We demonstrate that HLA-G is expressed by differentiated airway epithelial cells in culture. Both G1 and G5 transcripts were demonstrated by qPCR, and soluble G5 and very modest membrane-bound G1 protein expression, were demonstrated by confocal microscopy. These data corroborate the finding of HLA-G in normal and asthmatic airways in vivo
[22, 26] and thus provide a useful model for studies of regulation and secretion. Further, we demonstrate that the Th2-associated cytokines IL-13, IL-4, and IL-5, and the immunoregulatory cytokine IL-10, do not regulate the expression of HLA-G in dAEC.
Epithelial cell secretion of HLA-G may have a role in regulating inflammation in asthmatic airways. Soluble isoforms of HLA-G secreted either into the airway lumen or into the local circulation may have a paracrine role in down-regulating inflammatory cells within airways, and in extending its influence beyond the epithelial layer where it can directly contact lymphocytes and dendritic cells in and near the mucosa. Although there is no direct evidence yet for this hypothesis in asthma, in other contexts HLA-G has been shown to suppress dendritic cells and T cells
[27, 33] that participate in inflammation, and to activate CD4+CD25+FoxP3+ regulatory T cells
 that can suppress cells that participate in airway inflammation
. We have recently demonstrated increased presence of soluble HLA-G in bronchoalveolar lavage fluid recovered from subjects with mild asthma versus control, non-asthmatic subjects
; this may represent on-going attempts to suppress (incompletely or unsuccessfully) airway inflammation.
Because of this one could hypothesize that Th2-associated cytokines, demonstrated in increased abundance in asthmatic airways, would regulate HLA-G expression and abundance in airway epithelial cells that would in turn down-regulate certain aspects of airway inflammation would be counter-productive. The counter-hypothesis is that IL-4 and IL-13 drive airway inflammation and thus would not stimulate a counter-regulatory mediator such as HLA-G. The latter is correct: in our study, no Th2-associated cytokine stimulated HLA-G expression.
The overall role of Th2-associated cytokines in airway inflammation is clearly context-specific. Effector mechanisms by which these cytokines may defend the host appropriately (e.g., against infection) are the same that cause inappropriate or prolonged inflammation. Stimulation of AEC by IL-4 elicits production of several cytokines and chemokines
[60–62] that contribute to airway inflammation in asthma, yet IL-4 also stimulates AEC migration, a pro-reparative process
. Stimulation of AEC by IL-13 elicits cytokine expression
[61, 62] as well as mucoid metaplasia
[14, 63], both of which are pro-inflammatory processes.
Our data also show that the IL-10R1 receptor subunit is expressed at low concentration in both airway epithelium in situ and in our dAEC culture model, and stand in contrast to the observations of Lim et al.
. However, despite the presence of both receptor subunits, treatment of dAEC with IL-10 did not elicit either STAT3 or STAT1 phosphorylation and thus, not surprisingly, we were unable to demonstrate significant HLA-G5 expression in response to this cytokine. Our data suggest that in contrast to expression seen in macrophages
, IL-10 does not stimulate HLA-G expression in airway epithelial cells.
As our data are derived using an in vitro cell culture model, important influences on HLA-G expression may be absent, including local and circulating factors and the influence of cells beneath the basement membrane such as fibroblasts. Thus, the response to Th2-associated cytokines in the intact human airway may be different. However, the use of ALI culture permits examination of cytokine influences in isolation. Future experiments can examine the role of paracrine and circulating factors added to this system. HLA-G expression may also change as a function of disease state and inflammation in airways. Our experiments utilized cells collected from normal subjects, and it is possible that asthmatic epithelium may respond differently, either in a culture system or in the intact, inflamed airway. Careful comparison and correlation then will be required to understand specific receptor expression changes in a developing or repairing epithelium in vivo.
In conclusion, our study demonstrates the presence of HLA-G in differentiated airway epithelium in culture and its lack of regulation by Th2-associated cytokines such as IL-4 and IL-13 and by the immunomodulatory cytokine IL-10. While the role of HLA-G in airway inflammation and asthma remains to be defined, its presence, particularly in greater abundance in asthmatic airways (54) and in plasma of asthmatic subjects
[24, 25], suggests that epithelial cell expression may be of importance in airway inflammation.