DYSREGULATION OF THE ILT7/BST2 PDC NEGATIVE FEEDBACK BY HIV-1:IMPLICATIONS FOR HIV-1 TRANSMISSION AND IMMUNOPATHOGENESIS

Introduction: The immunoglobulin like transcript 7 (ILT7) is a surface molecule selectively expressed by human plasmacytoid dendritic cell (pDC). ILT7 cross-linking inhibits Toll like receptor (TLR) 7/9-mediated pDC activation and type I interferon (IFNI) production. The bone marrow stromal cell antigen 2 (BST2) is a natural ligand for ILT7, is expressed on several cell types and encoded by an IFN I-stimulated gene. BST2/ ILT7 interaction may provide a negative feedback for pDC activation. Alterations of the BST2/ILT7 negative feedback may contribute to HIV1-induced pDC over-activation and pathogenesis. We tested: 1) if BST2/ILT7 expression inperipheral blood mononuclear cell (PBMC) correlates with TLR-mediated pDC activation; 2) which stimuli can influence BST2 expression and IFN-I production by PBMC; 3) if TLR-induced pDC activation is directly modulated by BST2-expressing cells in vitro. Methods: PBMC from healthy donors were cultured overnight with or without imiquimod (TLR7L), CpG ODN (TLR9L), AT2-HIV1, TNF\u3b1, IFN\u3b3, IL4 IL10, Anti-Human Interferon Alpha/Beta Receptor Chain 2 (IFNAR2), BST2-GST fusion protein. T cells were stimulated using CD3 antibody. The effect of BST2 blockade and ILT7 cross-linking were tested using an anti-BST2 and cross linking-ILT7 monoclonal antibodies (mAbs), respectively. ILT7, BST2, CD83 and CCR7 expression was analyzed by flow cytometry. 293T cell lines transfected with BST2WT, or BST2 mutants, were used to test anti-BST2 mAb efficiency of binding or co-cultured with purified pDC to test the biologic effect of BST2. IFN\u3b1 production was quantified by ELISA. Statistical analyses were performed using SPSS 19.0. Results:pDC exclusively expressed ILT7, which was rapidly downregulated in vitro as part of a first step of pDC differentiation, characterized by an increase of the pDC morphological complexity and CCR7 expression. CD83 expression, indicative of full pDC activation and maturation, occurred only following TLR stimulation. Conversely, BST2 expression was not affected by in vitro culture; it was highest in monocytes, mDC and B cells compared to pDC and T cells and it was modulated by TLR7/9L-induced IFN\u3b1 production. BST2 expression on pDC was highest at intermediate stimuli concentrations but modestly increased at maximum concentrations; a profile which correlated with CD83 expression and IFN\u3b1 production but not with indoleamine 2,3dioxygenase (IDO) activity after HIV stimulation. PBMC pre-treatment with ILT7 cross-linking mAbs reduced both TLR9L/HIV-induced IFN\u3b1 production and HIV-induced IDO activity. In contrast, pre-treatment with blocking BST2 Abs did not increase IFN\u3b1 production or IDO activity. The lack of biological effect of BST2 was not due to inefficient \u3b1BST2 Ab binding, as PE-labelled \u3b1BST2 Ab efficiently stained BST2-transfected 293T cell lines. No change in IFN\u3b1 production were observed using either a soluble BST2 protein or a co-culture system based on purified pDC and BST2WT transfected 293T cells. T cell receptor engagement resulted in maximum BST2 expression on T cells, but BST2-blocking mAbs did not affect IFN\u3b1 release even when BST2 expression on T cell was enhanced. IL10 and TNF\u3b1 inhibited TLR9L-induced IFN\u3b1 production but BST2 blockade did not restored IFN\u3b1 responses. Conclusions: Our data suggest that ILT7 cross-linking may acts as homeostatic mechanism on circulating pDC rather than a negative feedback for activated mature pDC, and argue against the role of BST2 as a biologically active ILT7 ligand

Neural entrainment via perceptual inferences

Entrainment depends on sequential neural phase reset by regular stimulus onset, a temporal parameter. Entraining to sequences of identical stimuli also entails stimulus feature predictability, but this component is not readily separable from temporal regularity. To test if spectral regularities concur with temporal regularities in determining the strength of auditory entrainment, we devised sound sequences that varied in conditional perceptual inferences based on deviant sound repetition probability: strong inference (100% repetition probability: If a deviant appears, then it will repeat), weak inference (75% repetition probability) and no inference (50%: A deviant may or may not repeat with equal probability). We recorded EEG data from 15 young human participants pre-attentively listening to the experimental sound sequences delivered either isochronously or anisochronously (±20% jitter), at both delta (1.67 Hz) and theta (6.67 Hz) stimulation rates. Strong perceptual inferences significantly enhanced entrainment at either stimulation rate and determined positive correlations between precision in phase distribution at the onset of deviant trials and entrained power. We conclude that both spectral predictability and temporal regularity govern entrainment via neural phase control

Effect of Immunoglobin-Like Transcript 7 Cross-Linking on Plasmacytoid Dendritic Cells Differentiation into Antigen-Presenting Cells

Plasmacytoid dendritic cells (pDC) are innate immunity effector cells which play a critical role in the transition from innate to adaptive immune response. Circulating blood pDC present an immature phenotype and can differentiate into either antigen-presenting cells (APC) or type I interferon (IFN-I)-producing cells (IPC). The immunoglobulin-like transcript (ILT)7 is a surface receptor expressed by immature pDC, and ILT7 cross-linking (XL-ILT7) inhibits IFN-I production by pDC in response to toll-like receptor (TLR)7 and 9 stimulation. We used peripheral blood mononuclear cells (PBMC) from healthy donors to test the effect of XL-ILT7 on 1) TLR7/9-mediated regulation of gut mucosal (α4β7 integrin) and lymph node (CCR7) migration markers; and 2) the maturation of pDC into APC. We found that XL-ILT7 mitigated the upregulation of CCR7 and enhanced that of β7 on TLR7/9-stimulated pDC. TLR7/9 stimulation induced upregulation of CD40, CD80 and CD86. CD40 expression was partially reduced by XL-ILT7, whereas CD86 was further enhanced. Plasmacytoid DC stimulated with TLR9 ligand in presence of XL-ILT7 retained the ability to induce T cell proliferation and activation in response to staphylococcal enterotoxin B (SEB) in pDC-T cell co-cultures. Our results suggest that XL-ILT7 favours the differentiation of immature pDC into APC rather than IPC

Host-Based Treatments for Severe COVID-19

COVID-19 has been a global health problem since 2020. There are different spectrums of manifestation of this disease, ranging from asymptomatic to extremely severe forms requiring admission to intensive care units and life-support therapies, mainly due to severe pneumonia. The progressive understanding of this disease has allowed researchers and clinicians to implement different therapeutic alternatives, depending on both the severity of clinical involvement and the causative molecular mechanism that has been progressively explored. In this review, we analysed the main therapeutic options available to date based on modulating the host inflammatory response to SARS-CoV-2 infection in patients with severe and critical illness. Although current guidelines are moving toward a personalised treatment approach titrated on the timing of presentation, disease severity, and laboratory parameters, future research is needed to identify additional biomarkers that can anticipate the disease course and guide targeted interventions on an individual basis

The Soluble Recombinant Neisseria meningitidis Adhesin NadAΔ351–405 Stimulates Human Monocytes by Binding to Extracellular Hsp90

The adhesin NadA favors cell adhesion/invasion by hypervirulent Neisseria meningitidis B (MenB). Its recombinant form NadAΔ351–405, devoid of the outer membrane domain, is an immunogenic candidate for an anti-MenB vaccine able to stimulate monocytes, macrophages and dendritic cells. In this study we investigated the molecular mechanism of NadAΔ351–405 cellular effects in monocytes. We show that NadAΔ351–405 (against which we obtained polyclonal antibodies in rabbits), binds to hsp90, but not to other extracellular homologous heat shock proteins grp94 and hsp70, in vitro and on the surface of monocytes, in a temperature dependent way. Pre-incubation of monocytes with the MenB soluble adhesin interfered with the binding of anti-hsp90 and anti-hsp70 antibodies to hsp90 and hsp70 at 37°C, a condition in which specific cell-binding occurs, but not at 0°C, a condition in which specific cell-binding is very diminished. Conversely, pre-incubation of monocytes with anti-hsp90 and anti-hsp70 antibodies did not affected NadAΔ351–405 cell binding in any temperature condition, indicating that it associates to another receptor on their plasma membrane and then laterally diffuses to encounter hsp90. Consistently, polymixin B interfered with NadAΔ351–405 /hsp90 association, abrogated the decrease of anti-hsp90 antibodies binding to the cell surface due to NadAΔ351–405 and inhibited adhesin-induced cytokine/chemokine secretion without affecting monocyte-adhesin binding. Co-stimulation of monocytes with anti-hsp90 antibodies and NadAΔ351–405 determined a stronger but polymixin B insensitive cell activation. This indicated that the formation of a recombinant NadA/hsp90/hsp70 complex, although essential for full monocyte stimulation, can be replaced by anti-hsp90 antibody/hsp90 binding. Finally, the activation of monocytes by NadAΔ351–405 alone or in the presence of anti-hsp90 antibodies were both inhibited by neutralizing anti-TLR4 antibodies, but not by anti-TLR2 antibodies. We propose that hsp90-dependent recruitment into an hsp90/hsp70/TLR4 transducing signal complex is necessary for the immune-stimulating activity of NadAΔ351–405 anti-MenB vaccine candidate

Changes in phytoplankton communities along the Northern Antarctic Peninsula: Causes, impacts and research priorities

The Northern Antarctic Peninsula (NAP), located in West Antarctica, is amongst the most impacted regions by recent warming events. Its vulnerability to climate change has already led to an accumulation of severe changes along its ecosystems. This work reviews the current findings on impacts observed in phytoplankton communities occurring in the NAP, with a focus on its causes, consequences, and the potential research priorities toward an integrated comprehension of the physical–biological coupling and climate perspective. Evident changes in phytoplankton biomass, community composition and size structure, as well as potential bottom-up impacts to the ecosystem are discussed. Surface wind, sea ice and meltwater dynamics, as key drivers of the upper layer structure, are identified as the leading factors shaping phytoplankton. Short- and long-term scenarios are suggested for phytoplankton communities in the NAP, both indicating a future increase of the importance of small flagellates at the expense of diatoms, with potential devastating impacts for the ecosystem. Five main research gaps in the current understanding of the phytoplankton response to climate change in the region are identified: (i) anthropogenic signal has yet to be disentangled from natural climate variability; (ii) the influence of small-scale ocean circulation processes on phytoplankton is poorly understood; (iii) the potential consequences to regional food webs must be clarified; (iv) the magnitude and risk of potential changes in phytoplankton composition is relatively unknown; and (v) a better understanding of phytoplankton physiological responses to changes in the environmental conditions is required. Future research directions, along with specific suggestions on how to follow them, are equally suggested. Overall, while the current knowledge has shed light on the response of phytoplankton to climate change, in order to truly comprehend and predict changes in phytoplankton communities, there must be a robust collaboration effort integrating both Antarctic research programs and the whole scientific community under a common research framework

Evidence for sub-haplogroup h5 of mitochondrial DNA as a risk factor for late onset Alzheimer's disease

BACKGROUND: Alzheimer's Disease (AD) is the most common neurodegenerative disease and the leading cause of dementia among senile subjects. It has been proposed that AD can be caused by defects in mitochondrial oxidative phosphorylation. Given the fundamental contribution of the mitochondrial genome (mtDNA) for the respiratory chain, there have been a number of studies investigating the association between mtDNA inherited variants and multifactorial diseases, however no general consensus has been reached yet on the correlation between mtDNA haplogroups and AD. METHODOLOGY/PRINCIPAL FINDINGS: We applied for the first time a high resolution analysis (sequencing of displacement loop and restriction analysis of specific markers in the coding region of mtDNA) to investigate the possible association between mtDNA-inherited sequence variation and AD in 936 AD patients and 776 cognitively assessed normal controls from central and northern Italy. Among over 40 mtDNA sub-haplogroups analysed, we found that sub-haplogroup H5 is a risk factor for AD (OR=1.85, 95% CI:1.04-3.23) in particular for females (OR=2.19, 95% CI:1.06-4.51) and independently from the APOE genotype. Multivariate logistic regression revealed an interaction between H5 and age. When the whole sample is considered, the H5a subgroup of molecules, harboring the 4336 transition in the tRNAGln gene, already associated to AD in early studies, was about threefold more represented in AD patients than in controls (2.0% vs 0.8%; p=0.031), and it might account for the increased frequency of H5 in AD patients (4.2% vs 2.3%). The complete re-sequencing of the 56 mtDNAs belonging to H5 revealed that AD patients showed a trend towards a higher number (p=0.052) of sporadic mutations in tRNA and rRNA genes when compared with controls. CONCLUSIONS: Our results indicate that high resolution analysis of inherited mtDNA sequence variation can help in identifying both ancient polymorphisms defining sub-haplogroups and the accumulation of sporadic mutations associated with complex traits such as AD

Threatened and extinct amphibians and reptiles in Italian natural history collections are useful conservation tools

Natural history museums are irreplaceable tools to study and preserve the biological diversity around the globe and among the primary actors in the recognition of species and the logical repositories for their type specimens. In this paper we surveyed the consistency of the preserved specimens of amphibians and reptiles housed in the major Italian scientific collections, and verified the presence of threatened species according to the IUCN Red List, includ-ing the Extinct (EX), Extinct in the Wild (EW), Critically Endangered (CR), Endangered (EN), and Vulnerable (VU) categories. Altogether, we analyzed 39 Italian zoological collections. We confirmed the presence of one extinct reptile (Chioninia coctei) and five extinct or extinct in the wild amphibian species (Atelopus longirostris, Nectophrynoides asperginis, Pseudophilautus leucorhinus, P. nasutus, and P. variabilis). Seven CR amphibians, fourteen CR reptile species and the extinct skink C. coctei are shared by more than one institution. Museums which host the highest number of threatened and extinct amphibian species are respectively Turin (17 CR and 1 EX), Florence (13 CR and 1 EX), and Trento (15 CR and 1 EW), while for reptiles the richest museums are those from Genoa (15 CR and 1 EX), Florence (11 CR and 1 EX), and Pisa (7 CR). Finally, we discussed the utility of natural history museums and the strategies to follow for the implementation of their functionality. © Firenze University Press

Segregation of Fluorescent Membrane Lipids into Distinct Micrometric Domains: Evidence for Phase Compartmentation of Natural Lipids?

Background: We recently reported that sphingomyelin (SM) analogs substituted on the alkyl chain by various fluorophores (e.g. BODIPY) readily inserted at trace levels into the plasma membrane of living erythrocytes or CHO cells and spontaneously concentrated into micrometric domains. Despite sharing the same fluorescent ceramide backbone, BODIPY-SM domains segregated from similar domains labelled by BODIPY-D-e-lactosylceramide (D-e-LacCer) and depended on endogenous SM. Methodology/Principal Findings. We show here that BODIPY-SM further differed from BODIPY-D-e-LacCer or -glucosylceramide (GlcCer) domains in temperature dependence, propensity to excimer formation, association with a glycosylphosphatidylinositol (GPI)-anchored fluorescent protein reporter, and lateral diffusion by FRAP, thus demonstrating different lipid phases and boundaries. Whereas BODIPY-D-e-LacCer behaved like BODIPY-GlcCer, its artificial stereoisomer, BODIPY-L-t-LacCer, behaved like BODIPY- and NBD-phosphatidylcholine (PC). Surprisingly, these two PC analogs also formed micrometric patches yet preferably at low temperature, did not show excimer, never associated with the GPI reporter and showed major restriction to lateral diffusion when photobleached in large fields. This functional comparison supported a three-phase micrometric compartmentation, of decreasing order: BODIPY-GSLs > -SM > -PC (or artificial L-t-LacCer). Co-existence of three segregated compartments was further supported by double labelling experiments and was confirmed by additive occupancy, up to ~70% cell surface coverage. Specific alterations of BODIPY-analogs domains by manipulation of corresponding endogenous sphingolipids suggested that distinct fluorescent lipid partition might reflect differential intrinsic propensity of endogenous membrane lipids to form large assemblies. Conclusions/Significance. We conclude that fluorescent membrane lipids spontaneously concentrate into distinct micrometric assemblies. We hypothesize that these might reflect preexisting compartmentation of endogenous PM lipids into non-overlapping domains of differential order: GSLs > SM > PC, resulting into differential self-adhesion of the two former, with exclusion of the latter