An Inflammation-Centric View of Neurological Disease: Beyond the Neuron

Inflammation is a complex biological response fundamental to how the body deals with injury and infection to eliminate the initial cause of cell injury and effect repair. Unlike a normally beneficial acute inflammatory response, chronic inflammation can lead to tissue damage and ultimately its destruction, and often results from an inappropriate immune response. Inflammation in the nervous system ("neuroinflammation"), especially when prolonged, can be particularly injurious. While inflammation per se may not cause disease, it contributes importantly to disease pathogenesis across both the peripheral (neuropathic pain, fibromyalgia) and central [e.g., Alzheimer disease, Parkinson disease, multiple sclerosis, motor neuron disease, ischemia and traumatic brain injury, depression, and autism spectrum disorder] nervous systems. The existence of extensive lines of communication between the nervous system and immune system represents a fundamental principle underlying neuroinflammation. Immune cell-derived inflammatory molecules are critical for regulation of host responses to inflammation. Although these mediators can originate from various non-neuronal cells, important sources in the above neuropathologies appear to be microglia and mast cells, together with astrocytes and possibly also oligodendrocytes. Understanding neuroinflammation also requires an appreciation that non-neuronal cell-cell interactions, between both glia and mast cells and glia themselves, are an integral part of the inflammation process. Within this context the mast cell occupies a key niche in orchestrating the inflammatory process, from initiation to prolongation. This review will describe the current state of knowledge concerning the biology of neuroinflammation, emphasizing mast cell-glia and glia-glia interactions, then conclude with a consideration of how a cell's endogenousmechanisms might be leveraged to provide a therapeutic strategy to target neuroinflammation

Recensione di M.H. Salmon, Introduction to Logic and Critical Thinking

Book-revie

Role of microglia and astrocytes in inflammatory processes involving neurological diseases, chronic pain, and psychiatric disorders, with emphasis on the purinergic P2X7 receptor

Under pathological conditions microglia (resident central nervous system (CNS) immune cells) become activated, and produce reactive oxygen and nitrogen species and pro-inflammatory cytokines: molecules that can contribute to disorders including stroke, traumatic brain injury, progressive neurodegenerative disorders such as Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, multiple sclerosis, and several retinal diseases. Given that ATP is frequently released from CNS neurons during tissue damage and inflammation, its actions on microglia-mediated toxicity are especially pertinent. For example, the ATP-gated P2X7 purinergic receptor (P2X7R) cation channel is up-regulated around amyloid beta-peptide plaques in transgenic mouse models of Alzheimer's disease and co-localizes to microglia and astrocytes. Upregulation of P2X7R on microglia occurs also following spinal cord injury and after brain ischemia. ATP, via activation of P2X7R, is one of the most powerful stimuli for secretion of the key pro-inflammatory cytokine interleukin-1β (IL-1β) in its mature form. This project investigates the pharmacological and biochemical behaviors of P2X7R on microglia and astrocytes cultured from rat cerebral cortex, spinal cord and cerebellum, and the relationship between these two glial cell types. ATP is an efficient stimulus for IL-1β secretion only after the cells have undergone a short 'priming' with endotoxin (lipopolysaccharide (LPS)). Indeed LPS, but not ATP caused release of IL-1β from cortical microglia. However, it is known that the greater part of the IL-1β thus released is the precursor (biologically inactive) form. Purified (>99%) cortical microglia and enriched (>95%) astrocytes were primed for 2 hours with LPS, followed by addition of ATP for 1 hour. Culture medium was then collected and the content of IL-1β quantified by ELISA. The effects of LPS and ATP were concentration-dependent; although LPS alone (but not ATP) modestly stimulated IL-1β release, levels of cytokine release were much higher from primed cells incubated with ATP. The ATP-dependent component was fully blocked by selective P2X7R antagonists, and followed their known rank order of target potency. The P2X7R priming response was also seen with spinal cord and cerebellar microglia, a finding not described in the literature until now. To rule out a contribution by the minor population of microglia in our astrocyte cultures, the latter were treated with the lysosomotropic agent L-leucyl-L-leucine methyl ester (L-LME) which selectively eliminates cells with cytotoxic potential (e.g. macrophages, microglia). Immunocytochemical and molecular biological evaluation showed L-LME-treated cortical and spinal cord astrocytes to be fully depleted of microglia. These purified astrocytes failed to respond to LPS, and did not show the ATP priming behavior. Responsiveness was recovered upon addition of microglia to the L-LME-treated astrocytes and, moreover, a far more robust release of IL-1β occurred than that achieved with the same numbers of microglia alone. This astrocyte-microglia interaction was also observed for LPS-stimulated release of nitric oxide and IL-6, and was not mediated by astrocyte-derived soluble factors. Lastly, the LPS/ATP priming behavior was studied by examining the ability of other agents, linked to neuropathology, to replace either LPS or ATP. Neither ethanol (ethanol intoxication; in place of LPS) nor amyloid beta-peptides (Alzheimer disease; in place of ATP) were able to provoke IL-1β release from microglia. However, both zymosan and poly(I:C), agonists of Toll-like receptors -2 and -3, respectively, were capable of substituting LPS (a Toll-like receptor 4 agonist) in the P2X7R priming response. Release of IL-1β in all these cases was antagonized by inhibitors of p38 mitogen-activated protein kinase (a stress response kinase). TLRs contribute to CNS immunocompetent cell activation and the resulting pro-inflammatory cascade producing pathological pain. TLR4 recognizes not only LPS, but also ligands called damage associated molecular patterns, released by the injured tissue The involvement of extracellular TLR4 and TLR2, as well as TLR3 in preclinical pain models has been demonstrated. The findings described here further support the notion of astrocyte/microglia interaction, which may improve our understanding in how these cells respond to CNS injury or inflammation, in particular where TLRs are involve

Solid state molecular rectifier based on self organized metalloproteins

Recently, great attention has been paid to the possibility of implementing hybrid electronic devices exploiting the self-assembling properties of single molecules. Impressive progress has been done in this field by using organic molecules and macromolecules. However, the use of biomolecules is of great interest because of their larger size (few nanometers) and of their intrinsic functional properties. Here, we show that electron-transfer proteins, such as the blue copper protein azurin (Az), can be used to fabricate biomolecular electronic devices exploiting their intrinsic redox properties, self assembly capability and surface charge distribution. The device implementation follows a bottom-up approach in which the self assembled protein layer interconnects nanoscale electrodes fabricated by electron beam lithography, and leads to efficient rectifying behavior at room temperature.Comment: 13 pages including two figures. Accepted for publication in Advanced Material

Changes in single K+ channel behavior through the lipid phase transition

We show that the activity of an ion channel is strictly related to the phase state of the lipid bilayer hosting the channel. By measuring unitary conductance, dwell times, and open probability of the K+ channel KcsA as a function of temperature in lipid bilayers composed of POPE and POPG in different relative proportions, we obtain that all those properties show a trend inversion when the bilayer is in the transition region between the liquid disordered and the solid ordered phase. These data suggest that the physical properties of the lipid bilayer influence ion channel activity likely via a fine tuning of its conformations. In a more general interpretative framework, we suggest that other parameters such as pH, ionic strength, and the action of amphiphilic drugs can affect the physical behavior of the lipid bilayer in a fashion similar to temperature changes resulting in functional changes of transmembrane proteins

Serum amyloid A primes microglia for ATP-dependent interleukin-1\u3b2 release

Acute-phase response is a systemic reaction to environmental/inflammatory insults and involves production of acute-phase proteins, including serum amyloid A (SAA). Interleukin-1\u3b2 (IL-1\u3b2), a master regulator of neuroinflammation produced by activated inflammatory cells of the myeloid lineage, in particular microglia, plays a key role in the pathogenesis of acute and chronic diseases of the peripheral nervous system and CNS. IL-1\u3b2 release is promoted by ATP acting at the purinergic P2X7 receptor (P2X7R) in cells primed with toll-like receptor (TLR) ligands

Bisdemethoxycurcumin and Its Cyclized Pyrazole Analogue Differentially Disrupt Lipopolysaccharide Signalling in Human Monocyte-Derived Macrophages

Several studies suggest that curcumin and related compounds possess antioxidant and anti-inflammatory properties including modulation of lipopolysaccharide- (LPS-) mediated signalling in macrophage cell models. We here investigated the effects of curcumin and the two structurally unrelated analogues GG6 and GG9 in primary human blood-derived macrophages as well as the signalling pathways involved. Macrophages differentiated from peripheral blood monocytes for 7 days were activated with LPS or selective Toll-like receptor agonists for 24 h. The effects of test compounds on cytokine production and immunophenotypes evaluated as CD80+/CCR2+ and CD206+/CD163+ subsets were examined by ELISA and flow cytometry. Signalling pathways were probed by Western blot. Curcumin (2.5–10 μM) failed to suppress LPS-induced inflammatory responses. While GG6 reduced LPS-induced IκB-α degradation and showed a trend towards reduced interleukin-1β release, GG9 prevented the increase in proinflammatory CD80+ macrophage subset, downregulation of the anti-inflammatory CD206+/CD163+ subset, increase in p38 phosphorylation, and increase in cell-bound and secreted interleukin-1β stimulated by LPS, at least in part through signalling pathways not involving Toll-like receptor 4 and nuclear factor-κB. Thus, the curcumin analogue GG9 attenuated the LPS-induced inflammatory response in human blood-derived macrophages and may therefore represent an attractive chemical template for macrophage pharmacological targeting

Neuroinflammation, Mast Cells, and Glia: Dangerous Liaisons

The perspective of neuroinflammation as an epiphenomenon following neuron damage is being replaced by the awareness of glia and their importance in neural functions and disorders. Systemic inflammation generates signals that communicate with the brain and leads to changes in metabolism and behavior, with microglia assuming a pro-inflammatory phenotype. Identification of potential peripheral-to-central cellular links is thus a critical step in designing effective therapeutics. Mast cells may fulfill such a role. These resident immune cells are found close to and within peripheral nerves and in brain parenchyma/meninges, where they exercise a key role in orchestrating the inflammatory process from initiation through chronic activation. Mast cells and glia engage in crosstalk that contributes to accelerate disease progression; such interactions become exaggerated with aging and increased cell sensitivity to stress. Emerging evidence for oligodendrocytes, independent of myelin and support of axonal integrity, points to their having strong immune functions, innate immune receptor expression, and production/response to chemokines and cytokines that modulate immune responses in the central nervous system while engaging in crosstalk with microglia and astrocytes. In this review, we summarize the findings related to our understanding of the biology and cellular signaling mechanisms of neuroinflammation, with emphasis on mast cell-glia interactions

Ligand engagement of Toll-like receptors regulates their expression in cortical microglia and astrocytes

BACKGROUND: Toll-like receptor (TLR) activation on microglia and astrocytes are key elements in neuroinflammation which accompanies a number of neurological disorders. While TLR activation on glia is well-established to up-regulate pro-inflammatory mediator expression, much less is known about how ligand engagement of one TLR may affect expression of other TLRs on microglia and astrocytes. METHODS: In the present study, we evaluated the effects of agonists for TLR2 (zymosan), TLR3 (polyinosinic-polycytidylic acid (poly(I:C)), a synthetic analogue of double-stranded RNA) and TLR4 (lipopolysaccaride (LPS)) in influencing expression of their cognate receptor as well as that of the other TLRs in cultures of rat cortical purified microglia (>99.5 %) and nominally microglia-free astrocytes. Elimination of residual microglia (a common contaminant of astrocyte cultures) was achieved by incubation with the lysosomotropic agent L-leucyl-L-leucine methyl ester (L-LME). RESULTS: Flow cytometric analysis confirmed the purity (essentially 100 %) of the obtained microglia, and up to 5 % microglia contamination of astrocytes. L-LME treatment effectively removed microglia from the latter (real-time polymerase chain reaction). The three TLR ligands robustly up-regulated gene expression for pro-inflammatory markers (interleukin-1 and interleukin-6, tumor necrosis factor) in microglia and enriched, but not purified, astrocytes, confirming cellular functionality. LPS, zymosan and poly(I:C) all down-regulated TLR4 messenger RNA (mRNA) and up-regulated TLR2 mRNA at 6 and 24 h. In spite of their inability to elaborate pro-inflammatory mediator output, the nominally microglia-free astrocytes (>99 % purity) also showed similar behaviours to those of microglia, as well as changes in TLR3 gene expression. LPS interaction with TLR4 activates downstream mitogen-activated protein kinase and nuclear factor-κB signalling pathways and subsequently causes inflammatory mediator production. The effects of LPS on TLR2 mRNA in both cell populations were antagonized by a nuclear factor-κB inhibitor. CONCLUSIONS: TLR2 and TLR4 activation in particular, in concert with microglia and astrocytes, comprise key elements in the initiation and maintenance of neuropathic pain. The finding that both homologous (zymosan) and heterologous (LPS, poly(I:C)) TLR ligands are capable of regulating TLR2 gene expression, in particular, may have important implications in understanding the relative contributions of different TLRs in neurological disorders associated with neuroinflammation