We investigated the influence of a Wnt5A-gut microbiota axis on gut B-cell repertoire and protection from infection, having previously demonstrated that Wnt5A in association with gut commensals helps shape gut T-cell repertoire. Accordingly, Wnt5A heterozygous mice, which express less than wild-type level of Wnt5A, and their isolated Peyer\'s patches (PPs) were studied in comparison with the wild-type counterparts. The percentages of IgM- and IgA-expressing B cells were quite similar in the PP of both sets of mice. However, the PP of the Wnt5A heterozygous mice harbored significantly higher than wild-type levels of microbiota-bound B cell-secreted IgA, indicating the prevalence of a microbial population therein, which is significantly altered from that of wild-type. Additionally, the percentage of PP IgG1-expressing B cells was appreciably depressed in the Wnt5A heterozygous mice in comparison to wild-type. Wnt5A heterozygous mice, furthermore, exhibited notably higher than the wild-type levels of morbidity and mortality following infection with Salmonella typhimurium, a common gut pathogen. Differences in morbidity/mortality correlated with considerable disparity between the PP-B-cell repertoires of the Salmonella-infected Wnt5A heterozygous and wild-type mice, in which the percentage of IgG1-expressing B1b cells in the PP of heterozygous mice remains significantly low as compared to wild-type. Overall, these results suggest that a gut Wnt5A-microbiota axis is intrinsically associated with the maintenance of gut B-cell repertoire and protection from infection.IMPORTANCEAlthough it is well accepted that B cells and microbiota are required for protection from infection and preservation of gut health, a lot remains unknown about how the optimum B-cell repertoire and microbiota are maintained in the gut. The importance of this study lies in the fact that it unveils a potential role of a growth factor termed Wnt5A in the safeguarding of the gut B-cell population and microbiota, thereby protecting the gut from the deleterious effect of infections by common pathogens. Documentation of the involvement of a Wnt5A-microbiota axis in the shaping of a protective gut B-cell repertoire, furthermore, opens up new avenues of investigations for understanding gut disorders related to microbial dysbiosis and B-cell homeostasis that, till date, are considered incurable.

The immunoglobulin heavy constant gamma (IGHG) gene cluster encoding immunoglobulin G (IgG) subclasses is highly polymorphic, resulting in amino acid variation along the antibody constant heavy chain referred to as allotypes. IGHG1 and IGHG3 are the two most polymorphic IgG subclasses in humans, with 4 classical IgG1 allotypes and 13 allotypes described for IgG3, though recent studies suggest greater allelic diversity, especially in underrepresented ethnic populations. Polymerase chain reaction (PCR) and Sanger sequencing of IGHG amplicons allow for the identification of the single nucleotide polymorphisms (SNPs) responsible for the observed amino acid substitutions. Here, we provide a detailed protocol for the amplification of IGHG1 and IGHG3 segments by PCR, sample preparation for Sanger sequencing, and analysis of sequencing data to identify SNPs associated with different IgG1 and IgG3 allotypes.

BACKGROUND: p53, the most frequently mutated gene in cancer, lacks effective targeted drugs.
METHODS: We developed monoclonal antibodies (mAbs) that target a p53 hotspot mutation E285K without cross-reactivity with wild-type p53. They were delivered using lipid nanoparticles (LNPs) that encapsulate DNA plasmids. Western blot, BLI, flow cytometry, single-cell sequencing (scRNA-seq), and other methods were employed to assess the function of mAbs in vitro and in vivo.
RESULTS: These LNP-pE285K-mAbs in the IgG1 format exhibited a robust anti-tumor effect, facilitating the infiltration of immune cells, including CD8+ T, B, and NK cells. scRNA-seq revealed that IgG1 reduces immune inhibitory signaling, increases MHC signaling from B cells to CD8+ T cells, and enriches anti-tumor T cell and B cell receptor profiles. The E285K-mAbs were also produced in the dimeric IgA (dIgA) format, whose anti-tumor activity depended on the polymeric immunoglobulin receptor (PIGR), a membrane Ig receptor, whereas that of IgG1 relied on TRIM21, an intracellular IgG receptor.
CONCLUSIONS: Targeting specific mutant epitopes using DNA-encoded and LNP-delivered mAbs represents a potential precision medicine strategy against p53 mutants in TRIM21- or PIGR-positive cancers.

IgE is induced by the presence of IL-4 by class switching from IgM through IgG1 to IgE. IL-21 inhibits the IgE class switch by induction of Blimp1 leading to Stat6 and AID downregulation, and plasmablast/plasma cell differentiation.

The use of infant formulas (IFs) based on hydrolyzed cow\'s milk proteins to prevent cow\'s milk allergy (CMA) is highly debated. The risk of sensitization to milk proteins induced by IFs may be affected by the degree of hydrolysis (DH) as well as other physicochemical properties of the cow\'s milk-based protein hydrolysates within the IFs. The immunogenicity (specific IgG1 induction) and sensitizing capacity (specific IgE induction) of 30 whey- or casein-based hydrolysates with different physicochemical characteristics were compared using an intraperitoneal model of CMA in Brown Norway rats. In general, the whey-based hydrolysates demonstrated higher immunogenicity than casein-based hydrolysates, inducing higher levels of hydrolysate-specific and intact-specific IgG1. The immunogenicity of the hydrolysates was influenced by DH, peptide size distribution profile, peptide aggregation, nano-sized particle formation, and surface hydrophobicity. Yet, only the surface hydrophobicity was found to affect the sensitizing capacity of hydrolysates, as high hydrophobicity was associated with higher levels of specific IgE. The whey- and casein-based hydrolysates exhibited distinct immunological properties with highly diverse molecular composition and physicochemical properties which are not accounted for by measuring DH, which was a poor predictor of sensitizing capacity. Thus, future studies should consider and account for physicochemical characteristics when assessing the sensitizing capacity of cow\'s milk-based protein hydrolysates.

OBJECTIVE: Tubulointerstitial nephritis (TIN) has various etiologies, including IgG4-related disease (IgG4-RD), autoimmune diseases, antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV), and others. IgG4-positive plasma cell infiltration can occasionally be found in TIN unrelated to IgG4-RD. Therefore, there may be problems with usage of IgG4 immunostaining to differentiate between TIN with and TIN without IgG4-RD. This study aimed to compare the proportion of plasma cells that are positive for each IgG subclass and to clarify the predominant IgG subclass trends and clinical characteristics associated with IgG4-RD and non-IgG4-related interstitial nephritis.
METHODS: The study enrolled 44 cases of TIN: 6 of IgG4-RD, 8 of autoimmune disease, 9 of AAV, and 21 of unknown disease group. In addition to clinical characteristics, IgG subclass composition of interstitial plasma cells was evaluated among 4 groups by immunohistochemistry.
RESULTS: IgG1 was the predominant IgG subclass in TIN unrelated to IgG4-RD. In the IgG4-RD group, the IgG subclass rate was high in both IgG1 and IgG4. The rate of average IgG4-positive cells was significantly lower in the autoimmune disease group and unknown disease group compared with the IgG4-RD group.
CONCLUSIONS: The present study revealed IgG1-dominant immune profiles of TIN unrelated to IgG4-RD. Further investigation is required to elucidate the clinicopathological differences between IgG1-dominant and IgG4-dominant groups in IgG4-RD.

Visceral leishmaniasis (VL) represents the most severe form of Leishmaniasis infection, often resulting in fatality without timely treatment. Previous studies have found that immunosuppression increases the risk of VL disease progression and mortality, and the total immunoglobulin G (IgG) levels in peripheral blood vary before and after treatment. However, the distinct levels and roles of IgG subclasses in VL have not been documented yet. This study aims to elucidate the characteristics and clinical significance of IgG subclasses in VL.
A total of 43 cases newly-diagnosed with VL were enrolled in the cohort. We measured the levels of IgG subclasses before and after standard treatment and conducted assessments of bone marrow features. In addition, we analysed other haematological indices and examined the variations in IgG subclasses, as well as their correlation with clinical and laboratory factors.
The levels of total IgG, IgG1, and the ratios of both IgG1/IgG and IgG1/IgG2 decreased significantly after treatment, whereas the ratios of IgG2/ IgG showed an obvious increase. The VL patients without hyperglobulinemia displayed significant lower IgG1/IgG2 ratios, but higher IgG2/IgG ratios compared with those with hyperglobulinemia. In addition, VL patients with positive bone marrow amastigotes had significant higher IgG1/IgG and IgG1/IgG2 ratios, but lower IgG2/IgG ratios. IgG subclasses were correlated with abnormal blood test results, particularly immunological elements including IgM and Complement 4 (C4).
IgG1 and IgG2 exhibited contrasting changes after treatment in VL patients. The features of bone marrow and laboratory tests indicated that IgG1 and IgG2 serve different roles in the progression of VL. The ratios of IgG subclasses may be more precise indicators to evaluate immune reaction in VL than traditional total IgG.

Serum proteomics has matured and is now able to monitor hundreds of proteins quantitatively in large cohorts of patients. However, the fine characteristics of some of the most dominant proteins in serum, the immunoglobulins, are in these studies often ignored, due to their vast, and highly personalized, diversity in sequences. Here, we focus exclusively on these personalized features in the serum proteome and distinctively chose to study individual samples from a low diversity population: elderly donors infected by severe acute respiratory syndrome corona virus 2 (SARS-CoV-2). By using mass spectrometry-based methods, immunoglobulin IgG1 and IgA1 clonal repertoires were monitored quantitatively and longitudinally in more than 50 individual serum samples obtained from 17 Corona virus disease 2019 patients admitted to intensive care units. These clonal profiles were used to examine how each patient reacted to a severe SARS-CoV-2 infection. All 17 donors revealed unique polyclonal repertoires and substantial changes over time, with several new clones appearing following the infection, in a few cases leading to a few, very high, abundant clones dominating their repertoire. Several of these clones were de novo sequenced through combinations of top-down, middle-down, and bottom-up proteomics approaches. This revealed sequence features in line with sequences deposited in the SARS-CoV-specific antibody database. In other patients, the serological Ig profiles revealed the treatment with tocilizumab, that subsequently dominated their serological IgG1 repertoire. Tocilizumab clearance could be monitored, and a half-life of approximately 6 days was established. Overall, our longitudinal monitoring of IgG1 and IgA1 repertoires of individual donors reveals that antibody responses are highly personalized traits of each patient, affected by the disease and the chosen clinical treatment. The impact of these observations argues for a more personalized and longitudinal approach in patients\' diagnostics, both in serum proteomics as well as in monitoring immune responses.

Autoimmune hepatitis (AIH) is a common and debilitating pathology that has acute, subacute, and chronic presentation, requiring prompt diagnosis and early intervention. Several serologic markers are found to be associated with the pathogenesis and progression of autoimmune hepatitis, most notably antinuclear antibodies and anti-smooth muscle antibodies [Front Immunol. 2018;9:609]. In addition, AIH is also characterized by the elevation of gamma globulin levels, mainly immunoglobulin G (IgG) [World J Gastroenterol. 2015;21(1):60-83]. Although the literature has well established the presence of increased IgG levels in AIH, few studies have evaluated the subtypes of IgG and their differential levels associated with AIH. Here, we present a rare case of AIH that lacks the common serologic markers but instead reveals an elevation in IgG1 level. Our patient was subsequently placed on corticosteroids, and her symptoms quickly resolved. We intend to introduce this case to the medical community in the hope of aiding in the proper diagnosis and timely intervention of subsequent cases with similar presentations.

We recently reported that multiple sclerosis (MS) plasma contains IgG aggregates and induces complement-dependent neuronal cytotoxicity (Zhou et al., 2023). Using ELISA, we report herein that plasma IgG levels in the aggregates can be used as biomarkers for MS. We enriched the IgG aggregates from samples of two cohorts (190 MS and 160 controls) by collecting flow-through after plasma binding to Protein A followed by detection of IgG subclass. We show that there are significantly higher levels of IgG1, IgG3, and total IgG antibodies in MS IgG aggregates, with an AUC >90%; higher levels of IgG1 distinguish secondary progressive MS from relapsing-remitting MS (AUC = 91%). Significantly, we provided the biological rationale for MS plasma IgG biomarkers by demonstrating the strong correlation between IgG antibodies and IgG aggregate-induced neuronal cytotoxicity. These non-invasive, simple IgG-based blood ELISA assays can be adapted into clinical practice for diagnosing MS and SPMS and monitoring treatment responses.