My interest on b-chemokines stems from our observation that characterized the chemokines RANTES, MIP-1a, and MIP-1b as inhibitors of HIV strains that preferentially infect macrophages; that observation was later explained by the discovery of the role of chemokine receptors as co-receptors for HIV infection. In particular CCR5, the receptor for RANTES, MIP-1a, and MIP-1b, plays a crucial role in primary transmission of HIV-1.

My studies on chemokines are aimed in two main directions a) understanding their role in immune responses b) evaluating their role in vivo in the course of HIV infection. These roles are likely to be inter-related, since CCR5 ligands are released in the course of memory immune responses and are one of the most sensitive and reproducible markers or response to recall antigens.

The potential importance of the production of CCR5 ligands upon antigen activation was highlighted in a previous publication, where we studied the production of these chemokines in subjects enrolled in the Baltimore site of the Multicenter AIDS Cohort Study (MACS). In these subjects, we observed a significant decrease of antigen-induced MIP-1a and MIP-1b production in subjects with AIDS symptoms, as compared to both HIV+ asymptomatic individuals and to seronegative controls. In addition, we also observed that cells from seronegative subjects from the same cohort, who had a history of unprotected sexual exposure to HIV infection but did not get infected, expressed high levels of CCR5 ligands, even in absence of antigen stimulation, as compared to both HIV+ subjects and to seronegative laboratory controls. These findings are consistent with the hypothesis that CCR5 ligands might protect from HIV infection and disease progression. We are interested in determining whether memory cells that express high levels of CCR5 ligands are protected from HIV infection. Therefore, we will perform a thorough phenotypic characterization by flow cytometry of antigen stimulated memory cells in the presence of HIV, measuring intracellular levels of the prototype CCR5 ligand MIP-1b production and levels of CCR5 on the cell membrane, to verify whether there is relationship between production of this chemokine (used as marker for CCR5 ligands), CCR5 expression, and production of the viral antigen p24.

More recently, our interests in host factors that limit HIV infection and spread have expanded to include a second family of proteins, i.e. b-defensins. Defensins are antimicrobial peptides expressed as part of the innate immune response. While a-defensins are expressed mostly by neutrophilic cells, b-defensins are expressed by epithelial cells. Interestingly, they are expressed at high levels in the oral mucosa, one mucosal site that is apparently pervious (compared to other mucosal surfaces) to HIV infection. Our results show that b-defensin 2 is particularly potent in inhibiting HIV replication in vitro, without toxic effects on cell metabolism. We are also learning that b-defensin 2 is likely to act through a receptor-mediated mechanism, and we are not about to embark on intracellular signaling studies. In particular, we have evidence that the antiviral effect is mediated by a cellular receptor, which we have tentatively identified. Accordingly, we intend to pursue three main research directions:

1) Study of the role of the cellular receptor in suppression of HIV mediated by hBD2 by blocking it with specific antibodies or siRNA

2) Identification of the signaling pathways downstream from the cellular receptor that influence HIV replication, using an array of compounds known to block select intracellular signaling pathways, and verify whether they revert hBD2 inhibition.

3) Identification of the process in HIV replication that is affected by b-defensins activity. In addition, thank to a collaborative study, we have started to study the expression of b-defensin 2 expression by immunohistochemistry in oral mucosa, using clinical samples from HIV+ subjects and seronegative controls. Our preliminary results show that b-defensins are expressed at high levels in the oral mucosa of seronegative subjects, in a continuous, laminar pattern. In HIV positive subjects, however, the levels of expression of b-defensin 2 is markedly decreased. This lower level of expression could be related to the occurrence of oral manifestation of HIV infection, such as oral candidiasis.