Henrik Wilms

Activation of microglia by human neuromelanin is NF-κB dependent and involves p38 mitogen-activated protein kinase: implications for Parkinson’s disease

It has been suggested that microglial inflammation augments the progression of Parkinsons disease (PD). However, endogenous factors initiating microglial activation are largely unknown. We therefore investigated the effects of human neuromelanin (NM) on the release of neurotoxic mediators and the underlying signaling pathways from rat microglia in vitro. The addition of NM to microglial cultures induced positive chemotactic effects, activated the proinflammatory transcription factor nuclear factor κB (NF‐κB) via phosphorylation and degradation of the inhibitor protein κB (IκB), and led to an up‐regulation of tumor necrosis factor α, interleukin‐6, and nitric oxide. The impairment of NF‐κB function by the IκB kinase inhibitor sulfasalazine was paralleled by a decline in neurotoxic mediators. NM also activated p38 mitogen‐activated protein kinase (MAPK), the inhibition of this pathway by SB203580 diminished phosphorylation of the transactivation domain of the p65 subunit of NF‐κB. These findings demonstrate a crucial role of NM in the pathogenesis of PD by augmentation of microglial activation, leading to a vicious cycle of neuronal death, exposure of additional neuromelanin, and chronification of inflammation. The antagonization of microglial activation by a pharmacological intervention targeting microglial NF‐κB or p38 MAPK could point to additional venues in the treatment of PD.

Neuromelanin of the substantia nigra: a neuronal black hole with protective and toxic characteristics

Neuromelanin accumulates in dopaminergic neurons during normal aging, and in Parkinsons disease, neurons with this pigment are those that selectively degenerate. Intraneuronal neuromelanin could play a protective role during its synthesis by preventing the toxic accumulation of cytosolic catechol derivatives and, in addition, by its ability to scavenge reactive metals, pesticides and other toxins to form stable adducts. However, dying neurons in Parkinsons disease that release neuromelanin might induce a vicious cycle of chronic neuroinflammation and neuronal loss.

The pathophysiology of parkinsonian tremor: a review.

Abstract Parkinsonian tremor is most likely due to oscillating neuronal activity within the CNS. Summarizing all the available evidence, peripheral factors only play a minor role in the generation, maintenance and modulation of PD tremor. Recent studies have shown that not a single but multiple oscillators and responsible. The most likely candidate producing these oscillations is the basal ganglia loop and its topographic organization might be responsible for the separation into different oscillators which, nevertheless, usually produce the same frquency. The neuronal mechanisms underlying these oscillations are not yet clear, but three hypotheses would be compatible with the presently available data from animal models and data recorded in patients. The first is a cortico-subthalamo-pallido-thalamic loop, the second is a pacemaker consisting of the external pallidum and the subthalamic nucleus, and the third is abnormal synchronization due to unknown mechanisms within the whole striato-pallido-thalamic pathway leading to a loss of segregation. Assuming the oscillator within the basal ganglia pathway, the mechanism of stereotactic surgery might be a desynchronization of the activity of the basal ganglia-thalamo-cortical or the cerebello-thalamo-cortical pathway.

Involvement of benzodiazepine receptors in neuroinflammatory and neurodegenerative diseases: evidence from activated microglial cells in vitro

Increased binding of a ligand for the peripheral benzodiazepine binding receptor is currently used in PET studies as an in vivo measurement of inflammation in diseases like multiple sclerosis and Alzheimers disease. Although peripheral-type benzodiazepin receptors (PBRs) are abundant in many cell types and expressed in the CNS physiologically only at low levels, previous reports suggest that after experimental lesions in animal models and in human neurodegenerative/-inflammatory diseases upregulated PBR expression with increased binding of its ligand PK11195 is confined mainly to activated microglia in vivo/in situ. Because the functional role of the PBR is unknown, we confirm by immunohistochemistry and PCR (I) that this receptor is expressed on microglia in vitro and (II) that benzodiazepines modulate proliferation of microglial cells and the release of the inflammatory molecules nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha) in cell culture supernatants of primary rat microglia. Compared to lipopolysaccharide-activated controls the release of NO was markedly decreased in cultures treated with benzodiazepines (clonazepam, midazolam, diazepam) and the PBR ligand PK11195. Moreover, release of TNF-alpha and proliferation was significantly inhibited in the benzodiazepine-treated groups. These findings link the in vivo data of elevated PBR levels in neurodegenerative/-inflammatory diseases to a functional role and opens up possible therapeutic intervention targeting the PBR in microglia.

Intrathecal synthesis of monocyte chemoattractant protein-1 (MCP-1) in amyotrophic lateral sclerosis: further evidence for microglial activation in neurodegeneration.

Autopsy studies and animal experiments suggest that microglial inflammation contributes to the pathogenesis of amyotrophic lateral sclerosis (ALS). Monocyte-chemoattractant protein (MCP-1) might play an important role in microglial recruitment. We studied MCP-1 levels in sera and cerebrospinal fluid of 29 ALS patients and compared the results with 11 control patients with tension headache. The MCP-1 level was determined using enzyme-linked immunosorbent assays (ELISA). A significant increase in cerebrospinal fluid MCP-1 level but not serum level was seen in the patients with ALS compared to the control subjects. These results suggest that cerebrospinal fluid MCP-1 activity may be a sensitive marker for neuroinflammation in ALS useful for monitoring treatment trials in ALS.

Ramification of microglia, monocytes and macrophages in vitro: influences of various epithelial and mesenchymal cells and their conditioned media*

Abstract.Microglial cells are able to switch between an ”active” amoeboid and a ramified ”resting” morphology during development and after experiencing lesions. We have previously shown that in vitro microglial morphology is controlled by their cellular environment, i.e. cells become ramified in astrocyte coculture but amoeboid on monolayers of fibroblasts. In the present study we have extended the analysis of the control of macrophage morphology by maintaining macrophages of different origins in coculture with different epithelial or mesenchymal cells and their conditioned media. Microglia, monocytes and spleen macrophages seeded onto monolayers of astrocytes, kidney epithelia or hepatoma cells developed the ramified morphology but remained amoeboid in fibroblast coculture. Ramification was also induced by media conditioned by these cells as well as by phorbolic esters, i.e. activators of protein kinase C. In double coculture assays, even small numbers of fibroblasts were able to override the ”epithelial” influence. Likewise, microglia remained amoeboid, when incubated on several constituents of the extracellular matrix. These results indicate that macrophage ramification is an active process initiated by diffusible factors secreted by various epithelial cells, possibly acting upon a protein-kinase-C-related receptor. We interprete the modification of macrophage morphology as a functional adaptation to the surrounding type of tissue that is enforced by its constituent cells. Thus, the specific morphologies of microglia, hepatic von Kupffer’s cells or peritubular kidney macrophages could be explained by similar epithelium–macrophage interaction.

Clinical spectrum and physiology of palatal tremor

Keywords: palatal tremor; palatal myoclonus; olivery pweudohypertrophy; inferior olive; brainstem

ATP and adenosine induce ramification of microglia in vitro

Microglial cells in the healthy adult brain possess a characteristic ramified morphology with multiple branched processes, small somata and down-regulated inflammatory properties. In contrast, microglial cells isolated from new-born rat brain inevitably show a non-ramified amoeboid phenotype, which is observed in vivo after pathologic activation or during development. To identify factors that control microglial morphology we investigated the effects of purines alone or in combination with astrocyte-conditioned medium (ACM). Under optimized culture conditions postnatal rat microglial cells developed an amoeboid to ovoid phenotype. Addition of 0.6-1 mM ATP or adenosine induced the outgrowth of numerous processes after 2-3 days that could be observed also in the presence of ACM as previously reported. Culture in ACM plus ATP or adenosine yielded an optimized ramified phenotype. ATP or adenosine, but not ACM alone, also prevented the formation of a flat, amoeboid morphology induced by lipopolysaccharide (LPS); however, at 0.6-1 mM they did not reduce the initial LPS-induced activation of the transcription factor NF-kappaB. By using specific agonists or antagonists the morphological transformations could not be confined to a distinct purinoreceptor subtype, but appeared to be mediated by long-term presence of adenosine in the medium to which phosphorylated purines were rapidly hydrolyzed by microglial cells. Since ACM did not contain sufficient concentrations of ATP or adenosine, purines are not the only ramification-inducing factors present in ACM; however, they are a valuable tool to induce microglial ramification in vitro.

Role of Glial Cells in the Functional Expression of LL-37/Rat Cathelin-Related Antimicrobial Peptide in Meningitis

Abstract Antimicrobial peptides are intrinsic to the innate immune system in many organ systems, but little is known about their expression in the central nervous system. We examined cerebrospinal fluid (CSF) and serum from patients with active bacterial meningitis to assess antimicrobial peptides and possible bactericidal properties of the CSF. We found antimicrobial peptides (human cathelicidin LL-37) in the CSF of patients with bacterial meningitis but not in control CSF. We next characterized the expression, secretion, and bactericidal properties of rat cathelin-related antimicrobial peptide, the homologue of the human LL-37, in rat astrocytes and microglia after incubation with different bacterial components. Using real-time polymerase chain reaction and Western blotting, we determined that supernatants from both astrocytes and microglia incubated with bacterial component supernatants had antimicrobial activity. The expression of rat cathelin-related antimicrobial peptide in rat glial cells involved different signal transduction pathways and was induced by the inflammatory cytokines interleukin 1&bgr; and tumor necrosis factor. In an experimental model of meningitis, infant rats were intracisternally infected with Streptococcus pneumoniae, and rat cathelin-related antimicrobial peptide was localized in glia, choroid plexus, and ependymal cells by immunohistochemistry. Together, these results suggest that cathelicidins produced by glia and other cells play an important part in the innate immune response against pathogens in central nervous system bacterial infections.

Neuroprotection with angiotensin receptor antagonists: a review of the evidence and potential mechanisms.

The peptide hormone angiotensin (A)-II, the major effector peptide of the renin-angiotensin system (RAS), is well established to play a pivotal role in the systemic regulation of blood pressure, fluid, and electrolyte homeostasis. Recent biochemical and neurophysiologic studies have documented local intrinsic angiotensin-generating systems in organs and tissues such as the brain, retina, bone marrow, liver, and pancreas. The locally generated angiotensin peptides have multiple and novel actions including stimulating cell growth and anti-proliferative and/or antiapoptotic actions. In the mammalian brain, all components of the RAS are present including angiotensin receptor subtypes 1 (AT1) and 2 (AT2). A-II exerts most of its well defined physiologic and pathophysiologic actions, including those on the central and peripheral nervous system, through its AT1 receptor subtype. While the AT1 receptor is responsible for the classical effects of A-II, it has been found that the AT2 receptor is linked to totally different signalling mechanisms and this has revealed hitherto unknown functions of A-II. AT2 receptors are expressed at low density in many healthy adult tissues, but are upregulated in a variety of human diseases. This receptor not only contributes to stroke-related pathologic mechanisms (e.g. hypertension, atherothrombosis, and cardiac hypertrophy) but may also be involved in post-ischemic damage to the brain. It has been reported that the AT2 receptor regulates several functions of nerve cells, e.g. ionic fluxes, cell differentiation, and neuronal tissue regeneration, and also modulates programmed cell death. In this article, we review the experimental evidence supporting the notion that blockade of brain AT1receptors can be beneficial with respect to stroke incidence and outcome. We further delineate how AT2 receptors could be involved in neuronal regeneration following brain injury such as stroke or CNS trauma. The current review is focussed on some of the new functions arising from the locally formed A-II with particular attention to its emerging neuroprotective role in the brain.