Scarcity of autoreactive human blood IgA+ memory B cells

Class‐switched memory B cells are key components of the “reactive” humoral immunity, which ensures a fast and massive secretion of high‐affinity antigen‐specific antibodies upon antigenic challenge. In humans, IgA class‐switched (IgA⁺) memory B cells and IgA antibodies are abundant in the blood. Although circulating IgA⁺ memory B cells and their corresponding secreted immunoglobulins likely possess major protective and/or regulatory immune roles, little is known about their specificity and function. Here, we show that IgA⁺ and IgG⁺ memory B‐cell antibodies cloned from the same healthy humans share common immunoglobulin gene features. IgA and IgG memory antibodies have comparable lack of reactivity to vaccines, common mucosa‐tropic viruses and commensal bacteria. However, the IgA⁺ memory B‐cell compartment contains fewer polyreactive clones and importantly, only rare self‐reactive clones compared to IgG⁺ memory B cells. Self‐reactivity of IgAs is acquired following B‐cell affinity maturation but not antibody class switching. Together, our data suggest the existence of different regulatory mechanisms for removing autoreactive clones from the IgG⁺ and IgA⁺ memory B‐cell repertoires, and/or different maturation pathways potentially reflecting the distinct nature and localization of the cognate antigens recognized by individual B‐cell populations.     

Tissue resident memory T cells in the respiratory tract

Owing to their capacity to rapidly spread across the population, airborne pathogens represent a significant risk to global health. Indeed, several of the past major global pandemics have been instigated by respiratory pathogens. A greater understanding of the immune cells tasked with protecting the airways from infection will allow for the development of strategies that curb the spread and impact of these airborne diseases. A specific subset of memory T-cell resident in both the upper and lower respiratory tract, termed tissue-resident memory (Trm), have been shown to play an instrumental role in local immune responses against a wide breadth of both viral and bacterial infections. In this review, we discuss factors that influence respiratory tract Trm development, longevity, and immune surveillance and explore vaccination regimes that harness these cells, such approaches represent exciting new strategies that may be utilized to tackle the next global pandemic.     

Oral administration of a recombinant cholera toxin B subunit promotes mucosal healing in the colon

Cholera toxin B subunit (CTB) is a component of a licensed oral cholera vaccine. However, CTB has pleiotropic immunomodulatory effects whose impacts on the gut are not fully understood. Here, we found that oral administration in mice of a plant-made recombinant CTB (CTBp) significantly increased several immune cell populations in the colon lamina propria. Global gene expression analysis revealed that CTBp had more pronounced impacts on the colon than the small intestine, with significant activation of TGFβ-mediated pathways in the colon epithelium. The clinical relevance of CTBp-induced impacts on colonic mucosa was examined. In a human colon epithelial model using Caco2 cells, CTBp, but not the non-GM1-binding mutant G33D-CTBp, induced TGFβ-mediated wound healing. In a dextran sodium sulfate (DSS) acute colitis mouse model, oral administration of CTBp protected against colon mucosal damage as manifested by mitigated body weight loss, decreased histopathological scores, and blunted escalation of inflammatory cytokine levels while inducing wound healing-related genes. Furthermore, biweekly oral administration of CTBp significantly reduced disease severity and tumorigenesis in the azoxymethane/DSS model of ulcerative colitis and colon cancer. Altogether, these results demonstrate CTBp's ability to enhance mucosal healing in the colon, highlighting its potential application in ulcerative colitis therapy besides cholera vaccination.     

Intranasal vaccination of humans with recombinant cholera toxin B subunit induces systemic and local antibody responses in the upper respiratory tract and the vagina.

Forty-five volunteers were vaccinated twice intranasally with 10, 100, or 1,000 microg of cholera toxin B subunit (CTB). Blood and nasal and vaginal secretions were collected before and 1 week after the second vaccination from all volunteers, and the specific and total immunoglobulin A (IgA) and IgG titers were determined by enzyme-linked immunosorbent assay. Samples were also taken 6 months (n = 16) and 1 year (n = 14) after the vaccination. The 10- and 100-microg doses were well tolerated by the volunteers, but the 1,000-microg dose induced increased secretions from the nose and repetitive sneezings for several hours. The CTB-specific serum IgA and IgG increased 21- and 7-fold, respectively, 1 week after vaccination with the medium dose and increased 61- and 37-fold, respectively, after the high dose. In nasal secretions the specific IgA and IgG increased 2- and 6-fold after the medium dose and 2- and 20-fold after the high dose, respectively. In vaginal secretions the specific IgA and IgG increased 3- and 5-fold after the medium dose and 56- and 74-fold after the high dose, respectively. The lowest dose did not induce any significant antibody titer increases in serum or in secretions. The specific IgA and IgG levels in secretions were still elevated after 6 months but were decreasing 1 year after the vaccination. These results show that intranasal vaccination of humans with CTB induces strong systemic and mucosal antibody responses and suggest that CTB may be used as a carrier for antigens that induce protective immunity against systemic as well as respiratory and genital infections.     

IgG and IgA subclass distribution of antitoxin antibody responses after oral cholera vaccination or cholera disease

The immunoglobulin subclass distribution of cholera antitoxin antibody responses in serum was studied in Swedish volunteers after different routes of immunization with a cholera B subunit-whole cell vaccine (B + WCV) and in Bangladeshi patients convalescing from cholera disease. Both oral and parenteral immunization induced antitoxin antibodies of all the different IgG subclasses (IgG1, IgG2, IgG3 and IgG4) whilst the IgA antibodies were restricted to the IgA1 subclass. A single oral dose of B + WCV induced proportionally higher levels of IgG4 antitoxin in previously cholera-immunized volunteers than in a matched group who had not been cholera-vaccinated before, suggesting that repeated immunization preferentially stimulate formation of IgG4 antibodies. The IgG and IgA subclass distribution of antitoxin antibodies in orally vaccinated Swedes closely resembled that in Bangladeshi cholera patients.     

Toll-like receptor 2 induces mucosal homing receptor expression and IgA production by human B cells

There is a need for developing vaccines that elicit mucosal immunity. Although oral or nasal vaccination methods would be ideal, current strategies have yielded mixed success. Toll-like receptor 2 (TLR2) ligands are effective adjuvants and are currently used in the Haemophilus influenzae type B vaccine. Induction of humoral immunity in the mucosa is critical for effective vaccination; thus, we sought to determine the effects of TLR2 ligands on human mucosal B cell differentiation. We demonstrate that TLR2 ligands induce CCR9 and CCR10 expression by circulating B cells and increased chemotaxis to cognate chemokines CCL25 and CCL28 suggesting that TLR2 induces B cell homing to the gastrointestinal tract. TLR2 stimulation of B cells also induced J chain and IgA production demonstrating the induction of mucosal-like antibody secreting cells. These observations suggest that vaccines containing TLR2-ligands as adjuvants could induce mucosal B cell immunity even when delivered in a non-mucosal manner.     

Mast cells crosstalk with B cells in the gut and sustain IgA response in the inflamed intestine

B lymphocytes are among the cell types whose effector functions are modulated by mast cells (MCs). The B/MC crosstalk emerged in several pathological settings, notably the colon of inflammatory bowel disease (IBD) patients is a privileged site in which MCs and IgA⁺ cells physically interact. Herein, by inducing conditional depletion of MCs in red MC and basophil (RMB) mice, we show that MCs control B cell distribution in the gut and IgA serum levels. Moreover, in dextran sulfate sodium (DSS)-treated RMB mice, the presence of MCs is fundamental for the enlargement of the IgA⁺ population in the bowel and the increase of systemic IgA production. Since both conventional B-2 and peritoneal-derived B cells populate the intestine and communicate with MCs in physiological conditions and during inflammation, we further explored this interplay through the use of co-cultures. We show that MCs finely regulate different aspects of splenic B cell biology while peritoneal B cells are unresponsive to the supporting effects provided by MCs. Interestingly, peritoneal B cells induce a pro-inflammatory skewing in MCs, characterized by increased ST2 and TNF-α expression. Altogether, this study uncovers the versatility of the B/MC liaison and highlights key aspects for the resolution of intestinal inflammation.     

Differential expression of chemokine receptors on human IgA+ and IgG+ B cells

Organ-specific lymphocyte homing is dependent on the expression of tissue-specific homing receptors and selected chemokine receptors. During the effector phase of an immune response, IgA and IgG antibody-secreting cells (ASC) are differently distributed in the body. Still, B cell expression of L-selectin and the mucosal homing receptor integrin alpha4beta7 is not related to the isotype produced, but only to the site of antigen encounter. In this study, we examined if differences in chemokine responsiveness between IgA+ and IgG+ B cells could explain their different tissue localization. Circulating CD19+ B cells were isolated and their expression of IgA, IgG, and selected chemokine receptors was determined by flow cytometry. Few Ig+ cells expressed CCR2, CCR3, or CCR9, and there was no difference in the expression of these receptors between IgA+ and IgG+ cells. In contrast, CCR4, CCR5, and CXCR3 was expressed on significantly more IgG+ than IgA+ cells. The function of chemokine receptors on memory B cells and ASC was then tested in the transwell system. IgG+ memory cells migrated to a higher extent than IgA+ cells towards the CXCR3 ligand CXCL11/I-TAC, while there was only a small migration towards the CCR4 ligand CCL17/TARC and the CCR9 ligand CCL25/TECK. ASC migrated poorly to all chemokines tested. In conclusion, this study shows that IgG+ and IgA+ memory B cells have a differential expression of the Th1 associated chemokine receptor CXCR3, as well as of CCR4 and CCR5. In contrast, none of the studied chemokine receptors was preferentially expressed by IgA+ cells.     

Human germinal centres engage memory and naive B cells after influenza vaccination

正Influenza viruses remain a major public health threat.Seasonal influenza vaccination in humans primarily stimulates pre-existing memory B cells, leading to a transient wave of circulating antibody-secreting plasmablasts1-3.This recall response contributes to original antigenic sin, the selective boosting of antibody specificities from prior exposures to influenza virus antigens4.It remains unclear whether such vaccination can also induce germinal centre(GC) reactions in the draining lymph     

Fast Na+ current in circular smooth muscle cells of the large intestine

Whole-cell voltage clamp was carried out on freshly dispersed single smooth muscle cells from adult rat and human colons to investigate the regulation of the Ca²⁺ channels. In this study, we unexpectedly discovered the existence of a fast Na⁺ channel current. With normal physiological salt solution (PSS) plus 4-amino-pyridine (3 mM) in the bath and high-Cs⁺ solution in the pipette to inhibit outward K⁺ currents, an inward current possessing fast and slow components was observed when the cell membrane was depolarized to a value more positive than –20 mV from a holding potential of –100 mV. When Ca²⁺ ions were removed from the PSS, or when nifedipine (10 M) and Ni²⁺ (30 M) were simultaneously applied, the slow component disappeared and the fast component remained. The fast current component became almost completely inactivated within 10 ms. This fast component was dependent on extracellular Na⁺ concentration and was inhibited by tetrodotoxin (TTX) dose dependently (IC₅₀ of 130 nM in rat and 14 nM in human). These results suggest that the slow component of inward current was a Ca²⁺ channel current, whereas the fast component was a TTX-sensitive fast Na⁺ channel current. The threshold voltage, the voltage for peak current, and the reversal potential for the fast Na⁺ current were, respectively, about –50, –20, and + 50 mV in rats, and –40, 0, and + 60 mV in humans. The incidence of cells possessing fast Na⁺ currents depended on the region of the colon. In rat proximal colon, the incidence was 64% (14 out of 22 cells tested); in distal colon, it was 10% (2 out of 21 cells tested). In humans, the incidence in the ascending colon was 73% (16 out of 22 cells tested), and in the descending colon was 22% (7 out of 32 cells tested). The densities of fast Na⁺ and Ca²⁺ currents were 3.2 and 4.5 pA/pF in rats and 1.0 and 1.4 pA/pF in humans, respectively. The ratio of both current densities (Na⁺ vs Ca²⁺) was 0.71, in both rats and humans. We conclude that the major ion channels associated with the generation of inward currents in the circular smooth muscle cells of rat and human colon are voltage-dependent Ca²⁺ channels and TTX-sensitive Na⁺ channels. The fast Na⁺ current may facilitate propagation of excitation.     

小鼠脂肪间充质干细胞的分离培养及肠道归巢

背景：脂肪来源间充质干细胞因其取材安全、创伤小、易纯化、增殖快等优点而备受关注。 目的：建立一种有效、快速分离培养较高纯度小鼠脂肪间充质干细胞的方法,荧光标记后探索其是否可在体内向肠道归巢。 方法：取小鼠腹股沟及附睾脂肪,用0.1%Ⅰ型胶原酶消化得到单细胞,并与剩余未消化脂肪组织块一起接种,贴壁培养获得脂肪间充质干细胞,通过细胞形态、细胞表型、生长动力学、成骨成脂分化潜能4个方面进行鉴定,并将脂肪间充质干细胞用PKH67标记后经尾静脉注射移植到溃疡性结肠炎模型小鼠体内,观察其向肠道的归巢情况。 结果与结论：脂肪间充质干细胞镜下呈梭形,快速增殖呈漩涡状排列,细胞高表达 CD29、CD44、CD90,不表达CD45。成骨诱导碱性磷酸酶染色显示有黑色颗粒生成,茜素红S染色呈红色结节,成脂诱导油红O染色显示有大量脂滴。细胞生长曲线显示第3-5天为对数增长期,细胞生长活力良好。结肠冰冻切片在荧光显微镜下可见绿色荧光光点,并随时间推移有增加趋势。以上结果表明采用胶原酶消化加组织块贴壁法体外分离培养的脂肪间充质干细胞增殖快、纯度高,可向成骨成脂细胞分化,在体内可向结肠归巢并增殖。中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程全文链接：     

Effect of aflatoxin B1 on IgA+ cell number and immunoglobulin mRNA expression in the intestine of broilers

Abstract

Aflatoxin B₁ (AFB₁) is the most toxic group of mycotoxins produced by two species of the Aspergillus, common contaminants of food and animal feed. The purpose of our study was to determine the effect of AFB₁ on the number of IgA⁺ cell and immunoglobulin mRNA expression in the intestine of broilers. One hundred and fifty six one-day-old healthy Cobb broilers were randomly divided into the control group (the dosage of 0?mg/kg AFB₁) and AFB₁ group (the dosage of 0.6?mg/kg AFB₁) with three replicates per group and 26 birds per replicate for 21 days, respectively. After necropsy at 7, 14 and 21 days of age, duodenum, jejunum and ileum samples were taken for analyzing IgA⁺ cell by immunohistochemistry and IgA, pIgR, IgM and IgG mRNA expression by qRT-PCR. IgA⁺ cells were mainly distributed in the lamina propria of small intestinal mucosa in both groups at 14 and 21 days of age. A significant decrease in the number of IgA⁺ cells in the duodenum, jejunum and ileum was revealed in the AFB₁ group compared with that of the control group. The expression levels of IgA, pIgR, IgM and IgG mRNA in the intestinal mucosa were lower in the AFB₁ group than those in the control group at 14 and 21 days of age. Our data demonstrated that the dosage of 0.6?mg/kg AFB₁ in broiler diet reduced the number of IgA⁺ cell and the expression of IgA, pIgR, IgM and IgG mRNA in the small intestine.     

Enterotoxigenic Escherichia coli diarrhea in an endemic area prepares the intestine for an anamnestic immunoglobulin A antitoxin response to oral cholera B subunit vaccination.

We examined whether infection with enterotoxigenic Escherichia coli (ETEC) producing the heat-labile enterotoxin (LT) can prime the gut immune system to respond more efficiently to the immunologically related cholera B subunit component of a recently developed oral B subunit-whole-cell cholera vaccine (B-WCV). Nine Bangladeshi adults who had been hospitalized for watery diarrhea caused by LT-producing ETEC were given a single oral immunization with B-WCV on day 28 after hospital admission. The vaccine preparation used was adjusted to contain a lower-than-usual dose of B subunit, which had been found in previous studies to elicit a significant gut mucosal immunoglobulin A antitoxin response mainly in individuals with previous toxin-specific priming of their gut immune system. For comparison, nine patients convalescing from severe cholera disease and eight healthy subjects with no recent history of either cholera or ETEC infection were given the same oral vaccination with B-WCV. Vaccination in the ETEC convalescents induced an immunoglobulin A antitoxin response in intestinal lavage fluid which was comparable with that in the vaccinated cholera convalescents and superior to that in the vaccinated, previously uninfected controls. By contrast, only the cholera patients responded with anamnestic-type anti-cholera lipopolysaccharide antibody titer rises in the intestine after vaccination. These data support the specificity of the anamnestic anti-cholera toxin response in the ETEC patients after vaccination with cholera B-WCV.     

Antibodies to food and environmental antigens in serum, respiratory tract and intestine of pigs

Serum antibodies to food, bedding and dust were detected in pigs by precipitation and ELISA. There was an age-related decrease in the number of animals with precipitating antibodies and in the specific activity of IgG and IgM antibodies. Antibodies to food, bedding and dust were detected in respiratory and intestinal secretions, but there were differences in the distribution and isotype of antibodies to these three antigens.     

Effects of pneumococcal vaccination on tonsillo-pharyngitis and upper respiratory tract flora

The effect of a 14-valent pneumococcal vaccine on upper respiratory tract infections and carriage of beta-haemolytic streptococci was studied in a double-blind prospective study of 405 children 0.5-5 years of age. In the children under 2 years of age at vaccination, the cases of acute tonsillitis were more frequent among the vaccinees than among the controls (p less than 0.01). In contrast, among the children over 2 years of age at vaccination, fewer episodes of acute tonsillitis were reported among the vaccinees than among the controls (p less than 0.01). In the older age group, the total rate of upper respiratory tract infections was also significantly reduced. Similarly, an increase in asymptomatic carriage of group A streptococci was registered in children vaccinated at 2-5 years of age (p less than 0.01). Inversely, the carriership of group C streptococci diminished among the vaccinated children (p less than 0.05). Some possible mechanisms underlying these unexpected findings, including non-specific mitogenic features and cross-protection due to antigenic similarities, were explored.