Publications by authors named "Anna Vilalta"

24 Publications

  • Page 1 of 1

The brain penetrant PPARγ agonist leriglitazone restores multiple altered pathways in models of X-linked adrenoleukodystrophy.

Sci Transl Med 2021 06;13(596)

Minoryx Therapeutics S.L., Barcelona 08302, Spain.

X-linked adrenoleukodystrophy (X-ALD), a potentially fatal neurometabolic disorder with no effective pharmacological treatment, is characterized by clinical manifestations ranging from progressive spinal cord axonopathy [adrenomyeloneuropathy (AMN)] to severe demyelination and neuroinflammation (cerebral ALD-cALD), for which molecular mechanisms are not well known. Leriglitazone is a recently developed brain penetrant full PPARγ agonist that could modulate multiple biological pathways relevant for neuroinflammatory and neurodegenerative diseases, and particularly for X-ALD. We found that leriglitazone decreased oxidative stress, increased adenosine 5'-triphosphate concentration, and exerted neuroprotective effects in primary rodent neurons and astrocytes after very long chain fatty acid-induced toxicity simulating X-ALD. In addition, leriglitazone improved motor function; restored markers of oxidative stress, mitochondrial function, and inflammation in spinal cord tissues from AMN mouse models; and decreased the neurological disability in the EAE neuroinflammatory mouse model. X-ALD monocyte-derived patient macrophages treated with leriglitazone were less skewed toward an inflammatory phenotype, and the adhesion of human X-ALD monocytes to brain endothelial cells decreased after treatment, suggesting the potential of leriglitazone to prevent the progression to pathologically disrupted blood-brain barrier. Leriglitazone increased myelin debris clearance in vitro and increased myelination and oligodendrocyte survival in demyelination-remyelination in vivo models, thus promoting remyelination. Last, leriglitazone was clinically tested in a phase 1 study showing central nervous system target engagement (adiponectin increase) and changes on inflammatory biomarkers in plasma and cerebrospinal fluid. The results of our study support the use of leriglitazone in X-ALD and, more generally, in other neuroinflammatory and neurodegenerative conditions.
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http://dx.doi.org/10.1126/scitranslmed.abc0555DOI Listing
June 2021

Wild-type sTREM2 blocks Aβ aggregation and neurotoxicity, but the Alzheimer's R47H mutant increases Aβ aggregation.

J Biol Chem 2021 Jan-Jun;296:100631. Epub 2021 Apr 3.

Department of Biochemistry, University of Cambridge, Cambridge, United Kingdom. Electronic address:

TREM2 is a pattern recognition receptor, expressed on microglia and myeloid cells, detecting lipids and Aβ and inducing an innate immune response. Missense mutations (e.g., R47H) of TREM2 increase risk of Alzheimer's disease (AD). The soluble ectodomain of wild-type TREM2 (sTREM2) has been shown to protect against AD in vivo, but the underlying mechanisms are unclear. We show that Aβ oligomers bind to cellular TREM2, inducing shedding of the sTREM2 domain. Wild-type sTREM2 bound to Aβ oligomers (measured by single-molecule imaging, dot blots, and Bio-Layer Interferometry) inhibited Aβ oligomerization and disaggregated preformed Aβ oligomers and protofibrils (measured by transmission electron microscopy, dot blots, and size-exclusion chromatography). Wild-type sTREM2 also inhibited Aβ fibrillization (measured by imaging and thioflavin T fluorescence) and blocked Aβ-induced neurotoxicity (measured by permeabilization of artificial membranes and by loss of neurons in primary neuronal-glial cocultures). In contrast, the R47H AD-risk variant of sTREM2 is less able to bind and disaggregate oligomeric Aβ but rather promotes Aβ protofibril formation and neurotoxicity. Thus, in addition to inducing an immune response, wild-type TREM2 may protect against amyloid pathology by the Aβ-induced release of sTREM2, which blocks Aβ aggregation and neurotoxicity. In contrast, R47H sTREM2 promotes Aβ aggregation into protofibril that may be toxic to neurons. These findings may explain how wild-type sTREM2 apparently protects against AD in vivo and why a single copy of the R47H variant gene is associated with increased AD risk.
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http://dx.doi.org/10.1016/j.jbc.2021.100631DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8113883PMC
April 2021

Id1 and PD-1 Combined Blockade Impairs Tumor Growth and Survival of -mutant Lung Cancer by Stimulating PD-L1 Expression and Tumor Infiltrating CD8 T Cells.

Cancers (Basel) 2020 Oct 28;12(11). Epub 2020 Oct 28.

Department of Oncology, Clínica Universidad de Navarra, 31008 Pamplona, Spain.

The use of PD-1/PD-L1 checkpoint inhibitors in advanced NSCLC is associated with longer survival. However, many patients do not benefit from PD-1/PD-L1 blockade, largely because of immunosuppression. New immunotherapy-based combinations are under investigation in an attempt to improve outcomes. (inhibitor of differentiation 1) is involved in immunosuppression. In this study, we explored the potential synergistic effect of the combination of inhibition and pharmacological PD-L1 blockade in three different syngeneic murine -mutant lung adenocarcinoma models. TCGA analysis demonstrated a negative and statistically significant correlation between PD-L1 and expression levels. This observation was confirmed in vitro in human and murine -driven lung cancer cell lines. In vivo experiments in -mutant syngeneic and metastatic murine lung adenocarcinoma models showed that the combined blockade targeting and PD-1 was more effective than each treatment alone in terms of tumor growth impairment and overall survival improvement. Mechanistically, multiplex quantification of CD3/CD4/CD8 T cells and flow cytometry analysis showed that combined therapy favors tumor infiltration by CD8 T cells, whilst in vivo CD8 T cell depletion led to tumor growth restoration. Co-culture assays using CD8 cells and tumor cells showed that T cells present a higher antitumor effect when tumor cells lack expression. These findings highlight that blockade may contribute to a significant immune enhancement of antitumor efficacy of PD-1 inhibitors by increasing PD-L1 expression and harnessing tumor infiltration of CD8 T lymphocytes.
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http://dx.doi.org/10.3390/cancers12113169DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7693788PMC
October 2020

TET2 Regulates the Neuroinflammatory Response in Microglia.

Cell Rep 2019 10;29(3):697-713.e8

Instituto de Biomedicina de Sevilla (IBiS), Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, 41013 Sevilla, Spain; Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad de Sevilla, 41012 Sevilla, Spain; Blizard Institute, Barts and The London School of Medicine and Dentistry, QMUL, London E1 2AT, UK. Electronic address:

Epigenomic mechanisms regulate distinct aspects of the inflammatory response in immune cells. Despite the central role for microglia in neuroinflammation and neurodegeneration, little is known about their epigenomic regulation of the inflammatory response. Here, we show that Ten-eleven translocation 2 (TET2) methylcytosine dioxygenase expression is increased in microglia upon stimulation with various inflammogens through a NF-κB-dependent pathway. We found that TET2 regulates early gene transcriptional changes, leading to early metabolic alterations, as well as a later inflammatory response independently of its enzymatic activity. We further show that TET2 regulates the proinflammatory response in microglia of mice intraperitoneally injected with LPS. We observed that microglia associated with amyloid β plaques expressed TET2 in brain tissue from individuals with Alzheimer's disease (AD) and in 5xFAD mice. Collectively, our findings show that TET2 plays an important role in the microglial inflammatory response and suggest TET2 as a potential target to combat neurodegenerative brain disorders.
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http://dx.doi.org/10.1016/j.celrep.2019.09.013DOI Listing
October 2019

Galectin-3, a novel endogenous TREM2 ligand, detrimentally regulates inflammatory response in Alzheimer's disease.

Acta Neuropathol 2019 08 20;138(2):251-273. Epub 2019 Apr 20.

Department of Experimental Medical Science, Experimental Neuroinflammation Laboratory, Lund University, 221 84, Lund, Sweden.

Alzheimer's disease (AD) is a progressive neurodegenerative disease in which the formation of extracellular aggregates of amyloid beta (Aβ) peptide, fibrillary tangles of intraneuronal tau and microglial activation are major pathological hallmarks. One of the key molecules involved in microglial activation is galectin-3 (gal3), and we demonstrate here for the first time a key role of gal3 in AD pathology. Gal3 was highly upregulated in the brains of AD patients and 5xFAD (familial Alzheimer's disease) mice and found specifically expressed in microglia associated with Aβ plaques. Single-nucleotide polymorphisms in the LGALS3 gene, which encodes gal3, were associated with an increased risk of AD. Gal3 deletion in 5xFAD mice attenuated microglia-associated immune responses, particularly those associated with TLR and TREM2/DAP12 signaling. In vitro data revealed that gal3 was required to fully activate microglia in response to fibrillar Aβ. Gal3 deletion decreased the Aβ burden in 5xFAD mice and improved cognitive behavior. Interestingly, a single intrahippocampal injection of gal3 along with Aβ monomers in WT mice was sufficient to induce the formation of long-lasting (2 months) insoluble Aβ aggregates, which were absent when gal3 was lacking. High-resolution microscopy (stochastic optical reconstruction microscopy) demonstrated close colocalization of gal3 and TREM2 in microglial processes, and a direct interaction was shown by a fluorescence anisotropy assay involving the gal3 carbohydrate recognition domain. Furthermore, gal3 was shown to stimulate TREM2-DAP12 signaling in a reporter cell line. Overall, our data support the view that gal3 inhibition may be a potential pharmacological approach to counteract AD.
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http://dx.doi.org/10.1007/s00401-019-02013-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6660511PMC
August 2019

Effective Knockdown of Gene Expression in Primary Microglia With siRNA and Magnetic Nanoparticles Without Cell Death or Inflammation.

Front Cell Neurosci 2018 21;12:313. Epub 2018 Sep 21.

Department of Clinical Neurosciences, Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom.

Microglia, the resident immune cells of the brain, have multiple functions in physiological and pathological conditions, including Alzheimer's disease (AD). The use of primary microglial cell cultures has proved to be a valuable tool to study microglial biology under various conditions. However, more advanced transfection methodologies for primary cultured microglia are still needed, as current methodologies provide low transfection efficiency and induce cell death and/or inflammatory activation of the microglia. Here, we describe an easy, and effective method based on the Glial-Mag method (OZ Biosciences) using magnetic nanoparticles and a magnet to successfully transfect primary microglia cells with different small interfering RNAs (siRNAs). This method does not require specialist facilities or specific training and does not induce cell toxicity or inflammatory activation. We demonstrate that this protocol successfully decreases the expression of two key genes associated with AD, the triggering receptor expressed in myeloid cells 2 (TREM2) and CD33, in primary microglia cell cultures.
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http://dx.doi.org/10.3389/fncel.2018.00313DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6161539PMC
September 2018

Neurophagy, the phagocytosis of live neurons and synapses by glia, contributes to brain development and disease.

FEBS J 2018 10 29;285(19):3566-3575. Epub 2017 Nov 29.

Department of Biochemistry, University of Cambridge, UK.

It was previously thought that neurons were phagocytosed only when dead or dying. However, it is increasingly clear that viable synapses, dendrites, axons and whole neurons can be phagocytosed alive (defined here as neurophagy), and this may contribute to a wide range of developmental, physiological and pathological processes. Phagocytosis of live synapses, dendrites and axons by glia contributes to experience-dependent sculpting of neuronal networks during development, but excessive phagocytosis of synapses may contribute to pathology in Alzheimer's disease, schizophrenia and ageing. Neurons can expose phosphatidylserine or calreticulin, which act as 'eat me' signals provoking phagocytosis via microglial receptors, whereas sialylation of neuronal surfaces acts as a 'don't eat me' signal that inhibits phagocytosis and desialylation can provoke phagocytosis. Opsonins, such as complement components and apolipoproteins, are released during inflammation and enhance engulfment. Phagocytosis of neurons is seen in multiple human diseases, but it is as yet unclear whether inhibition of phagocytosis will be beneficial in treating neurological diseases. Here we review the signals regulating glial phagocytosis of live neurons and synapses, and the involvement of this phagocytosis in development and disease.
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http://dx.doi.org/10.1111/febs.14323DOI Listing
October 2018

Anti-CD47 antibodies induce phagocytosis of live, malignant B cells by macrophages the Fc domain, resulting in cell death by phagoptosis.

Oncotarget 2017 Sep 15;8(37):60892-60903. Epub 2017 Jun 15.

Department of Biochemistry, University of Cambridge, Cambridge, UK.

When expressed on the surface of cells, CD47 inhibits phagocytosis of these cells by phagocytes. Most human cancers overexpress CD47, and antibodies to CD47 have shown a remarkable ability to clear a range of cancers in animal models. However, the mechanism by which these antibodies cause cancer cell death is unclear. We find that CD47 is expressed on the surface of three B-cell lines from human malignancies: 697 (pre-B-ALL lymphoblasts), Ramos and DG-75 (both mature B-cells, Burkitt's lymphoma), and anti-CD47 antibodies greatly increase the phagocytosis of all three cell line by macrophages. In the presence of macrophages, the antibodies cause clearance of the lymphoblasts within hours, but in the absence of macrophages, the antibodies have no effect on lymphoblast viability. Macrophages engulf viable lymphoblasts containing mitochondria with a normal membrane potential, but following engulfment the mitochondrial membrane potential is lost indicating a loss of viability. Inhibition of phagocytosis protects lymphoblasts from death indicating that phagocytosis is required for anti-CD47 mediated cell death. Blocking either the antibody Fc domain or Fc receptors inhibits antibody-induced phagocytosis. Antibodies against cell surface markers CD10 or CD19 also induced Fc-domain-dependent phagocytosis, but at a lower level commensurate with expression. Thus, phagoptosis may contribute to the efficacy of a number of therapeutic antibodies used in cancer therapy, as well as potentially endogenous antibodies. We conclude that anti-CD47 antibodies induce phagocytosis by binding CD47 on lymphoblast and Fc receptors on macrophages, resulting in cell death by phagocytosis, i.e. phagoptosis.
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http://dx.doi.org/10.18632/oncotarget.18492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5617392PMC
September 2017

Activated Microglia Desialylate and Phagocytose Cells via Neuraminidase, Galectin-3, and Mer Tyrosine Kinase.

J Immunol 2017 06 12;198(12):4792-4801. Epub 2017 May 12.

Department of Biochemistry, University of Cambridge, Cambridge CB2 1QW, United Kingdom

Activated microglia can phagocytose dying, stressed, or excess neurons and synapses via the phagocytic receptor Mer tyrosine kinase (MerTK). Galectin-3 (Gal-3) can cross-link surface glycoproteins by binding galactose residues that are normally hidden below terminal sialic acid residues. Gal-3 was recently reported to opsonize cells via activating MerTK. We found that LPS-activated BV-2 microglia rapidly released Gal-3, which was blocked by calcineurin inhibitors. Gal-3 bound to MerTK on microglia and to stressed PC12 (neuron-like) cells, and it increased microglial phagocytosis of PC12 cells or primary neurons, which was blocked by inhibition of MerTK. LPS-activated microglia exhibited a sialidase activity that desialylated PC12 cells and could be inhibited by Tamiflu, a neuraminidase (sialidase) inhibitor. Sialidase treatment of PC12 cells enabled Gal-3 to bind and opsonize the live cells for phagocytosis by microglia. LPS-induced microglial phagocytosis of PC12 was prevented by small interfering RNA knockdown of Gal-3 in microglia, lactose inhibition of Gal-3 binding, inhibition of neuraminidase with Tamiflu, or inhibition of MerTK by UNC569. LPS-induced phagocytosis of primary neurons by primary microglia was also blocked by inhibition of MerTK. We conclude that activated microglia release Gal-3 and a neuraminidase that desialylates microglial and PC12 surfaces, enabling Gal-3 binding to PC12 cells and their phagocytosis via MerTK. Thus, Gal-3 acts as an opsonin of desialylated surfaces, and inflammatory loss of neurons or synapses may potentially be blocked by inhibiting neuraminidases, Gal-3, or MerTK.
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http://dx.doi.org/10.4049/jimmunol.1502532DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5458330PMC
June 2017

Galectin-3 released in response to traumatic brain injury acts as an alarmin orchestrating brain immune response and promoting neurodegeneration.

Sci Rep 2017 01 27;7:41689. Epub 2017 Jan 27.

Centre for Neuroscience and Trauma. Blizard Institute. Queen Mary University of London, E1 2AT London, United Kingdom.

Traumatic brain injury (TBI) is currently a major cause of morbidity and poor quality of life in Western society, with an estimate of 2.5 million people affected per year in Europe, indicating the need for advances in TBI treatment. Within the first 24 h after TBI, several inflammatory response factors become upregulated, including the lectin galectin-3. In this study, using a controlled cortical impact (CCI) model of head injury, we show a large increase in the expression of galectin-3 in microglia and also an increase in the released form of galectin-3 in the cerebrospinal fluid (CSF) 24 h after head injury. We report that galectin-3 can bind to TLR-4, and that administration of a neutralizing antibody against galectin-3 decreases the expression of IL-1β, IL-6, TNFα and NOS2 and promotes neuroprotection in the cortical and hippocampal cell populations after head injury. Long-term analysis demonstrated a significant neuroprotection in the cortical region in the galectin-3 knockout animals in response to TBI. These results suggest that following head trauma, released galectin-3 may act as an alarmin, binding, among other proteins, to TLR-4 and promoting inflammation and neuronal loss. Taking all together, galectin-3 emerges as a clinically relevant target for TBI therapy.
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http://dx.doi.org/10.1038/srep41689DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5269662PMC
January 2017

Activated microglia cause reversible apoptosis of pheochromocytoma cells, inducing their cell death by phagocytosis.

J Cell Sci 2016 Jan 13;129(1):65-79. Epub 2015 Nov 13.

Department of Biochemistry, University of Cambridge, Cambridge CB2 1QW, UK

Some apoptotic processes, such as phosphatidylserine exposure, are potentially reversible and do not necessarily lead to cell death. However, phosphatidylserine exposure can induce phagocytosis of a cell, resulting in cell death by phagocytosis: phagoptosis. Phagoptosis of neurons by microglia might contribute to neuropathology, whereas phagoptosis of tumour cells by macrophages might limit cancer. Here, we examined the mechanisms by which BV-2 microglia killed co-cultured pheochromocytoma (PC12) cells that were either undifferentiated or differentiated into neuronal cells. We found that microglia activated by lipopolysaccharide rapidly phagocytosed PC12 cells. Activated microglia caused reversible phosphatidylserine exposure on and reversible caspase activation in PC12 cells, and caspase inhibition prevented phosphatidylserine exposur and decreased subsequent phagocytosis. Nitric oxide was necessary and sufficient to induce the reversible phosphatidylserine exposure and phagocytosis. The PC12 cells were not dead at the time they were phagocytised, and inhibition of their phagocytosis left viable cells. Cell loss was inhibited by blocking phagocytosis mediated by phosphatidylserine, MFG-E8, vitronectin receptors or P2Y6 receptors. Thus, activated microglia can induce reversible apoptosis of target cells, which is insufficient to cause apoptotic cell death, but sufficient to induce their phagocytosis and therefore cell death by phagoptosis.
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http://dx.doi.org/10.1242/jcs.174631DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4732292PMC
January 2016

How microglia kill neurons.

Brain Res 2015 Dec 2;1628(Pt B):288-297. Epub 2015 Sep 2.

Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QW, UK.

Microglia are resident brain macrophages that become inflammatory activated in most brain pathologies. Microglia normally protect neurons, but may accidentally kill neurons when attempting to limit infections or damage, and this may be more common with degenerative disease as there was no significant selection pressure on the aged brain in the past. A number of mechanisms by which activated microglia kill neurons have been identified, including: (i) stimulation of the phagocyte NADPH oxidase (PHOX) to produce superoxide and derivative oxidants, (ii) expression of inducible nitric oxide synthase (iNOS) producing NO and derivative oxidants, (iii) release of glutamate and glutaminase, (iv) release of TNFα, (v) release of cathepsin B, (vi) phagocytosis of stressed neurons, and (vii) decreased release of nutritive BDNF and IGF-1. PHOX stimulation contributes to microglial activation, but is not directly neurotoxic unless NO is present. NO is normally neuroprotective, but can react with superoxide to produce neurotoxic peroxynitrite, or in the presence of hypoxia inhibit mitochondrial respiration. Glutamate can be released by glia or neurons, but is neurotoxic only if the neurons are depolarised, for example as a result of mitochondrial inhibition. TNFα is normally neuroprotective, but can become toxic if caspase-8 or NF-κB activation are inhibited. If the above mechanisms do not kill neurons, they may still stress the neurons sufficiently to make them susceptible to phagocytosis by activated microglia. We review here whether microglial killing of neurons is an artefact, makes evolutionary sense or contributes in common neuropathologies and by what mechanisms. This article is part of a Special Issue entitled SI: Neuroprotection.
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http://dx.doi.org/10.1016/j.brainres.2015.08.031DOI Listing
December 2015

Tumour necrosis factor alpha-induced neuronal loss is mediated by microglial phagocytosis.

FEBS Lett 2014 Aug 6;588(17):2952-6. Epub 2014 Jun 6.

Department of Biochemistry, University of Cambridge, UK. Electronic address:

Tumour necrosis factor-α (TNF-α) is a pro-inflammatory cytokine, expressed in many brain pathologies and associated with neuronal loss. We show here that addition of TNF-α to neuronal-glial co-cultures increases microglial proliferation and phagocytosis, and results in neuronal loss that is prevented by eliminating microglia. Blocking microglial phagocytosis by inhibiting phagocytic vitronectin and P2Y6 receptors, or genetically removing opsonin MFG-E8, prevented TNF-α induced loss of live neurons. Thus TNF-α appears to induce neuronal loss via microglial activation and phagocytosis of neurons, causing neuronal death by phagoptosis.
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http://dx.doi.org/10.1016/j.febslet.2014.05.046DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4158418PMC
August 2014

Deoxyglucose prevents neurodegeneration in culture by eliminating microglia.

J Neuroinflammation 2014 Mar 26;11:58. Epub 2014 Mar 26.

Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QW, UK.

Background: 2-Deoxy-D-glucose is an inhibitor of glycolysis, which is protective in animal models of brain pathology, but the mechanisms of this protection are unclear. We examined whether, when and how deoxyglucose protects neurons in co-culture with astrocytes and microglia. Microglia are brain macrophages, which can damage neurons in inflammatory conditions.

Methods: Deoxyglucose was added to primary cultures of microglia and astrocytes from rat cortex, or neurons and glia from rat cerebellum, or the BV-2 microglial cell line, and cell death and cell functions were evaluated.

Results: Surprisingly, addition of deoxyglucose induced microglial loss and prevented spontaneous neuronal loss in long-term cultures of neurons and glia, while elimination of microglia by L-leucine-methyl ester prevented the deoxyglucose-induced neuroprotection. Deoxyglucose also prevented neuronal loss induced by addition of amyloid beta or disrupted neurons (culture models of Alzheimer's disease and brain trauma respectively). However, deoxyglucose greatly increased the neuronal death induced by hypoxia. Addition of deoxyglucose to pure microglia induced necrosis and loss, preceded by rapid ATP depletion and followed by phagocytosis of the microglia. Deoxyglucose did not kill astrocytes or neurons.

Conclusions: We conclude that deoxyglucose causes microglial loss by ATP depletion, and this can protect neurons from neurodegeneration, except in conditions of hypoxia. Deoxyglucose may thus be beneficial in brain pathologies mediated by microglia, including brain trauma, but not where hypoxia is involved.
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http://dx.doi.org/10.1186/1742-2094-11-58DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3986974PMC
March 2014

Caspase inhibitors protect neurons by enabling selective necroptosis of inflamed microglia.

J Biol Chem 2013 Mar 5;288(13):9145-52. Epub 2013 Feb 5.

Department of Biochemistry, University of Cambridge, Cambridge, CB2 1QW, United Kingdom.

Microglia are resident brain macrophages, which can cause neuronal loss when activated in infectious, ischemic, traumatic, and neurodegenerative diseases. Caspase-8 has both prodeath and prosurvival roles, mediating apoptosis and/or preventing RIPK1-mediated necroptosis depending on cell type and stimulus. We found that inflammatory stimuli (LPS, lipoteichoic acid, or TNF-α) caused an increase in caspase-8 IETDase activity in primary rat microglia without inducing apoptosis. Inhibition of caspase-8 with either Z-VAD-fmk or IETD-fmk resulted in necrosis of activated microglia. Inhibition of caspases with Z-VAD-fmk did not kill non-activated microglia, or astrocytes and neurons in any condition. Necrostatin-1, a specific inhibitor of RIPK1, prevented microglial caspase inhibition-induced death, indicating death was by necroptosis. In mixed cerebellar cultures of primary neurons, astrocytes, and microglia, LPS induced neuronal loss that was prevented by inhibition of caspase-8 (resulting in microglial necroptosis), and neuronal death was restored by rescue of microglia with necrostatin-1. We conclude that the activation of caspase-8 in inflamed microglia prevents their death by necroptosis, and thus, caspase-8 inhibitors may protect neurons in the inflamed brain by selectively killing activated microglia.
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http://dx.doi.org/10.1074/jbc.M112.427880DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3610987PMC
March 2013

Normobaric hyperoxia in traumatic brain injury: does brain metabolic state influence the response to hyperoxic challenge?

J Neurotrauma 2011 Jul 30;28(7):1139-48. Epub 2011 Jun 30.

Neurotraumatology and Neurosurgery Research Unit, Vall d'Hebron University Hospital, Universitat Autònoma de Barcelona, Paseo Vall d'Hebron 119-129, Barcelona, Spain.

This study sought to investigate whether normobaric hyperoxia (NH) improves brain oxygenation and brain metabolism in the early phase of severe and moderate traumatic brain injury (TBI) and whether this effect occurs uniformly in all TBI patients. Thirty patients (9 women and 21 men) with a median initial Glasgow Coma Score (GCS) of 6 (range, 3-12) were monitored using a brain microdialysis (MD) catheter with a brain tissue oxygen sensor (PtiO(2)) placed in the least-injured hemisphere. The inspired oxygen fraction was increased to 100% for 2 h. Patients were divided into two groups: Group 1: patients with baseline brain lactate ≤3 mmol/L and Group 2: patients with baseline brain lactate >3 mmol/L, and therefore increased anaerobic metabolism in the brain. In Group 1, no significant changes in brain metabolic parameters were found after hyperoxic challenge, whereas a significant increase in glucose and a decrease in the lactate-pyruvate ratio (LPR) were found in Group 2. In this latter group of patients, brain glucose increased on average by 17.9% (95% CI, +9.2% to +26.6%, p<0.001) and LPR decreased by 11.6% (95% CI, -16.2% to -6.9%, p<0.001). The results of our study show that moderate and severe TBI may induce metabolic alterations in the brain, even in macroscopically normal brain tissue. We observed that NH increased PaO(2) and PtiO(2) and significantly decreased LPR in patients in whom baseline brain lactate levels were increased, suggesting that NH improved the brain redox state. In patients with normal baseline brain lactate levels, we did not find any significant changes in the metabolic variables after NH. This suggests that the baseline metabolic state should be taken into account when applying NH to patients with TBI. This maneuver may only be effective in a specific group of patients.
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http://dx.doi.org/10.1089/neu.2010.1720DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3136740PMC
July 2011

Brain perihematoma genomic profile following spontaneous human intracerebral hemorrhage.

PLoS One 2011 Feb 2;6(2):e16750. Epub 2011 Feb 2.

Neurovascular Research Laboratory and Department of Neurology, Universitat Autònoma de Barcelona, Institut de Recerca, Hospital Vall d'Hebron, Barcelona, Spain.

Background: Spontaneous intracerebral hemorrhage (ICH) represents about 15% of all strokes and is associated with high mortality rates. Our aim was to identify the gene expression changes and biological pathways altered in the brain following ICH.

Methodology/principal Findings: Twelve brain samples were obtained from four deceased patients who suffered an ICH including perihematomal tissue (PH) and the corresponding contralateral white (CW) and grey (CG) matter. Affymetrix GeneChip platform for analysis of over 47,000 transcripts was conducted. Microarray Analysis Suite 5.0 was used to process array images and the Ingenuity Pathway Analysis System was used to analyze biological mechanisms and functions of the genes. We identified 468 genes in the PH areas displaying a different expression pattern with a fold change between -3.74 and +5.16 when compared to the contralateral areas (291 overexpressed and 177 underexpressed). The top genes which appeared most significantly overexpressed in the PH areas codify for cytokines, chemokines, coagulation factors, cell growth and proliferation factors while the underexpressed codify for proteins involved in cell cycle or neurotrophins. Validation and replication studies at gene and protein level in brain samples confirmed microarray results.

Conclusions: The genomic responses identified in this study provide valuable information about potential biomarkers and target molecules altered in the perihematomal regions.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0016750PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3032742PMC
February 2011

Brain extracellular fluid protein changes in acute stroke patients.

J Proteome Res 2011 Mar 18;10(3):1043-51. Epub 2011 Jan 18.

Biomedical Proteomics Group, Department of Structural Biology and Bioinformatics, Faculty of Medicine, University of Geneva, Geneva, Switzerland.

In vivo human brain extracellular fluids (ECF) of acute stroke patients were investigated to assess the changes in protein levels associated with ischemic damages. Microdialysates (MDs) from the infarct core (IC), the penumbra (P), and the unaffected contralateral (CT) brain regions of patients suffering an ischemic stroke (n = 6) were compared using a shotgun proteomic approach based on isobaric tagging and mass spectrometry. Quantitative analysis showed 53 proteins with increased amounts in the IC or P with respect to the CT samples. Glutathione S-transferase P (GSTP1), peroxiredoxin-1 (PRDX1), and protein S100-B (S100B) were further assessed with ELISA on the blood of unrelated control (n = 14) and stroke (n = 14) patients. Significant increases of 8- (p = 0.0002), 20- (p = 0.0001), and 11-fold (p = 0.0093) were found, respectively. This study highlights the value of ECF as an efficient source to further discover blood stroke markers.
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http://dx.doi.org/10.1021/pr101123tDOI Listing
March 2011

[Matrix metaloproteinases in neurological brain lesions: a new therapeutic target?].

Rev Neurol 2010 Jul;51(2):95-107

Unidad de Investigación de Neurotraumatología y Neurocirugía, Servicio de Neurocirugía, Hospital Universitari Vall d'Hebron, 08035 Barcelona, España.

Introduction: Matrix metalloproteinases (MMP) are a family of proteolytic enzymes that degrade the extracellular matrix and are found in a large amount of human tissues. Their main functions are to maintain the integrity of the extracellular matrix, to modulate the interaction of the cells during development, to contribute to tissue remodeling, to directly participate in angiogenesis and to facilitate cellular migration. Due to the importance of its maintaining extracellular matrix function, MMP expression is tightly regulated at transcriptional level, through proform activation and with the binding to tissular inhibitors. Despite this complex regulation system, MMP regulation can be altered, producing an overexpression of these proteolytic proteins that alter the tissular structure, possibly destroying the tissue, as observed in some neurologic pathologies such as multiple sclerosis, aneurism formation and cerebral ischemia. The role that MMP have in traumatic brain lesions is almost unknown and is derived mainly from in vivo and in vitro experimental studies, and only from three papers performed in humans. There are some experimental studies that relate the brain alterations produced after traumatic brain injury with an increase in the concentration of various MMP.

Aim: To review the role of these proteases in human brain lesions, emphasizing on the function of these proteases in traumatic brain injury lesions and their possible therapeutic target.

Development: A bibliographic search was performed on Medline database.

Conclusions: Some MMP could be related to blood-brain barrier alteration and postraumatic edema formation, turning them into promising therapeutic targets.
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July 2010

Intravascular cooling for rapid induction of moderate hypothermia in severely head-injured patients: results of a multicenter study (IntraCool).

Intensive Care Med 2009 May 26;35(5):890-8. Epub 2008 Nov 26.

Department of Neurosurgery, Vall d'Hebron University Hospital, Universitat Autonoma de Barcelona (UAB), Passeig Vall d'Hebron 119-129, 08035 Barcelona, Spain.

Objectives: To evaluate the feasibility, safety and effectiveness of a new method of intravascular temperature management for inducing moderate hypothermia (MHT).

Design And Settings: Prospective, international-multicenter clinical trial conducted in four university hospitals.

Patients: In a 2-year period 24 patients with severe head injury and refractory high ICP were treated with MHT (32.5 degrees C) by intravascular methods.

Results: Seventeen were males and seven females, with a median age of 25 years (range 15-60). The median Glasgow Coma Scale upon admission was 7 (range 3-13) and the median Injury Severity Score was 22 (range 13-43). A total of 75% of patients presented a diffuse lesion in the pre-enrollment computed tomography. Median time from injury until reaching refractory high ICP was 71.5 h after injury (minimum 14 h, maximum 251 h). Twelve patients (50%) reached this situation within the first 72 h after injury. MHT was attained in a median time of 3 h. Pre-enrollment median ICP was 23.8 mmHg and was reduced to 16.8 mmHg upon reaching target temperature. At 6 months after injury, nine patients had died (37.5%), six were severely disabled (25%), two moderately disabled (8.3%) and seven had a good recovery (29.2%). Of the nine patients who died, in four the cause was rebound ICP during rewarming, one death was attributed to accidental potassium overload, two to septic shock, one to cardiac arrest of unknown origin and the ninth to a pulmonary embolism.

Conclusion: Intravascular methods to induce MHT combined with precooling with cold saline at 4 degrees C appear to be feasible and effective in reducing ICP in patients with high ICP refractory to first-line therapeutic measures.
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http://dx.doi.org/10.1007/s00134-008-1357-4DOI Listing
May 2009

Moderate and severe traumatic brain injury induce early overexpression of systemic and brain gelatinases.

Intensive Care Med 2008 Aug 19;34(8):1384-92. Epub 2008 Mar 19.

Neurotraumatology and Neurosurgery Research Unit , Vall d'Hebron University Hospital, Universitat Autònoma de Barcelona , Passeig Vall d'Hebron 119-129, 08035, Barcelona, Spain.

Objective: Recent experimental evidence suggests that matrix metalloproteinases (MMPs) are implicated in the pathophysiology of traumatic brain injury (TBI) by increasing blood-brain barrier permeability and exacerbating posttraumatic edema. We examined the acute profile of MMP-2 and MMP-9 in the plasma of patients with moderate or severe TBI and in the brain extracellular fluid (ECF).

Design: Prospective observational study.

Setting: Neurotraumatology intensive care unit of a tertiary university hospital.

Patients: Twenty patients with moderate or severe TBI were included and three groups were used as controls: 20 patients with a mild head injury and normal CT scan, 15 moderate polytrauma patients without TBI, and 20 healthy volunteers.

Interventions: Plasma samples were collected within the first 12[Symbol: see text]h and at 24[Symbol: see text]h post-injury. Simultaneous brain microdialysate and plasma samples were obtained in four moderate-severe TBI patients at additional timepoints: 48, 72, and 96[Symbol: see text]h post-TBI.

Measurements And Main Results: Gelatinases (MMP-2 and MMP-9) were measured by gelatin zymography. A significant increase in plasma gelatinases was observed at baseline when compared with healthy volunteers in the study group. This early increase was followed by a significant decrease at 24[Symbol: see text]h post-injury. Brain microdialysis samples presented a similar time profile as plasma samples for both gelatinases.

Conclusions: High levels of gelatinases were found in plasma and brain ECF in the early phase of TBI, indicating that both local and systemic trauma-induced upregulation of gelatinases in the acute phase might play an important role in the pathophysiology of TBI and could be a future therapeutic target.
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http://dx.doi.org/10.1007/s00134-008-1056-1DOI Listing
August 2008

Cooling the injured brain: how does moderate hypothermia influence the pathophysiology of traumatic brain injury.

Curr Pharm Des 2007 ;13(22):2310-22

Department of Neurosurgery Vall d'Hebron University Hospital, Barcelona, Spain.

Neither any neuroprotective drug has been shown to be beneficial in improving the outcome of severe traumatic brain injury (TBI) nor has any prophylactically-induced moderate hypothermia shown any beneficial effect on outcome in severe TBI, despite the optimism generated by preclinical studies. This contrasts with the paradox that hypothermia still is the most powerful neuroprotective method in experimental models because of its ability to influence the multiple biochemical cascades that are set in motion after TBI. The aim of this short review is to highlight the most recent developments concerning the pathophysiology of severe TBI, to review new data on thermoregulation and induced hypothermia, the regulation of core and brain temperature in mammals and the multiplicity of effects of hypothermia in the pathophysiology of TBI. Many experimental studies in the last decade have again confirmed that moderate hypothermia confers protection against ischemic and non-ischemic brain hypoxia, traumatic brain injury, anoxic injury following resuscitation after cardiac arrest and other neurological insults. Many posttraumatic adverse events that occur in the injured brain at a cellular and molecular level are highly temperature-sensitive and are thus a good target for induced hypothermia. The basic mechanisms through which hypothermia protects the brain are clearly multifactorial and include at least the following: reduction in brain metabolic rate, effects on cerebral blood flow, reduction of the critical threshold for oxygen delivery, blockade of excitotoxic mechanisms, calcium antagonism, preservation of protein synthesis, reduction of brain thermopooling, a decrease in edema formation, modulation of the inflammatory response, neuroprotection of the white matter and modulation of apoptotic cell death. The new developments discussed in this review indicate that, by targeting many of the abnormal neurochemical cascades initiated after TBI, induced hypothermia may modulate neurotoxicity and, consequently, may play a unique role in opening up new therapeutic avenues for treating severe TBI and improving its devastating effects. Furthermore, greater understanding of the pathophysiology of TBI, new data from both basic and clinical research, the good clinical results obtained in randomized clinical trials in cardiac arrest and better and more reliable cooling methods have given hypothermia a second chance in treating TBI patients. A critical evaluation of hypothermia is therefore mandatory to elucidate the reasons for previous failures and to design further multicenter randomized clinical trials that would definitively confirm or refute the potential of this therapeutic modality in the management of severe traumatic brain injuries.
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http://dx.doi.org/10.2174/138161207781368756DOI Listing
August 2007

Lack of utility of arteriojugular venous differences of lactate as a reliable indicator of increased brain anaerobic metabolism in traumatic brain injury.

J Neurosurg 2007 Apr;106(4):530-7

Department of Neurosurgery, Vail d'Hebron University Hospital and Vall d'Hebron Research Institute, Autonomous University of Barcelona, Spain.

Object: Ischemic lesions are highly prevalent in patients with traumatic brain injuries (TBIs) and are the single most important cause of secondary brain damage. The prevention and early treatment of these lesions is the primary aim in the modem treatment of these patients. One of the most widely used monitoring techniques at the bedside is quantification of brain extracellular level of lactate by using arteriojugular venous differences of lactate (AVDL). The purpose of this study was to determine the sensitivity, specificity, and predictive value of AVDL as an indicator of increases in brain lactate production in patients with TBIs.

Methods: Arteriojugular venous differences of lactate were calculated every 6 hours using samples obtained though a catheter placed in the jugular bulb in 45 patients with diffuse head injuries (57.8%) or evacuated brain lesions (42.2%). Cerebral lactate concentration obtained with a 20-kD microdialysis catheter implanted in undamaged tissue was used as the de facto gold standard. Six hundred seventy-three AVDL determinations and cerebral microdialysis samples were obtained simultaneously; 543 microdialysis samples (81%) showed lactate values greater than 2 mmol/L, but only 21 AVDL determinations (3.1%) showed an increase in brain lactate. No correlation was found between AVDL and cerebral lactate concentration (p = 0.014, p = 0.719). Arteriojugular venous differences of lactate had a sensitivity and specificity of 3.3 and 97.7%, respectively, with a false-negative rate of 96.7% and a false-positive rate of 2.3%.

Conclusions: Arteriojugular venous differences of lactate do not reliably reflect increased cerebral lactate production and consequently are not reliable in ruling out brain ischemia in patients with TBIs. The clinical use of this monitoring method in neurocritical care should be reconsidered.
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http://dx.doi.org/10.3171/jns.2007.106.4.530DOI Listing
April 2007

Percutaneous implantation of cerebral microdialysis catheters by twist-drill craniostomy in neurocritical patients: description of the technique and results of a feasibility study in 97 patients.

J Neurotrauma 2006 Oct;23(10):1510-7

Department of Neurosurgery, Vall d'Hebron University Hospital, Barcelona, Spain.

Cerebral microdialysis is increasingly used to monitor several types of neurocritical patients. This study presents the technique used in our unit for percutaneous implantation of cerebral microdialysis catheters using a small twist-drill craniostomy that can be performed in the intensive care unit (ICU). We also present the results of this technique in 89 head-injured patients and in eight patients with a malignant middle cerebral artery (MCA) infarction. One hundred and twenty-two cerebral microdialysis catheters were implanted in the 97 patients included in this study. One cerebral microdialysis catheter was implanted in the less damaged hemisphere of 67 head-injured patients with a diffuse brain injury. An additional microdialysis catheter was inserted in the pericontusional parenchyma of 22 patients with brain contusions. In five of the eight patients with a malignant MCA infarction, only one microdialysis probe was inserted in the penumbral zone. In the remaining three patients, two cerebral microdialysis catheters were implanted in the same hemisphere (one in the ischemic core and the other in the penumbra). Technical problems were detected in 18 (15%) of the 122 microdialysis catheters implanted and were more frequent during the initial period of using microdialysis in our unit. In four patients (3% of implanted catheters), follow-up computed tomography (CT) scans showed a small intracerebral blood collection (always
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http://dx.doi.org/10.1089/neu.2006.23.1510DOI Listing
October 2006
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