HIV-1 Proteins Preferentially Activate Anti-Inflammatory M2-Type Macrophages

Abstract

HIV-1 proteins, including Tat, gp120, and Nef, activate macrophages (MΦ), which is consistent with the fact that HIV-1 infection is characterized by sustained immune activation. Meanwhile, MΦ are functionally classified into two types: proinflammatory M1-MΦ and anti-inflammatory M2-MΦ. We show that HIV-1 proteins, particularly Nef, preferentially activate M2-MΦ. Extracellular Tat, gp120, and Nef activated MAPK and NF-κB pathways in human peripheral blood monocyte-derived MΦ. However, the activation was marked in M-CSF–derived M2-MΦ but not GM-CSF–derived M1-MΦ. Nef was the most potent activator, and its signaling activation was comparable to that by TNF-α. Indeed, Nef was internalized more rapidly by M2-MΦ than by M1-MΦ. The myristoylation and proline-rich motif of Nef were responsible for the observed signaling activation. Consistent with the activation of MAPK/NF-κB pathways, Nef stimulated the production of a number of proinflammatory cytokines/chemokines by M2-MΦ. However, Nef reduced the expression of CD163 and phagocytosis, the characteristic markers of M2-MΦ, indicating that Nef drives an M2-like to M1-like phenotypic shift. Because the differentiation of most tissue MΦ depends on M-CSF and its receptor, which is the essential axis for the anti-inflammatory M2-MΦ phenotype, the current study reveals an efficient mechanism by which HIV-1 proteins, such as Nef, induce the proinflammatory MΦ.

M-CSF Inhibits Anti–HIV-1 Activity of IL-32, but They Enhance M2-like Phenotypes of Macrophages

“…The approval for this study was obtained from the Kumamoto University medical ethics committee. Macrophages were prepared in the following several methods (3,10). PBMCs obtained using PANCOLL reagent (PAN Biotech, Aidenbach, Germany) were suspended in RPMI 1640 medium, 1% FCS at a density of 1 3 10 6 cells/ml, and seeded in multiwell plates or dishes.…”

“…Western blotting was performed essentially as described previously (3,10). In brief, macrophages were lysed on ice with Nonidet P-40 lysis buffer (1% Nonidet P-40, 50 mM Tris, and 150 mM NaCl) containing protease inhibitors (1 mM EDTA, 1 mM PMSF, 1 mg/ml aprotinin, 1 mg/ml leupeptin, and 1 mg/ml pepstatin) and phosphatase inhibitors (1 mM Na 3 VO 4 and 1 mM NaF).…”

“…The relative levels of multiple cytokines and chemokines in the supernatants of macrophages were analyzed using a human cytokine array (R&D Systems) according to the manufacturer's instructions (3). In brief, macrophage culture supernatants (100 ml), which were collected after centrifugation, were added to dot blots onto which the capture Abs had been spotted in duplicate.…”

“…Because the phenotypes of polarized M1 and M2 macrophages can be reversed to some extent by various signals, the pathologies of many diseases are associated with dynamic changes in the balance between M1 and M2 macrophages (2). For instance, we recently demonstrated that Nef, a pathogenic HIV-1 protein, drives an M2-like to M1-like phenotypic shift (3), which might contribute to the sustained activation of the immune system observed during HIV-1 infections (4,5). Macrophages are also one of the major cellular targets of HIV-1; however, they are more resistant to the cytopathic effects of HIV-1 than CD4 + T cells, and hence are able to produce and accumulate replication-competent viruses for long periods, even in patients receiving highly active antiretroviral therapy (6).…”

Potential Role of the Formation of Tunneling Nanotubes in HIV-1 Spread in Macrophages

“…Human peripheral blood monocyte-derived macrophages (MDMs) were prepared as described previously (25,26). Approval for this study was obtained from the Kumamoto University medical ethics committee.…”