Fluids and Barriers of the CNS最新文献

Background: Heterozygous CWH43 loss-of-function (LOF) variants have been identified as iNPH risk factors, with 10-15% of iNPH patients carrying these variants in cohorts from the US. Mouse model harboring CWH43 LOF variants display a hydrocephalic phenotype with ventricular cilia alterations. Our aim was to study the effect of CWH43 variants on disease risk and clinical phenotype in Finnish and Norwegian iNPH cohorts.

Methods: We analyzed CWH43 LOF frameshift deletions (4:49032652 CA/C, Leu533Ter and 4:49061875 CA/C, Lys696AsnfsTer23) in Finnish iNPH patients from the Kuopio NPH registry (n = 630) and FinnGen (iNPH n = 1 131, controls n = 495 400), and Norwegian iNPH patients from EADB (n = 306). The Kuopio and Norwegian cohorts included possible and probable iNPH patients based on the American-European iNPH guidelines. FinnGen cohort included iNPH patients based on ICD-10 G91.2 with the exclusion of secondary etiologies, and controls having no diagnosis of hydrocephalus.

Results: In the Kuopio cohort of Finnish iNPH patients, 2.9% carried CWH43 variants (Leu533Ter 2.1%, Lys696AsnfsTer23 0.8%), with one homozygous Leu533Ter carrier. In FinnGen, 3.1% of iNPH patients carried heterozygous variants (Leu533Ter 2.6%, Lys696AsnfsTer23 0.5%) compared to 2.5% of controls (p = 0.219, OR = 1.23, 95% CI 0.85-1.72), with no effect on disease risk or onset age. Importantly in the FinnGen cohort, none of the 23 compound heterozygote or 59 homozygote individuals had hydrocephalus diagnosis. In the Norwegian iNPH cohort, 5.2% of patients were heterozygous variant carriers (Leu533Ter 3.3%, Lys696AsnfsTer23 2.0%). No differences in clinical phenotype (age, triad symptoms, shunt response, vascular comorbidities) were found between carriers and noncarriers in any cohort. However, 74% of variant-carrying iNPH patients in FinnGen were female, compared to 47% of noncarriers (p = 0.002). Pedigrees indicated no autosomal dominant co-inheritance of iNPH and the CWH43 variants.

Conclusions: We studied the iNPH-associated CWH43 LOF variants for the first time on a population-scale. Contrary to previously reported findings in smaller cohorts, our study revealed a low prevalence of these variants in the population-scale Finnish iNPH cohort, with no effect on disease risk of iNPH. The prevalence in the Norwegian iNPH cohort was also low compared to previous studies.

Hydrocephalus, characterized by ventriculomegaly due to cerebrospinal fluid accumulation in the cerebral ventricles, is a co-morbidity factor in several neurodevelopmental, psychiatric and neurodegenerative diseases. Aquaporin-4 (AQP4) is crucial for brain water homeostasis, with Aqp4 knockout mice showing sporadic ventriculomegaly and increased brain water content. Kinase D interacting substrate of 220 kDa (Kidins220), a transmembrane protein involved in neuronal survival, synaptic activity and neurogenesis, controls AQP4 levels in ependymocytes and brain astrocytes. Indeed, Kidins220 deficiency in mice leads to hydrocephalus by downregulating VPS35, a key component of the retromer complex, and targeting AQP4 to lysosomal degradation. Importantly, the ependymal barrier of idiopathic normal pressure hydrocephalus patients shows a similar downregulation of KIDINS220 and AQP4. In addition, pathogenic variants in the KIDINS220 gene are linked to SINO syndrome, a rare disorder characterized by spastic paraplegia, intellectual disability, nystagmus, and obesity associated with hydrocephalus and ventriculomegaly. Given the retina's structural and functional similarities to the brain, we hypothesized that Kidins220 deficiency would affect retinal water regulation. However, the diminished expression of Kidins220 and VPS35 in the retina of Kidins220-deficient hydrocephalus mice, did not cause edema or downregulate AQP4 in Müller cells. Surprisingly, there was an increase in AQP4 levels within this glial cell population. Conversely, AQP4 expression in the optic nerve astrocytes was reduced, as observed in brain astrocytes, suggesting a distinctive adaptive response to hydrocephalus in Müller glia within the Kidins220-deficient retina. Furthermore, we observed phenotypic modifications in retinal glia in Kidins220-deficient hydrocephalus mice. However, we did not find any signs of neuronal damage in the retina. Future studies using OCT and OCTA in SINO syndrome patients with ventriculomegaly will be essential in elucidating the relationship between KIDINS220 pathogenic variants, retinal alterations, papilledema, and visual function.

Background: The choroid plexus (CP) has been recently implicated in the pathogenesis of the neuropsychiatric manifestations of systemic lupus erythematosus (NPSLE). Lupus patients demonstrate increased serum and cerebrospinal fluid (CSF) concentrations of interleukin-6 (IL-6), which can disrupt vital blood-CSF barrier (B-CSFB) functions performed by the CP. However, difficulty accessing this tissue has largely precluded dynamic imaging or evaluation of CP barrier function in vivo.

Methods: In this study, explant CP spheroids which replicate the functional and structural properties of the B-CSFB were generated from 12 + week old female MRL/lpr (IL-6 wildtype; IL-6 WT) lupus mice, IL-6 knockout (IL-6 KO) MRL/lpr mice, and congenic control MRL/mpj mice. CP spheroids derived from IL-6 WT MRL/lpr mice were found to synthesize and secrete IL-6, similar to the CP in vivo, whereas the IL-6 KO spheroids did not produce IL-6. Accumulation of different fluorescent tracers within the central CSF-like fluid vacuole of spheroids, modeling brain ventricles, was measured to probe transcellular permeability, paracellular diffusion, and clearance functions of the CP.

Results: As shown by blocking the IL-6 receptor in IL-6 WT spheroids or comparing them to IL-6 KO spheroids, IL-6 signaling decreased spheroid clearance of methotrexate, a chemotherapeutic drug employed in the therapy of lupus, and lucifer yellow. This suppression occurred without altering CP epithelial morphology and ultrastructure. Methotrexate and lucifer yellow efflux can occur through ATP-binding cassette (ABC) transporters, including BCRP and MRP1. Cytoplasmic accumulation of the ABC-specific dye fluorescein diacetate was also increased by IL-6. Pharmacologic inhibition of either BCRP or MRP1 in IL-6 KO spheroids was sufficient to recreate the clearance deficits observed in IL-6 WT spheroids. Moreover, CP expression of BCRP was significantly lower in IL-6 WT mice.

Conclusions: In this study, we establish, validate, and apply a CP spheroid model to the study of B-CSFB function in lupus. Our results show that IL-6, a key cytokine increased in NPSLE, can potentially suppress the CP-specific function and expression of BCRP and MRP1. Therefore, IL-6 could affect the CSF clearance of inflammatory substrates (e.g., leukotrienes), the accumulation of which would incite neurotoxicity and promote progression of NPSLE.

Streptococcus suis is a globally emerging zoonotic pathogen that can cause invasive disease commonly associated with meningitis in pigs and humans. To cause meningitis, S. suis must invade the central nervous system (CNS) by crossing the neurovascular unit, also known as the blood-brain barrier (BBB), or vascularized choroid plexus (ChP) epithelium known as the blood-cerebrospinal fluid barrier (BCSFB). Recently developed ChP organoids have been shown to accurately replicate the cytoarchitecture and physiological functions of the ChP epithelium in vivo. Here, we used human induced pluripotent stem cells (iPSC)-derived ChP organoids as an in vitro model to investigate S. suis interaction and infection at the BCSFB. Our study revealed that S. suis is capable of translocating across the epithelium of ChP organoids without causing significant cell death or compromising the barrier integrity. Plasminogen (Plg) binding to S. suis in the presence of tissue plasminogen activator (tPA), which converts immobilized Plg to plasmin (Pln), significantly increased the basolateral to apical translocation across ChP organoids into the CSF-like fluid in the lumen. S. suis was able to replicate at the same rate in CSF and laboratory S. suis culture medium but reached a lower final density. The analysis of transcriptomes in ChP organoids after S. suis infection indicated inflammatory responses, while the addition of Plg further suggested extracellular matrix (ECM) remodeling. To our knowledge, this is the first study using ChP organoids to investigate bacterial infection of the BCSFB. Our findings highlight the potential of ChP organoids as a valuable tool for studying the mechanisms of bacterial interaction and infection of the human ChP in vitro.

Background: Patients with the genetic disorder Niemann-Pick type C2 disease (NP-C2) suffer from lysosomal accumulation of cholesterol causing both systemic and severe neurological symptoms. In a murine NP-C2 model, otherwise successful intravenous Niemann-Pick C2 protein (NPC2) replacement therapy fails to alleviate progressive neurodegeneration as infused NPC2 cannot cross the blood-brain barrier (BBB). Genetic modification of brain endothelial cells (BECs) is thought to enable secretion of recombinant proteins thereby overcoming the restrictions of the BBB. We hypothesized that an adeno-associated virus (AAV-BR1) encoding the Npc2 gene could cure neurological symptoms in Npc2-/- mice through transduction of BECs, and possibly neurons via viral passage across the BBB.

Methods: Six weeks old Npc2-/- mice were intravenously injected with the AAV-BR1-NPC2 vector. Composite phenotype scores and behavioral tests were assessed for the following 6 weeks and visually documented. Post-mortem analyses included gene expression analyses, verification of neurodegeneration in Purkinje cells, determination of NPC2 transduction in the CNS, assessment of gliosis, quantification of gangliosides, and co-detection of cholesterol with NPC2 in degenerating neurons.

Results: Treatment with the AAV-BR1-NPC2 vector improved motor functions, reduced neocortical inflammation, and preserved Purkinje cells in most of the mice, referred to as high responders. The vector exerted tropism for BECs and neurons resulting in a widespread NPC2 distribution in the brain with a concomitant reduction of cholesterol in adjacent neurons, presumably not transduced by the vector. Mass spectrometry imaging revealed distinct lipid alterations in the brains of Npc2-/- mice, with increased GM2 and GM3 ganglioside accumulation in the cerebellum and hippocampus. AAV-BR1-NPC2 treatment partially normalized these ganglioside distributions in high responders, including restoration of lipid profiles towards those of Npc2+/+ controls.

Conclusion: The data suggests cross-correcting gene therapy to the brain via delivery of NPC2 from BECs and neurons.

Primary ciliary dyskinesia (PCD) is a genetic disorder causing motile ciliary dysfunction primarily affecting the respiratory and reproductive systems. However, the impact of PCD on the central nervous system remains poorly understood. Rodent models of PCD exhibit marked hydrocephalus leading to early animal mortality, however, most humans with PCD do not develop hydrocephalus for unknown reasons. We hypothesized that patients with PCD exhibit sub-clinical ventriculomegaly related to ependymal motile ciliary dysfunction. We demonstrated highly specific expression levels of known PCD-related genes in human brain multiciliated ependymal cells (p < 0.0001). To assess ventricular size, computed tomography sinus images from patients with PCD (n = 33) and age/sex-matched controls (n = 64) were analysed. Patients with PCD displayed significantly larger ventricular areas (p < 0.0001) and Evans index (p < 0.01), indicating ventriculomegaly that was consistent across all genetic subgroups. Ventricular enlargement correlated positively with increasing age in patients with PCD compared to controls (p < 0.001). Additionally, chart review demonstrated a high prevalence (39%) of neuropsychiatric/neurological disorders in adult PCD patients that did not correlate with degree of ventriculomegaly. Our findings suggest that patients with PCD may have unrecognized, mild ventriculomegaly which correlates with ageing, potentially attributable to ependymal ciliary dysfunction. Further study is required to determine causality, and whether ventricular enlargement contributes to neuropsychiatric/neurological or other morbidity in PCD.

Background: Therapeutic antibodies for the treatment of neurological disease show great potential, but their applications are rather limited due to limited brain exposure. The most well-studied approach to enhance brain influx of protein therapeutics, is receptor-mediated transcytosis (RMT) by targeting nutrient receptors to shuttle protein therapeutics over the blood-brain barrier (BBB) along with their endogenous cargos. While higher brain exposure is achieved with RMT, the timeframe is short due to rather fast brain clearance. Therefore, we aim to increase the brain half-life of antibodies by binding to myelin oligodendrocyte glycoprotein (MOG), a CNS specific protein.

Methods: Alpaca immunization with mouse/human MOG, and subsequent phage selections and screenings for MOG binding single variable domain antibodies (VHHs) were performed to find mouse/human cross-reactive VHHs. Their ability to increase the brain half-life of antibodies was evaluated in healthy wild-type mice by coupling two different MOG VHHs (low/high affinity) in a mono- and bivalent format to a β-secretase 1 (BACE1) inhibiting antibody or a control (anti-SARS-CoV-2) antibody, fused to an anti-transferrin receptor (TfR) VHH for active transport over the BBB. Brain pharmacokinetics and pharmacodynamics, CNS and peripheral biodistribution, and brain toxicity were evaluated after intravenous administration to balb/c mice.

Results: Additional binding to MOG increases the C_max and brain half-life of antibodies that are actively shuttled over the BBB. Anti-SARS-CoV-2 antibodies coupled with an anti-TfR VHH and two low affinity anti-MOG VHHs could be detected in brain 49 days after a single intravenous injection, which is a major improvement compared to an anti-SARS-CoV-2 antibody fused to an anti-TfR VHH which cannot be detected in brain anymore one week post treatment. Additional MOG binding of antibodies does not affect peripheral biodistribution but alters brain distribution to white matter localization and less neuronal internalization.

Conclusions: We have discovered mouse/human/cynomolgus cross-reactive anti-MOG VHHs which have the ability to drastically increase brain exposure of antibodies. Combining MOG and TfR binding leads to distinct PK, biodistribution, and brain exposure, differentiating it from the highly investigated TfR-shuttling. It is the first time such long brain antibody exposure has been demonstrated after one single dose. This new approach of adding a binding moiety for brain specific targets to RMT shuttling antibodies is a huge advancement for the field and paves the way for further research into brain half-life extension.

Background: Acute and critical neurological diseases are often accompanied with elevated intracranial pressure (ICP), leading to insufficient cerebral perfusion, which may cause severe secondary lesion. Existing ICP monitoring techniques often fail to effectively meet the demand for real-time noninvasive ICP monitoring and warning. This study aimed to explore the use of electrical impedance tomography (EIT) to provide real-time early warning of elevated ICP by observing cerebral perfusion.

Methods: An intracranial hypertension model was prepared by injecting autologous un-anticoagulated blood into the brain parenchyma of twelve Landrace swine. Invasive ICP monitoring was used as a control method, and a high-precision EIT system was used to acquire and analyze the changing patterns of cerebral perfusion EIT image parameters with respect to ICP. Four EIT parameters related to cerebral perfusion were extracted from the images, and their potential application in detecting ICP elevation was analyzed.

Results: When ICP increased, all EIT perfusion parameters decreased significantly (P < 0.05). When the subjects were in a state of intracranial hypertension (ICP > 22 mmHg), the correlation between EIT perfusion parameters and ICP was more significant (P < 0.01), with correlation coefficients ranging from -0.72 to -0.83. We tested the objects when they were in baseline ICP and in ICP of 15-40 mmHg. Under both circumstances, ROC curve analysis showed that the comprehensive model of perfusion parameters based on the random forest algorithm had a sensitivity and specificity of more than 90% and an area under the curve (AUC) of more than 0.9 for detecting ICP increments of both 5 and 10 mmHg.

Conclusion: This study demonstrates the feasibility of using perfusion EIT to detect ICP increases in real time, which may provide a new method for real-time non-invasive monitoring of patients with increased ICP.

Background: The approval of new disease-modifying therapies by the U.S. Food and Drug Administration and the European Medicine Agency makes it necessary to optimize non-invasive and cost-effective tools for the identification of subjects at-risk of developing Alzheimer's Disease (AD). Plasma biomarkers are excellent candidates. However, their ability to reflect the cerebrospinal fluid (CSF) profile - that remains to date the gold standard for the biochemical diagnosis of AD - needs to be confirmed and validated before their implementation in clinical practice. The aims of this study are to analyse the correlation between CSF and plasma Aβ40, Aβ42, Aβ42/Aβ40 and pTau181, and to assess the diagnostic performance of plasma biomarkers in a cohort of subjects affected by Mild Cognitive Impairment (MCI).

Methods: The study was performed on 306 subjects affected by MCI, enrolled in the context of the Italian Interceptor Project. Aβ40, Aβ42 and pTau181 were analysed in plasma and CSF, and pTau217 was measured in plasma. The fully automated chemiluminescence enzyme immunoassay and the Lumipulse^® G600II (Fujirebio) instrument were used for all measurements. We analysed the correlations between CSF and plasma biomarkers and the differences of plasma biomarker concentrations after grouping MCI cases according to AT classification of CSF AD biomarker profiles.

Results: We found statistically significant positive correlations between CSF and plasma Aβ42, Aβ42/Aβ40 ratio and pTau181. All the biomarkers, except Aβ40, showed differences in A+ vs. A-, A+T+ vs. A-T- and A+T- vs. A-T- patients. Moreover, Aβ42 and Aβ42/Aβ40 plasma levels were lower in A+T- compared to A-T- and A-T+ groups, and pTau181 and pTau217 plasma levels were higher in A+T+ compared to A+T-. Aβ42/Aβ40 and pTau217 showed a robust performance in distinguishing A+ from A- (AUC = 0.857 and 0.862, respectively) and A+T+ from A-T- (AUC = 0.866 and 0.911) subjects.

Conclusions: Our results suggest that plasma biomarkers, and especially Aβ42/Aβ40 ratio and pTau217, are promising candidates for the early detection of AD pathology.

Oxidative stress and neuronal apoptosis could be an important factor leading to post-hemorrhagic consequences after germinal matrix hemorrhage (GMH). Previously study have indicated that relaxin 2 receptor activation initiates anti-oxidative stress and anti-apoptosis in ischemia-reperfusion injury. However, whether relaxin 2 activation can attenuate oxidative stress and neuronal apoptosis after GMH remains unknown. To investigate the beneficial effect of relaxin 2 on oxidative stress injury and neuronal apoptosis by GMH, a total of 150 rat pups were subjected to GMH by an intraparenchymal injection of bacterial collagenase. Recombinant human relaxin-2 (rh-relaxin-2) was administered intraperitoneally injections at 1 h and 13 h after GMH. Lenti-virus with sgRXFP1 and sgCtrl was administered intracerebroventricular (i.c.v.) on the left side of the brain to inhibit the RXFP1 at 2d prior to GMH induction, and LY321499, ERK inhibitor, was administered by i.c.v. injection at 1 h on the left side of the brain prior to GMH induction, respectively. Co-immunoprecipitation, immunofluorescence, TUNEL, Fluoro-Jade C, DHE staining, western blot, Nitrix Oxide (NO) quantification and side effect experiments were performed to evaluate post-GMH. We found endogenous relaxin-2 interacts with RXFP1 and both protein colocalized in neurons on the first day after GMH. Additionally, RXFP1 activation with rh-relaxin-2 significantly inhibited oxidative stress and neuronal apoptosis in GMH + rh-relaxin-2 group compared with GMH + vehicle group. Moreover, rh-relaxin-2 treatment significantly inhibited the phosphorylation of ERK and nNOS, as well as upregulated expression of Bcl2 and NO and downregulated expression of Bax and Romo 1. The beneficial effects of rh-relaxin-2 were reversed by i.c.v. injection of lenti-virus with sgRXFP1 and LY321499, respectively. Furthermore, the side effect experiment showed rh-relaxin-2 did not affect neurological behavior and the function of liver and kidney. In conclusion, our finding showed that rh-relaxin-2 attenuated oxidative stress and neuronal apoptosis after GMH through RXFP1-ERK-nNOS-NO signaling pathway.