0.96 ± 0.1 vs. 1.08 ± 0.2 mg/L, and 0.18 ± 0.1 vs. 0.25 ± 0.1 mg/L, respectively, p < 0.05); with much higher frequency rates of C3 and C4 deficiencies in athletes (31.3 vs. 14.5%, and 6 vs. 0%, p < 0.05). Simultaneously, athletes had much higher frequency rates of deficiencies of activation of classical and alternative complement pathways; while, deficiency of activation of the lectin pathway was similar in both cohorts. CONCLUSIONS: We confirmed a high frequency of defects in the complement system in elite athletes. Lower concentrations of C3 and C4 complement components, with high frequencies of deficiencies of the classical and alternative complement activation pathways were the most prevalent disorder of the complement system in elite athletes. Further studies are needed to uncover the functional impacts of these observations upon the susceptibility to infectious diseases.

Immunity following SARS-CoV-2 vaccination in autoimmune neurological disorders treated with rituximab or ocrelizumab.

Background: Rituximab (RTX) and ocrelizumab (OCR), B cell-depleting therapy targeting CD20 molecules, affect the humoral immune response after vaccination. How these therapies influence T-cell-mediated immune response against SARS-CoV-2 after immunization remains unclear. We aimed to evaluate the humoral and cellular immune response to the COVID-19 vaccine in a cohort of patients with multiple sclerosis (MS), neuromyelitis optica spectrum disorders (NMOSD), and myasthenia gravis (MG). Methods: Patients with MS (83), NMOSD (19), or MG (7) undergoing RTX (n=47) or OCR (n=62) treatment were vaccinated twice with the mRNA BNT162b2 vaccine. Antibodies were quantified using the SARS-CoV-2 IgG chemiluminescence immunoassay, targeting the spike protein. SARS-CoV-2-specific T cell responses were quantified by interferon γ release assays (IGRA). The responses were evaluated at two different time points (4-8 weeks and 16-20 weeks following the 2nd dose of the vaccine). Immunocompetent vaccinated individuals (n=41) were included as controls. Results: Almost all immunocompetent controls developed antibodies against the SARS-CoV-2 trimeric spike protein, but only 34.09% of the patients, without a COVID-19 history and undergoing anti-CD20 treatment (via RTX or OCR), seroconverted. This antibody response was higher in patients with intervals of longer than 3 weeks between vaccinations. The duration of therapy was significantly shorter in seroconverted patients (median 24 months), than in the non-seroconverted group. There was no correlation between circulating B cells and the levels of antibodies. Even patients with a low proportion of circulating CD19+ B cells (<1%, 71 patients) had detectable SARS-CoV-2 specific antibody responses. SARS-CoV-2 specific T cell response measured by released interferon γ was detected in 94.39% of the patients, independently of a humoral immune response. Conclusion: The majority of MS, MG, and NMOSD patients developed a SARS-CoV-2-specific T cell response. The data suggest that vaccination can induce SARS-CoV-2-specific antibodies in a portion of anti-CD20 treated patients. The seroconversion rate was higher in OCR-treated patients compared to those on RTX. The response represented by levels of antibodies was better in individuals, with intervals of longer than 3 weeks between vaccinations.

SARS-CoV-2 Specific Humoral Immune Responses after BNT162b2 Vaccination in Hospital Healthcare Workers.

BACKGROUND: COVID-19 pandemic has led to a loss of human life in millions and devastating socio-economic consequences worldwide. So far, vaccination is the most effective long-term strategy to control and prevent severe COVID-19 disease. The aim of the current study was to evaluate the humoral immune responses raised against the BNT162b2 vaccine in hospital healthcare workers. METHODS: Total number of 173 healthcare workers enrolled in the study. Their blood samples were collected in three different time intervals after the second SARS-CoV-2 vaccination and evaluated by the ELISA method to detect anti-spike protein IgM and IgG antibodies. The baseline characteristics of all participants were collected using questionnaires and were evaluated for finding any significant data. RESULTS: Our results demonstrated that the levels of antibodies were higher in the young group (21-30 years old) and also among male participants. Moreover, the highest levels of antibodies were detected from the group that received the third shot vaccination. CONCLUSIONS: Our results indicate that age, gender and third-dose vaccination can affect the levels of humoral immune responses against the BNT162b2 vaccine in healthcare workers.

Safety of the Geneva Cocktail, a Cytochrome P450 and P-Glycoprotein Phenotyping Cocktail, in Healthy Volunteers from Three Different Geographic Origins.

INTRODUCTION AND OBJECTIVE: Cytochrome P450 enzymes are the major drug-metabolizing enzymes in humans and the importance of drug transport proteins, in particular P-glycoprotein, in the variability of drug response has also been highlighted. Activity of cytochrome P450 enzymes and P-glycoprotein can vary widely between individuals and genotyping and/or phenotyping can help assess their activity. Several phenotyping cocktails have been developed. The Geneva cocktail is composed of a specific probe for six different cytochrome P450 enzymes and one for P-glycoprotein and was used in the context of a research aiming at exploring genotypes and phenotypes in distinct human populations (NCT02789527). The aim of the present study is to solely report the safety results of the Geneva cocktail in the healthy volunteers of these populations. MATERIALS AND METHODS: The Geneva cocktail is composed of caffeine, bupropion, flurbiprofen, omeprazole, dextromethorphan, midazolam, and fexofenadine. The volunteers fasted and avoided drinking caffeine-containing beverages or food and grapefruit juice overnight before receiving the cocktail orally. They provided blood spots for the probes' concentrations at 2, 3, and 6 h after ingestion and were asked about adverse events. RESULTS: A total of 265 healthy adult volunteers were included from Ethiopia, Oman, and the Czech Republic. The mean plasma concentrations at the 2-h sampling time of each probe drug in the total sample were: 1663 ng/mL for caffeine, 8 ng/mL for bupropion, 789 ng/mL for flurbiprofen, 6 ng/mL for dextromethorphan, 2 ng/mL for midazolam, 35 ng/mL for fexofenadine, and 103 ng/mL for omeprazole. Four adverse events were observed representing an occurrence of 1.5%. All these events were categorized as mild to moderate, non-serious, and resolved spontaneously. A causal link with the cocktail cannot be excluded because of the temporal relationship but is at most evaluated as possible according to the World Health Organization-Uppsala Monitoring Centre causal assessment system. CONCLUSIONS: In this research, healthy volunteers from three different human populations were phenotyped with the Geneva cocktail. Four adverse events were observed, confirming the safety of this cocktail that is given at lower than clinically relevant doses and therefore results in concentrations lower than those reported to cause adverse events.

Czech Hizentra Noninterventional Study With Rapid Push: Efficacy, Safety, Tolerability, and Convenience of Therapy With 20% Subcutaneous Immunoglobulin.

PURPOSE: Immunoglobulin substitution therapy is an essential therapeutic approach for patients with primary antibody deficiencies. Different methods of administration, including intravenous immunoglobulin (IVIG) or subcutaneous immunoglobulin (SCIG) preparations, provide effective and tolerable treatment and enable the adjustment of therapy to patients' needs. A new 20% SCIG represents a new therapeutic option and a new route of administration using rapid-push application. The aim of the Czech Hizentra Noninterventional Study With Rapid Push (CHHINSTRAP) is to evaluate patient satisfaction with as well as the tolerability and efficacy of nonmedical switch to 20% SCIG from previous treatment with IVIG or SCIG and rapid push as a new way to administer SCIG. CHHINSTRAP is the first Phase IV, noninterventional, open-label, prospective, multicentric study of this type conducted in Central and Eastern Europe. METHODS: Primary end points, including efficacy, adverse effects, convenience of use, and overall satisfaction, were evaluated by Treatment Satisfaction Questionnaire for Medication version II. Secondary end points, such as serum IgG trough levels, infusion duration, number of application sites, frequency of infections, related hospital admissions, and antibiotic consumption, were obtained from patients at each follow-up visit. FINDINGS: Together, 50 eligible patients with primary antibody deficiency were switched from SCIG or IVIG to an equivalent dose of 20% SCIG and were followed up for 12 months during 5 consecutive visits. The results indicate that patients switched from previous IVIG or SCIG preparations had significantly higher serum trough IgG levels and a lower incidence of infections and related events, such as hospital admissions or consumption of antibiotics. These findings were also reflected in gradually increasing convenience of use and overall satisfaction reported by patients. Apart from duration of application, no differences were found between patients previously receiving SCIG or IVIG. Moreover, our study found a high level of safety of 20% SCIG rapid push, which was comparable to other preparations and application methods. IMPLICATIONS: On the basis of the results of CHHINSTRAP study, we conclude that 20% SCIG is a tolerable and effective immunoglobulin preparation, representing a new therapeutic approach in patients with primary antibody deficiencies. Its efficacy and tolerability have been found in patients on nonmedical switch from previous treatment with IVIG or SCIG.

The effect of dipeptidyl peptidase-IV inhibition on circulating T cell subpopulations in patients with type 2 diabetes mellitus.

AIM: To assess intraindividually the effects of DPP-IV inhibition on the subpopulations of immune cells in type 2 diabetes mellitus (DM2) patients during the course of treatment with sitagliptin. METHODS: In this open label non-randomized observational study with a control group DM2 patients were examined before the initiation of the DPP-IV inhibitor administration (sitagliptin 100mg once daily) and then after 4weeks and 12months. Inhibition of the blood plasma DPP-IV enzymatic activity was determined by a chromogenic assay, the immunophenotyping of the blood cell subpopulations was performed using flow cytometry and blood plasma cytokine concentrations were quantified using an array-based multiplex ELISA. All parameters were evaluated in relation to the entry values in individual patients. RESULTS: The blood plasma DPP-IV enzymatic activity was effectively inhibited during the sitagliptin treatment. A significant decrease of the proportion of Treg cells (to 86±31% (median±SD) of entry values, p=0.001) and an increase of Th1 cells (to 120±103% (median±SD) of entry values, p=0.004) were observed after 4weeks but not after one year of the sitagliptin treatment. No changes were observed in the ratio of CD4(+)/CD8(+) cells, in the quantity of NK and Th2 cells and blood plasma cytokine levels. CONCLUSIONS: Sitagliptin treatment may cause temporary changes of the proportion of lymphocyte subpopulations in patients with DM2. The consequent deregulation of the immune system should be considered as a possible cause of the eventual side effects of long term DPP-IV inhibition.