MOLECULAR AND CELLULAR MECHANISMS IN ASTROCYTE-T CELL CROSS-TALK

Migration of encephalitogenic T cells into the brain parenchima through the blood\u2013brain barrier (BBB) is a crucial feature for initiating tissue injury in different neuroinflammatory diseases. The BBB is comprised of astrocyte processes and endothelial cells, which form the lumen of the brain microvasculature and help in maintaining immune quiescence through contact-dependent mechanisms as well as release of soluble factors. Activated CD4+ T cells may establish physical contacts with astrocytes, thereby reciprocally influencing cellular activity and functions. In addition, astrocytes and CD4 T cells may communicate through the secretion of soluble signaling molecules during contact. Among molecules secreted by astrocyte, ATP is a key messenger which can also signal to CD4 T cell through purinergic P2 receptors. Our results show that activated CD4+ T cells inhibit calcium oscillations in astrocytes through direct modulation of extracellular ATP levels. This effect correlated with the expression of plasma membrane ectonucleoside triphosphate diphosphohydrolase CD39, which is induced by contact of the activated T cell with astrocyte. In addition, T cell contact with astrocyte results in the upregulation of the ecto-5\u2019-nucleotidase CD73, which converts AMP to adenosine. This effect was peculiar of T cell contact with astrocyte since it did not occur with microglia or peritoneal macrophages. Pharmacological inhibition of Ca2+ oscillations in astrocyte completely prevented CD73 induction on T cell, thus suggesting that a gliostrasmitter released by astrocyte in a Ca2+-dependent fashion might be responsible of this effect. Since degradation of ATP to adenosine by CD73 regulates BBB permeability and leukocytes infiltration into the brain this regulatory circuit might have important pathogenetic implications in multiple sclerosis and other neuroinflammatory conditions. Finally, functional characterization of T cell upon contact with astrocyte allowed us to assess a proinflammatory phenotype and Th17 skewing albeit with important differences, such as CD39 and CD73 expression, with respect to conventionally activated cells. Thus, we characterized an astrocyte specific signature of T cell activation, which might be important in the pathogenesis of neuroinflammatory disorders

Ectonucleotidase activity and immunosuppression in astrocyte-CD4 T cell bidirectional signaling

Astrocytes play a crucial role in neuroinflammation as part of the glia limitans, which regulates infiltration of the brain parenchyma by leukocytes. The signaling pathways and molecular events, which result from the interaction of activated T cells with astrocytes are poorly defined. Here we show that astrocytes promote the expression and enzymatic activity of CD39 and CD73 ectonucleotidases in recently activated CD4 cells by a contact dependent mechanism that is independent of T cell receptor interaction with class II major histocompatibility complex (MHC). Transforming growth factor-\u3b2 (TGF-\u3b2) is robustly upregulated and sufficient to promote ectonucleotidases expression. T cell adhesion to astrocyte results in differentiation to an immunosuppressive phenotype defined by expression of the transcription factor Roryt, which characterizes the CD4 T helper 17 subset. CD39 activity in T cells in turn inhibits spontaneous calcium oscillations in astrocytes that correlated with enhanced and reduced transcription of CCL2 chemokine and Sonic hedgehog (Shh), respectively. We hypothesize this TCR-independent interaction promote an immunosuppressive program in T cells to control possible brain injury by deregulated T cell activation during neuroinflammation. On the other hand, the increased secretion of CCL2 with concomitant reduction of Shh might promote leukocytes extravasation into the brain parenchyma

Assessment of the antibiotic resistance profile, genetic heterogeneity and biofilm production of methicillin-resistant staphylococcus aureus (MRSA) isolated from the italian swine production chain

The main aim of the present study was to evaluate the level of antibiotic resistance, prevalence and virulence features of methicillin-resistant Staphylococcus aureus (MRSA) isolated from heavy swine at abattoir level and farming environments in Lombardy (Northern Italy). With this scope, 88 different heavy swine farms were surveyed, obtaining a total of n = 440 animal swabs and n = 150 environmental swabs. A total of n = 87 MRSA isolates were obtained, with an overall MRSA incidence of 17.50% (n = 77) among animal samples and a 6.67% (n = 10) among environmental. Molecular characterisation using multilocus sequence typing (MLST) plus spa-typing showed that sequence type ST398/t899 and ST398/t011 were the most commonly isolated genotypes, although other relevant sequence types such as ST1 or ST97 were also found. A lack of susceptibility to penicillins, tetracycline and ceftiofur was detected in >91.95, 85.05 and 48.28% of the isolates, respectively. Resistance to doxycycline (32.18%), enrofloxacin (27.59%) and gentamicin (25.29%) was also observed. Additionally, a remarkable level of antibiotic multiresistance (AMR) was observed representing a 77.01% (n = 67) of the obtained isolates. Genetic analysis revealed that 97.70% and 77.01% of the isolates harboured at least one antibiotic resistance or enterotoxin gene, respectively, pointing out a high isolate virulence potential. Lastly, 55.17% (n = 48) were able to produce measurable amounts of biofilm after 24 h. In spite of the current programmes for antibiotic reduction in intensively farming, a still on-going high level of AMR and virulence potential in MRSA was demonstrated, making this pathogen a serious risk in swine production chain, highlighting once more the need to develop efficient, pathogen-specific control strategies

Torque Teno Sus Virus (TTSuV) Prevalence in Wild Fauna of Northern Italy

Torque teno sus virus (TTSuV) is a non-enveloped circular ssDNA virus which frequently infects swine and has been associated with hepatic, respiratory, and autoimmune disorders. TTSuV’s pathogenic role is still uncertain, and clear data in the literature on virus reservoirs are lacking. The aims of this study were to investigate the presence of potentially zoonotic TTSuV in wild animals in Northern Italy and to evaluate their role as reservoirs. Liver samples were collected between 2016 and 2020 during four hunting seasons from wild boars (Sus scrofa), red deer (Cervus elaphus), roe deer (Capreolus capreolus), and chamois (Rupicapra rupicapra). Samples originated from areas in Northern Italy characterized by different traits, i.e., mountains and flatland with, respectively low and high farm density and anthropization. Viral identification was carried out by end-point PCR with specific primers for TTSuV1a and TTSuVk2a species. TTSuV prevalence in wild boars was higher in the mountains than in the flatland (prevalence of 6.2% and 2.3%, respectively). In wild ruminants only TTSuVk2a was detected (with a prevalence of 9.4%). Our findings shed light on the occurrence and distribution of TTSuV in some wild animal species, investigating their possible role as reservoirs

TREM2 is required for microglial instruction of astrocytic synaptic engulfment in neurodevelopment

Variants in the microglial receptor TREM2 confer risk for multiple neurodegenerative diseases. However, it remains unknown how this receptor functions on microglia to modulate these diverse neuropathologies. To understand the role of TREM2 on microglia more generally, we investigated changes in microglial function in Trem2−/− mice. We found that loss of TREM2 impairs normal neurodevelopment, resulting in reduced synapse number across the cortex and hippocampus in 1-month-old mice. This reduction in synapse number was not due directly to alterations in interactions between microglia and synapses. Rather, TREM2 was required for microglia to limit synaptic engulfment by astrocytes during development. While these changes were largely normalized later in adulthood, high fat diet administration was sufficient to reinitiate TREM2-dependent modulation of synapse loss. Together, this identifies a novel role for microglia in instructing synaptic pruning by astrocytes to broadly regulate appropriate synaptic refinement, and suggests novel candidate mechanisms for how TREM2 and microglia could influence synaptic loss in brain injury and disease

Morphologic Features of Invasion in Lung Adenocarcinoma: Diagnostic Pitfalls.

Reproducibility of pulmonary invasive adenocarcinoma diagnosis is poor when applying the World Health Organization (WHO) classification. In this article, we aimed first to explain by 3-dimensional morphology why simple pattern recognition induces pitfalls for the assessment of invasion as applied in the current WHO classification of pulmonary adenocarcinomas. The underlying iatrogenic-induced morphologic alterations in collapsed adenocarcinoma in situ overlap with criteria for invasive adenocarcinoma. Pitfalls in seemingly acinar and papillary carcinoma are addressed with additional cytokeratin 7 and elastin stains. In addition, we provide more stringent criteria for a better reproducible and likely generalizable classification