The mucosa is the first barrier against many pathogens, including HIV-1. Furthermore, GALT is the largest lymphoid organ in the body (25
) and is a major reservoir of HIV-1 replication during chronic infection (32
), due to constitutive activation of gut lymphocytes (4
). Results from studies of infected humans and nonhuman primates indicate an important role for mucosal immune responses, particularly mucosal CTL, in the prevention of HIV-1 and SIV infection (12
). However, the technical barrier of obtaining sufficient mucosal lymphocytes has limited the study of these responses in HIV-1-infected persons. Our previous work has shown that 20 sigmoid colon biopsies yield between 1 million and 3 million total CD3+
T lymphocytes (33
), a quantity insufficient to perform comprehensive mapping of HIV-1-specific CD8+
T-lymphocyte responses even with cell-sparing assays such as ELISpot.
Here we apply recently developed methods (33
) to expand mucosal CD8+
T lymphocytes to evaluate the HIV-1-specific CTL responses of several chronically infected individuals. In agreement with a prior study (16
), we found that there is a good correlation between CTL responses detected in fresh CD8+
PBMC and expanded CD8+
PBMC with this expansion protocol, suggesting that bias in PBMC expansion is minor. We therefore applied this method in parallel to MMC to allow comparisons of the blood and mucosal compartments. While we cannot entirely exclude that this nonspecific expansion induced bias in the MMC, the consistency of detected CTL responses between independent visits in both expanded PBMC and expanded MMC provides evidence that any bias is consistent. In addition, targeting of HIV-1 by CTL in our subjects is similar to that previously described in other studies of blood (1
) for both blood and mucosa. A further caveat is the use of clade B consensus sequence peptides for CTL detection and not autologous sequences. We cannot exclude that these peptides preferentially detect shared responses in both compartments while other CTL specific for autologous sequences differ between compartments; however, such compartment-specific bias would seem unlikely. Thus, overall, the high degree of qualitative and quantitative similarity between the blood and mucosa suggest that HIV-1-specific lymphocytes may traffic freely between these compartments.
Despite the observed similarity, it remains to be determined whether the interactions between HIV-1 and CTL reflected in the peripheral blood are identical to those in the gut. Studies addressing the effects of CTL on HIV-1 sequence have documented viral escape in the blood (10
), but the extent to which this interaction reflects that in the gut, the major reservoir of viral replication (32
), remains undefined. It is known that replication in GALT can persist even when virus is undetectable in blood during chronic infection (35
), indicating that the compartments are not always equivalent. While this probably reflects that the virus in blood is spilled over from inadequate containment in tissues, it remains to be determined whether immune pressure on HIV-1 in GALT and other tissues is reflected in blood.
Further supporting the concept that peripheral blood HIV-1 reflects spillover from tissues, studies of acute SIV infection in macaques have shown that there is a period of early local mucosal replication preceding systemic dissemination (6
), which is not observable in blood until days after initial exposure (42
). There is also strong evidence for asymmetrical trafficking of CTL from the mucosa to the peripheral blood with little traffic from the blood to the mucosa (reviewed in reference 6
). These observations suggest that efficient generation of HIV-1-specific CTL responses in both blood and mucosa may require antigenic stimulation within the GALT.
This principle may have pivotal implications in HIV-1 vaccine design. The SIV macaque model suggests that local mucosal CTL responses are important for protective immunity, and their absence correlates with uncontrolled infection after mucosal challenge (12
). Peripheral immunization induces virus-specific CTL in blood (6
) but does not protect against mucosal transmission (5
). In humans, intramuscular delivery of canarypox-based vectors induces HIV-1-specific immunoglobulin G in serum, while mucosal delivery is inefficient for inducing this serum response (27
). However, intrarectal but not specific intramuscular vaccination is required to provoke HIV-1-specific immunoglobulin A in the gut (39
). These data indicate the complexity of immunity elicited by vaccines and underscore the importance of understanding the role of various immune responses in protection from infection and/or disease by HIV-1.
In summary, this study provides a comprehensive analysis of the HIV-1-specific CTL responses in GALT, finding that they are mirrored by responses in the peripheral blood during chronic infection. The pivotal role of this compartment as a portal of entry in acute infection and reservoir for replication in chronic infection underscores the importance of understanding the relationship of immune responses in peripheral blood to those in gut mucosa. Elucidating such mechanisms may be important to efforts to optimize vaccine efficacy by eliciting HIV-1-specific immunity in the GALT.