Regulation of Lymphatic GM-CSF Expression by the E3 Ubiquitin Ligase Cbl-b

Genome-wide association studies as well as lymphatic expression analyses have linked both Cbl-b and GM-CSF to human multiple sclerosis as well as other autoimmune diseases. Both Cbl-b and GM-CSF have been shown to play a prominent role in the development of murine encephalomyelitis; however, no functional connection between the two has yet been established. In this study, we show that Cblb knockout mice demonstrated significantly exacerbated severity of experimental autoimmune encephalomyelitis (EAE), augmented T cell infiltration into the central nervous system (CNS) and strongly increased production of GM-CSF in T cells in vitro and in vivo.GM-CSF neutralization demonstrated that the increased susceptibility of Cblb−/− mice to EAE was dependent on GM-CSF. Mechanistically, p50 binding to the GM-CSF promoter and the IL-3/GM-CSF enhancer element “CNSa” was strongly increased in nuclear extracts from Cbl-b-deficient T cells. This study suggests that Cbl-b limits autoimmunity by preventing the pathogenic effects of GM-CSF overproduction in T cells

B7h triggering inhibits the migration of tumor cell lines

Vascular endothelial cells (ECs) and several cancer cells express B7h, which is the ligand of the ICOS T cell costimulatory molecule. We have previously shown that B7h triggering via a soluble form of ICOS (ICOS-Fc) inhibits the adhesion of polymorphonuclear and tumor cell lines to HUVECs; thus, we suggested that ICOS-Fc may act as an anti-inflammatory and antitumor agent. Because cancer cell migration and angiogenesis are crucial for metastasis dissemination, the aim of this work was to evaluate the effect of ICOS-Fc on the migration of cancer cells and ECs. ICOS-Fc specifically inhibited the migration of HUVECs, human dermal lymphatic ECs, and the HT29, HCT116, PC-3, HepG2, JR8, and M14 tumor cell lines expressing high levels of B7h, whereas it was ineffective in the RPMI7932, PCF-2, LM, and BHT-101 cell lines expressing low levels of B7h. Furthermore, ICOS-Fc downmodulated hepatocyte growth factor facilitated the epithelial-to-mesenchymal transition in HepG2 cells. Moreover, ICOS-Fc downmodulated the phosphorylation of focal adhesion kinase and the expression of \u3b2-Pix in both HUVECs and tumor cell lines. Finally, treatment with ICOS-Fc inhibited the development of lung metastases upon injection of NOD-SCID-IL2R\u3b3null mice with CF-PAC1 cells, as well as C57BL/6 mice with B16-F10 cells. Therefore, the B7h-ICOS interaction may modulate the spread of cancer metastases, which suggests the novel use of ICOS-Fc as an immunomodulatory drug. However, in the B16-F10-metastasized lungs, ICOS-Fc also increased IL-17A/RORc and decreased IL-10/Foxp3 expression, which indicates that it also exerts positive effects on the antitumor immune response

Role of anti-osteopontin antibodies in multiple sclerosis and experimental autoimmune encephalomyelitis

Osteopontin (OPN) is highly expressed in demyelinating lesions in multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE). OPN is cleaved by thrombin into N- (OPN-N) and C-terminal (OPN-C) fragments with different ligands and functions. In EAE, administering recombinant OPN induces relapses, whereas treatment with anti-OPN antibodies ameliorates the disease. Anti-OPN autoantibodies (autoAbs) are spontaneously produced during EAE but have never been detected in MS. The aim of the study was to evaluate anti-OPN autoAbs in the serum of MS patients, correlate them with disease course, and recapitulate the human findings in EAE. We performed ELISA in the serum of 122 patients collected cross-sectionally, and 50 patients with relapsing-remitting (RR) disease collected at diagnosis and followed longitudinally for 10 years. In the cross-sectional patients, the autoAb levels were higher in the RR patients than in the primary- and secondary-progressive MS and healthy control groups, and they were highest in the initial stages of the disease. In the longitudinal group, the levels at diagnosis directly correlated with the number of relapses during the following 10 years. Moreover, in patients with active disease, who underwent disease-modifying treatments, autoAbs were higher than in untreated patients and were associated with low MS severity score. The autoAb displayed neutralizing activity and mainly recognized OPN-C rather than OPN-N. To confirm the clinical effect of these autoAbs in vivo, EAE was induced using myelin oligodendrocyte glycoprotein MOG35-55 in C57BL/6 mice pre-vaccinated with ovalbumin (OVA)-linked OPN or OVA alone. We then evaluated the titer of antibodies to OPN, the clinical scores and in vitro cytokine secretion by spleen lymphocytes. Vaccination significantly induced antibodies against OPN during EAE, decreased disease severity, and the protective effect was correlated with decreased T cell secretion of interleukin 17 and interferon-\u3b3 ex vivo. The best effect was obtained with OPN-C, which induced significantly faster and more complete remission than other OPN vaccines. In conclusion, these data suggest that production of anti-OPN autoAbs may favor remission in both MS and EAE. Novel strategies boosting their levels, such as vaccination or passive immunization, may be proposed as a future strategy in personalized MS therapy

Targeted delivery of a vaccine protein to Langerhans cells in the human skin via the C-type lectin receptor Langerin

Human skin is a preferred vaccination site as it harbors multiple dendritic cell (DC) subsets, which display distinct C-type lectin receptors (CLR) that recognize pathogens. Antigens can be delivered to CLR by antibodies or ligands to boost antigen-specific immune responses. This concept has been established in mouse models but detailed insights into the functional consequences of antigen delivery to human skin DC in situ are sparse. In this study, we cloned and produced an anti-human Langerin antibody conjugated to the EBV nuclear antigen 1 (EBNA1). We confirmed specific binding of anti-Langerin-EBNA1 to Langerhans cells (LC). This novel LC-based vaccine was then compared to an existing anti-DEC-205-EBNA1 fusion protein by loading LC in epidermal cell suspensions before coculturing them with autologous T cells. After restimulation with EBNA1-peptides, we detected elevated levels of IFN-γ- and TNF-α-positive CD4+ T cells with both vaccines. When we injected the fusion proteins intradermally into human skin explants, emigrated skin DC targeted via DEC-205-induced cytokine production by T cells, whereas the Langerin-based vaccine failed to do so. In summary, we demonstrate that antibody-targeting approaches via the skin are promising vaccination strategies, however, further optimizations of vaccines are required to induce potent immune responses

Determinants of long COVID among adults hospitalized for SARS-CoV-2 infection: A prospective cohort study

Rationale: Factors associated with long-term sequelae emerging after the acute phase of COVID-19 (so called "long COVID") are unclear. Here, we aimed to identify risk factors for the development of COVID-19 sequelae in a prospective cohort of subjects hospitalized for SARS-CoV-2 infection and followed up one year after discharge. Methods: A total of 324 subjects underwent a comprehensive and multidisciplinary evaluation one year after hospital discharge for COVID-19. A subgroup of 247/324 who consented to donate a blood sample were tested for a panel of circulating cytokines. Results: In 122 patients (37.8%) there was evidence of at least one persisting physical symptom. After correcting for comorbidities and COVID-19 severity, the risk of developing long COVID was lower in the 109 subjects admitted to the hospital in the third wave of the pandemic than in the 215 admitted during the first wave, (OR 0.69, 95%CI 0.51-0.93, p=0.01). Univariable analysis revealed female sex, diffusing capacity of the lungs for carbon monoxide (DLCO) value, body mass index, anxiety and depressive symptoms to be positively associated with COVID-19 sequelae at 1 year. Following logistic regression analysis, DLCO was the only independent predictor of residual symptoms (OR 0.98 CI 95% (0.96-0.99), p=0.01). In the subgroup of subjects with normal DLCO (> 80%), for whom residual lung damage was an unlikely explanation for long COVID, the presence of anxiety and depressive symptoms was significantly associated to persistent symptoms, together with increased levels of a set of pro-inflammatory cytokines: interferon-gamma, tumor necrosis factor-alpha, interleukin (IL)-2, IL-12, IL-1β, IL-17. In logistic regression analysis, depressive symptoms (p=0.02, OR 4.57 [1.21-17.21]) and IL-12 levels (p=0.03, OR 1.06 [1.00-1.11]) 1-year after hospital discharge were independently associated with persistence of symptoms. Conclusions: Long COVID appears mainly related to respiratory sequelae, prevalently observed during the first pandemic wave. Among patients with little or no residual lung damage, a cytokine pattern consistent with systemic inflammation is in place

Il confine internazionale tra Stati Uniti e Messico: identità transnazionali e impatti sui luoghi

Il Parlamento Europeo (2016) e l’OECD (2013) hanno riconosciuto un potenziale economico delle regioni di frontiera che si può esprimere nell’ambito del mercato globale. Tuttavia, recenti fenomeni sembrano negare l’esplicarsi di questo potenziale, come la recente Brexit e le tensioni separatiste emerse in Catalogna. La tendenza populista che questi fenomeni sottendono alimenta processi disgregativi sia sul piano sociale che politico. In Nord America il nuovo contesto politico, successivo alle elezioni del 2016, ha fortemente cambiato gli scenari al confine con il vicino e meno sviluppato Messico. Le crescenti tensioni si scontrano con l’identità dei luoghi e delle comunità transfrontaliere come accade nella regione bi-nazionale tra San Diego e Tijuana al confine californiano. Investigando gli aspetti legati all’identità transfrontaliera di questi territori, l’articolo adotta una prospettiva multi-livello per identificare: I) come i confini vengano percepiti quali elementi limitanti dell’interazione tra comunità (Sohn, 2014); II) la governance del territorio transfrontaliero, svelando un ruolo diverso rispetto al panorama europeo giocato dagli investitori privati e gli operatori del terzo settore; III) la costituzione di nuovi luoghi di raccordo oltre il confine attraverso due esempi: “Chicano Park” e “Dos Puertas”

Extracellular Vesicle Protein Expression in Doped Bioactive Glasses: Further Insights Applying Anomaly Detection

Proteomic analysis of extracellular vesicles presents several challenges due to the unique nature of these small membrane-bound structures. Alternative analyses could reveal outcomes hidden from standard statistics to explore and develop potential new biological hypotheses that may have been overlooked during the initial evaluation of the data. An analysis sequence focusing on deviating protein expressions from donors' primary cells was performed, leveraging machine-learning techniques to analyze small datasets, and it has been applied to evaluate extracellular vesicles' protein content gathered from mesenchymal stem cells cultured on bioactive glass discs doped or not with metal ions. The goal was to provide additional opportunities for detecting details between experimental conditions that are not entirely revealed with classic statistical inference, offering further insights regarding the experimental design and assisting the researchers in interpreting the outcomes. The methodology extracted a set of EV-related proteins whose differences between conditions could be partially explainable with statistics, suggesting the presence of other factors involved in the bioactive glasses' interactions with tissues. Outlier identification of extracellular vesicles' protein expression levels related to biomaterial preparation was instrumental in improving the interpretation of the experimental outcomes

Exploiting PLGA-Based Biocompatible Nanoparticles for Next-Generation Tolerogenic Vaccines against Autoimmune Disease

Tolerogenic vaccines are aimed at inhibiting antigen-specific immune responses. Antigen-loaded nanoparticles (NPs) have been recently emerged as ideal tools for tolerogenic vaccination because their composition, size, and capability of loading immunomodulatory molecules can be readily exploited to induce peripheral tolerance. Among polymeric NPs, poly(lactic-co-glycolic acid) (PLGA) NPs have the advantage of currently holding approval for several applications in drug delivery, diagnostics, and other clinical uses by the Food and Drug Administration (FDA). PLGA-NPs are non-toxic and display excellent biocompatibility and biodegradability properties. Moreover, surface functionalization may improve their interaction with biological materials, thereby optimizing targeting and performance. PLGA-NPs are the most extensively studied in pre-clinical model in the field of tolerogenic vaccination. Thus, this review describes their potential applications in the treatment of autoimmune diseases