In this report, we examined the in vivo production, cellular sources and function of the regulatory cytokine IL-10 during acute pulmonary RSV infection. Importantly, the primary source of IL-10 is the anti-viral CD4+ and CD8+ effector T cells recruited to the RSV infected lungs. We demonstrate that this effector T cell-derived IL-10 plays a critical role in preventing excess inflammation during the innate and particularly the adaptive immune response to RSV. In the absence of IL-10 signaling, an enhanced inflammatory response and a concomitant alteration in pulmonary function ensues. Thus, IL-10 may play a critical role in maintaining lung function during infection. Of note, our findings suggest that effector T cells are not only the major source of IL-10 in the infected lungs but may also serve as important cellular targets for the action of this regulatory cytokine, reflecting a novel autocrine pathway for the action of IL-10 during infection.
Several lines of evidence in the current study implicate effector T cells as the primary source of IL-10 observed in the RSV infected respiratory tract in this model. Similar to our recently reported findings in the murine influenza infection model 
, we noted minimal IL-10 secretion in RSV infected Rag1-/-
mice and localized IL-10 production by effector T cells in the infected lungs using the IL-10 reporter Vert-X mice for infection. However, we cannot formally exclude a major contribution of the small percentage (2%–5%) of Thy-1– (negative)
cells detected to the amelioration of pathology. In this case, however, we demonstrated that IL-10 derived solely from T-cells are sufficient to control disease development during RSV infection () which also implicated T-cell-derived IL-10 in the control of excess pulmonary inflammation. The IL-10-producing effector T cells are, as we observed in this report, IL-4 and GATA-3 negative (data not shown) and thus show few features characteristic of type 2 lineage of T cells; but rather are T-bet positive type 1 effectors. Interestingly, we observed a minor fraction of T-bet+
IL-10 producers which co-express Foxp-3, a cell population that has been recently described in other systems 
. Of note, CD25+
T regulatory cells have been recently demonstrated to modulate RSV specific CD8+
T cell responses and pulmonary inflammation during experimental RSV infection 
. How these Tregs interact with the IL-10 secreting effector T cells responding in the respiratory tract to RSV infection to control excess inflammation will require further investigation.
It is of interest that RSV infection induced a lower level of IL-10 release into the infected airways relative to the release of effector T cells derived pro-inflammatory effector cytokine IFN-γ than detected in influenza infected lungs. The explanation for this discrepancy is unclear, but this could reflect an as yet unappreciated mechanism by which RSV infection results in an imbalance in the expression of pro-inflammatory (e.g. IFN-γ and regulatory (e.g. IL-10) cytokines by effector T-cells resulting in exaggerated inflammation/injury profile in the infected lungs out of proportion to the degree of RSV replication in the infected lungs.
Multiple lines of evidence both from human studies and murine models implicate myeloid cells of the monocyte/macrophage/dendritic cell lineage as the major targets of IL-10 action in vitro
and (in model systems) in vivo 
. Thus results from the current study suggest that effector T cell-derived IL-10 can act directly on these inflammatory mononuclear cells infiltrating the RSV infected lungs to decrease production of pro-inflammatory cytokines/chemokines by these inflammatory mononuclear cells as well as to modulate the expression of costimulatory ligands (e.g. CD40, 80, 86 etc.) and T-cell stimulatory cytokines (e.g. IL-12 etc.) 
. The latter effects of IL-10 would be expected to diminish the efficiency of effector T cell triggering in response to contact with these APC populations and thus down regulating the effector activity of T cells. While our results are consistent with such a mechanism, our findings on the impact of conditional deletion of the IL-10Rα selectively in T-cells suggests the novel possibility that the effector T cell-derived IL-10 may also act in an autocrine fashion to regulate effector T cell activity directly. Of note, using animals deficient in IL-10Rβ chain, several recent reports have provided evidence that IL-10 may act directly on and suppress the function of T cells including Treg cells 
T cells directly in vivo
during acute LCMV infection 
as well as inhibit robust memory CD8+
T cell development 
. It should be noted, however, that IL-10Rβ is the common subunit for receptors recognizing several other cytokines including IL-22, IL-26 and IL-28 etc 
. Our results using the conditional deletion of the IL-10 receptor a chain, which is unique to IL-10, firmly establish that effector T cells are able to respond to IL-10 directly in vivo
during infection. Furthermore, we described a novel autocirne function of IL-10; that effector T cell-derived IL-10 is able to signal back to effector T cells, especially CD8+
effector T cells, to restrict the proinflammatory cytokine production by these effector T-cells. Importantly, this autocrine regulatory function of IL-10 acting on effector T cells modulates pulmonary inflammation and thereby results in diminished host morbidity. A similar autocrine mechanism has been reported recently for regulation of IL-10 producing macrophages during endotoxin challenge 
. The mechanisms through which IL-10R engagement on activated effector T cells acts to suppress effector T cell functions at sites of infection are currently under investigation.
IL-10 has been detected both in respiratory tract secretions and in the serum of infants and young children during the acute phase of RSV infection 
. Although controversial 
, several lines of evidence suggest that the level of IL-10 secretion may correlate inversely with disease severity 
. This may be particularly evident among children hospitalized for RSV infection where those children with symptoms of severe RSV bronchiolitis requiring mechanical ventilation express lower levels of IL-10 than hospitalized children with less severe disease 
. Similarly, a recent study reported lower IL-10 levels in stimulated cord blood of children who were hospitalized for RSV infection before 6 months of age than in cord blood of infected infants who were treated as outpatients 
. Furthermore, homozygosity for certain IL-10 alleles correlates with a higher risk of severe RSV bronchiolitis 
. Such studies point to the importance of IL-10 in controlling the severity of acute infection with RSV.
The sources of IL-10 during acute primary RSV infection have not been clearly defined. While various cell types have been implicated as the source of this IL-10 
, our results point to effector T cells as a potential major source of IL-10 produced during the acute phase of RSV infection. In this connection, we did observe a small increase of IL-10 released into the airways (BAL fluid) prior to the dramatic increase in IL-10 production observed in the respiratory tract at the time of effector T cell infiltration at day 5 post RSV infection. Thus, while one or more additional cell types may contribute to the pool of IL-10 observed during acute RSV infection, our results suggest that effector T-cells (both CD4+
) may be a major source of IL-10 during human infection. Therefore the contribution of IL-10 from effector T cells should be rigorously evaluated in future studies of human RSV infection. Our analysis also suggests that the RSV specific effector T cells in the infected lungs serve as an important target of IL-10 action. Thus, the development and extent of immune mediated pathology in RSV infection may not only be linked to the level of IL-10 production by effector T cells (and potentially other cell types in the infected lungs) but also dependent upon the effectiveness of IL-10 signaling through the IL-10R on effector T cells leading to modulation of effector T cell function.
In summary, we have discovered a critical role of effector T cell-derived IL-10 in controlling the pulmonary inflammation and function during RSV infection. Furthermore, we established a previously unrecognized autocrine function of IL-10 in controlling proinflammatory activity of anti-viral effector cells. Our findings thus provide a cellular and mechanistic link to earlier clinical studies which implicate IL-10 in the pathogenesis of RSV disease and may provide the groundwork for future studies examining IL-10 as a therapeutic option in the treatment of RSV induced bronchiolitis in young infants.