Proceedings of the 10th International Colloquium on Paratuberculosis

#68
Neutralisation <strong>of</strong> interleukin-10 from CD14+ monocytes enhances gamma interferon
production in peripheral blood mononuclear cells from Mycobacterium avium
subsp. paratuberculosis infected goats
Kari Røste Lybeck, Anne Kristine Storset, Ingrid Olsen
Department <strong>of</strong> Animal Health, National Veterinary Institute, Norway; Department <strong>of</strong> Food Safety and Infection
Biology, Norwegian School <strong>of</strong> Veterinary Science, Norway
Previous studies in Norway have shown that <strong>the</strong> sensitivity <strong>of</strong> <strong>the</strong> IFN-γ test in goats subclinically infected with
Mycobacterium avium subsp. paratuberculosis was lower than expected. Aiming to increase <strong>the</strong> sensitivity <strong>of</strong>
<strong>the</strong> IFN-γ test, we wanted to look at regulatory immune mechanisms that could affect <strong>the</strong> IFN-γ response in
goats with paratuberculosis. Goats from a herd with naturally acquired paratuberculosis were examined.
IL-10 was neutralised by adding a monoclonal antibody (mAb) against IL-10 to PPDj stimulated PBMCs
from 14 infected and 6 subclinically infected goats. An increase in IFN-γ production, measured by ELISA,
was seen in all goats when <strong>the</strong> anti-IL10 mAb was added compared to stimulation with PPDj only. This is in
accordance with previous results from o<strong>the</strong>r groups, and it was suggested that IL-10 is produced by regulatory
T-cells.
Depletion <strong>of</strong> putative regulatory CD25high T cells, using magnetic beads and an anti-CD25 mAb, had no
clear effect on <strong>the</strong> number <strong>of</strong> IFN-γ producing cells. The cells producing IL-10 was subsequently identified by
intracellular staining and flow cytometry to be CD14 + MHC class II + monocytes both in PPDj stimulated and
control cultures. However, possible regulatory T cells in goats were seen as CD4 + CD25 + T cells produced
IL-10 in response to Concanavalin-A stimulation. Additional investigations revealed that adding <strong>the</strong> anti-IL10
mAb also increased IFN-γ production in unstimulated PBMCs and in PPDj stimulated PBMCs from 8 out <strong>of</strong> 17
non-infected goats. Fur<strong>the</strong>rmore, we observed increased numbers <strong>of</strong> both CD4 + , CD8 + and CD8 + γδTCR + cells
producing IFN-γ after neutralisation <strong>of</strong> IL-10.
In conclusion IL-10 from CD14 positive monocytes inhibits maximal IFN-γ production in goats with paratuberculosis.
However due to <strong>the</strong> unspecific reactions observed, this does not appear to be a useful way <strong>of</strong>
increasing <strong>the</strong> sensitivity <strong>of</strong> <strong>the</strong> IFN-γ test for diagnosing paratuberculosis in goats.
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