It has been found that

It has been found that WHI-P131, a JAK3 inhibitor, induced apoptosis of CD4+ T JDTic 2HCl (Cetkovic-Cvrlje et al., 2012). Also, other JAK3 inhibitors (AG490 and WHI-P154) caused apoptosis in anaplastic large cell lymphoma (Amin et al., 2003). Taking into account the proapoptotic action of JAK inhibitors, it was also considered that the anti-inflammatory and anti-allergic effects of OCL, apart from the principal mechanism responsible for these effects, can be additionally related to the induction of apoptosis of effector CD4+ and possibly CD8+ T cells because the latter cells are likely to contribute to the pathogenesis of canine AD (Jassies-Van Der Lee et al., 2014). Therefore, the effect of OCL on the apoptosis of canine CD4+ and CD8+ T cells was assessed. In order to verify the above hypotheses an in vitro effect of OCL was assessed on: (a) a number of selected CD4+ and CD8+ T cell subsets; (b) activation-induced CD25 expression on CD4+ T cells; (c) apoptosis of CD4+ and CD8+ T cells.
Material and methods
Results
Discussion To the best of our knowledge, the available literature does not contain any reports on the effects of OCL and other JAK inhibitors on canine CD4+ and CD8+ T cells. Thus, we here report for the first time the in vitro effect of OCL on these cells. Presently, Foxp3 is considered as the most definitive marker used to distinguish Treg cells from their thymic progenitors and activated Teff cells (Foxp3−CD25+CD4+ and Foxp3−CD25+CD8+); the CD25−CD4+ and CD25−CD8+ phenotypes represent mostly resting T (Trest) cells (Sondel et al., 2003). As mentioned earlier, JAK3 kinase is essential for a proper expression of Foxp3, because the IL-2/JAK3/STAT5 signaling pathway participates in the initiation and maintenance of the Foxp3 factor in Tregs (Goldstein et al., 2016). In a study on mice it was found that such inhibitors of JAK3 as WHI-P131 (Cetkovic-Cvrlje et al., 2012), ZM39923 (Goldstein et al., 2016) and tyrphostin/AG490 (Goldstein et al., 2016) reduced Foxp3 expression in CD4+ Treg cells under in vitro conditions. Reduction in the expression of Foxp3 caused by JAK3 inhibition was also found in human CD25+CD4+ T cells (Goldstein et al., 2016). Therefore, in the present study, we hypothesized that inhibition of JAK3 induced by OCL may down-regulate Foxp3 expression in canine Treg cells. However, we did not find any effect of OCL on the Foxp3 expression in CD4+ and CD8+ Treg cells. This indicates that OCL does not affect JAK3 to the extent to influence the expression of Foxp3. This supports results of some previous research indicating that OCL is a relatively weak JAK3 inhibitor (Gonzales et al., 2014; Roskoski, 2016). Thus, the current results strongly suggest that, in terms of the effect on the expression of Foxp3, OCL seems to be a safe drug. This does not mean that OCL lacks any negative effect on CD4+ and CD8+ Treg cells. The existing literature does not provide any data that would refer to the influence of this agent on counts of CD4+ and CD8+ T cells in in vivo and/or in vitro conditions. Nevertheless, according to the drug manufacturer (), clinical pathology changes observed in OCL-treated dogs were limited to a decrease in mean leukocyte count, which included all white blood cell counts except lymphocyte counts. This information does not agree with the results of our investigation, where it was shown that exposure of peripheral blood lymphocytes to OCL led to a dramatic loss of CD4+ and CD8+ T cells, both Treg cells and Teff cells. The study reported in this paper demonstrated that the sensitivity of particular T lymphocyte subpopulations to the cytoreductive effect of OCL may be different. Treg cells seem to be more sensitive than Teff cells, and CD8+ Teff cells appear to be more sensitive than CD4+ Teff cells to the impact of OCL. At the first time point, the mean loss of these cells was smaller by just a few per cent. Thus, the cytoreductive effect of OCL appeared quickly and showed a slight tendency toward becoming stronger.