Much that is understood regarding CD4+
T cell depletion, heightened T cell activation states, T cell dynamics, and HIV-specific T cells in HIV infection is derived from the analysis of peripheral blood lymphocytes. Although the importance of studies of how HIV infection affects lymphoid tissue is certainly appreciated (15
), there are few and only very recent studies of the GI tract (8
) due to the difficulty in obtaining tissue samples. Substantial ground has been gained through the SIV rhesus macaque model (5
). However, to date there have been no reports comparing CD4+
T cell populations in peripheral blood, GI tract, and LNs from HIV-infected and -uninfected individuals.
From our current study, the following five major points emerged: (a) the GI tract has the most substantial CD4+ T cell depletion at all stages of HIV disease; (b) this depletion occurs preferentially within the CCR5+ CD4+ T cell subset, which accounts for the majority of GI tract CD4+ T cells; (c) HIV-associated immune activation results in an accumulation of effector/TEM cells within LNs; (d) HIV-specific T cells residing in LNs do not, alone, account for the inflammatory T cell response within HIV-infected LNs; and (e) T cell activation in LNs is associated with collagen deposition.
Quantitative analysis of CD4+
T cell depletion in LNs and the GI tract is substantially more challenging than in peripheral blood. LNs and the GI tract are not homogeneous organs and histological sections sampled for analysis may not be representative; therefore, other parameters must be examined. Perhaps the most obvious variables to quantify are either CD4+
T cell ratios or CD4+
T cell percentages by flow cytometry. Indeed, such analysis suggests that the GI tract is substantially more depleted of CD4+
T cells than either peripheral blood (24
) or LNs. However, examination of the same anatomical compartments from healthy individuals demonstrates that even under normal circumstances, the GI tract contains a lower percentage of CD4+
T cells compared with either peripheral blood or LNs. Thus CD4+
T cell percentages cannot be used to estimate CD4+
T cell depletion in tissue. Therefore, we examined three other, independent parameters to demonstrate that the GI tract was preferentially depleted of CD4+
T cells. Measurement of CCR5+
T cells, endoscopic and histological examination of the GI tract, and measurement of activated T cells in each compartment suggested that the GI tract was significantly depleted of CD4+
T cells compared with either peripheral blood or LNs and that this occurred even at very early time points after infection.
Interpretation of the changes in T cell subsets that differ between lymphoid tissue compartments is complicated by the nature of the analysis—“snap-shots” of tissues—and by the potential mechanisms to explain these changes that include preferential infection and killing of T cell subsets, and the redistribution, proliferation, activation-induced T cell death, and alterations in the lymphoid tissue milieu that accompany immune activation. We believe that compartment-specific variations in these processes account for the differences we observed, particularly in the CD4+ T cell population in the GI tract.
The majority of GI tract CD4+
T cells expresses CCR5 and comprises a higher frequency of activated T cells than peripheral blood or LNs in HIV-uninfected individuals. Thus, they represent ideal targets for HIV replication. It is likely that direct infection of this population results in its profound depletion during the acute phase of HIV disease. Ongoing direct infection and sustained death might explain the continued depletion of GI tract CCR5+
T cells that is only partially offset by proliferation of CD4+
T cells in the GI tract, and resulting in apparently normal levels of CD4+
T cell activation. On the other hand, in LNs, there is a smaller proportion of activated CCR5+
T cell targets and thus proliferation and recruitment of CD4+
T cells might sufficiently exceed direct killing for the heightened state of immune activation to be evident in this compartment. In addition, it is possible that during the chronic phase of the disease, the disruption of the homeostatic processes that maintain total body T cell numbers (44
) would in itself hinder CD4+
T cell reconstitution in lymphoid tissue. The consequence of this would be particularly damaging in the GI tract as, in contrast to LNs, there is only a negligible resident naive CD4+
T cell pool available to become activated, expand, and supply the already profoundly depleted memory CD4+
T cell pool.
Alternatively, the decrease in the frequency of CCR5+
T cells in the GI tract in HIV infection might arise from altered migration of activated CCR5+
T cells into the GI tract, or from recruitment of CCR5−
T cells to the GI tract. However, the latter explanation requires specific infiltration of two unrelated T cell subsets, CCR5+
T cells and CCR5−
T cells, even though total GI tract lymphoid tissue appears to be dramatically decreased overall. Therefore, we believe the most likely explanation is that direct infection and killing, either by HIV or by HIV-specific T cells, of GI tract CCR5+
T cells leads to their profound depletion in acute infection, and that this depletion is maintained during the chronic phase of the disease. In addition, it is possible that the few CCR5+
T cells in GI tract that persist represent resting CCR5+
T cells that have never seen antigen and might be resistant to HIV-mediated lysis (45
). Importantly, we chose to biopsy terminal ileum as this site has the greatest concentration of GI tract T cells. Although our samples contained a combination of lamina propria and Peyer's patch lymphoid tissue, the majority of our biopsies contained a low frequency of naive T cells, suggesting that they consisted predominantly, but not exclusively, of lamina propria. Because we cannot distinguish between these lymphoid compartments, it is possible that CD4+
T cell depletion is more substantial in lamina propria compared with Peyer's patches, and we may have observed more dramatic CD4+
T cell depletion if we had also biopsied jejunum or colon, which lack Peyer's patches (45
). Hence, we may have even underestimated the extent of CD4+
T cell depletion from the entire GI tract.
It has been suggested that immune activation associated with HIV infection would lead to increased percentages of CCR5+
T cells in peripheral blood, as seen in acute EBV infection (33
). However, although HIV-infected individuals contain increased frequencies of peripheral blood CCR5+
T cells, a corresponding increase in peripheral blood CCR5+
T cells is not observed. One explanation for this observation is that there might be redistribution of CCR5+
T cells from peripheral blood to LNs or GI tract in HIV infection (48
). However, upon examination of CD4+
T cells in the LNs and GI tract, we found no increases in CCR5+
T cell percentages. Therefore, we believe our data suggest that accumulation of CCR5+
T cells resulting from immune activation associated with HIV infection is offset by their preferential death rather than redistribution, whereas activated CD8+
T cells accrue. Importantly, as the percentage of infected peripheral blood CD4+
T cells is usually <1% in chronic infection (39
), it seems unlikely that direct infection of activated CD4+
T cells is solely responsible for the death of CCR5+
T cells, and other factors likely contribute. Taken together, the findings of substantial CD4+
T cell depletion in the GI tract and lack of evidence for redistribution to LNs, suggest that peripheral blood CD4+
T cell counts may actually underestimate the degree of overall T cell depletion in acute and chronic HIV infection.
Another mechanism contributing to the loss of CD4+
T cells in HIV infection is likely to involve disturbance of normal lymphoid tissue homeostatic processes. Peripheral LNs are structurally organized to promote interaction between antigens, chemokines, growth factors, and lymphocytes to generate an immunologic response and maintain populations of CD4+
T cells (52
). It is likely that the inflammation and tissue remodeling that accompany local innate and adaptive immune responses to HIV replication lead to destruction of LN architecture observed in HIV disease (28
), which, because of the particular dependency of CD4+
T cells on the LN milieu (53
), contributes to decreased survival and depletion of the CD4+
T cell subset. We interpret our data on the sequestration or retention of TEM
cells that are not normally found in LNs as evidence of the chronic proinflammatory responses generated as a result of ongoing viral replication in the LN. Furthermore, B cell activation, characterized by germinal center hyperplasia, might add to LN architectural damage. Importantly, trafficking of T cells to the follicular dendritic cell network is likely to depend on an intact LN architecture, and such trafficking might be consequently impaired in a fibrosed LN. Hence, what was an organ of antigen presentation and homeostasis in health becomes an organ of inflammation and fibrosis in HIV infection. Furthermore, our data continue to suggest that in individuals with significant LN collagen deposition, there might be therapeutic benefit from antiinflammatory and/or antifibrotic agents, singly or in combination with antiretroviral therapy, to prevent further or reduce LN damage.
Finally, our data also demonstrate that the majority of TEM
cells sequestered to or retained in LNs are not HIV specific. One possibility is that they are elicited by subclinical opportunistic infections secondary to the immunosuppressive effects of HIV infection (46
). In fact, the LNs actually had a smaller breadth of HIV-specific T cells compared with peripheral blood, suggesting that HIV-specific T cells are not preferentially recruited to LNs. Indeed, although we have not determined the cytotoxic capacity of HIV-specific T cells in LNs against the resident viral quasispecies, the low frequency of T cells responding at a major site of virus replication may point to one more example of the failure of immune defenses in fully controlling HIV replication.
The results also imply that LNs are not a continuous source of peripheral blood HIV-specific T cells. Moreover, the breadth of HIV-specific T cells was not the only variable we measured that differed between anatomical compartments. There was no clear correlation between the GI tract, LNs, and peripheral blood for any of the parameters we measured. Hence, data obtained from peripheral blood cannot be used to predict or model immunological processes or HIV pathogenesis in the GI tract or LNs, and such compartments should be directly sampled and studied.
The substantial depletion of GI tract CCR5+ CD4+ T cells implies that total body CD4+ T cell numbers are severely reduced, even in very early infection. This, by itself, would impose a considerable homeostatic strain on the maintenance of the memory CD4+ T cell pool. Immune activation that defines the chronic phase of the infection results in destruction of lymphoid tissue architecture that, in turn, impacts upon the ability of lymphoid tissue to support normal lymphocyte homeostasis and antigen presentation. Taken together, our data suggest that HIV infection directly and indirectly causes damage to many immune compartments and a combination of such deleterious effects ultimately leads to immune failure and AIDS. Understanding the relative contribution of each of the factors that we have described can provide a rational framework upon which future therapeutic interventions can be based.