Glutamate production by HIV‐1 infected human macrophage is blocked by the inhibition of glutaminase

Abstract

Mononuclear phagocyte (macrophages and microglia) dysfunction plays a significant role in the pathogenesis of human immunodeficiency virus (HIV) associated dementia (HAD) through the production and release of soluble neurotoxic factors including glutamate. The mechanism of glutamate regulation by HIV-1 infection remains unclear. In this report, we investigated whether the enzyme glutaminase is responsible for glutamate generation by HIV-1 infected monocytederived macrophages. We tested the functionality of novel small molecule inhibitors designed to specifically block the activity of glutaminase. Glutaminase inhibitors were first characterized in a kinetic assay with crude glutaminase from rat brain revealing an uncompetitive mechanism of inhibition. The inhibitors were then tested in vitro for their ability to prevent glutamate generation by HIV-infected macrophages, their effect upon macrophage viability, and HIV infection. To validate these findings, glutaminase specific siRNA was tested for its ability to prevent glutamate increase during infection. Our results show that both glutaminase specific small molecule inhibitors and glutaminase specific siRNA were effective at preventing increases in glutamate by HIV-1 infected macrophage. These findings support glutaminase as a potential component of the HAD pathogenic process and identify a possible therapeutic avenue for the treatment of neuroinflammatory states such as HAD. HIV-1 associated dementia (HAD) is a significant consequence of HIV infection resulting in a chronic, progressive dementia. The dementia is a consequence of neuronal damage that results from prolonged inflammation in the CNS. Mononuclear phagocytes (MP) are critical to HAD pathogenesis and have been hypothesized to induce neuronal injury through the production and release of various soluble neurotoxic factors including glutamate (Giulian et al. 1993;Pulliam et al. 1994;Zink et al. 1999;Belmadani et al. 2001;Jiang et al. 2001). Glutamate mediates numerous physiological functions through activation of multiple receptors (Cutler and Dudzinski 1974;Fonnum 1984;Orrego and Villanueva 1993); however, high concentrations of extracellular glutamate induce neuronal damage (Olney 1971;McCall et al. 1979;Choi 1988;Newcomb et al. 1997). HIV-1 infected macrophages are an important cellular source of extracellular glutamate (Jiang et al. 2001), and HIV-1-infected patients have significantly higher concentrations of glutamate in plasma as compared with uninfected controls (Droge et al. 1987; Ollenschlager Received December 11, 2006; accepted February 2, 2007. Address correspondence and reprint requests to Jialin Zheng, Laboratory of Neurotoxicology, Center for Neurovirology and Neurodegenerative Disorders and Departments of Pharmacology/Experimental Neuroscience and Pathology/Microbiology, 985880 Nebraska Medical Center, Omaha, NE 68198-5880, USA. E-mail: jzheng@unmc.edu 1 These authors contributed equally to this work.

Glutamate metabolism in HIV-1 infected macrophages: Role of HIV-1 Vpr

“…17 A recent study also demonstrated that recombinant Vpr increases production of glutaminase C, a regulator of glutamate synthesis in murine macrophages. 35 Our results are in agreement with previous findings where the HIV-1 infection of human macrophages leads to increase in glutamate production mediated by the enzyme glutaminase, 17,36 . Increase in glutamate levels has been reported to be glutamine-dependent, and the glutamatemediated neurotoxicity has been shown to be augmented by the mitochondrial glutaminase release.…”

Evidence of reduced glutamate in the frontal lobe of HIV‐seropositive patients

“…A www.interscience.wiley.com/journal/nbm REDUCED BRAIN GLUTAMATE IN HIV INFECTIONS second important point arises; because the observed change in drug-naïve patients was the same as or slightly larger than in treated patients, early brain injury appears to be a characteristic of HIV infection rather than a toxic effect of the drugs used. HIV itself has recently been implicated as a mitochondrial inhibitor of the neuronal enzyme glutaminase (43,44). Glutaminase is an important constituent of the glial-neuronal Gln-Glu cycle, which accounts for up to 80% of the relevant neurotransmitter pathway.…”

Macrophage-induced neurotoxicity is mediated by glutamate and attenuated by glutaminase inhibitors and gap junction inhibitors

“…Inhibiting glutaminase decreases the production of glutamate by HIV-infected macrophages (Erdmann et al, 2007). Therefore, macrophages may produce glutamate by a mechanism similar to that of microglia.…”