Conference Agenda

Meningeal inflammation is a detrimental process involved in several dysfunctions of the central nervous system (CNS). Particularly, this pathological process has been recently described as a specific feature of patients affected by multiple sclerosis (MS) and characterized by a worse prognosis. The link between neutrophil contribution to leptomeningeal inflammation and MS pathogenesis has been demonstrated by pioneer studies either directly using samples from MS patients or indirectly exploiting the most common animal model of MS pathology, namely the experimental autoimmune encephalomyelitis (EAE). Specifically, neutrophil extravasation in the sub-arachnoid space correlates with a more severe disease outcome, but how neutrophils interact with other local immune cells and contribute to neuroinflammation in the context of MS is largely unknown.

Our work aimed at characterizing the crosstalk between neutrophils and leptomeningeal macrophages, potentially representing a driver of neuropathological changes during MS and EAE. Single-cell RNA-sequencing (scRNA-seq) data showed that spinal cord-infiltrating neutrophils at EAE onset were enriched in migratory and cell-cell interaction pathways compared to those from the chronic phase of the disease. Indeed, in vivo two-photon laser scanning microscopy (TPLSM) functional studies showed a highly migratory neutrophil phenotype with a predominant fraction of infiltrating neutrophils localized close to leptomeningeal blood vessels and establishing contacts with local macrophages. Wide-field live imaging confirmed the crosstalk between leptomeningeal neutrophils and macrophages and identified the molecular mechanisms of these cell-cell interactions during EAE. Immunohistological results suggested that neutrophils may also interact with leptomeningeal macrophages in MS patients. Our data indicate that myeloid cell crosstalk at CNS borders may contribute to inflammation amplification and disease development suggesting that interfering with meningeal neutrophil-macrophages crosstalk may have therapeutic relevance for neuroinflammatory and autoimmune diseases.

A bidirectional crosstalk between peripheral players of immunity and the central nervous system (CNS) has been demonstrated. The blood–brain barrier (BBB) protects neurons from factors present in the periphery and maintains the highly regulated CNS internal milieu, which is required for proper synaptic and neuronal functioning.

As BBB breakdown is a factor contributing to, or even anticipating, neuronal dysfunction(s), in the present study we identify the alterations of the different components of BBB– such as astrocytes, pericytes and the endothelial cells–in a 6-hydroxydopamine (6-OHDA)-lesioned hemiparkinsonian rat model. Furthermore, in the same experimental model we assessed the effects of a non-pharmacological therapeutic strategy, namely Transcranial magnetic stimulation (TMS), on the different components of BBB.

In the striatum of 6-OHDA animals, mRNA and protein levels of markers of pericytes, such as PDGFβ, decreased significantly after the lesion compared to control animals. Notably, in 6-OHDA-lesioned animals, the decrease of pericytes markers correlated with a reduction of PECAM-1 (also known as CD31), a marker of vessel endothelial cells, and these changes were associated with impairments in brain microvasculature. After the injury, treatment with TMS protected both pericytes and endothelial cells of the striatal microvasculature from 6-OHDA-induced damage. Similarly, treatment restored microvessels integrity by restoring lectin and PECAM-1 expression, thus improving microvasculature integrity, that was found similar to what was observed in unlesioned animals. In parallel, the treatment decreased astrocytes reactivity and improved the expression of a key glial transporter in the reuptake of glutamate, namely GLAST in the astrocytes. Collectively, these findings open new therapeutic scenarios of paramount importance, demonstrating the potential of TMS as non-pharmacological approach to restore periphery-CNS communication, altered in several CNS neurodegenerative diseases.

Alzheimer’s disease (AD) is the most common age-related neurodegenerative disorder associated with accumulation of amyloid-β and hyperphosphorylation of the tau protein in the brain. Dysregulation of inflammatory/immune response is regarded as one of the major contributors to AD pathogenesis. Chronic activation of the brain-resident innate immune cells, peripheral leukocytes access across the brain barriers, as well as the release of inflammatory and neurotoxic factors, shape the neuroinflammatory response that drives the progression of neurodegenerative processes in AD.

The proinflammatory cytokine Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), a member of the TNF superfamily formerly known as TNFSF10, is substantially expressed in the human AD brain, where it actively modulates the immune response. TNFSF10 is abundantly produced and released by activated glia, brain-infiltrating macrophages, and damaged neurons, acting as a cell death signal. In addition, TNFSF10 promotes the recruitment of peripheral regulatory T cells in the brain of 3xTg-AD mice, thereby limiting the potential of immune system to control the accumulation of anomalous proteins.

Consistently, pharmacological neutralization of TNFSF10 in 3xTg-AD mice implies significant functional recovery, associated with a reduced expression of various inflammatory mediators and re-balance of both central and peripheral immune response, paralleled by dramatically decreased burden of Aβ and p-Tau. TNFSF10-antagonistic treatment demonstrates also a remarkable ability to alleviate the immune suppression typical of AD and orchestrate a phenotypic switching of monocytes/macrophages, leading them towards an anti-inflammatory/neuroprotective state in 3xTg-AD mice. Notably, neutralization of TNFSF10 ameliorate tissue pathology and inflammatory parameters within the retina of AD animals.

In conclusion, it appears plausible to hypothesize that TNFSF10 system-targeted treatments effectively restrain excessive neuroinflammation through a rebalancing effect on the immune response, thereby contributing to the attenuation of pathological processes that drive uncontrolled and progressive neurodegeneration in AD.

Physiological or pathological peripheral stimuli determine the response of the central nervous system (CNS) circuitry. The interface between the body and the CNS is characterized by a dynamic cellular and molecular diversity with an elaborate vascular network. We are analyzing the early changes in the spinal cord neurovascular unit (NVU) after peripheral damage using a rat model of sural spared nerve injury (SNI). Following the SNI procedure we analyzed the dorsal horn transsynaptic deafferentation and the ventral horn motor neuron axotomy using 1-, 2- or 7-day(s) time points. The analyses are unraveling the pleiotropic effects of the coagulation protein thrombin, through its main receptor (PAR-1). PAR-1 is firstly localized to the vascular network and neurons and is progressively clustered in rafts close to the astrocytic end-feet while the extracellular milieu is deeply reshaped with the contribution of metalloproteinase 9 (MMP9). Moreover, we showed early modifications of the blood-spinal cord interface changing the expression of tight junctions, basal lamina, and channel proteins. The reorganization of astrocytic water channels (AQP4) and gap junctions or hemichannels (Cx43) is paralleled by a steady invasion of microglial/macrophagic elements, differently affecting the dorsal and ventral horn of the spinal cord. Our data are deepening the understanding of NVU early response following peripheral injury, beyond the simplistic integrity of the so-called blood-brain barrier.