| 1. |
- Ahmed, Sheik, et al.
(författare)
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Thalassemia Patients from Baluchistan in Pakistan Are Infected with Multiple Hepatitis B or C Virus Strains
- 2021
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Ingår i: American Journal of Tropical Medicine and Hygiene. - : American Society of Tropical Medicine and Hygiene. - 0002-9637 .- 1476-1645. ; 104:4, s. 1569-1576
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Tidskriftsartikel (refereegranskat)abstract
- There are an estimated 2,000 children with 6-thalassemia in the province Baluchistan of Pakistan. These children are at high risk of acquiring transfusion-transmitted infections (TTIs) due to their need of regular blood transfusions for survival. Therefore, we investigated the frequencies of TTIs among these multi-transfused patients in a region where the WHO guidelines for blood safety are not always followed. Sera from 400 children (mean age 7.7 +/- 4.70 years) treated at two thalassemia centers in Baluchistan were investigated for TTIs. Eleven (2.8%) were hepatitis B surface antigen positive, and 72 (18.3%) had anti-hepatitis C virus (HCV), two of which were infected with both viruses. Only 22% of the children had been reached by the program for universal hepatitis B virus (HBV) vaccination which started in 2004. Half (51%) of the HCV infected had also been HBV infected. The HBV- and HCV-infected patients were older and had received more blood transfusions than the uninfected patients (P < 0.001). Molecular characterization of the viral strains revealed the presence of several genetically different strains in at least three HBV- and seven HCV-infected children. This is the first study to demonstrate infections with multiple HBV or HCV strains simultaneously infecting thalassemia patients. These may become the source for new emerging recombinant viruses of unknown virulence. The high prevalence of anti-HCV-positive children, and the presence of HBV infections among children who should have been vaccinated, highlights an urgent need for improvements of blood safety in this region of Pakistan.
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| 2. |
- Aydin, Ebru, et al.
(författare)
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Role of NOX2-Derived Reactive Oxygen Species in NK Cell-Mediated Control of Murine Melanoma Metastasis
- 2017
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Ingår i: Cancer Immunology Research. - : American Association for Cancer Research (AACR). - 2326-6066 .- 2326-6074. ; 5:9, s. 804-811
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Tidskriftsartikel (refereegranskat)abstract
- The NADPH oxidase of myeloid cells, NOX2, generates reactive oxygen species (ROS) to eliminate pathogens and malignant cells. NOX2-derived ROS have also been proposed to dampen functions of natural killer (NK) cells and other antineoplastic lymphocytes in the microenvironment of established tumors. The mechanisms by which NOX2 and ROS influence the process of distant metastasis have only been partially explored. Here, we utilized genetically NOX2-deficient mice and pharmacologic inhibition of NOX2 to elucidate the role of NOX2 for the hematogenous metastasis of melanoma cells. After intravenous inoculation of B16F1 or B16F10 cells, lung metastasis formation was reduced in B6.129S6 Cybb(m1DinK) (Nox2-KO) versus Nox2-sufficient wild-type (WT) mice. Systemic treatment with the NOX2-inhibitor histamine dihydrochloride (HDC) reduced melanoma metastasis and enhanced the infiltration of IFN gamma-producing NK cells into lungs of WT but not of Nox2-KO mice. IFN gamma-deficient B6.129S7-Ifngt(m1Ts)/ J mice were prone to develop melanoma metastases and did not respond to in vivo treatment with HDC. We propose that NOX2-derived ROS facilitate metastasis of melanoma cells by downmodulating NK-cell function and that inhibition of NOX2 may restore IFN gamma-dependent, NK cell-mediated clearance of melanoma cells.
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| 3. |
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| 4. |
- Bergström, Petra, et al.
(författare)
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Amyloid precursor protein expression and processing are differentially regulated during cortical neuron differentiation
- 2016
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 6
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Tidskriftsartikel (refereegranskat)abstract
- Amyloid precursor protein (APP) and its cleavage product amyloid beta (A beta) have been thoroughly studied in Alzheimer's disease. However, APP also appears to be important for neuronal development. Differentiation of induced pluripotent stem cells (iPSCs) towards cortical neurons enables in vitro mechanistic studies on human neuronal development. Here, we investigated expression and proteolytic processing of APP during differentiation of human iPSCs towards cortical neurons over a 100-day period. APP expression remained stable during neuronal differentiation, whereas APP processing changed. alpha-Cleaved soluble APP (sAPP alpha) was secreted early during differentiation, from neuronal progenitors, while beta-cleaved soluble APP (sAPP beta) was first secreted after deep-layer neurons had formed. Short A beta peptides, including A beta 1-15/16, peaked during the progenitor stage, while processing shifted towards longer peptides, such as A beta 1-40/42, when post-mitotic neurons appeared. This indicates that APP processing is regulated throughout differentiation of cortical neurons and that amyloidogenic APP processing, as reflected by A beta 1-40/42, is associated with mature neuronal phenotypes.
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| 5. |
- Bergström, Petra, et al.
(författare)
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Herpes Simplex Virus 1 and 2 Infections during Differentiation of Human Cortical Neurons
- 2021
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Ingår i: Viruses-Basel. - : MDPI AG. - 1999-4915. ; 13:10
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Tidskriftsartikel (refereegranskat)abstract
- Herpes simplex virus 1 (HSV-1) and 2 (HSV-2) can infect the central nervous system (CNS) with dire consequences; in children and adults, HSV-1 may cause focal encephalitis, while HSV-2 causes meningitis. In neonates, both viruses can cause severe, disseminated CNS infections with high mortality rates. Here, we differentiated human induced pluripotent stem cells (iPSCs) towards cortical neurons for infection with clinical CNS strains of HSV-1 or HSV-2. Progenies from both viruses were produced at equal quantities in iPSCs, neuroprogenitors and cortical neurons. HSV-1 and HSV-2 decreased viability of neuroprogenitors by 36.0% and 57.6% (p < 0.0001), respectively, 48 h post-infection, while cortical neurons were resilient to infection by both viruses. However, in these functional neurons, both HSV-1 and HSV-2 decreased gene expression of two markers of synaptic activity, CAMK2B and ARC, and affected synaptic activity negatively in multielectrode array experiments. However, unaltered secretion levels of the neurodegeneration markers tau and NfL suggested intact axonal integrity. Viral replication of both viruses was found after six days, coinciding with 6-fold and 22-fold increase in gene expression of cellular RNA polymerase II by HSV-1 and HSV-2, respectively. Our results suggest a resilience of human cortical neurons relative to the replication of HSV-1 and HSV-2.
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| 6. |
- Nazir, Faisal Hayat, et al.
(författare)
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Antibodies from serum and CSF of multiple sclerosis patients bind to oligodendroglial and neuronal cell-lines
- 2023
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Ingår i: Brain Communications. - : Oxford University Press (OUP). - 2632-1297. ; 5:3
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Tidskriftsartikel (refereegranskat)abstract
- Multiple sclerosis is a highly complex and heterogeneous disease. At the onset it often presents as a clinically isolated syndrome. Thereafter relapses are followed by periods of remissions, but eventually, most patients develop secondary progressive multiple sclerosis. It is widely accepted that autoantibodies are important to the pathogenesis of multiple sclerosis, but hitherto it has been difficult to identify the target of such autoantibodies. As an alternative strategy, cell-based methods of detecting autoantibodies have been developed. The objective of this study was to explore differences in the binding of antibodies from sera and CSF of multiple sclerosis patients and controls to oligodendroglial and neuronal cell-lines, related to antibody type, immunoglobulin (IgG/IgM), matrix (serum/CSF) and disease course. The oligodendroglial and neuronal cell-lines were expanded in tissue culture flasks and transferred to 96-well plates at a concentration of 50 000 cells/well followed by fixation and blocking with bovine serum albumin. Sera and CSF samples, from healthy controls and multiple sclerosis patients, were incubated with the fixed cells. Epitope binding of immunoglobulins (IgG and IgM) in sera and CSF was detected using biotinylated anti-human IgM and IgG followed by avidin conjugated to horseradish peroxidase. Horseradish peroxidase activity was detected with 3,3 ',5,5 '-tetramethylbenzidine substrate. Serum from 76 patients and 30 controls as well as CSF from 62 patients and 32 controls were investigated in the study. The binding was similar between clinically isolated syndrome patients and controls, whereas the largest differences were observed between secondary progressive multiple sclerosis patients and controls. Antibodies from multiple sclerosis patients (all disease course combined) bound more to all investigated cell-lines, irrespectively of matrix type, but binding of immunoglobulin G from CSF to human oligodendroglioma cell-line discriminated best between multiple sclerosis patients and controls with a sensitivity of 93% and a specificity of 96%. The cell-based enzyme linked immunosorbent assay (ELISA) was able to discriminate between multiple sclerosis patients and controls with a high degree of accuracy. The disease course was the major determinant for the antibody binding. Nazir et al. measured antibody binding to neuronal and oligodendroglial cell-lines with a cell-based ELISA. They report that this method could discriminate between multiple sclerosis patients and healthy controls with a high degree of accuracy. The disease course was the major determinant for the antibody binding.
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| 7. |
- Nazir, Faisal Hayat, et al.
(författare)
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Expression and secretion of synaptic proteins during stem cell differentiation to cortical neurons.
- 2018
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Ingår i: Neurochemistry international. - : Elsevier BV. - 1872-9754 .- 0197-0186. ; 121, s. 38-49
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Tidskriftsartikel (refereegranskat)abstract
- Synaptic function and neurotransmitter release are regulated by specific proteins. Cortical neuronal differentiation of human induced pluripotent stem cells (hiPSC) provides an experimental model to obtain more information about synaptic development and physiology in vitro. In this study, expression and secretion of the synaptic proteins, neurogranin (NRGN), growth-associated protein-43 (GAP-43), synaptosomal-associated protein-25 (SNAP-25) and synaptotagmin-1 (SYT-1) were analyzed during cortical neuronal differentiation. Protein levels were measured in cells, modeling fetal cortical development and in cell-conditioned media which was used as a model of cerebrospinal fluid (CSF), respectively. Human iPSC-derived cortical neurons were maintained over a period of at least 150 days, which encompasses the different stages of neuronal development. The differentiation was divided into the following stages: hiPSC, neuro-progenitors, immature and mature cortical neurons. We show that NRGN was first expressed and secreted by neuro-progenitors while the maximum was reached in mature cortical neurons. GAP-43 was expressed and secreted first by neuro-progenitors and its expression increased markedly in immature cortical neurons. SYT-1 was expressed and secreted already by hiPSC but its expression and secretion peaked in mature neurons. SNAP-25 was first detected in neuro-progenitors and the expression and secretion increased gradually during neuronal stages reaching a maximum in mature neurons. The sensitive analytical techniques used to monitor the secretion of these synaptic proteins during cortical development make these data unique, since the secretion of these synaptic proteins has not been investigated before in such experimental models. The secretory profile of synaptic proteins, together with low release of intracellular content, implies that mature neurons actively secrete these synaptic proteins that previously have been associated with neurodegenerative disorders, including Alzheimer's disease. These data support further studies of human neuronal and synaptic development in vitro, and would potentially shed light on the mechanisms underlying altered concentrations of the proteins in bio-fluids in neurodegenerative diseases.
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| 8. |
- Nazir, Faisal Hayat
(författare)
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Increasing the interpretability of Alzheimer-related biomarkers: cell- and cerebrospinal fluid-based studies with focus on neurogranin
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Biomarkers for Alzheimer’s disease (AD) is a growing field of research. A particularly vibrant field during recent years has been biomarkers for synaptic dysfunction. Sensitive assays for a synaptic protein called neurogranin (NRGN) have produced very interesting results when applied on cerebrospinal fluid (CSF) from AD patients and there are several other biomarker candidates that are thought to reflect different aspects of AD pathophysiology. The aim of this thesis was to investigate the expression and secretion of selected Alzheimer-associated biomarkers in a newly developed model of stem cell-derived cortical neurons that recapitulate the in vivo time frames of cortical development. For NRGN, we further investigated the processing and detection of its various molecular forms in CSF. First, human induced pluripotent stem cell (hiPSC)-derived cortical neurons were used to determine the expression and processing of one of the core AD biomarkers, amyloid precursor protein (APP)-derived amyloid beta (Aβ). Our findings suggested that APP was expressed throughout the differentiation, but its processing shifted during neuronal stages. The AD-associated amyloidogenic pathway was activated in mature cortical neurons. Although amyloid and tau pathology are the defining neuropathological lesions, synaptic dysfunction and degeneration are thought to be the earliest events in AD. Thus, secreted synaptic proteins in CSF during neurodegeneration could serve as potential AD biomarkers; a notion that has been supported by several studies on changes in concentration of NRGN in CSF in AD during recent years. To learn more about this biomarker, its expression and secretion were investigated in hiPSC-derived cortical neurons. We also examined three additional markers, namely synaptotagmin-1, SNAP-25 and GAP-43. NRGN, synaptotagmin-1 and SNAP-25 expression peaked in mature neurons, while GAP-43 expression was highest in immature cortical neurons and its secretion peaked in mature cortical neurons. The increased expression of synaptic proteins coincided with neurite network formation, which suggests that secretion of these proteins to the extracellular space reflects synapse maturity. For one of the synaptic proteins, NRGN, C-terminal peptides have been detected at increased levels in CSF from AD patients. Nonetheless, the enzyme(s) that generate these peptides were not known. Here, we identified calpain 1 (CALP1) and prolyl endopeptidase (PREP) as enzymes that cleave NRGN and its fragments. The fragments generated through cleavage by human CALP1 and PREP may suggest an increase in the activation and/or expression of these enzymes in AD. Further, CSF analysis revealed the presence of several molecular forms of NRGN that may represent NRGN fragments, monomers and oligomeric forms, or complexes of NRGN with yet unidentified binding partners. Furthermore, we determined that the ratio of C-terminal fragments to total-NRGN was about 50% in a CSF pool. Taken together, the results of this thesis show that a human-derived neuronal model can teach us a great deal on biomarker processing and secretion into biofluids, which may increase the interpretability of the biomarker results and tell us more about the underlying disease processes, which they may reflect.
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| 9. |
- Nazir, Faisal Hayat, et al.
(författare)
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Molecular forms of neurogranin in cerebrospinal fluid.
- 2021
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Ingår i: Journal of neurochemistry. - : Wiley. - 1471-4159 .- 0022-3042. ; 157:3, s. 816-833
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Tidskriftsartikel (refereegranskat)abstract
- Neurogranin (Ng) is a 78 amino acid neuronal protein and a biomarker candidate for Alzheimer's disease (AD). Ng has been suggested to bind to calmodulin and phosphatidic acid via its centrally located IQ domain. Ng is cleaved within this functionally important domain, yielding the majority of fragments identified in cerebrospinal fluid (CSF), suggesting that cleavage of Ng may be a mechanism to regulate its function. Up to now, Ng has been shown to be present in CSF as both C-terminal fragments as well as full-length protein. To obtain an overview of the different molecular forms of Ng present in CSF, we show by size exclusion chromatography (SEC), immunoblotting, immunoprecipitation and MS that Ng is present in CSF as several molecular forms. Besides monomeric full-length Ng, also higher molecular weight forms of Ng, and C-terminal- and previously not identified N-terminal fragments were observed. We found by immunodepletion that C-terminal peptides contribute on average to ~50 % of the total-Ng ELISA signal in CSF samples. There were no differences in the overall C-terminal fragment/total-Ng ratio between samples from AD and control groups. In addition, we found that monomeric Ng and its C-terminal fragments bind to heparin via a heparin-binding motif, which might be of relevance for their export mechanism from neurons. Taken together, this study highlights the presence of several molecular forms of Ng in CSF, comprising monomeric full-length Ng, and N- and C-terminal truncations of Ng, as well as larger forms of still unknown composition.
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| 10. |
- Pavlovic, Ivan, et al.
(författare)
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Cerebrospinal fluid mtDNA concentrations are increased in multiple sclerosis and were normalized after intervention with autologous hematopoietic stem cell transplantation
- 2024
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Ingår i: Multiple Sclerosis and Related Disorders. - : Elsevier. - 2211-0348 .- 2211-0356. ; 84
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundMitochondrial DNA (mtDNA) is a pro-inflammatory damage-associated molecular pattern molecule and could be an early indicator for inflammation and disease activity in MS. Autologous hematopoietic stem cell transplantation (aHSCT) is a potent treatment for MS, but its impact on mtDNA levels in cerebrospinal fluid (CSF) remains unexplored.ObjectivesTo verify elevated CSF mtDNA concentrations in MS patients and assess the impact of aHSCT on mtDNA concentrations.MethodsMultiplex droplet digital PCR (ddPCR) was used to quantify mtDNA and nuclear DNA in 182 CSF samples. These samples were collected from 48 MS patients, both pre- and post-aHSCT, over annual follow-ups, and from 32 healthy controls.ResultsCSF ccf-mtDNA levels were higher in patients with MS, correlated to multiple clinical and analytical factors and were normalized after intervention with aHSCT. Differences before aHSCT were observed with regard to MRI-lesions, prior treatment and number of relapses in the last year prior to aHSCT.ConclusionOur findings demonstrate elevated CSF mtDNA levels in MS patients, which correlate with disease activity and normalize following aHSCT. These results position mtDNA as a potential biomarker for monitoring inflammatory activity and response to treatment in MS.
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| 11. |
- Satir, Tugce Munise, et al.
(författare)
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Accelerated neuronal and synaptic maturation by BrainPhys medium increases Aβ secretion and alters Aβ peptide ratios from iPSC-derived cortical neurons.
- 2020
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Ingår i: Scientific reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 10:1
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Tidskriftsartikel (refereegranskat)abstract
- One of the neuropathological hallmarks of Alzheimer's disease (AD) is cerebral deposition of amyloid plaques composed of amyloid β (Aβ) peptides and the cerebrospinal fluid concentrations of those peptides are used as a biomarker for AD. Mature induced pluripotent stem cell(iPSC)-derived cortical neurons secrete Aβ peptides in ratios comparable to those secreted to cerebrospinal fluid in human, however the protocol to achieve mature neurons is time consuming. In this study, we investigated if differentiation of neuroprogenitor cells (NPCs) in BrainPhys medium, previously reported to enhance synaptic function of neurons in culture, would accelerate neuronal maturation and, thus increase Aβ secretion as compared to the conventional neural maintenance medium. We found that NPCs cultured in BrainPhys displayed increased expression of markers for cortical deep-layer neurons, increased synaptic maturation and number of astroglial cells. This accelerated neuronal maturation was accompanied by increased APP processing, resulting in increased secretion of Aβ peptides and an increased Aβ38 to Aβ40 and Aβ42 ratio. However, during long-term culturing in BrainPhys, non-neuronal cells appeared and eventually took over the cultures. Taken together, BrainPhys culturing accelerated neuronal maturation and increased Aβ secretion from iPSC-derived cortical neurons, but changed the cellular composition of the cultures.
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| 12. |
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| 13. |
- Tolf, Andreas, et al.
(författare)
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Factors Associated With Serological Response to SARS-CoV-2 Vaccination in Patients With Multiple Sclerosis Treated With Rituximab
- 2022
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Ingår i: JAMA Network Open. - : American Medical Association (AMA). - 2574-3805. ; 5:5
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Tidskriftsartikel (refereegranskat)abstract
- Importance: B-cell-depleting monoclonal antibodies are widely used for treatment of multiple sclerosis but are associated with an impaired response to vaccines.Objective: To identify factors associated with a favorable vaccine response to tozinameran.Design, Setting, and Participants: This prospective cohort study was conducted in a specialized multiple sclerosis clinic at a university hospital from January 21 to December 1, 2021. Of 75 patients evaluated for participation who received a diagnosis of multiple sclerosis with planned or ongoing treatment with rituximab, 69 were included in the study, and data from 67 were analyzed.Exposures: Sex, age, number of previous rituximab infusions, accumulated dose of rituximab, previous COVID-19 infection, time since last rituximab treatment, CD19+ B-cell count before vaccination, CD4+ T-cell count, and CD8+ T-cell count were considered potential factors associated with the main outcome.Main Outcomes and Measures: Serological vaccine responses were measured by quantitation of anti-spike immunoglobulin G (IgG) antibodies, anti-receptor-binding domain (RBD) IgG antibodies, and their neutralizing capacities. Cellular responses to spike protein-derived SARS-CoV-2 peptide pools were assessed by counting interferon gamma spot-forming units in a FluoroSpot assay.Results: Among 60 patients with ongoing rituximab treatment (49 women [82%]; mean (SD) age, 43 [10] years), the median (range) disease duration was 9 (1-29) years, and the median (range) dose of rituximab was 2750 (500-10 000) mg during a median (range) time of 2.8 (0.5-8.3) years. The median (range) follow-up from the first vaccination dose was 7.3 (4.3-10.0) months. Vaccine responses were determined before vaccination with tozinameran and 6 weeks after vaccination. By using established cutoff values for anti-spike IgG (264 binding antibody units/mL) and anti-RBD IgG (506 binding antibody units/mL), the proportion of patients with a positive response increased with the number of B cells, which was the only factor associated with these outcomes. A cutoff for the B-cell count of at least 40/μL was associated with an optimal serological response. At this cutoff, 26 of 29 patients (90%) had positive test results for anti-spike IgG and 21 of 29 patients (72%) for anti-RBD IgG, and 27 of 29 patients (93%) developed antibodies with greater than 90% inhibition of angiotensin-converting enzyme 2. No factor associated with the cellular response was identified. Depending on the peptide pool, 21 of 25 patients (84%) to 22 of 25 patients (88%) developed a T-cell response with interferon gamma production at the B-cell count cutoff of at least 40/μL.Conclusions and Relevance: This cohort study found that for an optimal vaccine response from tozinameran, rituximab-treated patients with multiple sclerosis may be vaccinated as soon as possible, with rituximab treatment delayed until B-cell counts have reached at least 40/μL. An additional vaccination with tozinameran should be considered at that point.
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