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Bitter taste receptors are involved not only in taste perception but in various physiological functions as their anatomical location is not restricted to the gustatory system. We previously demonstrated expression and activity of the subtype hTAS2R46 in human airway smooth muscle and broncho-epithelial cells, and here we show its expression and functionality in human skeletal muscle cells. Three different cellular models were used: micro-dissected human skeletal tissues, human myoblasts/myotubes and human skeletal muscle cells differentiated from urine stem cells of healthy donors. We used qPCR, immunohistochemistry and immunofluorescence analysis to evaluate gene and protein hTAS2R46 expression. In order to explore receptor activity, cells were incubated with the specific bitter ligands absinthin and 3ß-hydroxydihydrocostunolide, and calcium oscillation and relaxation were evaluated by calcium imaging and collagen assay, respectively, after a cholinergic stimulus. We show, for the first time, experimentally the presence and functionality of a type 2 bitter receptor in human skeletal muscle cells. Given the tendentially protective role of the bitter receptors starting from the oral cavity and following also in the other ectopic sites, and given its expression already at the myoblast level, we hypothesize that the bitter receptor can play an important role in the development, maintenance and in the protection of muscle tissue functions.

Very Early Onset Inflammatory Bowel Disease (VEO-IBD) is potentially associated with genetic disorders of the intestinal epithelial barrier or inborn errors of immunity (IEI). Dual oxidase 2 (DUOX2), an H2O2-producing NADPH oxidase expressed at apical enterocyte membranes, plays a crucial role in innate defense response. Biallelic DUOX2 mutations have been described only in two patients with VEO-IBD to date. We report the case of a 1-month-old female infant who presented persistent high C-reactive protein (CRP) levels from birth and anemia. Positive occult blood and very high calprotectin in the stool were detected and abdominal ultrasound showed thickened last ileal loop. Full endoscopy evaluation revealed important colon stenosis with multiple pseudo-polyploidy formations that resulted refractory to steroid therapy, requiring a partial colic resection. Histological examination of biopsy samples showed morphological features of IBD. Whole Exome Sequencing (WES) disclosed compound heterozygous variants in the DUOX2 gene: the pathogenic c.2524C>T; p.Arg842Ter and the variant of uncertain significance (VUS) c.3175C>T; p.Arg1059Cys. Molecular and functional studies showed the presence of mutant DUOX2 in the intestinal epithelium of the patient, albeit with at least 50% decreased catalytic activity. In conclusion, we describe the third patient to date with compound heterozygous variants of DUOX2, responsible for monogenic neonatal-IBD. This case expands the knowledge about Mendelian causes of VEO-IBD and DUOX2 deficiency. We suggest that DUOX2 should be part of the diagnostic evaluation of patients with suspected monogenic VEO-IBD.

Inborn errors of immunity associated with atopy (IEIs-A) are a group of inherited monogenic disorders that occur with immune dysregulation and frequent skin involvement. Several pathways are involved in the pathogenesis of these conditions, including immune system defects, alterations of skin barrier and metabolism perturbations. Current technological improvements and the higher accessibility to genetic testing, recently allowed the identification of novel molecular pathways involved in IEIs-A, also informing on potential tailored therapeutic strategies. Compared to other systemic therapy for skin diseases, biologics have the less toxic and the best tolerated profile in the setting of immune dysregulation. Here, we review IEIs-A with skin involvement focusing on the tailored therapeutic approach according to their pathogenetic mechanism.

Primary Immunodeficiencies (PID) are a group of rare congenital disorders of the immune system. Autoimmune cytopenia (AIC) represents the most common autoimmune manifestation in PID patients. Treatment of AIC in PID patients can be really challenging, since they are often chronic, relapsing and refractory to first line therapies, thus requiring a broad variety of alternative therapeutic options. Moreover, immunosuppression should be fine balanced considering the increased susceptibility to infections in these patients. Specific therapeutic guidelines for AIC in PID patients are lacking. Treatment choice should be guided by the underlying disease. The study of the pathogenic mechanisms involved in the genesis of AIC in PID and our growing ability to define the molecular underpinnings of immune dysregulation has paved the way for the development of novel targeted treatments. Ideally, targeted therapy is directed against an overexpressed or overactive gene product or substitutes a defective protein, restoring the impaired pathway. Actually, the molecular diagnosis or a specific drug is not always available. However, defining the category of PID or the immunological phenotype can help to choose a semi-targeted therapy directed towards the suspected pathogenic mechanism. In this review we overview all the therapeutic interventions available for AIC in PID patients, according to different immunologic targets. In particular, we focus on T and/or B cells targeting therapies. To support decision making in the future, prospective studies to define treatment response and predicting/stratifying biomarkers for patients with AIC and PID are needed.

The immunological events leading to type 1 diabetes (T1D) are complex and heterogeneous, underscoring the necessity to study rare cases to improve our understanding. Here, we report the case of a 16-year-old patient who showed glycosuria during a regular checkup. Upon further evaluation, stage 2 T1D, autoimmune thrombocytopenic purpura (AITP), and common variable immunodeficiency (CVID) were diagnosed. The patient underwent low carb diet, losing > 8 kg, and was placed on Ig replacement therapy. Anti-CD20 monoclonal antibody (Rituximab, RTX) was administered 2 years after diagnosis to treat peripheral polyneuropathy, whereas an atypical mycobacteriosis manifested 4 years after diagnosis and was managed with prolonged antibiotic treatment. In the fifth year of monitoring, the patient progressed to insulin dependency despite ZnT8A autoantibody resolution and IA-2A and GADA autoantibody decline. The patient had low T1D genetic risk score (GRS = 0.22817) and absence of human leukocyte antigen (HLA) DR3/DR4-DQ8. Genetic analysis identified the monoallelic mutation H159Y in TNFRSF13C, a gene encoding B-cell activating factor receptor (BAFFR). Significant reduced blood B-cell numbers and BAFFR levels were observed in line with a dysregulation in BAFF–BAFFR signaling. The elevated frequency of PD-1+ dysfunctional Tfh cells composed predominantly by Th1 phenotype was observed at disease onset and during follow-up. This case report describes a patient progressing to T1D on a BAFFR-mediated immunodysregulatory background, suggesting a role of BAFF–BAFFR signaling in islet-specific tolerance and T1D progression.

Actin-related protein 2/3 complex subunit 1B (ARPC1B) deficiency is a recently described inborn error of immunity (IEI) presenting with combined immunodeficiency and characterized by recurrent infections and thrombocytopenia. Manifestations of immune dysregulation, including colitis, vasculitis, and severe dermatitis, associated with eosinophilia, hyper-IgA, and hyper-IgE are also described in ARPC1B-deficient patients. To date, hematopoietic stem cell transplantation seems to be the only curative option for patients. ARPC1B is part of the actin-related protein 2/3 complex (Arp2/3) and cooperates with the Wiskott–Aldrich syndrome protein (WASp) in the regulation of the actin cytoskeleton remodeling and in driving double-strand break clustering for homology-directed repair. In this study, we aimed to investigate radiosensitivity (RS) in ARPC1B-deficient patients to assess whether it can be considered an additional disease trait. First, we performed trio-based next-generation-sequencing studies to obtain the ARPC1B molecular diagnosis in our index case characterized by increased RS, and then we confirmed, using three different methods, an increment of radiosensitivity in all enrolled ARPC1B-deficient patients. In particular, higher levels of chromatid-type aberrations and γH2AX foci, with an increased number of cells arrested in the G2/M-phase of the cell cycle, were found in patients’ cells after ionizing radiation exposition and radiomimetic bleomycin treatment. Overall, our data suggest increased radiosensitivity as an additional trait in ARPC1B deficiency and support the necessity to investigate this feature in ARPC1B patients as well as in other IEI with cytoskeleton defects to address specific clinical follow-up and optimize therapeutic interventions.

We described the case of a patient affected by activated PI3K-kinase delta syndrome (APDS) and a long-lasting and pauci-symptomatic SARS-CoV-2 infection, treated with multiple therapeutic agents including remdesivir and SARS-CoV-2-neutralizing monoclonal antibodies. We detected the clearance of the virus 105 days from the first positive swab and 7 days after monoclonal antibody administration. At genotyping, the SARS-CoV-2 virus resulted as wild type on all samples tested. This case shows the monoclonal antibodies’ good tolerability and efficacy in reducing viral shedding in long-lasting infections refractory to other treatments.

Mass SARS-Cov-2 vaccination campaign represents the only strategy to defeat the global pandemic we are facing. Immunocompromised patients represent a vulnerable population at high risk of developing severe COVID-19 and thus should be prioritized in the vaccination programs and in the study of the vaccine efficacy. Nevertheless, most data on efficacy and safety of the available vaccines derive from trials conducted on healthy individuals; hence, studies on immunogenicity of SARS-CoV2 vaccines in such populations are deeply needed. Here, we perform an observational longitudinal study analyzing the humoral and cellular response following the BNT162b2 mRNA COVID-19 vaccine in a cohort of patients affected by inborn errors of immunity (IEI) compared to healthy controls (HC). We show that both IEI and HC groups experienced a significant increase in anti-SARS-CoV-2 Abs 1 week after the second scheduled dose as well as an overall statistically significant expansion of the Ag-specific CD4+CD40L+ T cells in both HC and IEI. Five IEI patients did not develop any specific CD4+CD40L+ T cellular response, with one of these patients unable to also mount any humoral response. These data raise immunologic concerns about using Ab response as a sole metric of protective immunity following vaccination for SARS-CoV-2. Taken together, these findings suggest that evaluation of vaccine-induced immunity in this subpopulation should also include quantification of Ag-specific T cells.

Activated PI3K-kinase Delta Syndrome (APDS) is an autosomal-dominant primary immunodeficiency (PID) caused by the constitutive activation of the PI3Kδ kinase. The consequent hyperactivation of the PI3K-Akt-mTOR pathway leads to an impaired T- and B-cells differentiation and function, causing progressive lymphopenia, hypogammaglobulinemia and hyper IgM. Patients with APDS show recurrent sinopulmonary and chronic herpes virus infections, immune dysregulation manifestations, including cytopenia, arthritis, inflammatory enteropathy, and a predisposition to persistent non-neoplastic splenomegaly/lymphoproliferation and lymphoma. The recurrence of the lymphoproliferative disorder and the difficulties in the proper definition of malignancy on histological examination represents the main challenge in the clinical management of APDS patients, since a prompt and correct diagnosis is needed to avoid major complications. Targeted therapies with PI3Kδ-Akt-mTOR pathway pharmacologic inhibitors (i.e., Rapamycin, Theophylline, PI3K inhibitors) represent a good therapeutic strategy. They can also be used as bridge therapies when HSCT is required in order to control refractory symptoms. Indeed, treated patients showed a good tolerance, improved immunologic phenotype and reduced incidence/severity of immune dysregulation manifestations. Here, we describe our experience in the management of four patients, one male affected with APDS1 (P1) and the other three, a male and two females, with APDS2 (P2, P3, P4) presenting with chronic EBV replication, recurrent episodes of immune dysregulation manifestations and lymphomas. These cases highlighted the importance of a tailored and close follow-up, including serial endoscopic and lymph nodes biopsies control to detect a prompt and correct diagnosis and offer the best therapeutic strategy.

Introduction: Activated phosphoinositide 3-kinase-δ syndrome 2 (APDS2) is a rare primary immune regulatory disorder caused by heterozygous gain of function mutation in the PIK3R1 gene encoding PI3Kδ regulatory p85α subunit and resulting in PI3Kδ hyperactivation. Clinical features range from recurrent infections to manifestations of immune dysregulation like autoimmunity, inflammation, systemic lymphoproliferation, and increased risk of cancer. We describe a new dominant PIK3R1 mutation causing APDS2 presenting with lymphoma and systemic refractory autoimmunity.Case Presentation: A 30-year-old woman was referred to the Immunology Unit of our hospital for uncontrolled systemic lupus erythematosus, including chilblains lesions, systemic lymphoproliferation and IgA deficiency. At 19 years of age, she was diagnosed with Hodgkin's lymphoma. Subsequently, she presented systemic lupus erythematosus onset, with episodes of severe exacerbation, including autoimmune hemolytic anemia and pleuro-pericarditis. Initial clinical response to conventional treatments was reported. Immunological investigations performed during our first observation showed severe lymphopenia, IgA deficiency, elevated IgM with reduced IgG2 levels, and low vaccination antibody titers. Quantitative real-time polymerase chain reaction (PCR) assay for Cytomegalovirus and Epstein-Barr virus showed low viral loads for both viruses in serum. An increase of serum inflammatory markers highlighted persistent systemic hyperinflammation. The next-generation sequencing (NGS)-based gene panel tests for primary immunodeficiency showed a heterozygous A>G substitution in the splice acceptor site at c.1300-2 position of PIK3R1, leading to exon-skipping.Conclusion: This case emphasizes the importance of suspecting primary immune regulatory disorders in young adults, predominantly showing a severe, aggressive, and refractory to treatment immune dysregulation phenotype, even in the absence of major infectious diseases at the onset. Different treatments can be promptly started, and a delayed diagnosis can highly impact the outcome. Targeted therapy against PI3Kδ pathway defect effectively improves drug-resistant autoimmunity, lymphoproliferation, and risk of progression to malignancy; eligible patients could benefit from its use even as a bridge therapy to transplantation, currently the only definitive curative treatment. Therefore, identifying genetic mutation and prompt targeted treatment are essential to control disease manifestations, prevent long-term sequelae, and enable curative HSCT in APDS2 patients.

Evans syndrome (ES) is a rare but challenging condition, characterized by recurrent and refractory cytopenia episodes. Recent discoveries highlighted that an appropriate diagnostic workup is fundamental to identify an underlying immune dysregulation such as primary immunodeficiencies or a rheumatological disease. We hereby describe clinical features and laboratory results of 12 pediatric patients affected by ES referred to the Pediatric Onco-Hematology Unit of Bologna. Patients experienced a median of four acute episodes of cytopenia with 9 years as median age at the onset of symptoms. In 8/12 (67%) patients an underlying etiology, primary immunodeficiencies, or rheumatological disease was identified. In 4/12 children, other immune manifestations were associated (Thyroiditis, Celiac disease, Psoriasis, Vitiligo, Myositis, Membranoproliferative Glomerulonephritis). ES remained the primary diagnosis in four patients (33%). At a median follow-up time of 4 years, 5/12 (42%) patients revealed a chronic ITP, partially responsive to second line therapy. Immunoglobulin Replacement Therapy (IRT) was effective with a good hematological values control in three patients with a secondary ES (ALPS, CVID, and a patient with Rubinstein Taybi Syndrome and a progressive severe B cell deficiency with hypogammaglobulinemia). Our experience highlights that, in pediatric patients, ES is often only the first manifestation of an immunological or rheumatological disease, especially when cytopenias are persistent or resistant to therapy, with an early-onset or when are associated with lymphadenopathy.