Mechanism of Monocyte Neuroinvasion: Unique to HIV/SIV or Shared by Other Pathogens?
Candice Clay, UC-Davis
Biomedical and Clinical Sciences
Monocytes are thought to transport human immunodeficiency virus (HIV) and its simian counterpart SIV across the blood-brain barrier (BBB) by a poorly defined mechanism, triggering events that can lead to neurological complications and development of neuro-AIDS. Other neuroinvasive pathogens, such as cytomegalovirus (CMV) and Listeria, similarly infect monocytes, however dissemination into the brain may occur by an alternate route. We hypothesize that SIV infection triggers a unique monocyte migratory program promoting enhanced neuroinvasion and CNS inflammation, distinct from monocyte trafficking characteristics initiated byinfection withCMV or Listeria as well as Salmonella, a non-neuroinvasive pathogen. The specific objectives of the newly funded study are 1) to identify unique phenotypic and functional monocyte trafficking characteristics in SIV infection and 2) to delineate autocrine regulatory mechanism(s) in monocytes, such as production of proinflammatory chemokines, that are likely to contribute to continued monocyte brain recruitment.
To address these specific aims, we will conduct in vitro infection experiments and contrast migratory characteristics of monocytes infected with neurovirulent strains SIV/17E-Fr and SIV DeltaB670 to monocytes infected with rhesus CMV expressing enhanced green fluorescent protein (EGFP), Listeria monocytogenes wildtype strain 10403S or Salmonella typhimurium wildtype derivative IR715 (both transformed with a GFP-expressing construct).We will recapitulate the optimized conditions previously described for HIV infection to enhance susceptibility of rhesus monocytes to SIV infection in vitro. Freshly isolated monocytes will be cultured short-term in the presence of monocyte colony-stimulating factor and SIV-infected, prior to their differentiation and upregulation of macrophage marker CD71. For phenotypic and functional analysis, we established multi-color staining panels to define activation markers, chemokine receptor profiles and intracellular cytokine/chemokine production using flow cytometry. Assays of monocyte function in vitro will be complemented with characterization of infiltrating monocytes in brain tissues derived from acutely SIV-infected rhesus macaques (Clay et al, 2005).
Our anticipated results include expression of distinct chemokine receptors in rhesus monocytes infected with neurovirulent SIV strains, correlating with a neuroinvasive phenotype and with monocytes' suggested role as 'Trojan horse' for viral entry into the brain. This is supported by preliminary findings demonstrating increased CNS infiltration of fluorescein dye+ monocytes in acutely SIV-infected, but not uninfected macaques, following autologous cell transfer of fluorescein dye+ peripheral blood mononuclear cells after a 2-day in vivo migratory period. We also expect production of brain-specific chemokine signals in SIV-infected monocytes, such as fractalkine/CX3CL1 and MCP1/CCL2, that are likely to promote continued monocyte brain recruitment in vivo, potentially shared by other brain-invading pathogens.
Determining unique monocyte migratory parameters triggered upon SIV infection (versus infection with other viral or bacterial pathogens) as proposed under this new award, will help delineate mechanism(s) of monocyte neuroinvasion. This may lead to the development of novel drug targeting options and treatment strategies that inhibit aberrant monocyte neuroinfiltration during HIV infection by targeting specific monocyte subsets and their trafficking signals.