Joseph M. Carlin

Synergistic Transcriptional Activation of Indoleamine Dioxygenase by IFN-γ and Tumor Necrosis Factor-α

Interferon-γ (IFN-γ)-induced indoleamine 2,3-dioxygenase (IDO) activity inhibits the growth of susceptible intracellular pathogens by catalyzing the oxidative cleavage of the indole ring of L-trypt...

Tumor Necrosis Factor-alpha and Lipopolysaccharide Enhance Interferon-Induced Antichlamydial Indoleamine Dioxygenase Activity Independently

In macrophages, interleukin-1 (IL-1) and lipopolysaccharide (LPS) enhance the antichlamydial effect of interferon-gamma (IFN-gamma) by increasing indoleamine 2,3-dioxygenase (IDO) activity in a dose-dependent manner. Our objectives were to characterize the antichlamydial effect of tumor necrosis factor-alpha (TNF-alpha) on IFN-induced IDO activity and to establish the relationship between LPS and TNF-alpha in IDO potentiation. TNF-alpha inhibited chlamydial growth in a dose-dependent manner only in IFN-treated macrophages. Furthermore, excess tryptophan reversed the effect of combined cytokine treatment, indicating that IDO alone was responsible for chlamydial inhibition. The promonocyte THP-1 cell line, previously used to model the effect of IL-1 on IDO mRNA expression, was treated with IFN-gamma and increasing concentrations of LPS or TNF-alpha. IDO mRNA was quantified by RT-PCR, and IDO activity was measured by HPLC at 24 and 48 h after treatment, respectively. Both LPS and TNF-alpha enhanced IDO activity and IDO mRNA expression, with maximal IDO induction at 100 ng/ml LPS or 5 ng/ml TNF-alpha. Anti-TNF-alpha failed to neutralize the effects of LPS treatment, and insufficient TNF-alpha or IL-1 was produced by LPS-treated THP-1 cells to account for the enhancing effect of LPS, indicating that the effect of LPS on IDO was independent of TNF-alpha and IL-1.

Upregulation of interferon-induced indoleamine 2,3-dioxygenase in human macrophage cultures by lipopolysaccharide, muramyl tripeptide, and interleukin-1

The tryptophan decyclizing enzyme indoleamine 2,3-dioxygenase (IDO) was induced in human monocyte-derived macrophages (MDM) treated with human recombinant interferon-beta (IFN-beta) or interferon-gamma (IFN-gamma). Treated cells exhibited dose-dependent increases in IDO when assayed 48 hr after treatment. Cells exposed to IFN-gamma were observed to exhibit consistently higher peak levels of IDO when compared with cells incubated in the presence of IFN-beta. When IFN-beta-treated cells were incubated in the presence of specified amounts of bacterial lipopolysaccharide (LPS) or liposome-encapsulated muramyl tripeptide (MTP), peak IDO activity increased such that enzyme activity was comparable to maximal activity observed with IFN-gamma-treated cells. LPS and MTP also upregulated IFN-gamma-mediated IDO activity when suboptimal amounts of IFN-gamma were used. When macrophages were costimulated with various concentrations of human recombinant interleukin 1 alpha (IL-1 alpha), along with either maximum-stimulating amounts of IFN-beta or suboptimal amounts of IFN-gamma, IDO activity was upregulated in a manner similar to results obtained using the microbial products as stimuli. While neither IL-1 alpha or IL-1 beta was detected in culture supernatants from macrophages treated with either LPS or MTP (alone or in combination with IFN), IL-1 alpha was detected in cell lysates of macrophages treated with these upregulators. Although neutralizing antibody to IL-1 alpha abolished the upregulatory effect of exogenous IL-1 alpha, it had no effect on upregulation by LPS or MTP. This suggests that although LPS and MTP may induce production of cell-associated IL-1 alpha, upregulation of IDO activity by these agents is independent of IL-1 alpha production and may be mediated through distinct pathways.

Up-Regulation of Gamma Interferon Receptor Expression Due to Chlamydia-Toll-Like Receptor Interaction Does Not Enhance Signal Transducer and Activator of Transcription 1 Signaling

ABSTRACT Gamma interferon (IFN-γ)-induced indoleamine dioxygenase (IDO), which inhibits chlamydial replication by reducing the availability of tryptophan, is up-regulated by interleukin-1β (IL-1β) and tumor necrosis factor alpha (TNF-α). The mechanisms by which this occurs include an increase in the synthesis of interferon regulatory factor-1 as well as a nuclear factor-κB (NF-κB)-dependent increase in the expression of IFN-γ receptors (IFN-γR). Although Chlamydia is susceptible to IDO, it up-regulates IFN-γR expression to a greater degree than either IL-1β or TNF-α, perhaps through interaction with Toll-like receptors (TLR). The purpose of this study was to determine the mechanism by which Chlamydia psittaci up-regulates IFN-γR expression and evaluate this effect on IDO induction. Infection of HEK 293 cells with C. psittaci increased IFN-γR expression only in cells expressing either TLR2 or TLR4 and the adaptor protein MD-2. In addition, up-regulation of IFN-γR expression in Chlamydia-infected HeLa cells could be blocked either by neutralizing TLRs with anti-TLR2 and/or anti-TLR4 or by inhibiting NF-κB transactivation with a proteasome inhibitor. Although the newly expressed IFN-γR in Chlamydia-infected cells were capable of binding IFN-γ, they did not enhance IFN-γ-induced IDO activity in a manner similar to those observed for IL-1β and TNF-α. Instead, IDO activation in Chlamydia-infected cells was no different than that induced in uninfected cells, despite the increase in IFN-γR expression. Furthermore, the amount of IFN-γ-induced signal transducer and activator of transcription 1 (STAT-1) activation in infected cells paralleled that observed in uninfected cells, suggesting that STAT-1 activation by these newly expressed receptors was impaired.

Chlamydiae modulate gamma interferon, interleukin-1β, and tumor necrosis factor alpha receptor expression in HeLa cells

ABSTRACT Chlamydia psittaci was found to modulate receptor expression for the cytokine receptors that are involved in the synergistic induction of indoleamine dioxygenase in epithelial cells. Increases in receptor expression were seen even with inactivated Chlamydia, suggesting that chlamydial antigens and not products of infection are important for up-regulating cytokine receptor expression.