We undertook this study to examine the role of differences in chemokine receptor expression on infection of MMCs and PBMCs by R5- and X4-tropic HIV-1. A greater percentage of MMCs than PBMCs are infected by both R5- and X4-tropic HIV-1. Significant differences exist in terms of chemokine receptor expression in the blood and gastrointestinal mucosa; mucosal cells are predominantly CCR5+ CXCR4+, whereas these cells make up less than 20% of the peripheral blood cells. This cell population is most susceptible to infection with both R5- and X4-tropic HIV-1 in both compartments. Regardless of whether viral isolates were derived from the blood or mucosa of HIV-1-infected patients, HIV-1 p24 production was greater in MMCs than in PBMCs. Further, the chemokine receptor tropism of these patient-derived viral isolates did not differ between compartments. We conclude based on these findings that the gastrointestinal mucosa represents a favored target for HIV-1, in part due to its large population of CXCR4+ CCR5+ target cells and not due to differences in the virus that it contains.
The gastrointestinal mucosa is a secondary lymphoid organ that contains the majority of the body's CD4+
lymphocyte population (6
) and appears to support enhanced HIV-1 replication compared to other body compartments (10
). Our studies indicate that one significant reason for the unique susceptibility of the gastrointestinal mucosal compartment is the greater infection of, and increased virus production from, CCR5+
cells. We have shown that the majority of gastrointestinal mucosal CD4+
cells express both CXCR4 and CCR5 and are therefore susceptible to both R5- and X4-tropic HIV-1. In comparison, the majority of PBMCs are CXCR4+
and resistant to infection with R5 virus. The data we present in this study suggest that the unique susceptibility of the mucosal compartment is due, in part, to the greater percentage of CCR5+
cells that it contains compared to blood. Still, we have not proven that mucosal CCR5+
cells are more prone to R5- or X4-tropic HIV or produce more virus than do CCR5+
Antigenic stimulation of lymphocytes drives HIV replication (36
). Therefore, secondary lymphoid organs are the main sites of HIV replication in vivo (8
). The majority of gastrointestinal mucosal T lymphocytes are memory cells and exhibit phenotypic and functional features of activation (31
). This likely reflects the vital immunologic role these cells play at the major boundary between humans and the external environment. The higher expression of CCR5 in the gastrointestinal mucosa likely reflects this state of inflammation since CCR5 is expressed on activated memory lymphocytes (5
). Given these characteristics, the enhanced susceptibility to and replication of HIV-1 in the gastrointestinal mucosa is expected. Compared to PBMCs, the greater percentage of CCR5-expressing MMCs permits higher levels of infection by R5-tropic virus. Since these cells are predominantly of a memory and/or activated phenotype, they support HIV replication. Since the majority of MMCs expressing CXCR4 also coexpress CCR5 and are therefore memory and/or activated cells, infection of these cells with X4-tropic virus will also result in virus production. In comparison, infected CXCR4+
PBMCs, which have a naive phenotype, should not support significant viral replication. Since both M- and T-tropic HIV-1 predominantly infected CCR5+
cells in both compartments, the higher expression of mCD24 on the infected MMCs may reflect a higher degree of activation in the mucosa of these phenotypically similar cells. We have previously shown that CCR5-bearing MMCs express higher amounts of CCR5 per cell than do CCR5-bearing PBMCs (3
), which could potentially reflect a higher degree of activation.
A number of viral factors have been proposed to influence the clinical course of HIV-1 infection. These include quasispecies diversity, coreceptor usage, cellular tropism, and replicative capacity. HIV-1 quasispecies constantly evolve in response to their dynamic relationship with their cellular targets and the host immune response. If distinct viral evolution occurs in the PBMC and mucosal compartment, then viral factors could potentially explain the high mucosal susceptibility to HIV and SIV. Since viral replication and therefore genotypic evolution are driven by the inflammatory state of the infected cell, the dynamic inflammatory environment of the gastrointestinal mucosa may drive viral evolution to a greater extent than is seen in the peripheral blood compartment (41
). Enhanced viral evolution at an inflamed mucosal site was suggested by Panther et al., who showed greater env
heterogeneity in the female genital tract compared to paired blood samples (33
). A number of other studies examining lymphoid and nonlymphoid tissue compartments have also suggested compartmentalization of viral evolution (9
). Though there have been few studies of HIV-1 evolution in the intestinal mucosa, genotypic differences in the env
, and reverse transcriptase genes of HIV-1 quasispecies in the intestinal mucosa and blood have been described (35
). HIV-1 isolates from the gastrointestinal mucosa have also been shown to differ from paired blood isolates in terms of their ability to induce cytopathology in infected cells and showed a greater sensitivity to serum neutralization in one study (4
). Although we did not find significant differences in the ability of mucosa- and PBMC-derived viruses to replicate in allogeneic MMCs or PBMCs, infection of MMCs supported greater replication of each viral isolate compared to infection of PBMCs. Therefore, cellular characteristics appeared to play a more vital role than did viral replicative ability.
Biological properties such as chemokine receptor tropism might differentiate mucosa-derived viruses from those isolated from other tissues and blood. Changes in the diversity of the viral envelope gene underlie changes in cellular tropism, coreceptor usage, and immune system evasion and therefore may determine the ability to infect and spread within cells in a given anatomic compartment (11
). Compartmental differences in viral phenotype have been described in various tissues, including lymph nodes, spleen, bone marrow, kidney, liver, testes, lung, and brain (25
). Al-Mulla et al. showed that virus isolated from the gastrointestinal mucosa and blood may differ phenotypically in terms of syncytium induction (2
), although these authors did not specifically examine chemokine receptor tropism. While most studies have suggested independent tissue-specific evolution, one study did show restricted sequence variability in the HIV env
gene among different tissues that included the colonic mucosa. We found that chemokine receptor usage and tropism did not differ between viral isolates from the mucosal and PBMC compartments. This may be explained by the similar chemokine receptor expression of the cells that were infected in both compartments. In addition, in tissues, such as the gastrointestinal mucosa, where trafficking of lymphocytes is common, equalization of viral quasispecies between compartments may occur (13
The results that we present may have important therapeutic implications. While the results of our in vitro assessments may not adequately mimic or reliably predict the biological complexity found in an HIV-infected person, these findings do suggest that further study of HIV immunopathogenesis in the mucosal compartment is necessary. Given the enhanced infection of mucosal lymphoid cells by HIV infection, perhaps therapies should be directed specifically toward this important compartment. Efforts to suppress mucosal inflammation could potentially decrease recruitment of CCR5+ CD4+ lymphocytes to the mucosa. Suppression of mucosal inflammation might decrease HIV replication in HIV-infected cells. Future work should further compare the activation state of the dual chemokine receptor-expressing cells in the blood and mucosa to determine whether a higher state of cellular activation accounts for the greater HIV-1 replication in MMCs. Additional experiments could also determine whether the CCR5+ CXCR4+ PBMCs home to, or derive from, the gastrointestinal mucosa.