Background:  Perivascular macrophages/microglia are considered to be the primary source and reservoir of productive HIV brain infection. The reasons for the persistence of HIV infection in such cells are unclear. Quinolinic acid is a neurotoxic product of the kynurenine pathway (KP) and is related to HIV brain infection. Activation of the KP leads to immune tolerance:  depletion of tryptophan by indoleamine-2, 3-dioxygenase the first enzyme of the KP and by KP products, including kynurenine and quinolinic acid. We hypothesized that KP activation would be important in HIV persistence in the brain. However, it is unknown whether the cells associated with perivascular macrophages/microglia have functional enzymes of the KP present to be able to lead to an immune tolerant microenvironment. The KP expression is only known in macrophages/microglia and astrocytes. We therefore sought to map KP expression in human primary-cultured blood–brain barrier (BBB) endothelial cells and pericytes.

Methods:  Cells were stimulated for 24 hours with vehicle alone, human cytokines interferon-gamma, tumour necrosis factor-alpha or both. To assess downstream KP catabolism at the BBB we used human brain microvascular endothelial cells and human pericytes. We compared these to endothelial cells from human umbilical vein and from human dermal microvessels. Cells were stimulated as above in the presence of the KP intermediates 3-hydroxyanthranilic acid or quinolinic acid. KP metabolites in culture supernatants were quantitated by high performance liquid chromatography (HPLC) and gas chromatography-mass spectroscopy (GC-MS). Real time polymerase chain reaction (RT-PCR) was performed on cDNA reverse-transcribed from total cell RNA using primer sets for 5 KP enzymes (indoleamine-2, 3-dioxygenase, kynurenine 3-monooxygenase, L-kynurenine hydrolase, 3-hydroxyanthranilate 3,4-dioxygenase, and quinolinate phosphoribosyltransferase).

Results:  By RT-PCR and HPLC/GC-MS, BBB endothelial cells have indoleamine-2, 3-dioxygenase and partial KP expression. They synthesise kynurenic acid constitutively, and kynurenine after immune activation. Pericytes also have indoleamine-2, 3-dioxygenase and partial KP expression. They produce small amounts of picolinic acid and, after immune activation, kynurenine. Human umbilical vein endothelial cells express only low levels of kynurenic acid after immune activation. Human dermal microvascular endothelial cell KP expression resembles that in BBB endothelial cells.

Conclusions:  Immune activation of the cells associated with perivascular macrophages/microglia at the BBB leads to unique and significant KP activation, resulting in immune tolerance. This is likely an important contributor to persistence of HIV in the brain.