Acute HIV and SIV infections are associated with significant perturbations in mucosal homeostasis that are characterized by a massive loss of CD4 T cells, viral replication, and damaged mucosal barrier (6
). These changes occur within the first few weeks of infection and lead to a chronic state of inflammation that persists throughout the course of infection. Our findings suggest that systemic translocation of microbial products likely occurs very early during the course of infection.
Elevated sCD14 and LBP levels significantly correlated with the suppression of IL-17 production in CD4 T cells, supporting earlier observations that the loss of Th17 cells is associated with the translocation of microbial products (5
). Brenchley et al. (8
) showed that peripheral blood CD4 T cells are skewed toward a Th1 phenotype, as opposed to a Th17 phenotype, during chronic HIV infection, whereas others have reported a loss of IL-17-producing CD4 T cells in peripheral blood during SIV infection (4
The suppression of IL-17 expression was not likely due to the paucity in IL-17-promoting cytokines, such as IL-6, IL-21, IL-23, and TGF-β, as we did not observe a suppression of these responses in PBMCs. Other studies have shown that plasma levels of IL-6, IL-21, and TGF-β were elevated during HIV and SIV infections (45
), though studies have reported a decrease in IL-21 levels in the plasma (50
) likely due to the loss of IL-21-producing Th17 cells (44
). On the other hand, Zhao et al. showed that circulating IL-21 levels were increased during simian-human immunodeficiency virus infection (51
). The relatively normal expression of IL-21 that we observed in total PBMCs may be due to the presence of CD8 T cells that were capable of producing IL-21, as some studies have reported (52
Though altered cytokine levels may play a role in suppressing the induction of Th17 cells, the significantly low level of IL-17 expression at day 35 p.i. relative to that at day 7 p.i. after short-term stimulation with rIL-6 pointed to a role for other mechanisms in suppressing Th17 responses. In fact, we observed a significantly high negative correlation between IL-17 expression and the negative regulators of IL-17 production in CD4 T cells, namely, PIAS3, SHP2, and SOCS3.
PIAS3 is a potent inhibitor of activated STAT3 signaling (53
). PIAS3 transcripts are absent in Th17 cells but are present Th1 and Th2 cells, and a downregulation of PIAS3 was found to induce Th17 cells and increase the severity of experimental autoimmune encephalomyelitis (EAE) in mice (54
). On the other hand, SHP2 is a tyrosine phosphatase that has been shown to interfere with the STAT3 signaling pathway, and the transduction of SHP2 was accompanied by a failure to induce Th17 cells (55
). Likewise, SOCS3 has been shown to inhibit cytokine-induced phosphorylation of STAT3 and IL-17 production (29
). Miller et al. (56
) showed that SOCS3 mRNA levels in CD4 T cells were higher in HIV-infected patients than healthy subjects, and the overexpression of SOCS3 was found to inhibit the JAK/STAT pathway. Others have shown that IL-6 stimulation induces SOCS3 expression via the activation of the STAT3 signaling pathway (57
). Taken together these observations suggest that the elevated levels of PIAS3, SHP2, and SOCS3 could potentially play a role in suppressing IL-17 expression during acute SIV infection.
Given that IL-6-mediated signaling requires the activation of STAT3, it was surprising that STAT3 mRNA was not upregulated following short-term stimulation with rIL-6 for 15 min. However, IL-6 primarily acts at the level of STAT3 activation and phosphorylation, and it is highly likely that longer periods (1 to 3 h) of stimulation are required for IL-6-mediated expression of STAT3 mRNA, as some studies have suggested (58
It is not clear from our studies if protein levels for PIAS3, SHP2, and SOCS3 are increased in CD4 T cells, as we were unable to perform these studies for the lack of samples. The upregulated expression of the PIAS3, SHP2, and SOCS3 genes, however, along with the suppression of IL-17 expression, suggests that these negative regulatory factors may contribute to the suppression of Th17 cells very early during the course of infection.
It is highly likely that the suppression of IL-17 expression is due to the SIV-mediated loss of Th17 cells, as has been reported previously (8
), or due to other mechanisms, such as those mediated by the induction of indoleamine 2,3-dioxygenase-1 (61
) or the paucity of IL-21-producing CD4 T cells (44
). Our studies do not rule out a role for these mechanisms but suggest that additional mechanisms, such as those that negatively regulate the IL-17 signaling pathway, may play a role in suppressing the expression of Th17 cells during SIV infection and that these suppressive mechanisms were apparent very early during the course of infection.
Increased translocation of microbial products correlated with IL-23 expression in monocytes and was associated with a significant upregulation in the density of CD14 expression, suggesting that translocated products could contribute to the activation of monocytes during acute infection. Ancuta et al. (34
) demonstrated that microbial translocation was associated with increased monocyte activation in HIV-infected patients, whereas Manuzak et al. (63
) showed that bacterial antigens increase IL-23 production by peripheral blood monocytes. Louis et al. (64
) showed that monocytes from patients with primary HIV infection significantly enhance IL-23 production in response to LPS stimulation. Kader et al. (16
) showed that IL-23 mRNA levels were upregulated in the mucosa during acute stages of SIV infection and stayed high even after the loss of CD4 T cells.
Interestingly, neither the suppression of IL-17 expression nor increased levels of sCD14 or LBP directly correlated with acute immune activation, suggesting that mechanisms other than translocated microbial products likely drive the immune activation observed very early during infection. Acute infection is characterized by a massive amplification of viral infection that is accompanied by immune activation during the early stages of infection (10
), likely due to the release of various proinflammatory mediators (65
). In line with this argument, we observed a significantly high positive correlation between plasma viral loads and CD8+
T cells in peripheral blood. Interestingly, immune activation appeared to precede elevated levels of plasma sCD14 and LBP; CD8+
T cell levels were significantly higher at day 7 p.i. than at day 0, without an apparent change in sCD14 and LBP levels.
Though sCD14 and LBP levels did not correlate with acute immune activation, they significantly correlated with IL-23 expression in monocytes and the MFI of CD14 expression on monocytes, suggesting that translocated products could potentially interact with the higher levels of CD14 on monocytes and contribute to the generalized state of inflammation during acute SIV infection. The levels of sCD14 and LPB that we observed during acute stages of infection were lower than what has been reported in chronically infected animals (2
) and in line with those described in previous reports showing low but detectable levels of LPS in the mucosa during early stages of SIV infection (6
). Given the low levels of translocated products during early infection, it is highly likely that the direct effect of translocated microbial products on acute immune activation is probably masked by the release of inflammatory mediators associated with massive viral replication. Breed et al. (66
) showed that rhesus macaques infected with the SIVmac239ΔGY mutant strain exhibited wild-type acute viremia and immune activation but maintained their mucosal CD4 T cells and displayed a lack of microbial translocation.
In conclusion, our studies show that acute SIV infection is characterized by a significant increase in markers of microbial translocation that correlate with suppressed Th17 responses. Elevated expression of various negative regulatory genes likely plays a role in suppressing IL-17 expression in CD4 T cells during acute stages of SIV infection. Active viral replication rather than translocated microbial products, however, correlated with Ki-67 expression on CD8 T cells, suggesting a primary role for viral replication in acute immune activation.