IgA protects resident commensal microbiota against competitors

This week journal Science published new study from Sarkis Mazmanian lab at Caltech describing role of IgA in providing strain-specific competitive advantage to certain resident commensal microbiota. 

His lab has been studying immunobiology of Bacteroides fragilis (B. fragilis), a gut commensal. In initial series of experiments they have compared germ-free mice mono-colonized with either wild-type B. fragilis or its mutant variants such as, Δccf, shown to modify biosynthesis of its capsular polysaccharides. They noticed that in co-housing experiments wild-type B. fragilis from one mouse could out compete mutant variant in another mouse in a horizontal transfer assay.   

Since B. fragilis polysaccharides are known to interact with host's immune system, the authors wanted to find out whether host's immune system influenced co-housing experiments. Not surprisingly, the authors found that mutant B. fragilis did not efficiently bind IgA (induced by wild-type B. fragilis) and that it in turn induced IgA repertoire that bound wild-type B. fragilis even less effectively, suggesting some kind of association between IgA and missing antigens on mutant B. fragilis.  

To verify these observations, the authors compared co-housing experiments between germ-free IgA+ and IgA-KO mice (or treated with B cell depleting antibody) mono-colonized with wild-type B. fragilis. Indeed and surprisingly this time, wild-type B. fragilis resident in IgA KO mice were easily overtaken by wild-type B. fragilis from mono-colonized wild-type mice. These results suggested that in absence of IgA wild-type B. fragilis has lost competitive advantage against wild-type B. fragilis resident in IgA+ mice.

What could these results mean in biological context: it appears that certain resident commensal microbiota benefit from interacting with IgA. The authors proposed that "during health, IgA fosters mucosal colonization of microbiota with beneficial properties....while disease states may induce (or be caused by) IgA responses to pathogens or pathobionts that disrupt healthy microbiome equilibria." This is an interpretation that does not provide clear mechanistic explanation as to how IgA response could make such discrimination at the level of antigens between which microbes to keep and which ones to eject from the host. 

posted by David Usharauli

Specific microbiota species induce serum IgA that protects against sepsis

Some 10 years ago the scientists made observation that microbiota difference between different mouse colonies is responsible for selective T_H17 expansion in the gut. Since then, field of immunology was flooded with numerous observations linking gut [and other tissue] microbiota to functional status of immune system.

One such study was recently published in journal Cell Host and Microbe. The researchers observed that serum IgA secreted by bone marrow residing plasma cells (BM PCs) were selectively enriched in mouse colony harboring members of Proteobacteria phylum. More importantly, these serum IgA protected mice during sepsis following gut damage.

Initially, the authors observed that their institute's B6 mouse colony (PENN-SPF) differed from commercial vendor B6 mice in their serum [but not small intestine] IgA status. Co-housing experiment indicated potential involvement of microbiota.

Indeed, 16S ribosomal gene sequencing showed enrichment of Proteobacteria phylum in local mouse colony (and also Deferribacteres).

Serum IgA bound microbiota and sequencing of serum IgA+/IgA- microbiota species confirmed selective enrichment of species within Proteobacteria phylum.

Development of microbiota-specific serum IgA were T cell-dependent.

Finally, serum IgA protected mice against sepsis following gut tissue damage and microbiota invasion (translocation).

In summary, serum IgA, but not intestinal IgA, is produced by bone marrow plasma cells in response to selective microbiota species, mostly from Proteobacteria phylum in mice. These serum IgA could bind microbiota, it developed in a T cell-dependent manner and protected host during gut flora invasion (translocation) in condition such as sepsis. However, it is not clear why serum IgG [in IgA KO mice] could not protect against sepsis in this study since one previous study already showed that serum IgG protected against gram negative bacteria such as E. coli. It is possible that serum IgA and IgG play non-redundant functions by protected against different microbial species.

posted by David Usharauli

Microbiota drives airway IgA class switch via DC-derived TGF-β

IgA is a class of antibody specifically designed for protection and tolerance at mucosal surfaces. Since vast majority of antigens [both infectious or noninfectious nature] interact with mucosal tissues, knowledge of signals controlling IgA production would help to develop more robust vaccination protocols.

So I decided to review this paper from Journal of Experimental Medicine where the authors have analysed IgA promoting capacity of lung DCs.

Most experiments were done ex vivo on sorted DC and B cells. Most cultures included α-IgM and α-CD40 antibody to "mimic" T-dependent Ab production. First, the authors noticed that lung CD103⁺ and CD24⁺ DCs (but not CD64⁺ Mφ) could provide necessary signals to naive B cells to switch to IgA (similar to MLN DCs).

Next, the authors found that this IgA switch capacity of lung CD103⁺ and CD24⁺ DCs  were dependent of their ability to produce TGF-β and Retinoic Acid (Vitamin A metabolites).

Next, the authors found that microbiota influenced lung DCs capacity to drive IgA switch in naive B cells [though it is not clear whether it is airway or gut microbiota that does it].

Follow up experiments revealed that MyD88/TRIF signaling [most likely from microbiota] in lung DC cells increased TGF-β production and their IgA switch potential.

Finally, the authors showed that i.n (intra-nasal) or i.t. (intra-tracheal) immunization [but not s.c. immunization] with small dose of cholera toxin (CT) provided system-wide protection against cholera toxin re-challenge, implying body-wide re-distribution of airway primed CT-specific B cells.

In summary, this study showed the role of microbiota/MyD88/TGF-β/CD103⁺ DCs axis in driving airway mucosal IgA class switch.

David Usharauli

microbiota influences non-genomic hereditary immune phenotype

The major advances in fundamental immunology for the past 25 years have to do with major advances in artificial manipulations of gene expressions (knock-out, knock-in, conditional knock-out, CRISPR-Cas9, etc).

Until recently very little attention was given to the extra-genomic influence of such gene manipulation. The scientists rarely bothered and still many don't to study gene effects in homozygous offspring derived from heterozygous parents to balance for exrta-genomic influence. 

For example, it is conceivable that homozygous gene effect are influenced wholly or in part by extra-genomic factors such as microbiota. This is exactly what a new study in journal Nature suggests.

In this study, the authors made initial observation that two colonies of genetically identical B6 mice kept in two different research facilities differed in their gut derived fecal IgA level (IgA^high and IgA^low). Interestingly two colonies did not differ in serum IgA level. The authors found that fecal IgA phenotype was vertically transmittable from parents to progeny.

Surprisingly, co-housing experiments revealed that IgA^low phenotype was dominant. This dominance of IgA^low phenotype was reversed when fecal materials were passed through 0.45 micron filter before transplantation into IgA^high hosts.

In addition, the authors found that pre-treatment of IgA^low mice with broad-spectrum antibiotics or ampicillin could reverse IgA^low phenotype as well, indicating that presence of ampicillin sensitive large microbes were responsible for this microbiota effect.

The authors found that IgA^low mice intestine were more sensitive to chemical injury compared to IgA^high mice.

However, intestinal sensitivity to DSS were independent of microbiota and dependent of intestinal IgA secretion.

To understand the mechanism behind IgA^low phenotype the authors tested components of fecal IgA complexes. These experiments revealed that IgA^low mice had profound deficiency in secretory component of IgA complexes in fecal samples. Secretory component is known to protect IgA from degradation.  
 

Finally, the authors showed that microbial pellets derived from IgA^low mice could degrade secretory component in a protease-dependent manner (use of germ-free host mice would have been beneficial to know exactly what microbe was responsible for this effect).

In summary, these results showed that non-genomic microbiota could influence hereditary immune phenotype. When studying gene-manipulated mice models, it is of critical importance to evaluate the model in proper manner (fecal transplantation, co-housing or heterozygous parents).

David Usharauli

   

When more is less: gut IgA prevents maturation of natural IgM immunity

This is a very interesting and thought-provoking study from journal Immunity. It focuses on immunobiology of gut associated follicular T helper (TFH) cells. This is a subset of helper T cells that is designed to provide necessary signals to germinal center (GC) B cells to produce high-affinity antibodies.

The authors study the role of P2rx7, a purinergic, ATP-activated receptor 7 on gut TFH function. Initially, the authors made an observation that P2rx7 is selectively and highly expressed on TFH. 

Interestingly, P2rx7-KO mice harbored increased number of TFH in peyer's patches (PP).

This increase in TFH cells in P2rx7-KO mice was cell-intrinsic since it was observed in CD3KO recipient mice adoptively transferred with 1:1 ratio of WT or P2rx7-KO PP T cells. 

Co-housing and co-(cross)-fostering experiments showed that effect of P2rx7 deficiency on TFH was independent of a any unique gut flora present in P2rx7-KO mice (though better experiments would have been to compare P2rx7-KO mice on Germ-Free (GF) background or test WT and P2rx7-KO mice from the same littermates).

Next, the authors found that greater presence of TFH in P2rx7-KO mice was linked to a reduced sensitivity of TFH to cell death in absence of P2rx7.

Additional experiments revealed that P2rx7-KO mice harbored more IgA producing cells in the gut (small intestine), but fewer IgM producing cells in the serum.

A scanning electron microscopic examination of small intestine revealed dramatic reduction of SFB colonization in P2rx7-KO mice.

Finally, series of in vivo experiments with cecal ligation and puncture (CLP) showed that P2rx7-KO mice were highly sensitive to death after sublethal CLP and they could be rescued with serum IgM injection from WT mice.

In summary, the data in this paper suggest that overzealous gut TFH function in P2rx7-KO mice contributes to the decline of the serum level of natural IgM and subsequent reduction of the host fitness in response to systemic inflammation.  

Some questions remains to be answered: 

(a) Why are gut TFH cells but not spleen or LN TFH cells sensitive to ATP? 

(b) Why gut IgA cannot compensate for serum IgM?

(c) If TCR signaling makes WT TFH refractory to ATP mediated cell death, does it mean that P2rx7-KO mice generate an excess of antigen non-specific TFH but not antigen-specific TFH compared to WT mice? In other word, does ATP controls antigen-(commensal)-specific TFH development?

David Usharauli