Cytokine and Inducible Nitric Oxide Synthase mRNA Expression during Experimental Murine Cryptococcal Meningoencephalitis

Abstract

The immune events that take place in the central nervous system (CNS) during cryptococcal infection are incompletely understood. We used competitive reverse transcription-PCR to delineate the time course of the local expression of mRNAs encoding a variety of cytokines and inducible nitric oxide synthase (iNOS) during progressive murine cryptococcal meningoencephalitis and assessed the CNS inflammatory response using immunohistochemistry. Interleukin 18 (IL-18), transforming growth factor ␤1, and IL-12p 40 mRNAs were constitutively expressed in the brains of infected and uninfected mice; IL-2 mRNA was not detected at any time. Increased levels of transcripts corresponding to IL-1␣, tumor necrosis factor alpha (TNF-␣), and iNOS were detected as early as day 1 postinfection, with TNF-␣ rising by ϳ30-fold and iNOS increasing by ϳ5-fold by day 7. Each remained at these levels thereafter. IL-4, IL-6, and gamma interferon transcripts were detected on day 5, and IL-1␤ and IL-10 transcripts were detected beginning on day 7. Once detected, each remained at a relatively constant level through 28 days of infection. This cytokine profile does not suggest a polarized Th1 or Th2 response. Immunohistochemistry did not reveal inflammatory infiltrates before day 7, despite the presence of cryptococci. Intraparenchymal abscesses with inflammatory cells in their peripheries were found beginning on day 10. The infiltrates were comprised primarily of cells expressing CD4, CD8, or CD11b; low numbers of cells expressing CD45R/B220 were also present. The persistence of Cryptococcus observed in the CNS may result from an ineffective immune response, perhaps owing to an insufficient anticryptococcal effector function of endogenous glial cells resulting from competing pro-and anti-inflammatory cytokines. These data detail the immune response in the brain and could be important for the future design of specific immunomodulatory therapies for this important opportunistic infection.

Enhancement by Tumor Necrosis Factor Alpha of Dengue Virus-Induced Endothelial Cell Production of Reactive Nitrogen and Oxygen Species Is Key to Hemorrhage Development

“…The PCR products were subjected to electrophoresis on a 2% agarose gel. The sequences of the iNOS primer set were 5Ј-TGGGAATGGAGA CTGTCCCAG-3Ј and 5Ј-GGGATCTGAATGTGATGTTTG-3Ј (44), and those for TNF-␣ primers were 5Ј-ATCCGCGACCTCGCCCTG-3Ј and 5Ј-ACCGCCTG GAGTTCTGGAA-3Ј. The sequence for the hypoxanthine phosphoribosyltransferase (HPRT) primer set was 5Ј-GTTGGATACAGGCCAGACTTTGTTG-3Ј and 5Ј-GAGGGTAGGCTGGCCTATGGCT-3Ј (9).…”

Chemokine Receptor 2-Mediated Accumulation of Fungicidal Exudate Macrophages in Mice That Clear Cryptococcal Lung Infection

“…The results of our kinetic analysis suggest that ExMs are the predominant macrophage source of these effector molecules because both iNOS and TNF-␣ are significantly diminished in CCR2-deficient mice in which ExMs are scarce. Previous investigations 12,18,23,[62][63][64][65][66][67] have defined iNOS and TNF-␣ expression as strong predictors of cryptococcal killing in vitro and clearance of the organism in vivo, although other effector mechanisms may also contribute to ExM fungicidal activity. Additional studies are also required to understand whether and how these cells interact with other critical leukocyte populations that we have previously identified, including DCs and IFN-␥-producing T cells.…”

“…Second, we evaluated the expression of two molecules iNOS and TNF-␣, that are critically important in mediating antifungal host defense. 12,18,23,[62][63][64][65][66][67] Whole lung mRNA expression of iNOS and TNF-␣ was similar in uninfected CCR2 ϩ/ϩ mice and CCR2 Ϫ/Ϫ mice (data not shown). However, the expression of iNOS and TNF-␣ was significantly increased in CCR2 ϩ/ϩ mice relative to CCR2 Ϫ/Ϫ mice at 14 dpi ( Figure 8A).…”

IFN‐γ‐induced IDO and WRS expression in microglia is differentially regulated by IL‐4

“…In pathologies where IL-4 or IL-13 would be present, IFN-γ induced IDO would be upregulated but WRS would be downregulated. Expression of IFN-γ and IL-4 are found simultaneously in the brain in human neurocysticercosis and mouse cryptocococcal meningoencephalitis (Maffei et al 2004;Restrepo et al 2001). IDO expressing cells are indeed present in granulomatous inflammation (Popov et al 2006).…”