Age-induced alterations of granulopoiesis generate atypical neutrophils that aggravate stroke pathology.

Aging accounts for increased risk and dismal outcome of ischemic stroke. Here, we investigated the impact of age-related changes in the immune system on stroke. Upon experimental stroke, compared with young mice, aged mice had increased neutrophil clogging of the ischemic brain microcirculation, leading to worse no-reflow and outcomes. Aged mice showed an enhanced granulopoietic response to stroke that led to the accumulation of CD101+CD62Llo mature and CD177hiCD101loCD62Llo and CD177loCD101loCD62Lhi immature atypical neutrophils in the blood, endowed with increased oxidative stress, phagocytosis and procoagulant features. Production of CXCL3 by CD62Llo neutrophils of the aged had a key role in the development and pathogenicity of aging-associated neutrophils. Hematopoietic stem cell rejuvenation reverted aging-associated neutropoiesis and improved stroke outcome. In elderly patients with ischemic stroke, single-cell proteome profile of blood leukocytes identified CD62Llo neutrophil subsets associated with worse reperfusion and outcome. Our results unveil how stroke in aging leads to a dysregulated emergency granulopoiesis impacting neurological outcome.

COVID-19-associated serum and cerebrospinal fluid cytokines in post- versus para-infectious SARS-CoV-2-related Guillain-Barré syndrome.

INTRODUCTION: Guillain-Barré syndrome associated with Coronavirus-2-related severe acute respiratory syndrome (COV-GBS) occurs as para- or post-infectious forms, depending on the timing of disease onset. In these two forms, we aimed to compare the cerebrospinal fluid (CSF) and serum proinflammatory cytokine profiles to evaluate differences that could possibly have co-pathogenic relevance. MATERIALS AND METHODS: We studied a retrospective cohort of 26 patients with either post-COV-GBS (n = 15), with disease onset occurring > 7 days after SARS-CoV-2 infection, or para-COV-GBS (n = 11), with disease onset 7 days or less. TNF-α, IL-6, and IL-8 were measured in the serum with SimplePlex™ Ella™ immunoassay. In addition to the para-/post-COV-GBS patients, serum levels of these cytokines were determined in those with non-COVID-associated-GBS (NC-GBS; n = 43), paucisymptomatic SARS-CoV-2 infection without GBS (COVID, n = 20), and in healthy volunteers (HV; n = 12). CSF cytokine levels were measured in patients with para-/post-COV-GBS, in those with NC-GBS (n = 29), or with Alzheimer's disease (AD; n = 24). RESULTS: Serum/CSF cytokine levels did not differ in para- vs post-COV-GBS. We found that SARS-CoV-2 infection raises the serum levels of TNF-α, IL-6, and IL-8, as well as an increase of IL-6 (in serum and CSF) and IL-8 (in CSF) in either NC-GBS or COV-GBS than controls. CSF and serum cytokine levels resulted independent one with another. CONCLUSIONS: The change of cytokines linked to SARS-CoV-2 in COV-GBS appears to be driven by viral infection, although it has unique characteristics in GBS as such and does not account for cases with para- or post-infectious onset.

Neuroprotective Potential of Dendritic Cells and Sirtuins in Multiple Sclerosis.

Myeloid cells, including parenchymal microglia, perivascular and meningeal macrophages, and dendritic cells (DCs), are present in the central nervous system (CNS) and establish an intricate relationship with other cells, playing a crucial role both in health and in neurological diseases. In this context, DCs are critical to orchestrating the immune response linking the innate and adaptive immune systems. Under steady-state conditions, DCs patrol the CNS, sampling their local environment and acting as sentinels. During neuroinflammation, the resulting activation of DCs is a critical step that drives the inflammatory response or the resolution of inflammation with the participation of different cell types of the immune system (macrophages, mast cells, T and B lymphocytes), resident cells of the CNS and soluble factors. Although the importance of DCs is clearly recognized, their exact function in CNS disease is still debated. In this review, we will discuss modern concepts of DC biology in steady-state and during autoimmune neuroinflammation. Here, we will also address some key aspects involving DCs in CNS patrolling, highlighting the neuroprotective nature of DCs and emphasizing their therapeutic potential for the treatment of neurological conditions. Recently, inhibition of the NAD+-dependent deac(et)ylase sirtuin 6 was demonstrated to delay the onset of experimental autoimmune encephalomyelitis, by dampening DC trafficking towards inflamed LNs. Thus, a special focus will be dedicated to sirtuins' role in DCs functions.

Mesenchymal stem cells instruct a beneficial phenotype in reactive astrocytes.

Transplanted mesenchymal stromal/stem cells (MSC) ameliorate the clinical course of experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis (MS), reducing inflammation and demyelination. These effects are mediated by instructive cross-talk between MSC and immune and neural cells. Astroglial reaction to injury is a prominent feature of both EAE and MS. Astrocytes constitute a relevant target to control disease onset and progression and, based on their potential to acquire stem cell properties in situ, to foster recovery in the post-acute phase of pathology. We have assessed how MSC impact astrocytes in vitro and ex vivo in EAE. Expression of astroglial factors implicated in EAE pathogenesis was quantified by real-time PCR in astrocytes co-cultured with MSC or isolated from EAE cerebral cortex; astrocyte morphology and expression of activation markers were analyzed by confocal microscopy. The acquisition of neural stem cell properties by astrocytes was evaluated by neurosphere assay. Our study shows that MSC prevented astrogliosis, reduced mRNA expression of inflammatory cytokines that sustain immune cell infiltration in EAE, as well as protein expression of endothelin-1, an astrocyte-derived factor that inhibits remyelination and contributes to neurodegeneration and disease progression in MS. Moreover, our data reveal that MSC promoted the acquisition of progenitor traits by astrocytes. These data indicate that MSC attenuate detrimental features of reactive astroglia and, based on the reacquisition of stem cell properties, also suggest that astrocytes may be empowered in their protective and reparative actions by MSC.

IFN-γ orchestrates mesenchymal stem cell plasticity through the signal transducer and activator of transcription 1 and 3 and mammalian target of rapamycin pathways.

BACKGROUND: Mesenchymal stem cells (MSCs) display a therapeutic plasticity because of their ability to modulate immunity, foster tissue repair, and differentiate into mesodermal cells. IFN-γ has been described to differently affect human mesenchymal stem cell (hMSC) and mouse mesenchymal stem cell (mMSC) immunomodulation and differentiation, depending on the inflammatory milieu. OBJECTIVE: We aimed at dissecting the relevant intracellular pathways through which IFN-γ affects MSC plasticity and the consequence of their manipulation on MSC functions. METHODS: Modification of relevant IFN-γ-dependent pathways in mMSCs was carried out in vitro through gene silencing or chemical inhibition of key components. Functional outcomes were assessed by means of Western blotting, real-time PCR, differentiation, and proliferation assays on MSCs. The effect on T cells was addressed by T-cell proliferation assays; the effect of mammalian target of rapamycin (mTOR) manipulation in MSCs was studied in vivo in a mouse model of delayed-type hypersensitivity assay. To address whether similar mechanisms are involved also in hMSCs on IFN-γ stimulation, the effect of chemical inhibition on the same intracellular pathways was assessed by means of Western blotting, and the final outcome on immunomodulatory properties was evaluated based on real-time PCR and T-cell proliferation. RESULTS: We revealed that in mMSCs IFN-γ-induced immunoregulation is mediated by early phosphorylation of signal transducer and activator of transcription (STAT) 1 and STAT3, which is significantly enhanced by an extracellular signal-regulated kinase 1/2-dependent mTOR inhibition, thereby promoting pSTAT1 nuclear translocation. Accordingly, after intracellular mTOR inhibition, MSCs augmented their ability to inhibit T-cell proliferation and control delayed-type hypersensitivity in vivo. Similarly, on mTOR blockade, hMSCs also enhanced their immunoregulatory features. A sustained exposure to IFN-γ led to inhibition of STAT3 activity, which in mMSCs resulted in an impaired proliferation and differentiation. CONCLUSION: These results provide new insights about MSC intracellular pathways affected by IFN-γ, demonstrating that pharmacologic or genetic manipulation of MSCs can enhance their immunomodulatory functions, which could be translated into novel therapeutic approaches.

Nerve-Glial antigen 2: unmasking the enigmatic cellular identity in the central nervous system.

Chondroitin sulfate proteoglycans (CSPGs) are fundamental components of the extracellular matrix in the central nervous system (CNS). Among these, the Nerve-Glial antigen 2 (NG2) stands out as a transmembrane CSPG exclusively expressed in a different population of cells collectively termed NG2-expressing cells. These enigmatic cells, found throughout the developing and adult CNS, have been indicated with various names, including NG2 progenitor cells, polydendrocytes, synantocytes, NG2 cells, and NG2-Glia, but are more commonly referred to as oligodendrocyte progenitor cells. Characterized by high proliferation rates and unique morphology, NG2-expressing cells stand apart from neurons, astrocytes, and oligodendrocytes. Intriguingly, some NG2-expressing cells form functional glutamatergic synapses with neurons, challenging the long-held belief that only neurons possess the intricate machinery required for neurotransmission. In the CNS, the complexity surrounding NG2-expressing cells extends to their classification. Additionally, NG2 expression has been documented in pericytes and immune cells, suggesting a role in regulating brain innate immunity and neuro-immune crosstalk in homeostasis. Ongoing debates revolve around their heterogeneity, potential as progenitors for various cell types, responses to neuroinflammation, and the role of NG2. Therefore, this review aims to shed light on the enigma of NG2-expressing cells by delving into their structure, functions, and signaling pathways. We will critically evaluate the literature on NG2 expression across the CNS, and address the contentious issues surrounding their classification and roles in neuroinflammation and neurodegeneration. By unraveling the intricacies of NG2-expressing cells, we hope to pave the way for a more comprehensive understanding of their contributions to CNS health and during neurological disorders.

Cerebrospinal fluid NPTX2 changes and relationship with regional brain metabolism metrics across mild cognitive impairment due to Alzheimer's disease.

BACKGROUND: Neuronal pentraxin-2 (NPTX2), crucial for synaptic functioning, declines in cerebrospinal fluid (CSF) as cognition deteriorates. The variations of CSF NPTX2 across mild cognitive impairment (MCI) due to Alzheimer's disease (AD) and its association with brain metabolism remain elusive, albeit relevant for patient stratification and pathophysiological insights. METHODS: We retrospectively analyzed 49 MCI-AD patients grouped by time until dementia (EMCI, n = 34 progressing within 2 years; LMCI, n = 15 progressing later/stable at follow-up). We analyzed demographic variables, cognitive status (MMSE score), and CSF NPTX2 levels using a commercial ELISA assay in EMCI, LMCI, and a control group of age-/sex-matched individuals with other non-dementing disorders (OND). Using [18F]FDG PET scans for voxel-based analysis, we explored correlations between regional brain metabolism metrics and CSF NPTX2 levels in MCI-AD patients, accounting for age. RESULTS: Baseline and follow-up MMSE scores were lower in LMCI than EMCI (p value = 0.006 and p < 0.001). EMCI exhibited significantly higher CSF NPTX2 values than both LMCI (p = 0.028) and OND (p = 0.006). We found a significant positive correlation between NPTX2 values and metabolism of bilateral precuneus in MCI-AD patients (p < 0.005 at voxel level, p < 0.05 with family-wise error correction at the cluster level). CONCLUSIONS: Higher CSF NPTX2 in EMCI compared to controls and LMCI suggests compensatory synaptic responses to initial AD pathology. Disease progression sees these mechanisms overwhelmed, lowering CSF NPTX2 approaching dementia. Positive CSF NPTX2 correlation with precuneus glucose metabolism links to AD-related metabolic changes across MCI course. These findings posit CSF NPTX2 as a promising biomarker for both AD staging and progression risk stratification.

Exploring the brain metabolic correlates of process-specific CSF biomarkers in patients with MCI due to Alzheimer's disease: preliminary data.

We explored the brain metabolism correlates of emergent cerebrospinal fluid (CSF) biomarkers in a group of 26 patients with prodromal Alzheimer's disease (AD). Distinct volumes of interest (VOIs) expressed the sites of correlation between CSF biomarkers and brain metabolism as determined on [18F]FDG-PET images, as well as of significant hypometabolism in patients compared to healthy controls. Neurogranin- and α-synuclein-VOIs included left precuneus and/or posterior cingulate cortex (PC and/or PCC) and partially overlapped hypometabolism at those sites. ß-synuclein- and neurofilament light chain (NfL)-VOIs regarded either left or right lateral temporal areas, respectively, with partial overlap with hypometabolism only for the ß-synuclein-VOI, whereas the NfL-VOI did not include hypometabolic regions. We speculate that CSF neurogranin and α-synuclein express an already established hippocampal damage leading to PC and/or PCC deafferentation and hypometabolism. ß-synuclein may represent the progression of synaptopathy in the temporal lobe, while NfL the axonal injury in right temporal regions where neuronal loss is not yet evident.

Longitudinal Cluster Analysis of Hemodialysis Patients with COVID-19 in the Pre-Vaccination Era.

Coronavirus disease 2019 (COVID-19) in hemodialysis patients (HD) is characterized by heterogeneity of clinical presentation and outcomes. To stratify patients, we collected clinical and laboratory data in two cohorts of HD patients at COVID-19 diagnosis and during the following 4 weeks. Baseline and longitudinal values were used to build a linear mixed effect model (LME) and define different clusters. The development of the LME model in the derivation cohort of 17 HD patients (66.7 ± 12.3 years, eight males) allowed the characterization of two clusters (cl1 and cl2). Patients in cl1 presented a prevalence of females, higher lymphocyte count, and lower levels of lactate dehydrogenase, C-reactive protein, and CD8 + T memory stem cells as a possible result of a milder inflammation. Then, this model was tested in an independent validation cohort of 30 HD patients (73.3 ± 16.3 years, 16 males) assigned to cl1 or cl2 (16 and 14 patients, respectively). The cluster comparison confirmed that cl1 presented a milder form of COVID-19 associated with reduced disease activity, hospitalization, mortality rate, and oxygen requirement. Clustering analysis on longitudinal data allowed patient stratification and identification of the patients at high risk of complications. This strategy could be suitable in different clinical settings.

Impact of Natural Killer (NK) Cells on Immune Reconstitution, and Their Potential as a Biomarker of Disease Activity, in Alemtuzumab-Treated Patients with Relapsing Remitting Multiple Sclerosis: An Observational Study.

BACKGROUND: Defining immune mechanisms leading to multiple sclerosis (MS) is difficult, due to the great inter-individual difference in immune system responses. The anti-CD52 antibody alemtuzumab transiently abolishes differences in immune parameters among individuals, allowing analysis of subsequent immune cell repopulation patterns, and their possible role in MS. OBJECTIVE: To evaluate the correlation between innate and adaptive immune cell subsets and disease activity in MS in the context of treatment with alemtuzumab. METHODS: A two-center observational cohort of patients treated with alemtuzumab underwent immune profiling of T, B, and natural killer (NK) cells, biomarker, clinical and radiological follow-up. RESULTS: After treatment, the percentage of NK and B cells increased; NK, T- and B-cell populations underwent a profound rearrangement. Within the effector T-cell compartment, treatment led to a transient decrease, followed by an increase, of T-helper 1 cells, and to a transient decrease of T-helper 17 cells. Within the T-regulatory compartment, naïve T-regulatory cells increased. Within the B-cell compartment, memory B cells and mature B cells decreased, whereas transitional B cells increased. Within the NK cell compartment, CD56bright NK cells increased. Subjects without disease activity had a greater decrease in serum NfL and greater NK cell/CD3+ T cell ratio. NK cell numbers at baseline and after treatment influenced reconstitution of T and B cells, being inversely correlated with the reconstitution of proinflammatory CD3+ T cells and mature B cells, and directly correlated to the increase in transitional B cells. CONCLUSIONS: The results of this study provide novel evidence that NK cells influence reconstitution of adaptive immune cells upon alemtuzumab and that patients with a successful response to alemtuzumab have an early immune reconstitution dominated by NK cells.