Search

The mechanism of killing of obligately intracellular Rickettsia conorii within human target cells, mainly endothelium and, to a lesser extent, macrophages and hepatocytes, has not been determined. It has been a controversial issue as to whether or not human cells produce nitric oxide. AKN-1 cells (human hepatocytes) stimulated by gamma interferon, tumor necrosis factor alpha, interleukin 1beta, and RANTES (regulated by activation, normal T-cell-expressed and -secreted chemokine) killed intracellular rickettsiae by a nitric oxide-dependent mechanism. Human umbilical vein endothelial cells (HUVECs), when stimulated with the same concentrations of cytokines and RANTES, differed in their capacity to kill rickettsiae by a nitric oxide-dependent mechanism and in the quantity of nitric oxide synthesized. Hydrogen peroxide-dependent intracellular killing of R. conorii was demonstrated in HUVECs, THP-1 cells (human macrophages), and human peripheral blood monocytes activated with the cytokines. Rickettsial killing in the human macrophage cell line was also mediated by a limitation of the availability of tryptophan in association with the expression of the tryptophan-degrading enzyme indoleamine-2,3-dioxygenase. The rates of survival of all of the cell types investigated under the conditions of activation and infection in these experiments indicated that death of the host cells was not the explanation for the control of rickettsial infection. This finding represents the first demonstration that activated human hepatocytes and, in some cases, endothelium can kill intracellular pathogens via nitric oxide and that RANTES plays a role in immunity to rickettsiae. Human cells are capable of controlling rickettsial infections intracellularly, the most relevant location in these infections, by one or a combination of three mechanisms involving nitric oxide synthesis, hydrogen peroxide production, and tryptophan degradation.

Infection with Chlamydia pneumoniae, a human respiratory pathogen, has been implicated as a potential risk factor in atherosclerosis, possibly because the pathogen can exist in a persistent form similar to that described for Chlamydia trachomatis. The present study investigated whether gamma interferon (IFN-gamma) can induce indoleamine 2,3-dioxygenase (IDO) activity in aortic smooth muscle cells, leading to a marked inhibition of C. pneumoniae growth. Our data indicate a stimulation of IDO mRNA expression and dose-dependent enzymatic activity following IFN-gamma treatment. IDO-mediated increase in tryptophan catabolism resulted in a dose-dependent marked inhibition of C. pneumoniae replication.

The most prominent gamma interferon (IFN-gamma)-induced antimicrobial effector mechanisms are the induction of nitric oxide (NO) synthase (NOS) and of indoleamine 2,3-dioxygenase (IDO) activity. We have recently found that human glioblastoma cells and human macrophages inhibit the growth of group B streptococci after stimulation with IFN-gamma. In this report, we show that in addition, human RT4 (uroepithelial) cells can inhibit the growth of enterococci. Murine macrophages (RAW cells) are unable to inhibit bacterial growth after IFN-gamma stimulation. Stimulation of human glioblastoma cells, macrophages, and RT4 cells with human IFN-gamma results in a strong expression of IDO activity; however, NO production remains undetectable. In strong contrast, murine RAW cells produce large amounts of NO when stimulated with murine IFN-gamma and IDO activity is not detectable. Interleukin-1 (IL-1) induces NO synthase in human RT4 cells when the cells are costimulated with IFN-gamma. We found that IL-1 inhibits IFN-gamma-stimulated IDO activity and antimicrobial effects in RT4 cells, while in human glioblastoma cells, which lack detectable NO synthase activity, neither of these effects was altered by costimulation with IFN-gamma and IL-1. The IL-1-mediated inhibition of IDO activity and of subsequent antibacterial effect is due to the production of NO. This conclusion was supported by evidence that N(G)-monomethyl-L-arginine, a competitive inhibitor of inducible NOS activity, is able to block the inhibitory action of IL-1 on IFN-gamma-induced bacteriostasis. We therefore conclude that NO production does not inhibit the growth of enterococci but might be involved in the regulation of IDO activity in some human cells.

Studies were carried out to evaluate the proposed role of indoleamine 2,3-dioxygenase (INDO) induction in the antimicrobial and antiproliferative effects of gamma interferon (IFN-gamma) in human fibroblasts. The INDO cDNA coding region was cloned in the pMEP4 expression vector, containing the metallothionein (MTII) promoter in the sense (+ve) or the antisense (-ve) orientation. Human fibroblasts (GM637) stably transfected with the sense construct expressed INDO activity after treatment with CdCl2 or ZnSO4, but cells transfected with the antisense construct did not. The growth of Chlamydia psittaci was strongly inhibited in INDO +ve cells but not in INDO -ve cells after treatment with Cd2+ or Zn2+. The inhibition correlated with the level of INDO activity induced and could be reversed by the addition of excess tryptophan to the medium. The growth of Toxoplasma gondii was also strongly inhibited in INDO +ve cells but not in INDO -ve cells after treatment with Cd2+. Expression of Cd(2+)-induced INDO activity also inhibited thymidine incorporation and led to cytotoxicity in INDO +ve cells but not in INDO -ve cells. Thus, the induction of INDO activity by IFN-gamma may be an important factor in the antimicrobial and antiproliferative effects of IFN-gamma in human fibroblasts.