Stable ubiquitin conjugation for biological interrogation of ubiquitinated tau repeat domain.

Protein semisynthesis approaches are key for gaining insights into the effects of post-translational modifications (PTMs) on the structure and function of modified proteins. Among PTMs, ubiquitination involves the conjugation of a small protein modifier to a substrate amino acid residue and is unique in controlling a variety of cellular processes. Interest has grown in understanding the role of ubiquitination in neurodegenerative conditions, including tauopathies. The latter are characterized by the accumulation of the intrinsically disordered protein tau in the form of neurofibrillary tangles in the brains of patients. The presence of ubiquitinated tau in the pathological aggregates suggests that ubiquitination might play a role in the formation of abnormal protein deposits. In this study, we developed a new strategy, based on dehydroalanine chemistry, to install wild type ubiquitin on a tau repeat domain construct with site-specificity. We optimized a three-step reaction which yielded a good amount of highly pure tau repeat domain ubiquitinated in position 353. The structural features of the conjugate were examined by circular dichroism and NMR spectroscopy. The ubiquitinated tau was challenged in a number of assays: fibrils formation under aggregating conditions in vitro, chemical stability upon exposure to a variety of biological media including cell extracts, and internalization into astrocytes. The results demonstrated the wide applicability of the new semisynthetic strategy for the investigation of ubiquitinated substrates in vitro or in cell, and in particular for studying if ubiquitination has a role in the molecular mechanisms that underlie the aberrant transition of tau into pathological aggregates.

Diurnal and day-to-day biological variation of salivary cortisol and cortisone.

OBJECTIVES: There is a growing interest in the relevance of salivary cortisol and cortisone concentrations in stress-related research. To correctly attribute the magnitude of salivary cortisol and cortisone variation as an effect of a stressful event, a coherent understanding of the day-to-day intra-individual and inter-individual variability across the diurnal cycle of the two steroids is required. However, such information is currently lacking. METHODS: This study aimed to overcome these existing limitations by performing an investigation of the biological variation (BV) of salivary cortisol and cortisone within one day and between five days using an LC-MS/MS method. Saliva samples were collected from 20 healthy volunteers immediately after waking up, at 8:00, 12:00, 15:00, 19:00 and 23:00 on each day over five days. All samples were analyzed in duplicate in one run. Nested ANOVA was used to calculate the sums of squares for analytical and biological components of variation. RESULTS: The within-subject BV of salivary cortisol and cortisone (CVI) ranged from a minimum of 29.3 and 19.0 % to a maximum of 56.5 and 49.1 %, respectively, while the between-subject biological variation (CVG) ranged from 29.7 and 29.0 % to 51.6 and 43.6 %. The reference change values (RCVs) ranged from 96 to 245 % for cortisol and from 55 to 194 % for cortisone. A medium index of individuality was observed for both compounds at all time points. CONCLUSIONS: This study provides updated BV estimates and RCVs for different times of day that can be used to assess the magnitude of change in biomarkers in future stress-related research.

Comparison of Performance in the Six-Minute Walk Test (6MWT) between Overweight/Obese and Normal-Weight Children and Association with Haemodynamic Parameters: A Cross-Sectional Study in Four Primary Schools.

Physical activity plays a pivotal role in preventing obesity and cardiovascular risks. The six-minute walk test (6MWT) is a tool to assess functional capacity and predict cardiovascular events. The aim of this cross-sectional study was to compare the performance and haemodynamic parameters before and after a 6MWT between obese/overweight vs. normal-weight children (average age 8.7 ± 0.7 years) participating in a project involving four primary schools in South Verona (Italy). Validated questionnaires for physical activity and diet, as well as blood drops, were collected. Overweight or obese children (OW&OB; n = 100) covered a shorter 6MWT distance compared to normal-weight children (NW, n = 194). At the test's conclusion, the OW&OB group exhibited a higher Rate Pulse Product (RPP = Systolic Blood Pressure × Heart Rate) as compared to the NW. Body Mass Index, waist-to-height ratio, fat mass by electrical impedance, and trans fatty acids showed direct correlations with pre and post-test haemodynamic parameters, such as RPP, and inverse correlations with oxygen saturation. OW&OB children demonstrated lower performance in this low-intensity exercise test, along with an elevated haemodynamic response. Excess fat in childhood can be considered a risk factor for haemodynamic stress, with potential deleterious consequences later in life. Efforts should be initiated early to break this cycle.

Analytical evaluation of the novel Mindray high sensitivity cardiac troponin I immunoassay on CL-1200i.

OBJECTIVES: The current study was designed to evaluate the analytical performance of the new Mindray highly sensitive cardiac troponin I (hs-cTnI) chemiluminescent immunoassay on Mindray CL-1200i, as a thorough validation of novel hs-cTnI methods is required before introduction into clinical practice. METHODS: The evaluation of the analytical performance of this hs-cTnI immunoassay encompassed the calculation of the limit of blank (LOB), limit of detection (LOD), functional sensitivity, imprecision, linearity, 99th percentile upper reference limit (URL) and concordance with another previously validated hs-cTnI chemiluminescent immunoassay. RESULTS: The LOB and LOD were 0.32 and 0.35 ng/L, whilst the functional sensitivity (expressed as cTnI value with <10 % imprecision), was 0.35 ng/L. The linearity was excellent throughout a wide range of clinically measurable values (r=1.00 between 0.8 and 9,726.9 ng/mL). The intra-assay, inter-assay and total imprecision were 1.1-1.3 %, 5.5-8.1 % and 5.6-8.2 %, respectively. The 99th percentile URL calculated using residual plasma from 246 ostensibly healthy blood donors was 9.2 ng/L (4.3 ng/L in women vs. 12.3 ng/L in men). The Spearman's correlation between Mindray hs-cTnI and Access hs-TnI was 0.97, with mean bias of 7.2 % (95 % CI, 2.6-11.9 %). CONCLUSIONS: Although we failed to confirm the very optimistic analytical characteristics previously reported for this method, our evaluation of the novel Mindray hs-cTnI immunoassay on CL-1200i demonstrated that the overall performance is comparable to that of other commercially available hs-cTnI techniques, making it a viable alternative to other methods.

Cholestatic HCV Cryoglobulinemia: A New Clinical and Pathological Entity before and after Direct-Acting Antiviral Therapies-A Case-Control Study.

Twenty-nine patients with HCV infection (HCV+) and mixed cryoglobulinemia (MC+) were retrospectively selected and matched for age and sex with 31 HCV+ MC- patients. Biomarkers of cholestasis (direct bilirubin, alkaline phosphatase, and gamma-glutamyl transferase), HCV-RNA and genotype, and plasma cryoprecipitates were measured before and after virus eradication; liver histology and plasma cells (aggregation and distribution), observed blinded by two pathologists, were analyzed. Sixty participants (mean age: 56.5; range: 35-77, males: 50%) with HCV infection were enrolled. Cholestasis (≥2 pathologically increased cholestasis biomarkers) was significantly higher in the MC group (p = 0.02) and correlated with cryoglobulinemia (OR 6.52; p = 0.02). At liver histological assessment, plasma cells were significantly increased in the MC+ group (p = 0.004) and tended to form aggregates more than the control group (p = 0.05). At multivariate analysis with MC, age, HCV-RNA, HBV diabetes, and cirrhosis, cholestasis was only significantly correlated to MC (OR 8.30; p < 0.05). In 25% patients, MC persisted after virus eradication with new antiviral treatment. Our study identified for the first time an association between MC, cholestasis, and an increased number of intrahepatic plasma cells in chronic hepatitis C (CHC) patients before virus eradication. Future studies are required to understand how MC contributes to liver damage and how its persistence affects the patients' follow-up after antiviral therapies.

Immunogenicity and Safety of the BNT162b2 COVID-19 Vaccine in Patients with Cystic Fibrosis with or without Lung Transplantation.

Cystic fibrosis (CF) is characterized by a progressive decline in lung function, which may be further impaired by viral infections. CF is therefore considered a comorbidity of coronavirus disease 2019 (COVID-19), and SARS-CoV-2 vaccine prioritization has been proposed for patients with (pw)CF. Poor outcomes have been reported in lung transplant recipients (LTR) after SARS-CoV-2 infections. LTR have also displayed poor immunization against SARS-CoV-2 after mRNA-based BNT162b2 vaccination, especially in those undergoing immunosuppressive treatment, mostly those receiving mycophenolate mofetil (MMF) therapy. We aimed to determine here the immunogenicity and safety of the BNT162b2 vaccine in our cohort of 260 pwCF, including 18 LTR. Serum levels of neutralizing anti-SARS-CoV-2 IgG and IgA antibodies were quantified after the administration of two doses. PwCF displayed a vaccine-induced IgG and IgA antiviral response comparable with that seen in the general population. We also observed that the immunogenicity of the BNT162b2 vaccine was significantly impaired in the LTR subcohort, especially in patients undergoing MMF therapy. The BNT162b2 vaccine also caused minor adverse events as in the general population, mostly after administration of the second dose. Overall, our results justify the use of the BNT162b2 vaccine in pwCF and highlight the importance of a longitudinal assessment of the anti-SARS-CoV-2 IgG and IgA neutralizing antibody response to COVID-19 vaccination.

Lifestyle and dietary factors, iron status and one-carbon metabolism polymorphisms in a sample of Italian women and men attending a Transfusion Medicine Unit: a cross-sectional study.

Iron (Fe) status among healthy male and female blood donors, aged 18-65 years, is estimated. General characteristics and lifestyle factors, dietary habits and major one-carbon metabolism-related polymorphisms were also investigated. An explorative cross-sectional study design was used to examine a sample of blood donors attending the Transfusion Medicine Unit of the Verona University Hospital, Italy. From April 2016 to May 2018, 499 subjects were enrolled (255 men, 244 women, 155 of whom of childbearing age). Major clinical characteristics including lifestyle, dietary habits and Fe status were analysed. The MTHFR 677C > T, cSHMT 1420C > T, DHFR 19bp ins/del and RFC1 80G > A polymorphisms were also assayed. Mean plasma concentrations of Fe and ferritin were 16·6 µmol/l (95 % CI 16·0, 17·2) and 33·8 µg/l (95 % CI 31·5, 36·2), respectively. Adequate plasma Fe concentrations (> 10·74 µmol/l) were detected in 84·3 % and adequate ferritin concentrations (20-200 µg/l) was found in 72·5 % of the whole cohort. Among the folate-related polymorphisms analysed, carriers of the DHFR 19bp del/del mutant allele showed lower ferritin concentration when compared with DHFR 19bp ins/del genotypes. In a sample of Italian healthy blood donors, adequate plasma concentrations of Fe and ferritin were reached in a large proportion of subjects. The relationship of Fe status with lifestyle factors and folate-related polymorphisms requires more investigation to clarify further gene-nutrient interactions between folate and Fe metabolism.

High Plasma Levels of Activated Factor VII-Antithrombin Complex Point to Increased Tissue Factor Expression in Patients with SARS-CoV-2 Pneumonia: A Potential Link with COVID-19 Prothrombotic Diathesis.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causal agent of coronavirus disease 2019 (COVID-19), in which coagulation abnormalities and endothelial dysfunction play a key pathogenic role. Tissue factor (TF) expression is triggered by endothelial dysfunction. Activated factor VII-antithrombin (FVIIa-AT) complex reflects indirectly FVIIa-TF interaction and has been proposed as a potential biomarker of prothrombotic diathesis. FVIIa-AT plasma concentration was measured in 40 patients (30 males and 10 females; 64.8 ± 12.3 years) admitted with SARS-CoV-2 pneumonia during the first pandemic wave in Italy. Two sex- and age-matched cohorts without COVID-19, with or without signs of systemic inflammation, were used to compare FVIIa-AT data. The FVIIa-AT plasma levels in COVID-19 patients were higher than those in non-COVID-19 subjects, either with or without inflammation, while no difference was observed among non-COVID-19 subjects. The association between COVID-19 and FVIIa-AT levels remained significant after adjustment for sex, age, C-reactive protein, renal function, fibrinogen, prothrombin time and activated partial thromboplastin time. Our results indicate that SARS-CoV-2 infection, at least during the first pandemic wave, was characterized by high FVIIa-AT levels, which may suggest an enhanced FVIIa-TF interaction in COVID-19, potentially consistent with SARS-CoV-2-induced endotheliopathy.

Heparin: The Journey from Parenteral Agent to Nasal Delivery.

Although the worldwide usage of direct oral anticoagulants has continuously increased over the past decade, heparin remains an important weapon in the current arsenal of anticoagulant drugs. Parenteral heparin administration (i.e., either intravenously or subcutaneously) has represented for decades the only possible route for generating a significant anticoagulant effect, although being notoriously associated with some important drawbacks such as discomfort and risk of low compliance, thus paving the way to searching for more amenable means of administration. We provide here an updated analysis of animal and human studies that have explored the feasibility, suitability, and efficiency of heparin administration through the unconventional nasal route, as a possible alternative to the more traditional parenteral injection. The major hurdles that contribute to impair intranasal absorption and systemic delivery of heparin are represented by its relatively high molecular weight and negative charge. Therefore, although pure drug administration would not be associated with efficient nasal adsorption, or by systemic biological activity (i.e., anticoagulant effect), the combination of low molecular weight heparins and absorption enhancers such as surfactants, mucoadhesive, cyclodextrins, polyethylenimines and encapsulation into (nano)carriers seems effective to at least partially improve drug transport through the nasal route and allow systemic delivery in animals. Besides generating anticoagulant effects, intranasal heparin administration can also produce local pleiotropic effects, mostly related to anti-inflammatory properties, such as attenuating airway allergic inflammation or inhibiting the binding of the spike protein of some coronaviruses (including severe acute respiratory syndrome coronavirus 2) to their host cell receptors. This preliminary evidence represents a valuable premise for planning future studies in humans aimed at establishing the pharmacokinetics and biological activity of locally and systemically delivered intranasal heparin formulations.

Homocysteine in coronavirus disease (COVID-19): a systematic literature review.

OBJECTIVES: Coronavirus disease 2019 (COVID-19) is a life-threatening infectious disorder characterized by a sustained prothrombotic state. Since homocysteine is a potential biomarker of thrombotic diseases, the aim of this article is to provide an updated overview on the possible role played by hyperhomocysteinemia in influencing an unfavorable COVID-19 progression. METHODS: We carried out an electronic search in Medline (PubMed interface) using the keywords ("COVID-19" OR "SARS-CoV-2") AND "homocysteine", between 2019 and the present time, with no language restrictions, to identify all articles which explored the concentration of homocysteine in COVID-19 patients with or without unfavorable disease progression. RESULTS: Three studies, totaling 694 hospitalized COVID-19 patients, were included in our systematic review. Overall, the differences between the mean homocysteine values in non-severe vs. severe COVID-19 patients were always positive (i.e., 15.1%, 24.1% and 22.8%, generating a positive weight mean difference of 1.75 µmol/L (95%CI, 1.26-2.25 µmol/L; p=0.011), which translates into a cumulative difference of approximately ∼1.2 µmol/L. CONCLUSIONS: Despite the limited evidence that has been garnered so far, increased homocysteine ​​levels may be a potentially useful marker for predicting the risk of unfavorable progression in patients with COVID-19.

Artificial intelligence at the time of COVID-19: who does the lion's share?

OBJECTIVES: The development and use of artificial intelligence (AI) methodologies, especially machine learning (ML) and deep learning (DL), have been considerably fostered during the ongoing coronavirus disease 2019 (COVID-19) pandemic. Several models and algorithms have been developed and applied for both identifying COVID-19 cases and for assessing and predicting the risk of developing unfavourable outcomes. Our aim was to summarize how AI is being currently applied to COVID-19. METHODS: We conducted a PubMed search using as query MeSH major terms "Artificial Intelligence" AND "COVID-19", searching for articles published until December 31, 2021, which explored the possible role of AI in COVID-19. The dataset origin (internal dataset or public datasets available online) and data used for training and testing the proposed ML/DL model(s) were retrieved. RESULTS: Our analysis finally identified 292 articles in PubMed. These studies displayed large heterogeneity in terms of imaging test, laboratory parameters and clinical-demographic data included. Most models were based on imaging data, in particular CT scans or chest X-rays images. C-Reactive protein, leukocyte count, creatinine, lactate dehydrogenase, lymphocytes and platelets counts were found to be the laboratory biomarkers most frequently included in COVID-19 related AI models. CONCLUSIONS: The lion's share of AI applied to COVID-19 seems to be played by diagnostic imaging. However, AI in laboratory medicine is also gaining momentum, especially with digital tools characterized by low cost and widespread applicability.

Performance of non-invasive respiratory function indices in predicting clinical outcomes in patients hospitalized for COVID-19 pneumonia in medical and sub-intensive wards: a retrospective cohort study.

Coronavirus disease 2019 (COVID-19) is a newly recognized infectious disease which can lead to acute respiratory distress syndrome requiring ventilatory support and intensive care unit admission. The aim of our study is to evaluate the performance of two non-invasive respiratory function indices (the ROX index and the SatO2/FiO2 ratio), as compared to the traditional PaO2/FiO2 ratio, in predicting a clinically relevant composite outcome (death or intubation) in hospitalized patients for COVID-19 pneumonia. Four hospital centers in Northern Italy conducted an observational retrospective cohort study during the first wave of COVID-19 pandemic. Four hundred and fifty-six patients with COVID-19 pneumonia admitted to medical or sub-intensive wards were enrolled. Clinical, laboratory, and respiratory parameters, for the calculation of different indices, were measured at hospital admission. In medical wards (Verona and Padua) the PaO2/FiO2 ratio, ROX index and SatO2/FiO2 ratio were able to predict intubation or death with good accuracy (AUROC for the PaO2/FiO2 ratio, ROX index and SatO2/FiO2 ratio of 75%, 75% and 74%, respectively). Regarding sub-intensive wards (Milan and Mantua), none of the three respiratory function indices was significantly associated with the composite outcome. In patients admitted to medical wards for COVID-19 pneumonia, the ROX index and the SatO2/FiO2 ratio demonstrated not only good performance in predicting intubation or death, but their accuracy was comparable to that of the PaO2/FiO2 ratio. In this setting, where repeated arterial blood gas tests are not always feasible, they could be considered a reliable alternative to the invasive PaO2/FiO2 ratio.

Psychological Stress and Salivary Cortisol Levels in Patients with Plaque Psoriasis.

Psychological stress has long been recognized as a trigger for plaque psoriasis, and preliminary evidence suggests that psoriasis could be associated with alterations in the hypothalamic-pituitary-adrenal (HPA) axis, resulting in impaired cortisol response to stress. This study aimed to investigate psychological stress, anxiety, depression and salivary cortisol in psoriatic patients. A cross sectional study involving 126 adult patients with plaque psoriasis and 116 adult healthy controls was conducted. Demographic, clinical data, Perceived Stress Scale (PSS) and Hospital Anxiety and Depression Scale (HADS) were collected. Cases and controls were asked whether they felt stressed in the last month, whilst psoriatic patients were also interrogated whether they found that psoriasis could have been worsened by stress. Moreover, 54 randomly selected subjects (27 psoriasis patients and 27 controls) underwent salivary cortisol testing at 8 am. PSS, HADS depression and anxiety subscales were significantly higher in psoriatic patients than in controls (17.2 ± 0.6 vs. 15.1 ± 0.8 p = 0.0289), (9.5 ± 0.3 vs. 6.2 ± 0.3 p < 0.001) and (8.2 ± 0.4 vs. 4.2 ± 0.3 p < 0.001), respectively. A higher rate of psoriatic patients reported feeling stress over the last month (45% vs. 19%, p < 0.001), and stress was considered a potential trigger for psoriasis flare-ups in 69% of cases. Psoriasis was strongly associated with higher PSS and HADS scores independently of sex, body mass index, diabetes, hypertension, dyslipidemia, and occupational status. Salivary cortisol was significantly lower in psoriatic patients compared to controls (9.6 ± 0.5 vs. 14.0 ± 1.1 nmol/L, p < 0.001). In conclusion, psoriasis was associated with higher psychological stress, anxiety and depressive symptoms, and with impaired cortisol response to stress.

Comparison of five commercial anti-SARS-CoV-2 total antibodies and IgG immunoassays after vaccination with BNT162b2 mRNA.

BACKGROUND: Since universal vaccinations represents the most effective strategy to mitigate coronavirus disease 2019 (COVID-19), baseline assessment and post-vaccine monitoring of anti-SARS-CoV-2 neutralizing antibodies are essential to vaccination programs. Therefore, this study aimed to compare data of five commercial anti-SARS-CoV2 immunoassays after administration of an mRNA vaccine. METHODS: Venous blood was collected from three healthcare workers, receiving a double (30 g) dose of BNT162b2 mRNA Covid-19 vaccine (Comirnaty, Pfizer), on the day of the first vaccine dose and then at fixed intervals for the following 2 months. Anti-SARS-CoV-2 neutralizing antibody response was assayed with Roche Total Ig anti-RBD (receptor binding domain), DiaSorin TrimericS IgG (spike trimer), Beckman Coulter IgG anti-RBD, SNIBE IgG anti-RBD and Technogenetics IgG anti-N/S1. RESULTS: A total number of 45 samples were drawn at the end of the 2-month study period. The Spearman's correlations of absolute anti-SARS-CoV-2 antibodies were always excellent (all p<0.001), comprised between 0.967-0.994. Satisfactory results were also observed when absolute antiSARS-CoV-2 antibodies values of the five methods were compared with the mean consensus value, with correlations always higher than 0.979 (all p<0.001). The agreement of anti-SARS-CoV-2 antibodies positivity versus the consensus median positivity ranged between 0.764 and 1.000 (always p<0.001), but become always >0.900 after readjustment of one assay cutoff. CONCLUSIONS: All the immunoassays evaluated in this study appear suitable for monitoring anti-SARS-CoV-2 neutralizing antibodies response in subjects undergoing mRNA COVID-19 vaccination.

Variation of Forehead Temperature during Routine Working Shift in Hospital Laboratory Personnel: Implications for SARS-CoV-2 Screening.

BACKGROUND: Scarce information is available on circadian body temperature fluctuation in healthy healthcare workers. METHODS: Forehead temperature was measured with an infrared thermometer in 33 ostensibly healthy laboratory professionals (mean age, 43 ± 13 years; 76% females) throughout a regular working shift, from 800 AM to 300 PM, at 1-hour intervals. RESULTS: A significant difference was found at different times of the day by 1-way analysis of variance (F statistics, 13.79; p < 0.001). The lowest mean forehead temperature was 36.2 ± 0.3℃, recorded at 100 PM, whilst the highest was 36.7 ± 0.3℃, at 900 AM. The mean difference between forehead temperature at acrophase and nadir was 0.5℃ (95% CI, 0.3-0.6℃; p < 0.001). The forehead temperature measured between 900-1200 AM was also significantly higher than that measured between 100-300 PM (0.3℃; 95% CI, 0.2-0.4℃; p < 0.001). The mean intra-individual variation of forehead temperature was higher but not significantly different in men (1.0 ± 0.2%) compared to women (0.8 ± 0.3%; p = 0.112). CONCLUSION: Fever screening protocols for purposes of coronavirus disease 2019 (COVID-19) and other infectious diseases monitoring should consider normal daily fluctuations in forehead temperature.

Changes in Cerebrospinal Fluid Balance of TNF and TNF Receptors in Naïve Multiple Sclerosis Patients: Early Involvement in Compartmentalised Intrathecal Inflammation.

An imbalance of TNF signalling in the inflammatory milieu generated by meningeal immune cell infiltrates in the subarachnoid space in multiple sclerosis (MS), and its animal model may lead to increased cortical pathology. In order to explore whether this feature may be present from the early stages of MS and may be associated with the clinical outcome, the protein levels of TNF, sTNF-R1 and sTNF-R2 were assayed in CSF collected from 122 treatment-naïve MS patients and 36 subjects with other neurological conditions at diagnosis. Potential correlations with other CSF cytokines/chemokines and with clinical and imaging parameters at diagnosis (T0) and after 2 years of follow-up (T24) were evaluated. Significantly increased levels of TNF (fold change: 7.739; p < 0.001), sTNF-R1 (fold change: 1.693; p < 0.001) and sTNF-R2 (fold change: 2.189; p < 0.001) were detected in CSF of MS patients compared to the control group at T0. Increased TNF levels in CSF were significantly (p < 0.01) associated with increased EDSS change (r = 0.43), relapses (r = 0.48) and the appearance of white matter lesions (r = 0.49). CSF levels of TNFR1 were associated with cortical lesion volume (r = 0.41) at T0, as well as with new cortical lesions (r = 0.56), whilst no correlation could be found between TNFR2 levels in CSF and clinical or MRI features. Combined correlation and pathway analysis (ingenuity) of the CSF protein pattern associated with TNF expression (encompassing elevated levels of BAFF, IFN-γ, IL-1ß, IL-10, IL-8, IL-16, CCL21, haptoglobin and fibrinogen) showed a particular relationship to the interaction between innate and adaptive immune response. The CSF sTNF-R1-associated pattern (encompassing high levels of CXCL13, TWEAK, LIGHT, IL-35, osteopontin, pentraxin-3, sCD163 and chitinase-3-L1) was mainly related to altered T cell and B cell signalling. Finally, the CSF TNFR2-associated pattern (encompassing high CSF levels of IFN-ß, IFN-λ2, sIL-6Rα) was linked to Th cell differentiation and regulatory cytokine signalling. In conclusion, dysregulation of TNF and TNF-R1/2 pathways associates with specific clinical/MRI profiles and can be identified at a very early stage in MS patients, at the time of diagnosis, contributing to the prediction of the disease outcome.

Comprehensive assessment of humoral response after Pfizer BNT162b2 mRNA Covid-19 vaccination: a three-case series.

OBJECTIVES: Since universal vaccination is a pillar against coronavirus disease 2019 (COVID-19), monitoring anti-SARS-CoV-2 neutralizing antibodies is essential for deciphering post-vaccination immune response. METHODS: Three healthcare workers received 30 µg BNT162b2 mRNA Covid-19 Pfizer Vaccine, followed by a second identical dose, 21 days afterwards. Venous blood was drawn at baseline and at serial intervals, up to 63 days afterwards, for assessing total immunoglobulins (Ig) anti-RBD (receptor binding domain), anti-S1/S2 and anti-RBD IgG, anti-RBD and anti-N/S1 IgM, and anti-S1 IgA. RESULTS: All subjects were SARS-CoV-2 seronegative at baseline. Total Ig anti-RBD, anti-S1/S2 and anti-RBD IgG levels increased between 91 and 368 folds until 21 days after the first vaccine dose, then reached a plateau. The levels raised further after the second dose (by ∼30-, ∼8- and ∼8-fold, respectively), peaking at day 35, but then slightly declining and stabilizing ∼50 days after the first vaccine dose. Anti-S1 IgA levels increased between 7 and 11 days after the first dose, slightly declined before the second dose, after which levels augmented by ∼24-fold from baseline. The anti-RBD and anti-N/S1 IgM kinetics were similar to that of anti-S1 IgA, though displaying substantially weaker increases and modest peaks, only 4- to 7-fold higher than baseline. Highly significant inter-correlation was noted between total Ig anti-RBD, anti-S1/S2 and anti-RBD IgG (all r=0.99), whilst other anti-SARS-CoV-2 antibodies displayed lower, though still significant, correlations. Serum spike protein concentration was undetectable at all-time points. CONCLUSIONS: BNT162b2 mRNA vaccination generates a robust humoral immune response, especially involving anti-SARS-Cov-2 IgG and IgA, magnified by the second vaccine dose.