SummaryExposure of BALB/c mice to mosquitoes infected with irradiated Plasmodium berghei confers protective immunity against subsequent sporozoite challenge. Immunized mice challenged with viable sporozoites develop parasitemia when treated orally with substrate inhibitors of nitric oxide synthase (NOS). This suggests that the production of nitric oxide (NO) prevents the development of exoerythrocytic stages of malaria in liver. Liver tissue from immunized mice expressed maximal levels of mKNA for inducible NOS (iNOS) between 12 and 24 h after challenge with sporozoites. Intraperitoneal injection of neutralizing monoclonal antibody against interferon 3/(IFN-3') or in vivo depletion of CD8 + T cells, but not CD4 + T cells, at the time of challenge blocked expression of iNOS mlLNA and ablated protection in immunized mice. These results show that both CD8 § T cells and IFN-~/are important components in the regulation of iNOS in liver which contributes to the protective response of mice immunized with irradiated malaria sporozoites. IFN-3,, likely provided by malaria-specific CD8 + T cells, induces liver cells, hepatocytes and/or Kupffer cells, to produce NO for the destruction of infected hepatocytes or the parasite within these cells. W ithin minutes after an infected Anopheles mosquito bites the vertebrate host, malaria sporozoites migrate to the liver and invade hepatocytes. There, the parasite matures, and after several days the infected hepatocytes lyse, releasing thousands of merozoites. Once in circulation, the parasite infects erythrocytes causing parasitemia. Prior exposure to irradiated sporozoites confers protective immunity (1, 2). This immunity is directed against liver stage malaria, and does not protect against the blood stage malaria.CD8 + T cells and IFN-3' are required for protective immunity to sporozoite challenge. In vivo depletion of CD8 § T cells or neutralization of IFN-3~ blocks induction of effector activity at the hepatic stage, resulting in parasitaemia (3-5). In vitro studies show that IFN-qr kills parasites by stimulating malaria-infected hepatocytes to produce nitric oxide (NO), and the addition of monomethyl-r-arginine (NGMMLA), a substrate inhibitor of nitric oxide synthase (NOS), to primary cultures of mouse hepatocytes reversed the antiparasitic effects of IFN-'y (6, 7). Human hepatocytes also respond to IFN-3' for NO production (8). As to whether human hepatocytes exhibit antimalaria activity when stimulated to produce NO, remains to be examined.At present, the antimalaria effector mechanism triggered by sporozoites in immunized animals is not fully understood. Presumably malaria-specific CD8 + T cells act directly against infected hepatocytes by recognizing malaria antigen on the cell surface (i.e., induction of CTLs) or malaria-specific lymphocytes release cytokines, such as IFN-3,, upon parasite stimulation, which induces an antimalarial response (3)(4)(5)(9)(10)(11)(12). The relationship between CD8 + T cells, IFN-% and NO-mediated protection in sporozoite-immunized mice was ...

To discover how nitric oxide (NO) synthesis is controlled in different tissues as cells within these tissues combat intracellular pathogens, we examined three distinctively different experimental murine models designed for studying parasite-host interactions: macrophage killing of Leishmania major; nonspecific protection against tularemia (Francisella tularensis) by Mycobacterium bovis (BCG); and specific vaccine-induced protection against hepatic malaria with Plasmodium berghei. Each model parasite and host system provides information on the source and role of NO during infection and the factors that induce or inhibit its production. The in vitro assay for macrophage antimicrobial activity against L. major identified cytokines involved in regulating NO-mediated killing of this intracellular protozoan. L. major induced the production of two competing cytokines in infected macrophages: (1) the parasite activated the gene for tumor necrosis factor (TNF), and production of TNF protein was enhanced by the presence of interferon-gamma (IFN-gamma). TNF then acted as a autocrine signal to amplify IFN-gamma-induced production of NO; and (2) the parasite upregulated production of transforming growth factor-beta (TGF-beta), which blocked IFN-gamma-induced production of NO. Whether parasite-induced TNF (parasite destruction) or TGF-beta (parasite survival) prevailed depended upon the presence and quantity of IFN-gamma at the time of infection. The relationship between NO production in vivo and host resistance to infection was demonstrated with M. bovis (BCG).(ABSTRACT TRUNCATED AT 250 WORDS)

The live vaccine strain (LVS) ofFrancisella tularensis caused lethal disease in several mouse strains. Lethality depended upon the dose and route of inoculation. The lethal dose for 50% of the mice (LD50) in four of six mouse strains (A/J, BALB/cHSD, C3H/HeNHSD, and SWR/J) given an intraperitoneal (i.p.) inoculation was less than 10 CFU. For the other two strains tested, C3H/HeJ and C57BL/6J, the i.p. log LD5* was 1.5 and 2.7, respectively. Similar susceptibility was observed in mice inoculated by intravenous (i.v.) and intranasal (i.n.) routes: in all cases the LD50 was less than 1,000 CFU. Regardless of the inoculation route (i.p., i.v., or i.n.), bacteria were isolated from spleen, liver, and lungs within 3 days of introduction of bacteria; numbers of bacteria increased in these infected organs over 5 days. In contrast to the other routes of inoculation, mice injected with LVS intradermally (i.d.) survived infection: the LD50 of LVS by this route was much greater than 105 CFU. This difference in susceptibility was not due solely to local effects at the dermal site of inoculation, since bacteria were isolated from the spleen, liver, and lungs within 3 days by this route as well. The i.d.-infected mice were immune to an otherwise lethal i.p. challenge with as many as 104 CFU, and immunity could be transferred with either serum, whole spleen cells, or nonadherent spleen cells (but not Ig+ cells). A variety of infectious agents induce different disease syndromes depending on the route of entry. Francisella LVS infection in mice provides a model system for analysis of locally induced protective effector mechanisms.

C3H/HeN mice that are naturally resistant to cutaneous disease and systemic infections with the protozoan parasite, Leishmania major, were treated at the time of infection, and weekly thereafter, with mouse anti-rat IFN-gamma mAb or an irrelevant antibody of similar isotype. Anti-IFN-gamma-treated mice developed cutaneous lesions; parasites spread to the regional lymph nodes and then metastasized to spleens and livers. The course of disease in these animals was similar to that of genetically susceptible BALB/c mice. Two exceptions in the pathology of L. major infections were noted between BALB/c and anti-IFN-gamma-treated C3H/HeN mice: 1) BALB/c mice died of systemic complications, whereas C3H/HeN mice did not, and 2) multinucleated giant cells were observed in lymph nodes and spleens of infected BALB/c mice, whereas these cells were not observed in infected C3H/HeN mice. Control mice, those treated with either saline or irrelevant antibody of the same isotype as the anti-IFN-gamma monoclonal, showed no evidence of cutaneous disease (development of footpad lesions) or systemic infection (by histopathology). Total abrogation of the natural resistance of C3H/HeN mice could be achieved by treatment with as little as 0.5 mg/mouse/wk of anti-IFN-gamma antibody, or by a single dose of 1 mg/mouse anti-IFN-gamma antibody administered at the time of parasite inoculation. If antibody treatment was delayed for as little as 1 wk after parasite inoculation, the infections in treated animals resembled that of untreated or control antibody-treated mice: no cutaneous lesions (by footpad swelling or viable counts of leishmania in footpad tissue) or systemic disease (by microscopic analysis of touch preparations of internal organs, and histopathology of same). The production of IFN-gamma during the initial interaction of the parasite and host cells appears to be a major component of genetic control of natural resistance to infection with L. major in C3H/HeN mice.