In our study population 60% of women using contraceptives choose MPA, which unlike other contraceptives possesses selective glucocorticoid activity and can alter GR regulated genes 
. Therefore it is extremely important to investigate the potential immune modulatory effects of this synthetic progestin.
This study aimed to determine whether MPA would alter the BCG-specific memory responses of PBMCs of recently exposed HHCs of TB patients. At high concentrations MPA generally mimicked cortisol and unlike progesterone suppressed the production of several cytokines including IL-1α, IL-6 and IL-17. Even at pharmacological concentrations MPA and cortisol but not progesterone, inhibited the production of TNFα, IL-1ra, IL-13 and GM-CSF. Our in vitro findings therefore suggest that in general MPA mimics the effect of cortisol rather than its intended analog progesterone.
GCs affect many aspects of immune cell function and are well known to inhibit the production of Th1 cytokines and cause a shift from a Th1 to a Th2 cytokine response pattern 
. However the inhibitory effect of GCs goes beyond the traditional pro- and anti-inflammatory division and it has been shown that GCs inhibit the production of IL-2, IFNγ, IL-1β, IL-8, TNFα, IL-4, IL-5, IL-6, IL-10, IL-13, IL-17 as well as G-CSF 
. Furthermore GCs are able to decrease the activity of the Th1 transcription factor T-box expressed in T cell (T-bet) 
as well as the Th2 transcription factor GATA-binding protein 3 (GATA-3) 
As part of the classical “genomic” GC signaling pathway, GCs mediate their effects by binding to cytosolic GRs which then translocate to the nucleus and bind to the promoter regions of genes regulated by the GR 
. The anti-inflammatory effects of GCs are mainly mediated by interaction between the ligand activated GR and transcription factors such as NF-κB and AP-1 
. Similarly to GCs, MPA has been shown to inhibit the secretion of IL-2 and IL-6 in T cells almost to the same extent as the glucocorticoid dexamethasone, whereas progesterone only slightly inhibited the secretion of these cytokines 
. During this study we have demonstrated that as with GCs the inhibitory effect of MPA is not restricted to Th1 cytokines alone, but MPA can also impact on Th2 as well as Th17 cytokine responses in vitro.
Ex vivo, we saw that in response to BCG PBMCs from MPA users produced significantly lower levels of IL1α, IL-12p40, IL-10, IL-13 and G-CSF than PBMCs from non-contraceptive users as determined by the Fisher LSD Post-Hoc test. IL-12 plays an important role in the generation of protective immune responses against TB as shown in murine models 
. T cells rely on IL-12 to differentiate into a Th1 phenotype and produce IFNγ at the site of infection 
. IL-12 is a heterodimeric cytokine consisting of two subunits, p35 and p40 
, and is required for optimal differentiation and expansion of activated lymphocytes. It has been suggested that the p40 subunit drives cellular responses 
as p40 knock-out (KO) mice are more susceptible to M.tb
than p35 KO mice. IL-1 is one of the most important proinflammatory cytokines and IL-1 type 1 receptor KO mice as well as IL-1α/β KO mice infected with M.tb
, have significantly larger granulomas with neutrophilic infiltrates in the lungs compared to WT mice 
. The IL-1 receptor KO mice were highly susceptible to M.tb
infection and had significantly more bacteria in their lungs, livers and spleens 
. Susceptibility to M.tb
was associated with a lack of Th1 responses and impaired granuloma formation. IL-1 is therefore crucial for inflammatory cell recruitment into granulomas and host defense against M.tb
. Lower levels of BCG-induced IL-12p40 and IL-1α observed in MPA users in our study could therefore suggest that they are more susceptible to M.tb
and might have a worse outcome of disease.
IL-10 regulates Th1 cell responses by inhibiting the production of IFNγ and IL-12 
. Mice deficient in IL-10 have an enhanced ability to control M.tb
infection as they have significantly lower numbers of bacteria in the lungs and spleens 
. The increase in protection observed in IL-10 deficient mice was associated with enhanced Th1 responses at the site of disease; however these mice succumb to the disease due to elevated IL-12 and IFNγ levels which result in severe tissue pathology 
. The increased mortality seen in mice during the absence of IL-10 is not a result of the uncontrolled growth of the bacteria but rather that of unregulated immune responses to the infection. A reduction in IL-10 production will result in a decreased number of circulating Th2 cells leading to less IL-13 being produced; therefore we hypothesize that a reduction of IL-13 could be a result of an indirect effect of the hormone. Lower levels of IL-10 and IL-13, could result in more sever immunopathology in individuals suffering from active TB and using MPA. Taken together it is therefore likely that MPA use will not only have an effect on susceptibility to TB but possibly also TB disease severity.
GC therapy not only inhibits the production of cytokines, but has also been shown to reduce the circulating monocyte population in patients receiving GC therapy 
. Thus the reduction in numbers of circulating monocytes in women using MPA shown in this study could be attributed to the glucocorticoid activity of this steroid. We could therefore speculate that at least in part the inhibition of IL-12, IL-1α, IL-10 and IL-13 in MPA users can be attributed to lower numbers of circulating monocytes. The reduction of circulating monocytes together with reduced production of IL-1 and IL-12 in response to mycobacterial antigens could increase the susceptibility of MPA users to TB and increase the risk of progression from latent to active disease. For future studies with larger sample numbers it would be particularly interesting to correlate the immunological measurements of MPA users with duration of MPA use.
Our research study is the first to show that MPA impacts on infections outside the genital tract due to a systemic effect on immune function. These findings warrant further investigations into the effect of MPA on susceptibility to TB, disease severity and treatment outcome. Furthermore MPA use could change the efficacy of new BCG-based vaccines and especially prime-boost vaccine strategies which may be administered to adult or adolescent women in the future.