Immunomodulatory Factors Galectin-9 and Interferon-Gamma Synergize to Induce Expression of Rate-Limiting Enzymes of the Kynurenine Pathway in the Mouse Hippocampus

Abstract

Elevated levels of circulating pro-inflammatory cytokines are associated with symptomology of several psychiatric disorders, notably major depressive disorder. Symptomology has been linked to inflammation/cytokine-dependent induction of the Kynurenine Pathway. Galectins, like pro-inflammatory cytokines, play a role in neuroinflammation and the pathogenesis of several neurological disorders but without a clearly defined mechanism of action. Their involvement in the Kynurenine Pathway has not been investigated. Thus, we searched for a link between galectins and the Kynurenine Pathway using in vivo and ex vivo models. Mice were administered LPS and pI:C to determine if galectins (Gal’s) were upregulated in the brain following in vivo inflammatory challenges. We then used organotypic hippocampal slice cultures (OHSCs) to determine if Gal’s, alone or with inflammatory mediators [interferon-gamma (IFNγ), tumor necrosis factor-alpha (TNFα), interleukin-1beta (IL-1β), polyinosine-polycytidylic acid (pI:C), and dexamethasone (Dex; synthetic glucocorticoid)], would increase expression of indoleamine/tryptophan-2,3-dioxygenases (DO’s: Ido1, Ido2, and Tdo2; Kynurenine Pathway rate-limiting enzymes). In vivo, hippocampal expression of cytokines (IL-1β, TNFα, and IFNγ), Gal-3, and Gal-9 along with Ido1 and Ido2 were increased by LPS and pI:C (bacterial and viral mimetics). Of the cytokines induced in vivo, only IFNγ increased expression of two Ido1 transcripts (Ido1-FL and Ido1-v1) by OHSCs. Although ineffective alone, Gal-9 accentuated IFNγ-induced expression of only Ido1-FL. Similarly, IFNγ induced expression of several Ido2 transcripts (Ido2-v1, Ido2-v3, Ido2-v4, Ido2-v5, and Ido2-v6). Gal-9 accentuated IFNγ-induced expression of only Ido2-v1. Surprisingly, Gal-9 alone, slightly but significantly, induced expression of Tdo2 (Tdo2-v1 and Tdo2-v2, but not Tdo2-FL). These effects were specific to Gal-9 as Gal-1 and Gal-3 did not alter DO expression. These results are the first to show that brain Gal-9 is increased during LPS- and pI:C-induced neuroinflammation. Increased expression of Gal-9 may be critical for neuroinflammation-dependent induction of DO expression, either acting alone (Tdo2-v1 and Tdo2-v2) or to enhance IFNγ activity (Ido1-FL and Ido2-v1). Although these novel actions of Gal-9 are described for hippocampus, they have the potential to operate as DO-dependent immunomodulatory processes outside the brain. With the expanding implications of Kynurenine Pathway activation across multiple immune and psychiatric disorders, this synergy provides a new target for therapeutic development.

Desipramine decreases expression of human and murine indoleamine-2,3-dioxygenases

“…Interestingly, Ido1 induction was blocked by Desip in the circulation (murine PBMCs, PBMC −T s, T cells and human PBMCs) and the brain (murine hippocampi, microglia and astrocytes). Using organotypic hippocampal slice cultures (OHSCs), we found that IFNγ-induced Ido1 and Ido2 expression was blocked by Desip, and furthermore, that our previously described synergy between IFNγ and either Dex (Brooks et al, 2016b) or galectin-9 (Brooks et al, 2016a) to accentuate specific Ido1 and Ido2 transcripts were blocked by Desip.…”

“…C57BL/6J pups were used to generate OHSCs as previously described (Brooks et al, 2016a; Brooks et al, 2016b). Hippocampi were sliced and placed onto membrane inserts in 6-well plates containing medium (50% Eagle’s MEM, 25% HBSS, 25% heat-inactivated horse serum, penicillin/streptomycin, HEPES, 0.5% D-glucose).…”

“…Such pre-clinical models of depression are associated with increased DO expression and/or DO activity, primarily attributed to Ido1 as diminishing DO activity by the administration of Ido1 inhibitors or using Ido1 knockout (KO) mice results in decreased inflammation- and stress-induced depression-like behaviors (Lawson et al, 2013; Liu et al, 2015; O’Connor et al, 2009b; Salazar et al, 2012). Peripheral immune challenges, largely via IFNγ, induce Ido1 and Ido2 (Brooks et al, 2016a; Brooks et al, 2016b; Browne et al, 2012; O’Connor et al, 2009a); whereas, stress hormones induce Tdo2 (Brooks et al, 2016b) in rodent models. Our recent work suggests that stress-inducible factors, glucocorticoids, synergize with IFNγ to further induce the DOs (Brooks et al, 2016a; Brooks et al, 2016b).…”

“…Peripheral immune challenges, largely via IFNγ, induce Ido1 and Ido2 (Brooks et al, 2016a; Brooks et al, 2016b; Browne et al, 2012; O’Connor et al, 2009a); whereas, stress hormones induce Tdo2 (Brooks et al, 2016b) in rodent models. Our recent work suggests that stress-inducible factors, glucocorticoids, synergize with IFNγ to further induce the DOs (Brooks et al, 2016a; Brooks et al, 2016b). Thus, the induction of the Kynurenine Pathway following inflammatory and stress-induced stimulation is intimately associated with depression (Capuron et al, 2002; Dantzer and Kelley, 2007; Dantzer et al, 2011; Gibney et al, 2013; Hoyo-Becerra et al, 2014; O’Connor et al, 2009a).…”

“…Similarly, Ido2-FL (comprised of exons 1,2 ,4,5,6,7,8,9,10,11) is poorly expressed and poorly induced by inflammatory mediators or stress in the mouse brain (Brooks et al, 2016b). Ido2-v1 (comprised of exons 3 ,4,5,6,7,8,9,10,11) and Ido2-v3 (exon structure unknown) are well expressed within the brain and are increased following inflammatory stimuli (Brooks et al, 2016a). Similar to Ido1, differential relative expression and regulation suggest the use of distinct enhancer/promoter sequences to control Ido2 expression(Ball et al, 2009).…”