Background:  HIV-1 infects cells by binding of the viral Env gp120 to the primary receptor CD4, followed by engagement of the co-receptors CCR5 (R5-tropic) or CXCR4 (X4-tropic). Cells may also be exposed to virus or Env protein in the absence of infection. It has been demonstrated that virion or gp120 binding to macrophages results in the secretion of pro-inflammatory cytokines (MIP-1b, MCP-1, and TNF-a) that are believed to play a role in AIDS pathogenesis. It has also been shown that virion or gp120 activates several protein tyrosine kinases in macrophages via the chemokine receptors, including Pyk2 and members of the MAP kinase family. However, the relationship between kinase activation and mediator secretion is not well defined.

Methods:  Here we have examined the signaling and secretory responses evoked in human monocyte-derived macrophages (MDM) following stimulation of CCR5. Engagement of CCR5 by both recombinant gp120 from the R5-tropic JRFL isolate and the natural ligand MIP-1b resulted in the rapid and transient phosphorylation of the MAP kinases ERK-1/2 and p38, as well as secretion of TNF-a.

Results:  TNF-a secretion in response to R5 gp120 was blocked by the CCR5 antagonist M657. Secretion of TNF-a in response to CCR5 stimulation was also completely ablated by specific inhibitors of p38 (SB202190, PD169316) and ERK-1/2 (PD98059, UO126). To investigate signals upstream of the MAP kinases, we discovered that pretreatment of cells with 2 PI-3 kinase inhibitors (Wortmannin and LY294002) abolished both MAP kinase phosphorylation and subsequent TNF-a secretion in response to gp120.

Conclusions:  Thus, our data suggest that TNF-a secretion in human monocyte-derived macrophages following gp120 exposure is mediated by the CCR5 receptor, is dependent on the activation of the MAP kinases p38 and ERK-1/2, and that their activation occurs through a PI3 kinase-dependent pathway. Our findings provide insight into the initial signaling events that occur on HIV-1 binding in macrophages that may contribute to macrophage dysfunction in AIDS, including processes such as AIDS dementia and inappropriate activation where TNF-a secretion plays an important role.

Keywords: MAP kinase; PI3 kinase; TNF-alpha