Innate immune activation is critical to control infections. In the case of HIV infection, however, innate immune activation also drives HIV replication via signaling pathways downstream of TLRs.26
Therefore, HIV is able to exploit the activation of innate immunity for its own advantage. This feature of HIV may have important implications for HIV-1-infected patients who are coinfected with opportunistic pathogens. Although it is known that OIs in HIV-1-infected individuals are frequently associated with increased viral loads,27
the mechanisms by which coinfecting pathogens facilitate AIDS progression remain poorly understood. Here, we evaluated the effect of coinfection on TLR2 and TLR4 expression in monocytes and DCs from in HIV-1-infected patients.
studies suggest that stimulation of TLR2, TLR4, and TLR9 causes up-regulation of viral replication14–18
by activating NF-κB in HIV-1-infected cells. However, activation of NF-κB through TLR4 alone is not sufficient to activate viral LTR and virus replication,28
while stimulation via TLR2 always induces LTR HIV activation. Presumably, this is because TLR4 signaling is also able to induce type I IFN release,29
which acts as an antiviral factor. On the other hand, the fungal zymosan, which stimulates TLR2 and dectin-1, was reported to inhibit HIV replication.30
Nevertheless, pure TLR2 and TLR5 ligands increase cellular HIV-1 integration, trigger reactivation of latent HIV-1 provirus in T cells, and activate virus gene expression in central memory CD4+
Thus, TLR activation may lead to modulation of HIV infection due to the effect of downstream signaling effectors on viral replication.
We observed a marked increase of TLR2 and TLR4 expression in mDCs of HIV-1 patients coinfected with different pathogens in vivo
. Similar results were observed in monocyte-derived macrophages and PBMCs from healthy donors and in vitro
infected with HIV-1, treated with TLR2 and TLR4 agonist. Consequently, with this increase in TLR expression, an effect on the functionality of TLRs was observed based in the high expression of proinflammatory cytokines such as IL-1β, IL-6, IL-8, and TNF-α, and the induction of pDC maturation, compared to mock infection (data to be published separately). These results extend previous findings that HIV-1 infection itself leads to up-regulation of TLR2.19
Likewise, HIV-1 and its products can also modulate TLR expression and functions. For instance, the ssRNA40 (HIV-1-derived RNA) increased TLR3 and TLR8 expression in activated T cells,25
as well as TLR4 mRNA expression,20,21
together with an increased proinflammatory response to TLR ligands. Increased TLR4 expression by T cell subsets was reported in HIV-1-infected patients.22
Chronic HIV-1-infected patients who failed to respond to HAART showed reduced expression of TLR3, TLR4, and TLR9 together with increased expression of TLR7, which correlated with high HIV-1 RNA levels.23
The results suggest that the virus-mediated immune-modulator pathway involved in regulating TLR expression could represent an important pathogenic event in chronic HIV-1 infection.
Consistent with this hypothesis, an association between HIV disease progression and polymorphisms in TLR432
or the levels of soluble TLR2,35
has been reported. Interestingly, a 3′-UTR polymorphism in the NLRP3 gene, a member of the cytosolic NOD-like receptors, was also associated with increased susceptibility to HIV-1 infection.36
Overall, these results point to an important role of the TLRs, or even other PRRs, in modulating AIDS progression.
Whether TLR signaling is beneficial for the host, or enhances virus replication and spread, depends on different factors: the kind of TLRs activated, the doses of TLR ligands involved, the duration of the stimulation, the cell types stimulated, and the stage of HIV-1 infection (acute vs. chronic). Based on our results, we propose a dual role of TLR during HIV-1 infection. During the initial phase of infection, TLR signaling could promote IFN-α/β release, which could have antiviral effects. However during chronic infection, TLR stimulation could induce a strong inflammatory response that would increase HIV-1 replication. This can explain the different results obtained by several research groups, showing an increase or decrease in HIV-1 replication,14,37,38
or even transmission from DCs to CD4+
after TLR stimulation when using different TLR ligands, virus strains, and cellular models (mastocytes,16
macrophages, lymphoid tissue,41
), or animal models (transgenic mice that incorporate the HIV-1 genome).15,43
The main goal of HAART is to block HIV-1 replication and achieve immune reconstitution in HIV-1-infected patients. In this study we report that mDCs of HIV-1-infected subjects with OI and without HAART present higher expression levels of TLR2 and TLR4 than those from coinfected patients on HAART therapy. This could be associated with a higher viral load since TLR2 and TLR4 are able to mediate the activation of HIV-1 LTR through the NF-κB pathway.14,15,18
Together, these results suggest that TLR expression levels could be influenced by viral factors, opportunistic pathogens, and the immunological state of the host.
We have also observed that the percentage of monocytes and mDCs decreased significantly in HIV-1 patients coinfected with OI and without HAART. In contrast, the percentage of pDCs was similar in all the groups of HIV-1-infected individuals studied (Supplementary Fig. S1
). Interestingly, despite a lower percentage of these cell types, the mDCs of HIV-1-infected subjects with OI and without HAART expressed higher levels of TLR2 and TLR4, the pDCs expressed higher levels of TLR4, and the monocytes expressed higher levels of TLR2. Previous reports have shown restoration in cellular subsets following HAART.44,45
HIV-1 patients coinfected with OIs in our study exhibited restoration of mDCs, as well as monocytes, after HAART.
Based on these observations, we propose that increased TLR expression, together with a higher inflammatory response, increases viral replication through NF-κB. In addition to increased TLR4 expression in pDCs, its activity might also, thereby, induce IFN-α secretion and increased loss of CD4+
T cells via TRAIL-mediated apoptosis.46
On the other hand, there is a second possibility, whereby patients with higher viral load, especially those without HAART, maintain a proinflammatory state that also increases TLR expression. In both cases, TLR up-regulation in HIV-1-infected patients could represent an immune-pathogenic event that would accelerate the progression to AIDS in patients not on HAART, primarily in the context of an opportunist infection.
TB is one of the most important OIs in HIV-1-infected patients, especially among Colombian subjects. Patients with TB and without HIV-1 infection showed no difference in TLR2 expression in monocytes compared to healthy donors,47
while TLR8 expression was up-regulated in patients with acute TB, as well as in differentiated macrophages upon infection with the M. bovis
Our results demonstrate that TLR2 and TLR4 are most strongly up-regulated in mDCs of patients coinfected with TB and HIV-1, which could lead to enhanced activation of cells by TB-derived products.
After HAART use, the responder patients (subjects with HAART in whom the viral load was significantly reduced) show an apparent “normalization” of TLR expression, despite low CD4 cell counts. Thus, the association between TLR expression and plasma HIV-1 viral load points toward the regulation of expression by viral products rather than large shifts in innate cell populations. Interestingly, in monocytes, expression of TLR2 was also positively correlated with the viral load, and as one might expect, it was lower in coinfected patients with HAART compared to patients without HAART. This is similar to previous reports in chronic HIV-1-infected patients with advanced disease (CD4+
T cell count less than 200 cells/ml).20,22
However, our data do not show a correlation between TLR expression and CD4+
T cell count (data not shown). Finally, the mechanisms by which HIV-1 infection and opportunistic pathogens increase TLR expression and possibly TLR functions need to be further examined.
Similarly, up-regulation of TLR2 and/or TLR4 during viral infections has been reported. For example, overexpression of TLR2 and TLR4 in monocytes of hepatitis B virus (HBV)-infected patients49
and dengue virus-infected patients was reported.50
However, down-modulation of TLR7 and TLR9 expression was reported in cells of HCV- or HBV-infected patients, which negatively correlated with viral load.51,52
Together, these results suggest an important role of TLRs during viral infection. On the one hand, they act to increase antiviral immunity, but they can also promote pathogenic events with altered TLR expression.
Remarkably, we have also observed increased expression of TLR4 in pDC (defined as Lin1−
PBMCs) at the protein level in HIV-1-infected patients. This is the first report of TLR2 and TLR4 expression (at the protein level) on pDCs (Supplementary Fig. S2
), although previous reports showed very low amounts of TLR2 transcripts in BDCA-4-purified pDCs, as well as minimal CD80 induction after 48
h of lipopolysaccharide (LPS) stimulation.53,54
Moreover, we also observed this phenomenon of TLR2 and TLR4 expression on pDCs derived from patients with other viral infections (unpublished data).