Common signatures of differential microRNA expression in Parkinson's and Alzheimer's disease brains.

Dysregulation of microRNA gene expression has been implicated in many neurodegenerative diseases, including Parkinson's disease. However, the individual dysregulated microRNAs remain largely unknown. Previous meta-analyses have highlighted several microRNAs being differentially expressed in post-mortem Parkinson's disease and Alzheimer's disease brains versus controls, but they were based on small sample sizes. In this study, we quantified the expression of the most compelling Parkinson's and Alzheimer's disease microRNAs from these meta-analyses ('candidate miRNAs') in one of the largest Parkinson's/Alzheimer's disease case-control post-mortem brain collections available (n = 451), thereby quadruplicating previously investigated sample sizes. Parkinson's disease candidate microRNA hsa-miR-132-3p was differentially expressed in our Parkinson's (P = 4.89E-06) and Alzheimer's disease samples (P = 3.20E-24) compared with controls. Alzheimer's disease candidate microRNAs hsa-miR-132-5p (P = 4.52E-06) and hsa-miR-129-5p (P = 0.0379) were differentially expressed in our Parkinson's disease samples. Combining these novel data with previously published data substantially improved the statistical support (α = 3.85E-03) of the corresponding meta-analyses, clearly implicating these microRNAs in both Parkinson's and Alzheimer's disease. Furthermore, hsa-miR-132-3p/-5p (but not hsa-miR-129-5p) showed association with α-synuclein neuropathological Braak staging (P = 3.51E-03/P = 0.0117), suggesting that hsa-miR-132-3p/-5p play a role in α-synuclein aggregation beyond the early disease phase. Our study represents the largest independent assessment of recently highlighted candidate microRNAs in Parkinson's and Alzheimer's disease brains, to date. Our results implicate hsa-miR-132-3p/-5p and hsa-miR-129-5p to be differentially expressed in both Parkinson's and Alzheimer's disease, pinpointing shared pathogenic mechanisms across these neurodegenerative diseases. Intriguingly, based on publicly available high-throughput sequencing of RNA isolated by cross-linking immunoprecipitation data, hsa-miR-132 may interact with SNCA messenger RNA in the human brain, possibly pinpointing novel therapeutic approaches in fighting Parkinson's disease.

Multisystem screening reveals SARS-CoV-2 in neurons of the myenteric plexus and in megakaryocytes.

SARS-CoV-2, the causative agent of COVID-19, typically manifests as a respiratory illness, although extrapulmonary involvement, such as in the gastrointestinal tract and nervous system, as well as frequent thrombotic events, are increasingly recognised. How this maps onto SARS-CoV-2 organ tropism at the histological level, however, remains unclear. Here, we perform a comprehensive validation of a monoclonal antibody against the SARS-CoV-2 nucleocapsid protein (NP) followed by systematic multisystem organ immunohistochemistry analysis of the viral cellular tropism in tissue from 36 patients, 16 postmortem cases and 16 biopsies with polymerase chain reaction (PCR)-confirmed SARS-CoV-2 status from the peaks of the pandemic in 2020 and four pre-COVID postmortem controls. SARS-CoV-2 anti-NP staining in the postmortem cases revealed broad multiorgan involvement of the respiratory, digestive, haematopoietic, genitourinary and nervous systems, with a typical pattern of staining characterised by punctate paranuclear and apical cytoplasmic labelling. The average time from symptom onset to time of death was shorter in positively versus negatively stained postmortem cases (mean = 10.3 days versus mean = 20.3 days, p = 0.0416, with no cases showing definitive staining if the interval exceeded 15 days). One striking finding was the widespread presence of SARS-CoV-2 NP in neurons of the myenteric plexus, a site of high ACE2 expression, the entry receptor for SARS-CoV-2, and one of the earliest affected cells in Parkinson's disease. In the bone marrow, we observed viral SARS-CoV-2 NP within megakaryocytes, key cells in platelet production and thrombus formation. In 15 tracheal biopsies performed in patients requiring ventilation, there was a near complete concordance between immunohistochemistry and PCR swab results. Going forward, our findings have relevance to correlating clinical symptoms with the organ tropism of SARS-CoV-2 in contemporary cases as well as providing insights into potential long-term complications of COVID-19. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Alzheimer's disease pathology explains association between dementia with Lewy bodies and APOE-ε4/TOMM40 long poly-T repeat allele variants.

INTRODUCTION: The role of TOMM40-APOE 19q13.3 region variants is well documented in Alzheimer's disease (AD) but remains contentious in dementia with Lewy bodies (DLB) and Parkinson's disease dementia (PDD). METHODS: We dissected genetic profiles within the TOMM40-APOE region in 451 individuals from four European brain banks, including DLB and PDD cases with/without neuropathological evidence of AD-related pathology and healthy controls. RESULTS: TOMM 40-L/APOE-ε4 alleles were associated with DLB (OR TOMM40 -L = 3.61; P value = 3.23 × 10-9; OR APOE -ε4 = 3.75; P value = 4.90 × 10-10) and earlier age at onset of DLB (HR TOMM40 -L = 1.33, P value = .031; HR APOE -ε4 = 1.46, P value = .004), but not with PDD. The TOMM40-L/APOE-ε4 effect was most pronounced in DLB individuals with concomitant AD pathology (OR TOMM40 -L = 4.40, P value = 1.15 × 10-6; OR APOE - ε 4 = 5.65, P value = 2.97 × 10-8) but was not significant in DLB without AD. Meta-analyses combining all APOE-ε4 data in DLB confirmed our findings (ORDLB = 2.93, P value = 3.78 × 10-99; ORDLB+AD = 5.36, P value = 1.56 × 10-47). DISCUSSION: APOE-ε4/TOMM 40-L alleles increase susceptibility and risk of earlier DLB onset, an effect explained by concomitant AD-related pathology. These findings have important implications in future drug discovery and development efforts in DLB.

Post mortem examination of Parkinson's disease brains suggests decline in mitochondrial biomass, reversed by deep brain stimulation of subthalamic nucleus.

Deep brain stimulation (DBS) of the subthalamic nucleus (STN) is the most commonly used surgical treatment for Parkinson's disease (PD). The disease-modifying aspects of DBS at a cellular level are not fully understood, and the key question of the effect of DBS on the degeneration of the dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc) remains to be answered. A major technical hurdle in determining any neuroprotective effect by DBS is its use in mid- to late-stage patients with PD when a majority of the DA neurons have been lost. In this work, we hypothesized that the long-term clinical benefits of DBS are, at least in part, due to a neuromodulatory effect on the SNpc neurons. These changes would affect cellular energetics and mitochondrial metabolism. We examined the number and volume of mitochondria as well as their vicinity to the DA presynaptic terminals postmortem caudate and putamen of 3 healthy individuals, 4 PD cases, and 3 DBS-treated patients. PD seems to have caused an increase in the mean distance between mitochondria and presynaptic terminals as well as a decrease in mean mitochondrial volume and numbers in DA projections. Although there was no difference in distance between mitochondria and presynaptic terminals of SNpc neurons in PD brains vs. DBS-treated brains, DBS treatment seemed to have inhibited or reversed the reduction in mitochondrial volume and numbers caused by PD. These results suggest enhanced metabolic plasticity leading to neuroprotection in the SNpc as a result of STN-DBS.-Mallach, A., Weinert, M., Arthur, J., Gveric, D., Tierney, T. S., Alavian, K. N. Post mortem examination of Parkinson's disease brains suggests decline in mitochondrial biomass, reversed by deep brain stimulation of subthalamic nucleus.

Unveiling the olfactory proteostatic disarrangement in Parkinson's disease by proteome-wide profiling.

Olfactory dysfunction is one of the earliest features in Lewy-type alpha-synucleinopathies (LTSs) such as Parkinson's disease (PD). However, the underlying molecular mechanisms associated to smell impairment are poorly understood. Applying mass spectrometry-based quantitative proteomics in postmortem olfactory bulbs across limbic, early-neocortical, and neocortical LTS stages of parkinsonian patients, a proteostasis impairment, was observed, identifying 268 differentially expressed proteins between controls and PD phenotypes. In addition, network-driven proteomics revealed a modulation in ERK1/2, MKK3/6, and PDK1/PKC signaling axes. Moreover, a cross-disease study of selected olfactory molecules in sporadic Alzheimer's disease (AD) cases revealed different protein derangements in the modulation of secretagogin (SCGN), calcyclin-binding protein (CACYBP), and glucosamine 6 phosphate isomerase 2 (GNPDA2) between PD and AD. An inverse correlation between GNPDA2 and α-synuclein protein levels was also reflected in PD cerebrospinal fluid. Interestingly, PD patients exhibited significantly lower serum GNPDA2 levels than controls (n = 82/group). Our study provides important avenues for understanding the olfactory bulb proteostasis imbalance in PD, deciphering mechanistic clues to the equivalent smell deficits observed in AD and PD pathologies.

Patient-reported outcomes and survival in multiple sclerosis: A 10-year retrospective cohort study using the Multiple Sclerosis Impact Scale-29.

BACKGROUND: There is increasing emphasis on using patient-reported outcomes (PROs) to complement traditional clinical outcomes in medical research, including in multiple sclerosis (MS). Research, particularly in oncology and heart failure, has shown that PROs can be prognostic of hard clinical endpoints such as survival time (time from study entry until death). However, unlike in oncology or cardiology, it is unknown whether PROs are associated with survival time in neurological diseases. The Multiple Sclerosis Impact Scale-29 (MSIS-29) is a PRO sensitive to short-term change in MS, with questions covering both physical and psychological quality of life. This study aimed to investigate whether MSIS-29 scores can be prognostic for survival time in MS, using a large observational cohort of people with MS. METHODS AND FINDINGS: From 15 July 2004 onwards, MSIS-29 questionnaires were completed by people with MS registered with the MS Society Tissue Bank (n = 2,126, repeated 1 year later with n = 872 of the original respondents). By 2014, 264 participants (12.4%) had died. Higher baseline MSIS-29 physical (MSIS-29-PHYS) score was associated with reduced survival time (subgroup with highest scores versus subgroup with lowest scores: hazard ratio [HR] 5.7, 95% CI 3.1-10.5, p < 0.001). Higher baseline MSIS-29 psychological score was also associated with reduced survival time (subgroup with highest scores versus subgroup with lowest scores: HR 2.8, 95% CI 1.8-4.4, p < 0.001). In those with high baseline MSIS-29 scores, mortality risk was even greater if the MSIS-29 score worsened over 1 year (HR 2.3, 95% CI 1.2-4.4, p = 0.02). MSIS-29-PHYS scores were associated with survival time independent of age, sex, and patient-reported Expanded Disability Status Scale score in a Cox regression analysis (per 1-SD increase in MSIS-29-PHYS score: HR 1.8, 95% CI 1.1-2.9, p = 0.03). A limitation of the study is that this cohort had high baseline age and disability levels; the prognostic value of MSIS-29 for survival time at earlier disease stages requires further investigation. CONCLUSIONS: This study reports that PROs can be prognostic for hard clinical outcomes in neurological disease, and supports PROs as a meaningful clinical outcome for use in research and clinical settings.

Considerations in establishing a post-mortem brain and tissue bank for the study of myalgic encephalomyelitis/chronic fatigue syndrome: a proposed protocol.

BACKGROUND: Our aim, having previously investigated through a qualitative study involving extensive discussions with experts and patients the issues involved in establishing and maintaining a disease specific brain and tissue bank for myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS), was to develop a protocol for a UK ME/CFS repository of high quality human tissue from well characterised subjects with ME/CFS and controls suitable for a broad range of research applications. This would involve a specific donor program coupled with rapid tissue collection and processing, supplemented by comprehensive prospectively collected clinical, laboratory and self-assessment data from cases and controls. FINDINGS: We reviewed the operations of existing tissue banks from published literature and from their internal protocols and standard operating procedures (SOPs). On this basis, we developed the protocol presented here, which was designed to meet high technical and ethical standards and legal requirements and was based on recommendations of the MRC UK Brain Banks Network. The facility would be most efficient and cost-effective if incorporated into an existing tissue bank. Tissue collection would be rapid and follow robust protocols to ensure preservation sufficient for a wide range of research uses. A central tissue bank would have resources both for wide-scale donor recruitment and rapid response to donor death for prompt harvesting and processing of tissue. CONCLUSION: An ME/CFS brain and tissue bank could be established using this protocol. Success would depend on careful consideration of logistic, technical, legal and ethical issues, continuous consultation with patients and the donor population, and a sustainable model of funding ideally involving research councils, health services, and patient charities. This initiative could revolutionise the understanding of this still poorly-understood disease and enhance development of diagnostic biomarkers and treatments.

Meningeal inflammation plays a role in the pathology of primary progressive multiple sclerosis.

The primary progressive form of multiple sclerosis is characterized by accrual of neurological dysfunction from disease onset without remission and it is still a matter of debate whether this disease course results from different pathogenetic mechanisms compared with secondary progressive multiple sclerosis. Inflammation in the leptomeninges has been identified as a key feature of secondary progressive multiple sclerosis and may contribute to the extensive cortical pathology that accompanies progressive disease. Our aim was to investigate the extent of perivascular and meningeal inflammation in primary progressive multiple sclerosis in order to understand their contribution to the pathogenetic mechanisms associated with cortical pathology. A comprehensive immunohistochemical analysis was performed on post-mortem brain tissue from 26 cases with primary progressive multiple sclerosis. A variable extent of meningeal immune cell infiltration was detected and more extensive demyelination and neurite loss in the cortical grey matter was found in cases exhibiting an increased level of meningeal inflammation. However, no tertiary lymphoid-like structures were found. Profound microglial activation and reduction in neuronal density was observed in both the lesions and normal appearing grey matter compared with control cortex. Furthermore, cases with primary progressive multiple sclerosis with extensive meningeal immune cell infiltration exhibited a more severe clinical course, including a shorter disease duration and younger age at death. Our data suggest that generalized diffuse meningeal inflammation and the associated inflammatory milieu in the subarachnoid compartment plays a role in the pathogenesis of cortical grey matter lesions and an increased rate of clinical progression in primary progressive multiple sclerosis.

The neuropathological basis of clinical progression in multiple sclerosis.

Multiple sclerosis is the major inflammatory condition affecting the central nervous system (CNS) and is characterised by disseminated focal immune-mediated demyelination. Demyelination is accompanied by variable axonal damage and loss and reactive gliosis. It is this pathology that is thought to be responsible for the clinical relapses that often respond well to immunomodulatory therapy. However, the later secondary progressive stage of MS remains largely refractory to treatment and it is widely suggested that accumulating axon loss is responsible for clinical progression. Although initially thought to be a white matter (WM) disease, it is increasingly apparent that extensive pathology is also seen in the grey matter (GM) throughout the CNS. GM pathology is characterised by demyelination in the relative absence of an immune cell infiltrate. Neuronal loss is also seen both in the GM lesions and in unaffected areas of the GM. The slow progressive nature of this later stage combined with the presence of extensive grey matter pathology has led to the suggestion that neurodegeneration might play an increasing role with increasing disease duration. However, there is a paucity of studies that have correlated the pathological features with clinical milestones during secondary progressive MS. Here, we review the contributions that the various types of pathology are likely to make to the increasing neurological deficit in MS.

Attenuation of proliferation in oligodendrocyte precursor cells by activated microglia.

Activated microglia can influence the survival of neural cells through the release of cytotoxic factors. Here, we investigated the interaction between Toll-like receptor 4 (TLR4)-activated microglia and oligodendrocytes or their precursor cells (OPC). Primary rat or N9 microglial cells were activated by exposure to TLR4-specifc lipopolysaccharide (LPS), resulting in mitogen-activated protein kinase activation, increased CD68 and inducible nitric oxide synthase expression, and release of the proinflammatory cytokines tumor necrosis factor (TNF) and interleukin-6 (IL-6). Microglial conditioned medium (MGCM) from LPS-activated microglia attenuated primary OPC proliferation without inducing cell death. The microglial-induced inhibition of OPC proliferation was reversed by stimulating group III metabotropic glutamate receptors in microglia with the agonist L-AP4. In contrast to OPC, LPS-activated MGCM enhanced the survival of mature oligodendrocytes. Further investigation suggested that TNF and IL-6 released from TLR4-activated microglia might contribute to the effect of MGCM on OPC proliferation, insofar as TNF depletion of LPS-activated MGCM reduced the inhibition of OPC proliferation, and direct addition of TNF or IL-6 attenuated or increased proliferation, respectively. OPC themselves were also found to express proteins involved in TLR4 signalling, including TLR4, MyD88, and MAL. Although LPS stimulation of OPC did not induce proinflammatory cytokine release or affect their survival, it did trigger JNK phosphorylation, suggesting that TLR4 signalling in these cells is active. These findings suggest that OPC survival may be influenced not only by factors released from endotoxin-activated microglia but also through a direct response to endotoxins. This may have consequences for myelination under conditions in which microglial activation and cerebral infection are both implicated. , Inc.

Phosphorylation and compactness of neurofilaments in multiple sclerosis: indicators of axonal pathology.

AIMS: Axonal pathology extends to the axonal cytoarchitecture leaving its signature on axoskeletal proteins. This study investigated whether neurofilament (NfH) phosphorylation would relate to the dynamics of axonal pathology in multiple sclerosis (MS). METHODS: NfH phosphoforms (SMI32, SMI34, SMI35) were quantified by ELISA from microdissected samples of control and MS brain and spinal cord. Individual axons were analysed by electron microscopy, densitometrically and morphologically in adjacent tissue sections. Experiments were carried out pre- and post enzymatic dephosphorylation. RESULTS: In control tissue a rostro-caudal gradient of NfH indicated an increase in axonal density from the brain gray matter towards the spinal cord. The highest levels of phosphorylated and hyperphosphorylated NfH were found in acute lesions of brain and spinal cord, in contrast to chronic lesions where levels were lower than in white matter, consistent with axonal loss. Dephosphorylated NfH was higher, but less densly packed in MS white matter axons compared to control tissue. CONCLUSIONS: The findings suggest that a less organised/compact axoskeleton or impaired axonal transport may represent an early sign of axonal pathology within the normal appearing white matter in MS. Subsequently a proportional increase of dephosphorylated NfH, aberrant phosphorylation and/or aggregation may occur whilst the protein is transported through the white matter towards the MS plaque, where hyperphosphorylated NfH dominates.

Production of IL-16 correlates with CD4+ Th1 inflammation and phosphorylation of axonal cytoskeleton in multiple sclerosis lesions.

BACKGROUND: Multiple sclerosis (MS) is a central nervous system-specific autoimmune, demyelinating and neurodegenerative disease. Infiltration of lesions by autoaggressive, myelin-specific CD4+Th1 cells correlates with clinical manifestations of disease. The cytokine IL-16 is a CD4+ T cell-specific chemoattractant that is biased towards CD4+ Th1 cells. IL-16 precursor is constitutively expressed in lymphocytes and during CD4+ T cell activation; active caspase-3 cleaves and releases C-terminal bioactive IL-16. Previously, we used an animal model of MS to demonstrate an important role for IL-16 in regulation of autoimmune inflammation and subsequent axonal damage. This role of IL-16 in MS is largely unexplored. Here we examine the regulation of IL-16 in relation to CD4+ Th1 infiltration and inflammation-related changes of axonal cytoskeleton in MS lesions. METHODS: We measured relative levels of IL-16, active caspase-3, T-bet, Stat-1 (Tyr 701), and phosphorylated NF(M+H), in brain and spinal cord lesions from MS autopsies, using western blot analysis. We examined samples from 39 MS cases, which included acute, subacute and chronic lesions, as well as adjacent, normal-appearing white and grey matter. All samples were taken from patients with relapsing remitting clinical disease. We employed two-color immunostaining and confocal microscopy to identify phenotypes of IL-16-containing cells in frozen tissue sections from MS lesions. RESULTS: We found markedly increased levels of pro- and secreted IL-16 (80 kD and 22 kD, respectively) in MS lesions compared to controls. Levels of IL-16 peaked in acute, diminished in subacute, and were elevated again in chronic active lesions. Compared to lesions, lower but still appreciable IL-6 levels were measured in normal-appearing white matter adjacent to active lesions. Levels of IL-16 corresponded to increases in active-caspase-3, T-bet and phosphorylated Stat-1. In MS lesions, we readily observed IL-16 immunoreactivity confined to infiltrating CD3+, T-bet+ and active caspase-3+ mononuclear cells. CONCLUSION: We present evidence suggesting that IL-16 production occurs in MS lesions. We show correlations between increased levels of secreted IL-16, CD4+ Th1 cell inflammation, and phosphorylation of axonal cytoskeleton in MS lesions. Overall, the data suggest a possible role for IL-16 in regulation of inflammation and of subsequent changes in the axonal cytoskeleton in MS.

A role for the plasminogen activator system in inflammation and neurodegeneration in the central nervous system during experimental allergic encephalomyelitis.

Early signs of inflammatory demyelination include entry of fibrin(ogen) into the central nervous system (CNS), which is normally excluded by the blood-brain barrier, and up-regulation of components of the plasminogen activator system. Using mice deficient in tissue-type plasminogen activator (tPA-/-) and urokinase plasminogen activator receptor (uPAR-/-), we investigated the involvement of the PA system on the clinical and pathological features of experimental allergic encephalomyelitis, an animal model of multiple sclerosis. tPA-/- mice suffered an early and a more severe acute disease characterized by incomplete recovery when compared to wild-type controls, with significantly higher CNS levels of plasminogen activator inhibitor-1. This correlated with fibrin accumulation, which co-localized with nonphosphorylated neurofilament on thickened axons in experimental allergic encephalomyelitis tissue. In contrast, uPAR-/- mice had a delayed, less acute disease reflected in delayed infiltration of inflammatory cells. These animals developed chronic disease as a result of steadily increased inflammation, increased levels of urokinase-type plasminogen activator (uPA), and greater degree of demyelination. Thus, the plasminogen activator system can modulate both inflammatory and degenerative events in the CNS through the respective effects of tPA and uPAR on fibrinolysis and cell adhesion/migration, manipulation of which may have therapeutic implications for multiple sclerosis.

Tenascin C induces a quiescent phenotype in cultured adult human astrocytes.

Astrocytic scar formation occurs subsequent to brain and spinal cord injury and impedes repair. The exact mechanisms of scar formation have yet to be elucidated but it is known that astrocytes within the scar have a different antigenic phenotype from normal or reactive astrocytes. Astrocyte cell culture offers a suitable system to identify factors that induce the scar phenotype as well as factors that reverse this process and that may help identify therapeutic strategies to treat astrogliosis. However, when placed in standard culture conditions, astrocytes become activated/reactive and express molecules characteristic of scar tissue in vivo. In the present study, we made use of this phenomenon to identify culture conditions that change the activated phenotype of cultured astrocytes into one characteristic of normal quiescent astrocytes. In particular, we examined the effect of extracellular matrix (ECM) proteins found in the human brain, on the phenotype of human adult astrocytes. Significantly fewer astrocytes expressed scar properties when grown on tenascin-C (TN-C) than those cultured on other ECM proteins or poly-L-lysine-coated dishes. TN-C also significantly reduced the proliferation rate of the astrocytes in vitro. In addition, further manipulation of culture conditions induced partial astrocyte reactivation. Our findings suggest that astrocytes grown on TN-C revert to a quiescent, nonactivated state that is partially reversible. This raises the possibility that therapeutic strategies aimed at manipulating TN-C levels during CNS injury may help reduce astrocytic scarring.

tPA receptors and the fibrinolytic response in multiple sclerosis lesions.

Axonal damage in multiple sclerosis (MS) lesions is associated with failure of fibrinolysis because of the inhibition of the plasminogen activator system. Plasma membrane receptors for tissue plasminogen activator (tPA) and plasminogen concentrate proteolytic activity on the cell surface and provide protection from inhibitors that in turn may locally enhance the fibrinolytic response. Therefore, we have investigated expression of two of these receptors in MS lesions, annexin II tetramer (AIIt) and low-density lipoprotein receptor-related protein (LRP). In acute MS lesions both AIIt and LRP were immunolocalized on macrophages and astrocytes while LRP was additionally found on neuronal cells in cortical gray matter. Western blot analysis confirmed a significant increase in AIIt in MS lesions and in a proportion of normal-appearing white matter samples, with a highly significant correlation between annexin II levels and factors associated with impeded fibrinolysis, such as plasminogen activator inhibitor-1. Immunoblotting analysis of plasmin(ogen) revealed increased levels of lysine-plasminogen in samples expressing high AIIt protein levels. Our results suggest that limited availability of tPA in MS lesions because of formation of tPA-plasminogen activator inhibitor-1 complexes reduces capability of tPA receptors to generate plasmin, which further diminishes fibrinolytic capacity in active MS lesions and possibly leads to axonal damage.

Impaired fibrinolysis in multiple sclerosis: a role for tissue plasminogen activator inhibitors.

Tissue plasminogen activator (tPA), a neuronal as well as the key fibrinolytic enzyme, is found concentrated on demyelinated axons in multiple sclerosis lesions together with fibrin(ogen) deposits. The decreased tPA activity in normal-appearing white and grey matter and lesions of multiple sclerosis is reflected in diminished fibrinolysis as measured by a clot lysis assay. Nonetheless, peptide products of fibrin, including D-dimer, accumulate on demyelinated axons-the result of fibrinogen entry through a compromised blood-brain barrier (BBB). Analysis of tissue samples on reducing and non-reducing polyacrylamide gels demonstrates complexes of tPA with plasminogen activator inhibitor-1 (PAI-1) but not with neuroserpin, a tPA-specific inhibitor concentrated in grey matter. As total tPA protein remains unchanged in acute lesions and the concentration of PAI-1 rises several fold, complex formation is a probable cause of the impaired fibrinolysis. Although the tPA-plasmin cascade promotes neurodegeneration in excitotoxin-induced neuronal death, in inflammatory conditions with BBB disruption it has been demonstrated to have a protective role in removing fibrin, which exacerbates axonal injury. The impaired fibrinolytic capacity resulting from increased PAI-1 synthesis and complex formation with tPA, which is detectable prior to lesion formation, therefore has the potential to contribute to axonal damage in multiple sclerosis.

Broad expression of Toll-like receptors in the human central nervous system.

The family of Toll-like receptors (TLRs) plays a key role in controlling innate immune responses to a wide variety of pathogen-associated molecules. In this study we investigated expression of TLRs in vitro by purified human microglia, astrocytes, and oligodendrocytes, and in vivo by immunohistochemical examination of brain and spinal cord sections. Cultured primary microglia were found to express mRNA encoding a wide range of different TLR family members while astrocytes and oligodendrocytes primarily express TLR2 and TLR3. Comparisons between microglia derived from a series of control subjects and neurodegenerative cases indicate distinct differences in levels of mRNA encoding the different TLRs indifferent microglia samples. Interestingly, expression of TLR proteins in cultured microglia as revealed by immunocytochemistry was restricted to intracellular vesicles, whereas in astrocytes they were exclusively localized on the cell surface. Finally, in vivo expression of TLR3 and TLR4 was examined by immunohistochemical analysis of brain and spinal cord sections from both control and multiple sclerosis brains, revealing enhanced expression of either TLR in inflamed CNS tissues. Together, our data reveal broad and regulated expression of TLRs both in vitro and in vivo by human glia cells.