Interleukin-21 receptor signaling promotes metabolic dysfunction-associated steatohepatitis-driven hepatocellular carcinoma by inducing immunosuppressive IgA+ B cells.

BACKGROUND: Dysregulation of immune surveillance is tightly linked to the development of metabolic dysfunction-associated steatohepatitis (MASH)-driven hepatocellular carcinoma (HCC); however, its underlying mechanisms remain unclear. Herein, we aimed to determine the role of interleukin-21 receptor (IL-21R) in MASH-driven HCC. METHODS: The clinical significance of IL-21R was assessed in human HCC specimens using immunohistochemistry staining. Furthermore, the expression of IL-21R in mice was assessed in the STAM model. Thereafter, two different MASH-driven HCC mouse models were applied between IL-21R-deficient mice and wild type controls to explore the role of IL-21R in MASH-driven HCC. To further elucidate the potential mechanisms by which IL-21R affected MASH-driven HCC, whole transcriptome sequencing, flow cytometry and adoptive lymphocyte transfer were performed. Finally, flow cytometry, enzyme-linked immunosorbent assay, immunofluorescent staining, chromatin immunoprecipitation assay and western blotting were conducted to explore the mechanism by which IL-21R induced IgA+ B cells. RESULTS: HCC patients with high IL-21R expression exhibited poor relapse-free survival, advanced TNM stage and severe steatosis. Additionally, IL-21R was demonstrated to be upregulated in mouse liver tumors. Particularly, ablation of IL-21R impeded MASH-driven hepatocarcinogenesis with dramatically reduction of lipid accumulation. Moreover, cytotoxic CD8+ T lymphocyte activation was enhanced in the absence of IL-21R due to the reduction of immunosuppressive IgA+ B cells. Mechanistically, the IL-21R-STAT1-c-Jun/c-Fos regulatory axis was activated in MASH-driven HCC and thus promoted the transcription of Igha, resulting in the induction of IgA+ B cells. CONCLUSIONS: IL-21R plays a cancer-promoting role by inducing IgA+ B cells in MASH-driven hepatocarcinogenesis. Targeting IL-21R signaling represents a potential therapeutic strategy for cancer therapy.

Bifidobacterium longum Subsp. infantis Promotes IgA Level of Growing Mice in a Strain-Specific and Intestinal Niche-Dependent Manner.

Throughout infancy, IgA is crucial for maintaining gut mucosal immunity. This study aims to determine whether supplementing newborn mice with eight different strains of Bifidobacterium longum subsp. infantis might regulate their IgA levels. The strains were gavaged to BALB/C female (n = 8) and male (n = 8) dams at 1-3 weeks old. Eight strains of B. longum subsp. infantis had strain-specific effects in the regulation of intestinal mucosal barriers. B6MNI, I4MI, and I10TI can increase the colonic IgA level in females and males. I8TI can increase the colonic IgA level in males. B6MNI was also able to significantly increase the colonic sIgA level in females. B6MNI, I4MI, I8TI, and I10TI regulated colonic and Peyer's patch IgA synthesis genes but had no significant effect on IgA synthesis pathway genes in the jejunum and ileum. Moreover, the variety of sIgA-coated bacteria in male mice was changed by I4MI, I5TI, I8TI, and B6MNI. These strains also can decrease the relative abundance of Escherichia coli. These results indicate that B. longum subsp. infantis can promote IgA levels but show strain specificity. Different dietary habits with different strains of Bifidobacterium may have varying effects on IgA levels when supplemented in early infancy.

Generation of circulating autoreactive pre-plasma cells fueled by naive B cells in celiac disease.

Autoantibodies against the enzyme transglutaminase 2 (TG2) are characteristic of celiac disease (CeD), and TG2-specific immunoglobulin (Ig) A plasma cells are abundant in gut biopsies of patients. Here, we describe the corresponding population of autoreactive B cells in blood. Circulating TG2-specific IgA cells are present in untreated patients on a gluten-containing diet but not in controls. They are clonally related to TG2-specific small intestinal plasma cells, and they express gut-homing molecules, indicating that they are plasma cell precursors. Unlike other IgA-switched cells, the TG2-specific cells are negative for CD27, placing them in the double-negative (IgD-CD27-) category. They have a plasmablast or activated memory B cell phenotype, and they harbor fewer variable region mutations than other IgA cells. Based on their similarity to naive B cells, we propose that autoreactive IgA cells in CeD are generated mainly through chronic recruitment of naive B cells via an extrafollicular response involving gluten-specific CD4+ T cells.

Astragalus polysaccharides ameliorates experimental colitis by regulating memory B cells metabolism.

It is well-established that the reduced Memory B cells (MBCs) play an important role in the pathogenesis of ulcerative colitis (UC), rendering them a potential therapeutic target for UC intervention. Astragalus polysaccharide (APS), a primary active constituent derived from the classic traditional Chinese medicine Astragalus membranaceus (AM), has been used for centuries in the treatment of UC in both human and animal subjects due to its renowned immunomodulatory properties. However, it is unknown whether APS can regulate MBCs to alleviate experimental colitis. In the present investigation, the murine colitis was successfully induced using dextran sulphate sodium (DSS) and subsequently treated with APS for a duration of 7 days. APS exhibited significant efficacy in reducing the disease activity index (DAI), colonic weight index, the index of colonic weight/colonic length. Furthermore, APS mitigated colonic pathological injuries, restored the colonic length, elevated the immunoglobulin A (IgA), transforming growth factor-ß1 (TGF-ß1) and interleukin (IL)-10 levels, while concurrently suppressing IgG, IgM, IL-6, tumor necrosis factor alpha (TNF-α) levels. Crucially, the quantities of MBCs, IgA+MBCs and forkhead box P3 (Foxp3+) MBCs were notably increased along with a concurrent decrease in IgG1+MBCs, IG2a+MBCs, IgG2b+MBCs after APS administration in colitis mice. Additionally, the Mitotracker red expressions of MBCs and their subgroups demonstrated a significantly up-regulation. Meanwhile, the transcriptomics analysis identified mitochondrial metabolism as the predominant and pivotal mechanism underlying APS-mediated mitigation of DSS-induced colitis. Key differentially expressed genes, including B-cell linker (BLNK), aldehyde dehydrogenase 1A1 (ALDH1A1), B-cell lymphoma 6 (BCL-6), B-lymphocyte-induced maturation protein 1 (Blimp-1), paired box gene 5 (PAX5), purinergic 2 × 7 receptor (P2X7R), B Cell activation factor (BAFF), B Cell activation factor receptor (BAFFR), CD40, nuclear factor kappa-B (NF-κB), IL-6 and so on were implicated in this process. These mRNA expressions were validated through quantitative polymerase chain reaction (qPCR) and immunohistochemistry. These findings revealed that APS effectively restored MBCs and their balance to ameliorate DSS-induced colitis, which was potentially realized via promoting mitochondrial metabolism to maintain MBCs activation.

Sparsentan ameliorates glomerular hypercellularity and inflammatory-gene networks induced by IgA1-IgG immune complexes in a mouse model of IgA nephropathy.

IgA nephropathy (IgAN) is characterized by glomerular deposition of immune complexes (ICs) consisting of IgA1 with O-glycans deficient in galactose (Gd-IgA1) and Gd-IgA1-specific IgG autoantibodies. These ICs induce kidney injury, and in the absence of disease-specific therapy, up to 40% of patients with IgAN progress to kidney failure. IgA1 with its clustered O-glycans is unique to humans, which hampered development of small-animal models of IgAN. Here, we used a model wherein engineered ICs (EICs) formed from human Gd-IgA1 and recombinant human IgG autoantibody are injected into nude mice to induce glomerular injury mimicking human IgAN. In this model, we assessed the protective effects of sparsentan, a single-molecule dual endothelin angiotensin receptor antagonist (DEARA) versus vehicle on EIC-induced glomerular proliferation and dysregulation of gene expression in the kidney. Oral administration of sparsentan (60 or 120 mg/kg daily) to mice intravenously injected with EIC attenuated the EIC-induced glomerular hypercellularity. Furthermore, analysis of changes in the whole kidney transcriptome revealed that key inflammatory and proliferative biological genes and pathways that are upregulated in this EIC model of IgAN were markedly reduced by sparsentan, including complement genes, integrin components, members of the mitogen-activated protein kinase family, and Fc receptor elements. Partial overlap between mouse and human differentially expressed genes in IgAN further supported the translational aspect of the immune and inflammatory components from our transcriptional findings. In conclusion, our data indicate that in the mouse model of IgAN, sparsentan targets immune and inflammatory processes leading to protection from mesangial hypercellularity.NEW & NOTEWORTHY The mechanisms by which deposited IgA1 immune complexes cause kidney injury during early phases of IgA nephropathy are poorly understood. We used an animal model we recently developed that involves IgA1-IgG immune complex injections and determined pathways related to the induced mesangioproliferative changes. Treatment with sparsentan, a dual inhibitor of endothelin type A and angiotensin II type 1 receptors, ameliorated the induced mesangioproliferative changes and the associated alterations in the expression of inflammatory genes and networks.

The biogenesis, identification, and functionality of circWWP2 in lipopolysaccharide stimulated macrophages.

CircRNA, a non-coding RNA, is an ideal biomarker and a suitable potential therapeutic target for various disease due to its high stability, species conservation and cell/tissue specificity. Our previous study has found a circular RNA WWP2 (circWWP2) was significantly decreased in chicken macrophages during bacterial infection. However, the function of circWWP2 in chicken macrophages remains unclear. In this study, it was demonstrated that circWWP2 was a stable circular RNA created by back-splicing of exons 2 to 4 of WWP2 via PCR amplification, Sanger sequencing, RNase R exonuclease digestion, and RT-qPCR. Moreover, bioinformatics analysis showed circWWP2 could interact with 13 miRNAs and target 3,264 genes, which were significantly enriched in lysosomes, IgA-producing intestinal immune networks for IgA production, and Notch signaling pathway. Furthermore, CCK8 and RT-qPCR indicated that overexpression of circWWP2 could promote lipopolysaccharide (LPS)-induced cellular injury by decreasing cell viability and increasing the expression levels of pro-inflammatory cytokines and pro-apoptosis genes, and NO production. CircWWP2 may exert a potential target for the treatment of bacterial infection. Further experiments are necessary to validate the specific mechanism that circWWP2 regulates LPS induced cellular immune responses.

Effects of crude protease produced by Bacillus subtilis (MAFIC Y7) on growth performance, immune indices, and anti-inflammatory responses of broilers fed soybean meal- or cottonseed meal-based diets.

A strain of Bacillus subtilis (MAFIC Y7) was isolated from the intestine of Tibetan pigs and was able to express high protease activity. The aim of this study was to characterize the proteases produced by MAFIC Y7, and to investigate the effects of protease addition on growth performance, ileal amino acid digestibility, and serum immunoglobulin and immune factors of broilers fed SBM-based diets, or on growth performance, carcass characteristics, and intestinal morphology of broilers fed CSM-based diets. B. subtilis (MAFIC Y7) expressed protease showed its optimal enzyme activity at 50 °C and pH 7.0. The coated crude enzyme (CCE) showed greater stability at pH 3.0 than its uncoated counterpart. Experiment 1 was conducted with six diets based on three levels of crude protein (CP)-CPlow, CPmedium, and CPhigh-with or without CCE. In CPlow, CCE increased gain:feed (G:F) (days 1 to 21, days 1 to 42) by 8%, 3%, respectively, and enhanced apparent ileal digestibility (AID) of crude protein and lysine (on day 42) by 8.8%, 4.6%, respectively, compared with diets containing no CCE (P < 0.05). CCE increased G:F from days 1 to 21 from 0.63 to 0.68, improved G:F and average daily gain (ADG) during days 1 to 42, and enhanced AID of crude protein, lysine, cysteine, and isoleucine on day 42 compared with the unsupplemented treatments (in CPmedium, P < 0.05). CCE increased serum IgA (on day 21), serum IgA and IgG and increased serum IL-10 (on day 42), but decreased serum tumor necrosis factor-α (TNF-α; on day 21), and serum IL-8 and TNF-α (on day 42) compared with unsupplemented treatments. At CPhigh, CCE decreased serum levels of IL-6 and TNF-α (on day 21), and IL-8 and TNF-α (on day 42) compared with unsupplemented treatments (in CPhigh, P < 0.05). In experiment 2, CSM-based diets with two lysine-to-protein ratios (5.2% or 5.5%) with or without CCE. In the high Lys diet (5.5% Lys:protein), CCE increased ADG and G:F, increased carcass, but decreased abdominal fat compared with the unsupplemented treatment (P < 0.05). In the 5.2% Lys:protein dietary treatment, CCE improved duodenal villus height compared with the unsupplemented treatment (P < 0.05). Supplementation of protease produced by MAFIC Y7 was associated with lower inflammatory responses in SBM diets (CPmedium or CPhigh) and improved ADG in broilers fed CPmedium or CPhigh. The proteases improved ADG and the efficiency of CSM use when the ratio of Lys to protein was 5.5%.

The aim of this study was to investigate the effects of Bacillus subtilis (MAFIC Y7)-expressed protease on reducing inflammatory responses of soybean meal (SBM) diets and improving the efficiency of cottonseed meal (CSM) in broilers. Experiment 1 was conducted with six dietary treatments based on three levels of crude protein (CP)­CPlow, CPmedium, and CPhigh­without or with proteases (0 or 4,000 U/kg). Supplementation of proteases significantly improved growth performance, gain:feed (G:F), and apparent ileal digestibility of crude protein and amino acids (cysteine, isoleucine, and histidine) in broilers fed CPmedium treatment (P < 0.05). Proteases inhibited inflammatory responses in SBM-based diets by decreasing serum tumor necrosis factor-α (TNF-α) (in CPmedium and CPhigh), and interleukin (IL)-6 (in CPhigh); and IL-8 and TNF-α (in CPmedium and CPhigh) on day 21. In experiment 2, broilers were fed with CSM-based diets with two ratios of lysine-to-protein (5.2% or 5.5%) with or without proteases. Proteases in the diet of 5.5% Lys to protein ratio increased growth performance and G:F compared to diets without proteases (P < 0.05). Proteases produced by MAFIC Y7 improved growth performance and G:F in CPmedium. Supplementation of protease was associated with lower inflammatory responses of broilers fed SBM-based diets (CPmedium or CPhigh) and improved the efficiency of CSM use when the ratio of lysine-to-protein was 5.5%.

Oral bacteria induce IgA autoantibodies against a mesangial protein in IgA nephropathy model mice.

IgA nephropathy (IgAN) is caused by deposition of IgA in the glomerular mesangium. The mechanism of selective deposition and production of IgA is unclear; however, we recently identified the involvement of IgA autoantibodies. Here, we show that CBX3 is another self-antigen for IgA in gddY mice, a spontaneous IgAN model, and in IgAN patients. A recombinant antibody derived from gddY mice bound to CBX3 expressed on the mesangial cell surface in vitro and to glomeruli in vivo. An elemental diet and antibiotic treatment decreased the levels of autoantibodies and IgAN symptoms in gddY mice. Serum IgA and the recombinant antibody from gddY mice also bound to oral bacteria of the mice and binding was competed with CBX3. One species of oral bacteria was markedly decreased in elemental diet-fed gddY mice and induced anti-CBX3 antibody in normal mice upon immunization. These data suggest that particular oral bacteria generate immune responses to produce IgA that cross-reacts with mesangial cells to initiate IgAN.

Concurrent Ingestion of Alkaline Water and L-Glutamine Enhanced Salivary α-Amylase Activity and Testosterone Concentration in Boxing Athletes.

Athletes often take sport supplements to reduce fatigue and immune disturbances during or after training. This study evaluated the acute effects of concurrent ingestion of alkaline water and L-glutamine on the salivary immunity and hormone responses of boxers after training. Twelve male boxing athletes were recruited in this study. During regular training, the participants were randomly divided into three groups and asked to consume 400 mL of alkaline water (Group A), 0.15 g/kg body weight of L-glutamine with 400 mL of water (Group G), and 0.15 g/kg of L-glutamine with 400 mL of alkaline water (Group A+G) at the same time each day for three consecutive weeks. Before and immediately after the training, saliva, heart rates, and the rate of perceived exertion were investigated. The activity of α-amylase and concentrations of lactoferrin, immunoglobulin A (IgA), testosterone, and cortisol in saliva were measured. The results showed that the ratio of α-amylase activity/total protein (TP) significantly increased after training in Group A+G but not in Group A or G, whereas the ratios of lactoferrin/TP and IgA/TP were unaffected in all three groups. The concentrations of salivary testosterone after training increased significantly in Group A+G but not in Group A or G, whereas the salivary cortisol concentrations were unaltered in all groups. In conclusion, concurrent ingestion of 400 mL of alkaline water and 0.15 g/kg of L-glutamine before training enhanced the salivary α-amylase activity and testosterone concentration of boxers, which would be beneficial for post-exercise recovery.

Pharmacological effects of biologically synthesized ginsenoside CK-rich preparation (AceCK40) on the colitis symptoms in DSS-induced Caco-2 cells and C57BL mice.

BACKGROUND: Despite the notable pharmacological potential of natural ginsenosides, their industrial application is hindered by low oral bioavailability. Recent research centers on the production of less-glycosylated minor ginsenosides. PURPOSE: This study aimed to explore the effect of a biologically synthesized ginsenoside CK-rich minor ginsenoside complex (AceCK40), on ameliorating colitis using DSS-induced colitis models in vitro and in vivo. METHODS: The ginsenoside composition of AceCK40 was determined by HPLC-ELSD and UHPLC-MS/MS analyses. In vitro colitis model was established using dextran sodium sulfate (DSS)-induced Caco-2 intestinal epithelial model. For in vivo experiments, DSS-induced severe colitis mouse model was established. RESULTS: In DSS-stimulated Caco-2 cells, AceCK40 downregulated mitogen-activated protein kinase (MAPK) activation (p < 0.05), inhibited monocyte chemoattractant protein-1 (MCP-1) production (p < 0.05), and enhanced MUC2 expression (p < 0.05), mediated via signaling pathway regulation. Daily AceCK40 administration at doses of 10 and 30 mg/kg/day was well tolerated by DSS-induced severe colitis mice. These doses led to significant alleviation of disease activity index score (> 36.0% decrease, p < 0.05), increased luminal immunoglobulin (Ig)G (> 37.6% increase, p < 0.001) and IgA (> 33.8% increase, p < 0.001), lowered interleukin (IL)-6 (> 65.7% decrease, p < 0.01) and MCP-1 (> 116.2% decrease, p < 0.05), as well as elevated serum IgA (> 51.4% increase, p < 0.001) and lowered serum IL-6 (112.3% decrease at 30 mg/kg, p < 0.001). Hematoxylin and eosin (H&E) and periodic acid-Schiff (PAS) staining revealed that DSS-mediated thickening of the muscular externa, extensive submucosal edema, crypt distortion, and decreased mucin droplets were significantly alleviated by AceCK40 administration. Additionally, daily administration of AceCK40 led to significant recovery of colonic tight junctions damaged by DSS through the elevation in the expression of adhesion molecules, including occludin, E-cadherin, and N-cadherin. CONCLUSION: This study presents the initial evidence elucidating the anti-colitis effects of AceCK40 and its underlying mechanism of action through sequential in vitro and in vivo systems employing DSS stimulation. Our findings provide valuable fundamental data for the utilization of AceCK40 in the development of novel anti-colitis candidates.

[The proportion of CD19+CD25+ Bregs in peripheral blood and the expression of PD-1 and PD-L1 on cell surface in patients with systemic lupus erythematosus and their correlation with clinical indicators].

Objective To investigate the expression and clinical significance of PD-1 and its ligand PD-L1 in peripheral blood CD19+CD25+ regulatory B cells (Bregs) in patients with systemic lupus erythematosus (SLE). Methods Peripheral blood samples were collected from 50 patients and 41 healthy controls (HCs). The proportion of CD19+CD25+Bregs in peripheral blood as well as the expression of PD-1+B and PD-L1+B cells on CD19+CD25+/-B cells, were detected by flow cytometry. At the same time, clinical information, such as clinical manifestations and laboratory indexes, was collected from patients. CD4+T cells and CD19+B cells were isolated by immunomagnetic beads and co-cultured in vitro to detect the differentiation of Bregs. Results The proportion of CD19+CD25+Bregs in the peripheral blood of SLE patients was lower than that in HC, while the expression of PD-1 and PD-L1 on Bregs was higher than that in HCs. SLE patients with pleural effusion, arthritis, and elevated CRP had a higher frequency of Bregs compared to the corresponding negative group. SLE patients with decreased immunoglobulin M (IgM) and positive anti-ribonuclear protein (RNP) antibodies had a lower frequency of Bregs compared to the corresponding negative group. SLE patients with infection, fever, arthritis, and elevated immunoglobulin A (IgA) had a higher frequency of CD19+CD25+PD-1+ cells compared to the corresponding negative group. SLE patients with infection, fever, and elevated IgA had a higher frequency of CD19+CD25+PD-L1+ cells compared to the corresponding negative group. And activated CD4+T cells were beneficial to the expression of CD25 on CD19+B cells. Conclusion The peripheral blood CD19+CD25+ Bregs are decreased in SLE patients, while the expression of PD-1 and PD-L1 on cell surface is increased, which is correlated with clinical manifestations and laboratory parameters. Activation of CD4+T cells promotes the differentiation of Bregs.

Dietary supplementation with pseudostellaria heterophylla polysaccharide enhanced immunity and changed mRNA expression of spleen in chicks.

In recent years, increasing interest has focused on natural components extracted from plants, among which plant polysaccharides as natural immunomodulators that can promote animal immunity. The present study was performed to investigate the effect of feed supplement Pseudostellaria Heterophylla Polysaccharide (PHP) on serum Immunoglobulins, T lymphocyte subpopulations, Cytokines and Lysozyme (LZM) activity in chicks. In addition, the influence of PHP on splenic gene expression was investigated by transcriptome sequencing. Four hundred 7-day-old Gushi cocks were randomly divided into four groups in a completely randomized design. The chicks were fed with a basal diet supplemented with 0 (CON-A), 100 (PHP-L), 200 (PHP-M) and 400 (PHP-H) mg/kg PHP. Blood and spleen samples were collected from 6 randomly selected chicks in each group at 14, 21, 28, and 35 days of age. The results showed that compared to the CON-A group, the PHP-M group exhibited significant increases in the levels of IgA, IgG, IgM, CD3, and LZM in the serum at 14, 21, 28, and 35 days (P < 0.05), and at 28 d, there was a significant quadratic relationship between the levels of dietary PHP and the levels of IgG, IgM, IFN-γ, IL-2, CD3, and LZM. Furthermore, a total of 470 differentially expressed genes (DEGs) were identified in spleen from PHP-M and CON-A at 28 d. These DEGs were significantly enriched in the Phagosome, Intestinal immune network for IgA production and Cytokine-cytokine receptor interaction pathways. The present investigation highlights the ameliorating effect of dietary PHP on immunological variables and spleen of chicks, the study suggests that PHP supplementation can enhance immunity and positively impact spleen mRNA expression in chicks.

Porphyromonas gingivalis infection in the oral cavity is associated with elevated galactose-deficient IgA1 and increased nephritis severity in IgA nephropathy.

BACKGROUND: The relationship between the major periodontal bacteria, Porphyromonas gingivalis, and the pathogenesis of IgA nephropathy (IgAN)-particularly with respect to galactose-deficient IgA1 (Gd-IgA1)-has not been fully elucidated. METHODS: Saliva samples from 30 IgAN patients and 44 patients with chronic kidney disease (CKD) were subjected to analysis of P. gingivalis status via polymerase chain reaction using a set of P. gingivalis-specific primers. The associations between P. gingivalis presence and clinical parameters, including plasma Gd-IgA1, were analyzed in each group. RESULTS: Compared with the CKD group, the IgAN group demonstrated significantly higher plasma Gd-IgA1 levels (p < 0.05). Compared with the P. gingivalis-negative subgroup, the P. gingivalis-positive subgroup exhibited significantly higher plasma Gd-IgA1 levels in both IgAN and CKD patients (p < 0.05). Additionally, among IgAN patients, the P. gingivalis-positive subgroup displayed significantly higher plasma Gd-IgA1 and urine protein levels, compared with the P. gingivalis-negative subgroup (p < 0.05). With respect to renal biopsy findings, the frequencies of segmental glomerulosclerosis and tubular atrophy/interstitial fibrosis were significantly greater in the P. gingivalis-positive subgroup than in the P. gingivalis-negative subgroup, according to the Oxford classification of IgAN (p < 0.05). CONCLUSION: Our findings suggest an association between the presence of P. gingivalis in the oral cavity and the pathogenesis of IgAN, mediated by increased levels of Gd-IgA1.

IL-17 receptor A functions to help maintain barrier integrity and limit activation of immunopathogenic response to H. pylori infection.

Activation of Th17 cell responses, including the production of IL-17A and IL-21, contributes to host defense and inflammatory responses by coordinating adaptive and innate immune responses. IL-17A and IL-17F signal through a multimeric receptor, which includes the IL-17 receptor A (IL-17RA) subunit and the IL-17RC subunit. IL-17RA is expressed by many cell types, and data from previous studies suggest that loss of IL-17 receptor is required to limit immunopathology in the Helicobacter pylori model of infection. Here, an Il17ra-/- mouse was generated on the FVB/n background, and the role of IL-17 signaling in the maintenance of barrier responses to H. pylori was investigated. Generating the Il17ra-/- on the FVB/n background allowed for the examination of responses in the paragastric lymph node and will allow for future investigation into carcinogenesis. While uninfected Il17ra-/- mice do not develop spontaneous gastritis following H. pylori infection, Il17ra-/- mice develop severe gastric inflammation accompanied by lymphoid follicle production and exacerbated production of Th17 cytokines. Increased inflammation in the tissue, increased IgA levels in the lumen, and reduced production of Muc5ac in the corpus correlate with increased H. pylori-induced paragastric lymph node activation. These data suggest that the cross talk between immune cells and epithelial cells regulates mucin production, IgA production, and translocation, impacting the integrity of the gastric mucosa and therefore activating of the adaptive immune response.

LPS/TLR4 Pathway Regulates IgA1 Secretion to Induce IgA Nephropathy.

Context: Studies have reported that the incidence and severity of IgA nephropathy (IgAN) are closely related to the imbalance of the intestinal flora. Imbalance of the intestinal flora may cause abnormalities, such as intestinal mucosal immunity or mesenteric B1 lymphocyte subsets. These can lead to an increase in immunoglobulin A (IgA) production and IgA structural changing, which can eventually cause IgA1 deposition in the glomerular mesangial area and nephritis. Objective: The study intended to explore whether the LPS/TLR4 pathway regulates mesenteric B cells, secreting Gd-IgA1 to induce IgA nephropathy. Design: The research team designed an animal study. Setting: The study took place at Department of Nephrology, Minhang Hospital, Fudan University. Animals: The animals were 60 specific pathogen free (SPF) C57BL/6 (B6, H-2b) male mice from that were 6-8 weeks old and weighed 20-25 grams. Intervention: The research team established a mouse model of IgA nephropathy. The team created five groups of mice: (1) the NC group, a normal negative control group without induced nephropathy and with no treatments; (2) the IgA nephropathy (IgAN) group, a positive control group with induced nephropathy and with no treatments; (3) the IgAN+anti-TLR4 group, an intervention group, with induced nephropathy and with a TLR4-antibody (anti-TLR4) treatment; (4) the IgAN+GEC group, an intervention group, with induced nephropathy and with treatment with glutamine enteric-coated capsules (GEC); and (5) the IgAN+anti-TLR4+GEC group, an intervention group, with induced nephropathy and with treatment with anti-TLR4 and GEC. Outcome Measures: The research team collected the blood and urine of all the mice and used an enzyme-linked immunoassay (ELISA) to analyze the levels of blood creatinine, urine protein, and urea nitrogen (BUN). The team also used the ELISA to analyze signal molecules for serum inflammation: interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α), monocyte chemotactic protein 1 (MCP-1), cyclooxygenase-2 (COX2), and galactose-deficient IgA1(Gd-IgA1). The team analyzed the distribution and content of IgA+B220+B lymphocytes in the intestinal tissues of all the mice, using tissue immunofluorescence tracking technology, and used hematoxylin-eosin (HE) staining to analyze the pathological damage in the kidney tissue. For analysis of glomerular IgA deposition, the team used a tissue immunofluorescence technique, and for detection of protein expression-toll-like receptor 4 (TLR4), B-cell activating factor (BAFF), and a proliferation-inducing ligand (APRIL)-in mesenteric lymphoid tissues, the team used western blot analysis. Results: For the five groups of mice, the amount or degree of the physiological indicators and inflammatory factors that ELISA detected, the B lymphocytes and IgA sedimentation that immunofluorescence tracing measured, the kidney pathological that HE staining detected, and the expression of immune-related proteins that western blotting measured, all showed a common trend: IgAN group> IgAN+ glomerular endothelial cells (GEC) group> IgAN+anti-TLR4 group> IgAN+anti-TLR4+GEC group> NC group. Conclusions: The TLR4 antibody and GEC for the treatment of the intestinal tract can regulate and repair intestinal function, so that IgAN can also be relieved at the same time. The results supported the hypothesis that a relationship exists between IgAN and the LPS/TLR4 pathway that regulates mesenteric B cells to secrete low-glycosylated poly-IgA1, which provides a new potential therapeutic plan for IgA nephritis.

Hydroxychloroquine ameliorates immune functionality and intestinal flora disorders of IgA nephropathy by inhibition of C1GALT1/Cosmc pathway.

BACKGROUND: Hydroxychloroquine (HCQ) has emerged as a potential and secure antiproteinuric agent in IgA nephropathy (IgAN). This study endeavored to explore the impact of HCQ on the immune functionality and intestinal flora disorders in IgAN rats, as well as to elucidate the underlying mechanisms through in vivo and in vitro experiments. METHODS: IgAN model was established in Sprague-Dawley rats through the administration of BSA, LPS, and CCl4, and the IgAN rats received a continuous 8-week treatment with HCQ. Moreover, the human glomerular mesangial cells (HMCs) were incubated with IgA1 to establish an in vitro cellular model of IgAN. At the end of experimental period, samples were collected for further analysis. RESULTS: HCQ ameliorated the elevated levels of 24hUTP, SCr, BUN, the number of urinary RBC, and the activation of inflammation-related proteins within the TLR4/NF-κB signaling pathway. In the IgAN rat group, there was a pronounced escalation in IgA deposition, mesangial matrix hyperplasia, and glomerular inflammatory cell infiltration, while the administration of HCQ effectively mitigated these pathological changes. In addition, the reduced production of CD4+CD25+Foxp3+ Treg in the IgAN group was effectively reversed by HCQ. Furthermore, HCQ has the capacity to restore the compromised state of the intestinal mucosal barrier induced by IgAN and mitigate the circumstances of intestinal permeability and disruption in the intestinal flora. CONCLUSION: HCQ diminishes IgA aberrant glycosylation levels, ameliorates renal and intestinal histopathological damage, and attenuates intestinal flora disorders and immune dysfunction in IgAN rats by means of activating the C1GALT1/Cosmc pathway.

Targeting intracellular oncoproteins with dimeric IgA promotes expulsion from the cytoplasm and immune-mediated control of epithelial cancers.

Dimeric IgA (dIgA) can move through cells via the IgA/IgM polymeric immunoglobulin receptor (PIGR), which is expressed mainly on mucosal epithelia. Here, we studied the ability of dIgA to target commonly mutated cytoplasmic oncodrivers. Mutation-specific dIgA, but not IgG, neutralized KRASG12D within ovarian carcinoma cells and expelled this oncodriver from tumor cells. dIgA binding changed endosomal trafficking of KRASG12D from accumulation in recycling endosomes to aggregation in the early/late endosomes through which dIgA transcytoses. dIgA targeting of KRASG12D abrogated tumor cell proliferation in cell culture assays. In vivo, KRASG12D-specific dIgA1 limited the growth of KRASG12D-mutated ovarian and lung carcinomas in a manner dependent on CD8+ T cells. dIgA specific for IDH1R132H reduced colon cancer growth, demonstrating effective targeting of a cytoplasmic oncodriver not associated with surface receptors. dIgA targeting of KRASG12D restricted tumor growth more effectively than small-molecule KRASG12D inhibitors, supporting the potential of this approach for the treatment of human cancers.

Polymeric immunoglobulin receptor (pIgR) in cancer.

BACKGROUND: The polymeric immunoglobulin receptor (pIgR) is a transmembrane transporter of polymeric IgA through the intestinal epithelium. Its overexpression has been reported in several cancers, but its role as a diagnostic and prognostic biomarker of oncogenesis is currently unclear. METHOD: A literature search was conducted to summarize the functions of pIgR, its expression levels, and its clinical implications. RESULTS: pIgR expression has previously been investigated by proteomic analysis, RNA sequencing, and tissue microarray at the level of both RNA and protein in various cancers including pancreatic, esophageal, gastric, lung, and liver. However, studies have reported inconsistent results on how pIgR levels affect clinical outcomes such as survival rate and chemotherapy resistance. Possible explanations include pIgR mRNA levels being minimally correlated with the rate of downstream pIgR protein synthesis, and the diversity of antibodies used in immunohistochemistry studies further magnifying this ambiguity. In ovarian cancer cells, the transcytosis of IgA accompanied a series of transcriptional changes in intracellular inflammatory pathways that inhibit the progression of cancer, including the upregulation of IFN-gamma and downregulation of tumor-promoting ephrins. These findings suggest that both the levels of pIgR and secreted IgA from tumor-infiltrating B cells affect clinical outcomes. CONCLUSION: Overall, no direct correlation was observed between the levels of pIgR inside tumor tissue and the clinical features in cancer patients. Measuring pIgR protein levels with a more specific and possibly chemically defined antibody, along with tumoral IgA, is a potential solution to better understand the pathways and consequences of pIgR overexpression in cancer cells.

Dioscin Mediated IgA Nephropathy Alleviation by Inhibiting B Cell Activation In Vivo and Decreasing Galactose-Deficient IgA1 Production In Vitro.

The increase of circulating galactose-deficient IgA1 (Gd-IgA1) is caused by excessive activation of IgA-positive secretory cells in the process of mucosal immune responses, which is a critical link in the pathogenesis of IgA nephropathy (IgAN). Peyer's patch, the prominent place where B lymphocytes are transformed into IgA-secreting plasma cells, is the primary source of IgA. In addition, the lower expression of core 1ß-1,3-galactosyltransferase (C1GalT1) and its molecular chaperone, C1GalT1-specific molecular chaperone (Cosmc), is related to abnormal glycosylation of IgA1 in IgAN patients. Our clinical experience shows that Dioscoreae Nipponicae Rhizoma's (DNR) herbal medicine can relieve proteinuria and hematuria and improve renal function in IgAN patients. Dioscin (DIO) is one of the main active ingredients of DNR, which has various pharmacological activities. This study explores DIO's possible mechanism in treating IgAN.The IgAN model mouse was established by mucosal immune induction. The mice were divided into the control, model, and DIO gavage groups. The glomerular IgA deposition in mice, renal pathological changes, and B cell markers CD20 and CXCR5 expression in Peyer's patch were detected by immunofluorescence and immunohistochemistry. After lipopolysaccharide (LPS) stimulation, DIO's effects on DAKIKI cells proliferation, IgA and Gd-IgA1 secretion, C1GalT1, and Cosmc expression were studied by cell counting kit-8 (CCK-8) assay, enzyme-linked immunosorbent assay (ELISA) test, quantitative real-time polymerase chain reaction (QRT-PCR), and western blotting (WB). In in vivo studies, IgA deposition accompanied by glomerular mesangial hyperplasia and increased expression of CD20 and CXCR5 in Peyer's patch in the IgAN model mouse was alleviated by DIO. In vitro studies showed 0.25 µg/mL to 1.0 µg/mL DIO inhibited LPS-induced DAKIKI cell proliferation, IgA and Gd-IgA1 secretion, and up-regulated the mRNA and protein expression of C1GalT1 and Cosmc. This study demonstrates that DIO may reduce Gd-IgA1 production by inhibiting excessive activation of IgA-secreting cells and up-regulating C1GALT1/Cosmc expression.

RhoA vesicle trafficking-mediated transglutaminase 2 membrane translocation promotes IgA1 mesangial deposition in IgA nephropathy.

Transglutaminase 2 (TGase2) has been shown to contribute to the mesangial IgA1 deposition in a humanized mouse model of IgA nephropathy (IgAN), but the mechanism is not fully understood. In this study, we found that inhibition of TGase2 activity could dramatically decrease the amount of polymeric IgA1 (pIgA1) isolated from patients with IgAN that interacts with human mesangial cells (HMC). TGase2 was expressed both in the cytosol and on the membrane of HMC. Upon treatment with pIgA1, there were more TGase2 recruited to the membrane. Using a cell model of mesangial deposition of pIgA1, we identified 253 potential TGase2-associated proteins in the cytosolic fraction and observed a higher concentration of cellular vesicles and increased expression of Ras homolog family member A (RhoA) in HMC after pIgA1 stimulation. Both the amount of pIgA1 deposited on HMC and membrane TGase2 level were decreased by inhibition of the vesicle trafficking pathway. Mechanistically, TGase2 was found to be coprecipitated with RhoA in the cellular vesicles. Membrane TGase2 expression was greatly increased by overexpression of RhoA, while it was reduced by knockdown of RhoA. Our in vitro approach demonstrated that TGase2 was transported from the cytosol to the membrane through a RhoA-mediated vesicle-trafficking pathway that can facilitate pIgA1 interaction with mesangium in IgAN.