A co-ultramicronized palmitoylethanolamide/luteolin composite mitigates clinical score and disease-relevant molecular markers in a mouse model of experimental autoimmune encephalomyelitis

Background: Persistent and/or recurrent inflammatory processes are the main factor leading to multiple sclerosis (MS) lesions. The composite ultramicronized palmitoylethanolamide, an endogenous N-acylethanolamine, combined with the flavonoid luteolin, PEALut, have been found to exert neuroprotective activities in experimental models of spinal and brain injury and Alzheimer disease, as well as a clinical improvement in human stroke patients. Furthermore, PEALut enhances the expression of different myelin proteins in oligodendrocyte progenitor cells suggesting that this composite might have protective effects in MS experimental models. Methods: The mouse model of experimental autoimmune encephalomyelitis (EAE) based on active immunization with a fragment of myelin oligodendrocyte glycoprotein (MOG35-55) was used. The daily assessment of clinical score and the expression of serum amyloid A (SAA1), proinflammatory cytokines TNF-\u3b1, IL-1\u3b2, IFN-\u3b3, and NLRP3 inflammasome, as well as TLR2, Fpr2, CD137, CD3-\u3b3, and TCR-\u3b6 chain, heterodimers that form T cell surface glycoprotein (TCR), and cannabinoid receptors CB1, CB2, and MBP, were evaluated in the brainstem and cerebellum at different postimmunization days (PIDs). Results: Vehicle-MOG35-55-immunized (MOG35-55) mice developed ascending paralysis which peaked several days later and persisted until the end of the experiment. PEALut, given intraperitoneally daily starting on day 11 post-immunization, dose-dependently improved clinical score over the range 0.1-5 mg/kg. The mRNA expression of SAA1, TNF-\u3b1, IL-1\u3b2, IFN-\u3b3, and NLRP3 were significantly increased in MOG35-55 mice at 14 PID. In MOG35-55 mice treated with 5 mg /kg PEALut, the increase of SAA1, TNF- \u3b1, IL-1\u3b2, and IFN-\u3b3transcripts at 14 PID was statistically downregulated as compared to vehicle-MOG35-55 mice (p < 0.05). The expression of TLR2, Fpr2, CD137, CD3-\u3b3, TCR-\u3b6 chain, and CB2 receptors showed a significant upregulation in vehicle-MOG35-55 mice at 14 PID. Instead, CB1 and MBP transcripts have not changed in expression at any time. In MOG/PEALut-treated mice, TLR2, Fpr2, CD137, CD3-\u3b3, TCR-\u3b6 chain, and CB2 mRNAs were significantly downregulated as compared to vehicle MOG35-55 mice. Conclusions: The present results demonstrate that the intraperitoneal administration of the composite PEALut significantly reduces the development of clinical signs in the MOG35-55 model of EAE. The dose-dependent improvement of clinical score induced by PEALut was associated with a reduction in transcript expression of the acute-phase protein SAA1, TNF-\u3b1, IL-1\u3b2, IFN-\u3b3, and NLRP3 proinflammatory proteins and TLR2, Fpr2, CD137, CD3-\u3b3, TCR-\u3b6 chain, and CB2 receptors

Nanoscale mechanical properties of lipid bilayers and their relevance in biomembrane organization and function

The mechanical properties of biological systems are emerging as fundamental in determining their functional activity. For example, cells continuously probe their environment by applying forces and, at the same time, are exposed to forces produced by the same environment. Also in biological membranes, the activity of membrane related proteins are affected by the overall mechanical properties of the hosting environment. Traditionally, the mesoscopic mechanical properties of lipid bilayers have been studied by micropipette aspiration techniques. In recent years, the possibility of probing mechanical properties of lipid bilayers at the nanoscale has been promoted by the force spectroscopy potentiality of Atomic Force Microscopes (AFM). By acquiring force-curves on supported lipid bilayers (SLBs) it is possible to probe the mechanical properties on a scale relevant to the interaction between membrane proteins and lipid bilayers and to monitor changes of these properties as a result of a changing environment. Here, we review a series of force spectroscopy experiments performed on SLBs with an emphasis on the functional consequences the measured mechanical properties can have on membrane proteins. We also discuss the force spectroscopy experiments on SLBs in the context of theories developed for dynamic force spectroscopy experiments with the aim to extract the kinetic and energetic description of the process of membrane rupture

Demonstration of an electrostatic-shielded cantilever

The fabrication and performances of cantilevered probes with reduced parasitic capacitance starting from a commercial Si3N4 cantilever chip is presented. Nanomachining and metal deposition induced by focused ion beam techniques were employed in order to modify the original insulating pyramidal tip and insert a conducting metallic tip. Two parallel metallic electrodes deposited on the original cantilever arms are employed for tip biasing and as ground plane in order to minimize the electrostatic force due to the capacitive interaction between cantilever and sample surface. Excitation spectra and force-to-distance characterization are shown with different electrode configurations. Applications of this scheme in electrostatic force microscopy, Kelvin probe microscopy and local anodic oxidation is discussed.Comment: 4 pages and 3 figures. Submitted to Applied Physics Letter

Quantum Zeno and anti-Zeno effects by indirect measurement with finite errors

We study the quantum Zeno effect and the anti-Zeno effect in the case of `indirect' measurements, where a measuring apparatus does not act directly on an unstable system, for a realistic model with finite errors in the measurement. A general and simple formula for the decay rate of the unstable system under measurement is derived. In the case of a Lorentzian form factor, we calculate the full time evolutions of the decay rate, the response of the measuring apparatus, and the probability of errors in the measurement. It is shown that not only the response time but also the detection efficiency plays a crucial role. We present the prescription for observing the quantum Zeno and anti-Zeno effects, as well as the prescriptions for avoiding or calibrating these effects in general experiments.Comment: 4 pages, 3 figure

Adhesion mechanisms of the contact interface of TiO2 nanoparticles in films and aggregates

Fundamental knowledge about the mechanisms of adhesion between oxide particles with diameters of few nanometers is impeded by the difficulties associated with direct measurements of contact forces at such a small size scale. Here we develop a strategy based on AFM force spectroscopy combined with all-atom molecular dynamics simulations to quantify and explain the nature of the contact forces between 10 nm small TiO2 nanoparticles. The method is based on the statistical analysis of the force peaks measured in repeated approaching/retracting loops of an AFM cantilever into a film of nanoparticle agglomerates and relies on the in-situ imaging of the film stretching behavior in an AFM/TEM setup. Sliding and rolling events first lead to local rearrangements in the film structure when subjected to tensile load, prior to its final rupture caused by the reversible detaching of individual nanoparticles. The associated contact force of about 2.5 nN is in quantitative agreement with the results of molecular dynamics simulations of the particle–particle detachment. We reveal that the contact forces are dominated by the structure of water layers adsorbed on the particles’ surfaces at ambient conditions. This leads to nonmonotonous force–displacement curves that can be explained only in part by classical capillary effects and highlights the importance of considering explicitly the molecular nature of the adsorbates

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

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

Electron Transport Properties of Single-Molecule-Bearing Multiple Redox Levels Studied by EC-STM/STS

Multielectron systems as possible components of molecular electronics devices are attracting compelling experimental and theoretical interest. Here we studied by electrochemical scanning tunneling techniques (EC-STMicroscopy and EC-STSpectroscopy) the electron transport properties of a redox molecule endowed with two redox levels, namely, the hydroquinone/quinone (H2Q/Q) couple. By forming self-assembled monolayers on Au(111) of oligo-phenylene-vinylene (OPV) derivatized H2Q/Q moieties, we were able to explore the features of the tunneling current/overpotential relation in the EC-STS setup. The behavior of the tunneling current sheds light onto the mechanism of electron transport involving the redox levels of the H2Q/Q redox pair coupled to tip and substrate electrodes

Expression and Differential Responsiveness of Central Nervous System Glial Cell Populations to the Acute Phase Protein Serum Amyloid A

Acute-phase response is a systemic reaction to environmental/inflammatory insults and involves hepatic production of acute-phase proteins, including serum amyloid A (SAA). Extrahepatically, SAA immunoreactivity is found in axonal myelin sheaths of cortex in Alzheimer's disease and multiple sclerosis (MS), although its cellular origin is unclear. We examined the responses of cultured rat cortical astrocytes, microglia and oligodendrocyte precursor cells (OPCs) to master pro-inflammatory cytokine tumour necrosis factor (TNF)-\u3b1 and lipopolysaccaride (LPS). TNF-\u3b1 time-dependently increased Saa1 (but not Saa3) mRNA expression in purified microglia, enriched astrocytes, and OPCs (as did LPS for microglia and astrocytes). Astrocytes depleted of microglia were markedly less responsive to TNF-\u3b1 and LPS, even after re-addition of microglia. Microglia and enriched astrocytes showed complementary Saa1 expression profiles following TNF-\u3b1 or LPS challenge, being higher in microglia with TNF-\u3b1 and higher in astrocytes with LPS. Recombinant human apo-SAA stimulated production of both inflammatory mediators and its own mRNA in microglia and enriched, but not microglia-depleted astrocytes. Co-ultramicronized palmitoylethanolamide/luteolin, an established anti-inflammatory/neuroprotective agent, reduced Saa1 expression in OPCs subjected to TNF-\u3b1 treatment. These last data, together with past findings suggest that co-ultramicronized palmitoylethanolamide/luteolin may be a novel approach in the treatment of inflammatory demyelinating disorders like MS