Clonal Expansions of CD8(+) T Cells with IL-10 Secreting Capacity Occur during Chronic Mycobacterium tuberculosis Infection

Cyktor, Joshua C.; Carruthers, Bridget; Beamer, Gillian L.; Turner, Joanne

We determined the IL-10 producing capacity of CD8 + Vb8 + T cells using magnetic bead separation, which purified the dominant Vb8 expressing CD8 + population. CD8 + Vb8 + and CD8 + Vb8 neg T cells were isolated from Mtb-infected CBA/J mice or C57BL/6 mice (controls) at day 150 post Mtb infection and cultured in IL-10 ELISpot plates with autologous IL-10 deficient BMDCs in the presence of anti-CD3/CD28. Significantly more CD8 + Vb8 + T cells from Mtb-infected CBA/J mice were capable of producing IL-10 than CD8 + Vb8 neg T cells from CBA/J mice or from CD8 + Vb8 + and CD8 + Vb8 neg T cells from C57BL/6 mice (Fig. 4f) . These data indicate a dominant role for CD8 + Vb8 + expressing T cells in the IL-10 production we previously observed in purified CD8 + T cell cultures. Culture supernatants were also assayed for IFN-c by ELISA (Fig. 4g) and our data indicate that CD8 + Vb8 + T cells can have dual secretion of IL-10 and IFN-c, or represent a mixed population, as has been described by others [44] . In vivo depletion of IL-10 producing CD8 + T cells during chronic Mtb infection alters pro-inflammatory responses but fails to modify the bacterial load CD8 + T cells or Vb8 + cells were depleted from wild-type CBA/ J mice from day 90-120 after Mtb infection, a time when IL-10 and Vb8 + CD8 + T cells were increasing in the lungs of Mtbinfected CBA/J mice [5] . Following CD8 + depletion, Mtb CFU (Fig. 5a) , total CD4 + T cell numbers (Fig. 5b) and IFN-cproducing CD4 + T cells (Fig. 5c) were all moderately altered but data did not reach statistical significance. Interestingly, CD8 + T cell depletion led to a significant decrease in the total amount of (Fig. 5d) , reflecting the IL-10secreting capacity of CD8 + T cells we observed in vitro. Depletion of CD4 + T cells led to significantly increased Mtb burden and mortality before day 120 (not shown) showing that depletion of a known protective T cell subset increased susceptibility. Specific depletion of Vb8 + cells also failed to significantly impact the Mtb burden (Fig. 5e) , although a modest reduction in CFU was similarly observed. This modest reduction, similar to our findings with CD8 + T cell depletion, suggests that although IL-10 producing CD8 + T cells may not negatively impact the growth of Mtb during the timeframe we investigated, they do not provide protection.