Ph.D. Projects


 M1/M2 macrophage polarization and human immunodeficiency virus (HIV) infection

Background and rationale
Although mononuclear phagocytes are an important target of HIV infection and replication the potential role of cytokine-directed polarization into either pro-inflammatory (M1) or anti-inflammatory (M2) macrophages has not been thoroughly investigated in the context of this viral disease. We have already defined the fundamental features of human M1 and M2 macrophages by exposing conventional monocyte-derived macrophages (MDM) to either tumor necrosis factor-a plus interferon-g (M1) or interleukin-4 (IL-4; M2) in terms of differential expression and modulation of cell surface determinants as well as of secretion of chemokines and cytokines that may play a relevant role in HIV infection. Of interest, both M1 and M2 polarization resulted in a transient decrease of CCR5-dependent (R5) HIV-1 replication in comparison to control unpolarized autologous MDM, an inhibitory pattern supported, at least in part, by a transient downregulation of CD4, the primary viral receptor for entry, in contrast to the viral co-receptor CCR5 that is not decreased in M1-MDM and it is even upregulated under M2-polarizing conditions at the time of infection. Kinetic analyses indicated that M1 polarization induces a stronger but less durable inhibitory effect on HIV-1 replication compared to M2 conditioning. M1-MDM show an increased secretion of the CCR5-binding chemokine CCL3 as well as of CXCL10 and IL-6. Conversely, M2-MDM are characterized by a modest up-regulation of the HIV inhibitory chemokine CCL22 and of IL-10. Of note, M1 polarization induces a delayed downregulation of M2-related chemokine and cytokines and vice versa. These preliminary findings suggest that polarization of tissue macrophages in either M1 or M2 cells in response to cytokines or bacterial products may favor a state of latent over productive infection and, therefore, impair the capacity of the adaptive T cells response to eliminate macrophage viral reservoirs.

Specific goals
While the pro-inflammatory (M1) pathway of differentiation and activation of macrophages is well characterized, there are different modalities to induce M2 polarization in addition to IL-4, including cell stimulation by IL-10 and Toll-like receptor ligands whereas, in addition, deactivating factors such as transforming growth factor-b can also profoundly affect macrophage function. The regulatory effect of these cytokines on HIV infection and replication has been previously investigated, but not in the context of macrophage polarization. Furthermore, the possibility to exploit cell lines, such as promonocytic U937 cells, as inducible models of M1/M2 polarization will be part of the project. In addition to learn how to perform experiments under biosafety level-3 (BSL-3) conditions and the basic fundamentals to study live HIV infections, this project will provide expertise on the immunology of human macrophages, and on molecular assays related to characterize viral transcription, protein expression, and living cell imaging.

Key references
E. Cassol, M. Alfano, P. Biswas, & G. Poli. Monocyte-derived macrophages and myeloid cell lines as targets of HIV-1 replication and persistence.  J. Leuk. Biol. 80:1018-1030, 2006.

E. Cassol, C. Rizzi, L. Cassetta, M. Alfano, & G. Poli. M1 and M2a polarization of human monocyte-derived macrophages inhibits HIV-1 replication by distinct mechanisms. J. Immunol., , 182: 6237-6246, 2009.