Antibodies in breast milk: Pro-bodies designed for healthy newborn development.

This manuscript sheds light on the impact of maternal breast milk antibodies on infant health. Milk antibodies prepare and protect the newborn against environmental exposure, guide and regulate the offspring's immune system, and promote transgenerational adaptation of the immune system to its environment. While the transfer of IgG across the placenta ceases at birth, milk antibodies are continuously replenished by the maternal immune system. They reflect the mother's real-time adaptation to the environment to which the infant is exposed. They cover the infant's upper respiratory and digestive mucosa and are perfectly positioned to control responses to environmental antigens and might also reach their circulation. Maternal antibodies in breast milk play a key role in the immune defense of the developing child, with a major impact on infectious disease susceptibility in both HIC and LMIC. They also influence the development of another major health burden in children-allergies. Finally, emerging evidence shows that milk antibodies also actively shape immune development. Much of this is likely to be mediated by their effect on the seeding, composition and function of the microbiota, but not only. Further understanding of the bridge that maternal antibodies provide between the child and its environment should enable the best interventions to promote healthy development.

Persistently active interferon-γ pathway and expansion of T-bet+ B cells in a subset of patients with childhood-onset systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune disease causing significant morbidity and mortality, despite important improvements in its management in the last decades. The objective of this work is to investigate the role of IFN-γ in the pathogenesis of childhood-onset systemic lupus erythematosus (cSLE), evaluating the crosstalk between IFN-α and IFN-γ and the expression of T-bet, a transcription factor induced by IFN-γ, in B cells of patients with cSLE. Expression levels of both IFN-α and IFN-γ-induced genes were upregulated in patients with cSLE. We found increased serum levels of CXCL9 and CXCL10 in patients with cSLE. Type I IFN score decreased with initiation of immunosuppressive treatment; conversely, type II IFN score and levels of CXCL9 were not significantly affected by immunosuppressive treatment. Type II IFN score and CXCL9 were significantly higher in patients with lupus nephritis. We observed the expansion of a population of naïve B cells expressing T-bet in a cluster of patients with cSLE. IFN-γ, but not IFN-α, induced the expression of T-bet in B cells. Our data suggest that IFN-γ is hyperactive in cSLE, especially in patients with lupus nephritis, and it is not modulated by therapy. Our data reinforce the potential of IFN-γ as a therapeutic target in SLE.

Age and memory B cells at baseline are associated with risk of relapse and memory B-cell reappearance following anti-CD20 treatment in pediatric frequently-relapsing/steroid-dependent nephrotic syndrome.

B-cell depleting anti-CD20 monoclonal antibodies, such as rituximab, have proven efficacy in children with frequently-relapsing/steroid-dependent nephrotic syndrome (FR/SDNS). However, drug-free remission is variable and specific baseline markers predictive of relapse after anti-CD20 treatment are still being defined. To clarify these, we performed a bicentric observational study in a large cohort of 102 children and young adults with FR/SDNS treated with anti-CD20 monoclonal antibodies (rituximab and ofatumumab). Sixty-two patients (60.8%) relapsed during a 24-month period (median [interquartile range] relapse-free survival, 14.4 months [7.9-24.0]). A lower risk of relapse was significantly associated with an older age (over 9.8 years, hazard ratio, 0.44; 95% confidence interval, 0.26-0.74) and a higher risk of relapse was significantly associated with higher circulating levels of memory B cells (1.14; 1.09-1.32) at time of anti-CD20 infusion, independent of time elapsed from onset, previous anti-CD20 treatment, type of administered anti-CD20 monoclonal antibodies, and previous or maintenance oral immunosuppression. Patients younger than 9.8 years at anti-CD20 infusion had a subsequent higher recovery of total, transitional, mature-naïve and memory B-cell subsets independent of previous anti-CD20 treatment and maintenance immunosuppression. Significantly, younger age and higher circulating levels of memory B cells at time of anti-CD20 infusion were also independently associated with the recovery of memory B cells by linear mixed-effects modelling. Thus, both younger age and higher circulating levels of memory B cells at time of infusion are independently associated with a higher risk of relapse and an earlier recovery of memory B cells following anti-CD20 treatment in children with FR/SDNS.

Severe Acute Respiratory Syndrome Coronavirus 2 Monoclonal Antibody Combination Therapy in Patients With Coronavirus Disease 2019 and Primary Antibody Deficiency.

BACKGROUND: Previous reports highlighted the efficacy of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific monoclonal antibodies (mAbs) against coronavirus disease 2019. METHODS: We conducted a prospective study on the clinical outcome and antiviral effects of mAbs added to standard of care therapy in SARS-CoV-2-infected patients with primary antibody defects. RESULTS: Median time of SARS-CoV-2 quantitative polymerase chain reaction (qPCR) positivity was shorter in 8 patients treated with mAbs (22 days) than in 10 patients treated with standard of care therapy only (37 days, P=.026). Median time of SARS-CoV-2 qPCR positivity from mAb administration was 10 days. CONCLUSIONS: The SARS-CoV-2 mAbs treatment was effective and well tolerated in patients with primary antibody defects.

Severe Acute Respiratory Syndrome Coronavirus 2 Infection Versus Vaccination in Pregnancy: Implications for Maternal and Infant Immunity.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has been associated with adverse maternal and neonatal outcomes, yet uptake of SARS-CoV-2 vaccines during pregnancy and lactation has been slow. As a result, millions of pregnant and lactating women and their infants remain susceptible to the virus. METHODS: We measured spike-specific immunoglobulin G (anti-S IgG) and immunoglobulin A (anti-S IgA) in serum and breastmilk (BM) samples from 3 prospective mother-infant cohorts recruited in 2 academic medical centers. The primary aim was to determine the impact of maternal SARS-CoV-2 immunization vs infection and their timing on systemic and mucosal immunity. RESULTS: The study included 28 mothers infected with SARS-CoV-2 in late pregnancy (INF), 11 uninfected mothers who received 2 doses of the BNT162b2 vaccine in the latter half of pregnancy (VAX-P), and 12 uninfected mothers who received 2 doses of BNT162b2 during lactation. VAX dyads had significantly higher serum anti-S IgG compared to INF dyads (P < .0001), whereas INF mothers had higher BM:serum anti-S IgA ratios compared to VAX mothers (P = .0001). Median IgG placental transfer ratios were significantly higher in VAX-P compared to INF mothers (P < .0001). There was a significant positive correlation between maternal and neonatal serum anti-S IgG after vaccination (r = 0.68, P = .013), but not infection. CONCLUSIONS: BNT161b2 vaccination in late pregnancy or lactation enhances systemic immunity through serum anti-S immunoglobulin, while SARS-CoV-2 infection induces mucosal over systemic immunity more efficiently through BM immunoglobulin production. Next-generation vaccines boosting mucosal immunity could provide additional protection to the mother-infant dyad. Future studies should focus on identifying the optimal timing of primary and/or booster maternal vaccination for maximal benefit.

The immune response as a double-edged sword: The lesson learnt during the COVID-19 pandemic.

The COVID-19 pandemic has represented an unprecedented challenge for the humanity, and scientists around the world provided a huge effort to elucidate critical aspects in the fight against the pathogen, useful in designing public health strategies, vaccines and therapeutic approaches. One of the first pieces of evidence characterizing the SARS-CoV-2 infection has been its breadth of clinical presentation, ranging from asymptomatic to severe/deadly disease, and the indication of the key role played by the immune response in influencing disease severity. This review is aimed at summarizing what the SARS-CoV-2 infection taught us about the immune response, highlighting its features of a double-edged sword mediating both protective and pathogenic processes. We will discuss the protective role of soluble and cellular innate immunity and the detrimental power of a hyper-inflammation-shaped immune response, resulting in tissue injury and immunothrombotic events. We will review the importance of B- and T-cell immunity in reducing the clinical severity and their ability to cross-recognize viral variants.

Comprehensive phenotyping of human peripheral blood B lymphocytes in pathological conditions.

Several diseases are associated with alterations of the B-cell compartment. Knowing how to correctly identify by flow cytometry the distribution of B-cell populations in the peripheral blood is important to help in the early diagnosis. In the accompanying article we describe how to identify the different B-cell subsets in the peripheral blood of healthy donors. Here we show a few examples of diseases that cause dysregulation of the B-cell compartment.

Comprehensive phenotyping of human peripheral blood B lymphocytes in healthy conditions.

The B cell compartment provides innate and adaptive immune defenses against pathogens. Different B cell subsets, reflecting the maturation stages of B cells, have noninterchangeable functions and roles in innate and adaptive immune responses. In this review, we provide an overview of the B cell subsets present in peripheral blood of healthy individuals. A specific gating strategy is also described to clearly and univocally identify B cell subsets based on the their phenotypic traits by flow cytometric analysis.

Circulating plasmablasts in children with steroid-sensitive nephrotic syndrome.

BACKGROUND: The therapeutic efficacy of B cell-depleting anti-CD20 treatment in both pediatric and adult steroid-sensitive nephrotic syndromes (SSNS) suggests that B cells play a pathogenic role in the disease. In adults with minimal change disease (MCD), only circulating plasmablasts are increased during the active phase of the disease, among B cell subsets. These cells have not been studied yet in children with SSNS. METHODS: We retrospectively quantified by flow cytometry analysis circulating plasmablasts in 107 pediatric patients with SSNS (51 at disease onset, 27 during relapse, and 29 in remission). Data were compared with an equal number of age- and sex-matched healthy donors (HD). RESULTS: Circulating plasmablast levels, expressed as percentage of total CD19+ B cells or as percentage of total lymphocytes, were normal in all SSNS subgroups, compared to HD. Patients in remission had significantly fewer circulating plasmablasts compared to patients at disease onset. No significant correlation was observed between plasmablast levels and proteinuria or serum proteins, at onset. Treatment with prednisone and mycophenolate mofetil significantly reduced circulating levels of plasmablasts, unlike treatment with prednisone and calcineurin inhibitors. CONCLUSIONS: The B cell phenotype of children with SSNS differs from that of adults with MCD. This may justify different therapeutic approaches.

COVID-19-Associated Multisystem Inflammatory Syndrome in a Neonate with Atypical Coronary Artery Involvement.

OBJECTIVE: The study aimed to report a novel coronavirus disease 2019 (COVID-19)-associated multisystem inflammatory syndrome in children (MIS-C) in a neonate found to have an atypical diffuse thickening in coronary artery walls whose diagnosis required a multi-imaging approach. STUDY DESIGN: A neonate presented at birth with multiple organ involvement and coronary artery anomalies. A diagnosis of MIS-C associated with COVID-19 was supported by maternal severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection during pregnancy, and by the presence of both immunoglobulin (Ig)-G against SARS-CoV-2 and spike-specific memory B-cells response in the neonatal blood. Other plausible causes of the multiple organ involvement were excluded. RESULT: At admission, a severe coronary artery dilatation was identified on echocardiography, supporting the diagnosis of the MIS-C Kawasaki-like disease; however, coronary artery internal diameters were found to be normal using cardiac computed tomography angiography. At discharge, comparing the two imaging techniques each other, the correct diagnosis resulted to be an abnormal thickening in coronary arterial walls. These findings suggest that the inflammatory process affecting the coronary arterial wall in MIS-C could result not only in typical coronary artery lesions such as dilatation of the lumen or aneurysms development but also in abnormal thickening of the coronary artery wall. CONCLUSION: Our case provides an alert for pediatric cardiologists about the complexity to assess coronary artery involvement in MIS-C and raises the question that whether an abnormal vascular remodeling, with normal inner diameters, is to be considered like coronary artery dilatation for risk stratification. KEY POINTS: · COVID-19 associated MIS-C can present in neonates with multiple organ involvement.. · Coronary artery assessment in neonatal MIS-C could be complex, and a multi-imaging approach could be required.. · Beside the typical coronary artery lesions, such as dilatation of the lumen or aneurysms, also abnormal thickening of the coronary artery wall can occur..

SARS-CoV-2 Vaccine Induced Atypical Immune Responses in Antibody Defects: Everybody Does their Best.

BACKGROUND: Data on immune responses to SARS-CoV-2 in patients with Primary Antibody Deficiencies (PAD) are limited to infected patients and to heterogeneous cohorts after immunization. METHODS: Forty-one patients with Common Variable Immune Deficiencies (CVID), six patients with X-linked Agammaglobulinemia (XLA), and 28 healthy age-matched controls (HD) were analyzed for anti-Spike and anti-receptor binding domain (RBD) antibody production, generation of Spike-specific memory B-cells, and Spike-specific T-cells before vaccination and one week after the second dose of BNT162b2 vaccine. RESULTS: The vaccine induced Spike-specific IgG and IgA antibody responses in all HD and in 20% of SARS-CoV-2 naive CVID patients. Anti-Spike IgG were detectable before vaccination in 4 out 7 CVID previously infected with SARS-CoV-2 and were boosted in six out of seven patients by the subsequent immunization raising higher levels than patients naïve to infection. While HD generated Spike-specific memory B-cells, and RBD-specific B-cells, CVID generated Spike-specific atypical B-cells, while RBD-specific B-cells were undetectable in all patients, indicating the incapability to generate this new specificity. Specific T-cell responses were evident in all HD and defective in 30% of CVID. All but one patient with XLA responded by specific T-cell only. CONCLUSION: In PAD patients, early atypical immune responses after BNT162b2 immunization occurred, possibly by extra-follicular or incomplete germinal center reactions. If these responses to vaccination might result in a partial protection from infection or reinfection is now unknown. Our data suggests that SARS-CoV-2 infection more effectively primes the immune response than the immunization alone, possibly suggesting the need for a third vaccine dose for patients not previously infected.

COVID-19 - pathogenesis and immunological findings across the clinical manifestation spectrum.

PURPOSE OF REVIEW: The wide spectrum of COVID-19 clinical manifestations demonstrates the determinant role played by the individual immune response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the course of the disease. Thanks to the large number of published data, we are beginning to understand the logic of the human response to a virus adapted to bat immunity. RECENT FINDINGS: Impairment of types I and III interferon responses may facilitate the occurrence of severe COVID-19 with reduced antiviral activity associated to potent inflammation. The human T and B-cell germline repertoire contain the specificities able to react against SARS-CoV-2 antigens. Although inflammation disrupts the structure of germinal centers, memory T and B cells can be found in the blood of patients after mild and severe COVID 19. SUMMARY: Further studies are indispensable to better understand the human immune response to SARS-CoV-2. The diversity of the individual reaction may contribute to explain the clinical manifestation spectrum. Immunological memory can be demonstrated in patients, convalescent from mild, moderate, or severe COVID-19, but we do not know whether asymptomatic individuals have memory of the virus. Tailored vaccination protocols may be needed for individuals with previous SAS-CoV-2 infection.

IgM on the surface of T cells: a novel biomarker of pediatric-onset systemic lupus erythematosus.

BACKGROUND: Children with systemic lupus erythematosus (SLE) frequently have kidney involvement. Lupus nephritis sometimes presents alone, without systemic SLE features, representing the so-called full-house nephropathy (FHN). Distinguishing patients with SLE or FHN has therapeutic and prognostic implications. METHODS: In this retrospective observational study, we determined the presence of IgM on the surface of T cells (T cell IgM) by flow cytometry and characterized its ability in distinguishing SLE and FHN patients in a large pediatric cohort (n = 84). Fifty-seven patients with SLE (≥ 4 SLICC criteria at disease onset or during the follow-up) and 27 patients with FHN (3 or less SLICC criteria) were enrolled. RESULTS: Elevated T cell IgM levels were found in 24/25 SLE patients in active phase of disease and in 29/45 SLE patients in remission. In contrast, among FHN patients, only 1/9 presented this characteristic in active phase of disease and 0/20 in remission. Compared with standardized SLICC laboratory parameters, i.e., autoantibody titers and hypocomplementemia, T cell IgM positivity showed an extremely high sensitivity and specificity for the diagnosis of SLE, with the highest area under the curve (0.97, p < 0.001) by receiver operating characteristic analysis, similar to ANA (0.96, p < 0.001) and anti-dsDNA (0.90, p < 0.001) autoantibodies. CONCLUSIONS: Altogether, our data indicate that T cell IgM intensity may be a useful tool to correctly classify patients with lupus nephritis as SLE or FHN since disease onset.

Atypical IgM on T cells predict relapse and steroid dependence in idiopathic nephrotic syndrome.

The clinical heterogeneity of idiopathic nephrotic syndrome in childhood may reflect different mechanisms of disease that are as yet unclear. Here, we evaluated the association between an atypical presence of IgM on the surface of T cells (T-cell IgM) and the response to steroid therapy in a total of 153 pediatric patients with idiopathic nephrotic syndrome in different phases of disease. At disease onset, T-cell IgM median levels were significantly elevated and predictive of risk of relapse in 47 patients. They were also significantly increased comparing 58 steroid-dependent to 8 infrequently relapsing and 14 frequently relapsing patients, especially during relapse, whereas they were within the normal range in 7 genetic steroid-resistant patients. T-cell IgM in vivo was not affected by the amount of total circulating IgM, nor by concomitant acute infections or oral immunosuppression. However, it was affected by rituximab treatment in 21 steroid-dependent patients. By in vitro experiments, elevated T-cell IgM was not influenced by total circulating IgM levels or by the presence of other circulating factors, and there was no distinctive antigen-specificity or atypical IgM polymerization. Rather, we found that increased T-cell IgM correlates with reduced IgM sialylation, which influences T-cell response to steroid inhibition and T-cell production of podocyte-damaging factors. Thus, the atypical presence of IgM on the surface of T cells may predispose a subset of steroid-sensitive pediatric patients with idiopathic nephrotic syndrome to a poor response to steroid therapy since disease onset.

B cell phenotype in pediatric idiopathic nephrotic syndrome.

BACKGROUND: A pathogenic role of B cells in non-genetic nephrotic syndrome has been suggested by the efficacy of rituximab, a B cell depleting antibody, in maintaining a prolonged remission. However, little information is available on B cell homeostasis in nephrotic syndrome patients. METHODS: We retrospectively analyzed by flow cytometry the distribution of different B cell subpopulations in 107 steroid-sensitive and in 6 genetic steroid-resistant nephrotic syndrome pediatric patients, compared with age- and sex-matched controls. RESULTS: Fifty-one steroid-sensitive patients at disease onset, before starting immunosuppression, presented significantly increased levels of total, transitional, memory, and switched memory B cells compared to controls. Oral immunosuppression strongly affected transitional and mature B cell levels in 27 patients in relapse and also in 29 patients in remission, whereas memory B cells were significantly higher compared to controls during relapse, despite the immunosuppressive treatment, and were normalized only in patients in remission. Children with genetic forms of steroid-resistant nephrotic syndrome presented no differences in B cell profile from controls. CONCLUSIONS: Our study indicates that memory B cells, more than other B cell subsets, are increased and appear to be pathogenically relevant in steroid-sensitive nephrotic syndrome pediatric patients.

B-cell activation with CD40L or CpG measures the function of B-cell subsets and identifies specific defects in immunodeficient patients.

Around 65% of primary immunodeficiencies are antibody deficiencies. Functional tests are useful tools to study B-cell functions in vitro. However, no accepted guidelines for performing and evaluating functional tests have been issued yet. Here, we report our experience on the study of B-cell functions in infancy and throughout childhood. We show that T-independent stimulation with CpG measures proliferation and differentiation potential of memory B cells. Switched memory B cells respond better than IgM memory B cells. On the other hand, CD40L, a T-dependent stimulus, does not induce plasma cell differentiation, but causes proliferation of naïve and memory B cells. During childhood, the production of plasmablasts in response to CpG increases with age mirroring the development of memory B cells. The response to CD40L does not change with age. In patients with selective IgA deficiency (SIgAD), we observed that switched memory B cells are reduced due to the absence of IgA memory B cells. In agreement, IgA plasma cells are not generated in response to CpG. Unexpectedly, B cells from SIgAD patients show a reduced proliferative response to CD40L. Our results demonstrate that functional tests are an important tool to assess the functions of the humoral immune system.

Human B-cell memory is shaped by age- and tissue-specific T-independent and GC-dependent events.

Switched and IgM memory B cells execute different and noninterchangeable functions. We studied memory B cells in children of different ages, in peripheral blood and spleen and compared them with those of children born asplenic or unable to build germinal centers. We show that, whereas switched memory B cells are mostly generated in the germinal centers at all ages, IgM memory B cells can be distinct in three types with different developmental history. Innate IgM memory B cells, the largest pool in infants, are generated in the spleen by a germinal center-independent mechanism. With age, if the spleen is present and germinal centers are functional, innate IgM memory B cells are remodelled and accumulate somatic mutations. The third type of IgM memory B cell is a by-product of the germinal center reaction. Our data suggest that the B-cell memory developmental program is implemented during the first 5-6 years of life.