Remote control of tissue tolerance by regulatory T cells

This week journal PNAS published an interesting study on Foxp3+ regulatory T cells (Tregs). It showed that presence of Tregs within secondary lymphoid organs (in lymph nodes) was sufficient to prevent peripheral tissue immunopathology.

As a starting point for this study, the authors showed that mice with Tregs-specific deficiency of kruppel-like factor 2 (KLF2) develop non-fatal peripheral tissue immunopathology, even though in vitro such Tregs [Foxp3-cre; Klf2^fl/fl] displayed normal suppressive activity).

Interestingly, in vivo adoptive transfer model, KLF2-KO Tregs also failed to prevent colitis and weight loss when co-transferred with WT naive T cells [but not with KLF2-KO naive T cells].

These results suggested that Tregs re-circulation between tissue and lymphoid tissues could have been involved. Surprisingly, the authors found no difference for Tregs presence [%-wise, no data about #] between KLF2-KO mice or WT (this is probably why this article ended up in PNAS).

Finally, the authors showed that combined deficiency of KLF2 and CCR7 in Tregs accelerated tissue immunopathology [by preventing Tregs access to lymph nodes].

In summary, the authors speculated that Tregs access to lymphoid tissue played a crucial role in preventing tissue immunopathology.

David Usharauli

Brain generates an autonomous immune protection against infection

Vaccines save lives. However even for most of vaccines that work we don't know why they work, i.e. we have no clue about protection correlates. They just work. Only source of information in human vaccine studies that are readily available is antibody titre analysis in serum [in blood]. But more recent studies in mice question the whole premise of vaccination strategy and analysis of its [correlates of] effectiveness. 

I am talking about tissue resident memory T cells. Several studies now clearly showed that most of the protection against infection is driven by local tissue-resident memory T cells rather than memory T cells found in circulation. For example, new study in Journal of Experimental Medicine showed that protection against brain viral infection is due to local brain tissue resident memory CD8 T cells, CD8 T_RM cells.

To test the role of brain CD8 T_RM cells in protection against viral infection the authors used recombinant, attenuated strain of LCMV virus (rLCMV). Initially they showed that unlike i.v. injection, when rLCMV  strain was administered directly in the brain (intrathecally) it led to robust development brain CD8 T_RM cells.  

Moreover, when these mice were later challenged with WT virulent LCMV strain, mice with brain CD8 T_RM cells showed superior control of virus (development of protective function of brain CD8 T_RM cells were Ag-specific, IFN-γ and perforin-dependent, but CD4 and NK cell independent).

To further test the role of brain CD8 T_RM cells in viral protection, the authors depleted circulating CD8 T cells with antibody. Still, these mice were fully protected against viral challenge with just brain CD8 T_RM cells (Ab depletion does not affect tissue resident T cells). These data suggested that brain CD8 T_RM cells were providing superior and independent protection against viral infection (though I don't think the authors have tested CD8 T cell depletion on mice injected with rLCMV intrathecally alone, because in most experiments on mice were done either with i.v. or i.v.+ i.th injections).

In summary, This study supports idea that correlates of some of viral [vaccine] protections could be found in tissues rather than in the blood.

David Usharauli

Infection-induced cell apoptosis activates self-epitope reactive T cells but barely

This week journal Nature Immunology published a study that had great title but poor data and conclusions. In fact, I rarely read such weak study for a long time, especially from top subject-matter journal. 

Basically, the authors tried to show that infection-induced cell apoptosis leads to self-peptide presentation and self-reactivity or even autoimmunity. Data are however misleading.

For this study the authors used the rodent pathogen Citrobacter rodentium that infects intestinal epithelial cells and induces their apoptosis. As a control, they have used infection with ΔEspF Citrobacter rodentium, a variant that lacks the secreted protein EPEC that mediates apoptosis. Initially, they showed that infection with WT Citrobacter rodentium, but not ΔEspF Citrobacter rodentium, induces Th17 response from large intestinal lamina propria (LI LP).

Next, the authors tried to examine whether infection-induced apoptotic cells will also provide self peptides for T cell activation (alongside of Citrobacter rodentium peptides). To do it, they have used so called double transgenic (DTg) mice derived from crossing OT-II mice with Act-mOVA mice. Now, these DTg mice delete absolute majority of OVA-specific OT-II cells in the thymus (from 1.5x10^6 to ~1,000 cells, i.e >1000X fold reduction of auto-reactive cells). The authors noted that DTg mice did not spontaneously develop autoimmunity and were healthy.

Next, when DTg mice were infected with Citrobacter rodentium, some portion of those OT-II cells left in DTg mice responded to it by up-regulating IL-17. The authors did not quantify the number of responding self-reactive OT-II cells and dot plot analysis reveals that their numbers seemed extremely low (on contour plot analysis). Moreover, it is not even clear whether self-reactive OT-II were responding to self-antigen or simply to inflammatory cytokine milieu [homeostatically] since even un-infected DTg mice showed proliferation and IL-17 expression in LI LP self-reactive OT-II cells.

The authors also showed that when infected with Citrobacter rodentium DTg mice showed little spike in anti-OVA IgA response driven by OT-II cells. However, it is not clear whether this anti-OVA IgA response has any pathogenic role.

Still, the authors believed that Th17 OT-II cells generated in DTg mice upon Citrobacter rodentium infection played pathogenic role in gut inflammation. As a "proof" they provided H&E staining of sections of large intestine from wild-type and DTg mice on day 40 after infection. Now, if scale bar on this H&E staining is 250 μm on both sections, then it is obvious DTg mice intestine is almost 2x more swollen or inflamed. But the authors noted that "DTg mice did not exhibit altered susceptibility to C. rodentium relative to that of wild-type or OT-II mice" and OT-II depletion did not significantly modify gut inflammation. So it is not clear from these data whether anti-OVA IgA or Th17 response after Citrobacter rodentium infection were in fact driving those observed pathogenic changes in the DTg mice guts (use of IL-17KO OT-II cells would have provided some guidance on this matter).

In summary, in my view this study only showed that WT Citrobacter rodentium infection induces little Th17 response from self-reactive T cells, however it failed to show that such Th17 response had any consequential effect.

David Usharauli