Molecular survival strategies of Echinococcus multilocularis in the murine host.
Larval infection with Echinococcus multilocularis starts with the intrahepatic postoncospheral development of a metacestode that-at its mature stage-consists of an inner germinal and an outer laminated layer (GL & LL). In certain cases, an appropriate host immune response may inhibit parasite proliferation. Several lines of evidence obtained in vivo and in vitro indicate the important bio-protective role of the LL. For instance, the LL has been proposed to protect the GL from nitric oxide produced by periparasitic macrophages and dendritic cells, and also to prevent immune recognition by surrounding T cells. On the other hand, the high periparasitic NO production by peritoneal exsudate cells contributes to periparasitic immunosuppression, explaining why iNOS deficienct mice exhibit a significantly lower susceptibility towards experimental infection. The intense periparasitic granulomatous infiltration indicates a strong host-parasite interaction, and the involvement of cellular immunity in control of the metacestode growth kinetics is strongly suggested by experiments carried out in T cell deficient mouse strains. Carbohydrate components of the LL, such as Em2(G11) and Em492, as well as other parasite metabolites yield immunomodulatory effects that allow the parasite to survive in the host. I.e., the IgG response to the Em2(G11)-antigen takes place independently of alpha-beta+CD4+T cells, and in the absence of interactions between CD40 and CD40 ligand. Such parasite molecules also interfere with antigen presentation and cell activation, leading to a mixed Th1/Th2-type response at the later stage of infection. Furthermore, Em492 and other (not yet published) purified parasite metabolites suppress ConA and antigen-stimulated splenocyte proliferation. Infected mouse macrophages (AE-MØ) as antigen presenting cells (APC) exhibited a reduced ability to present a conventional antigen (chicken ovalbumin, C-Ova) to specific responder lymph node T cells when compared to normal MØ. As AE-MØ fully maintain their capacity to appropriately process antigens, a failure in T cell receptor occupancy by antigen-Ia complex or/and altered co-stimulatory signals can be excluded. Studying the status of accessory molecules implicated in T cell stimulation by MØ, it could be shown that B7-1 (CD80) and B7-2 (CD86) remained unchanged, whereas CD40 was down-regulated and CD54 (=ICAM-1) slightly up-regulated. FACS analysis of peritoneal cells revealed a decrease in the percentage of CD4+ and CD8+T cells in AE-infected mice. Taken together the obstructed presenting-activity of AE-MØ appeared to trigger an unresponsiveness of T cells leading to the suppression of their clonal expansion during the chronic phase of AE infection. Interesting information on the parasite survival strategy and potential can be obtained upon in vitro and in vivo treatment. Hence, we provided very innovative results by showing that nitazoxanide, and now also, respectively, new modified compounds may represent a useful alternative to albendazole. In the context of chemotherapeutical repression of parasite growth, we searched also for parasite molecules, whose expression levels correlate with the viability and growth activity of E. multilocularis metacestode. Expression levels of 14-3-3 and II/3-10, relatively quantified by realtime reverse transcription-PCR using a housekeeping gene beta-actin, were studied in permissive nu/nu and in low-permissive wild type BALB/c mice. At 2 months p.i., the transcription level of 14-3-3 was significantly higher in parasites actively proliferating in nu/nu mice compared to parasites moderately growing in wild type mice. Immunoblotting experiments confirmed at the protein level that 14-3-3 was over-expressed in parasites derived from nu/nu mice at 2 months p.i. In vitro-treatment of E. multilocularis with an anti-echinococcal drug nitazoxanide for a period of 8 days resulted in a significant decrease of both 14-3-3 and II/3-10 transcription levels, which correlated with the kinetics of a housekeeping gene, beta-actin. This indicates that 14-3-3-exhibits a good potential as a molecular marker to assess viability and growth activity of the parasite.
Gottstein
Bruno
B
Institute of Parasitology, Vetsuisse Faculty and Faculty of Medicine, University of Bern, Switzerland. bruno.gottstein@ipa.unibe.ch
Haag
Karen
K
Walker
Mirjam
M
Matsumoto
Jun
J
Mejri
Naceur
N
Hemphill
Andrew
A
D048948:14-3-3 Proteins / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D000923:Anticestodal Agents / Q000008:administration & dosage / Q000627:therapeutic use*; D004444:Echinococcosis, Hepatic / Q000188:drug therapy* / Q000276:immunology / Q000469:parasitology; D048210:Echinococcus multilocularis / Q000187:drug effects / Q000254:growth & development* / Q000472:pathogenicity; D006790:Host-Parasite Interactions; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D009574:Nitro Compounds; D013844:Thiazoles / Q000008:administration & dosage / Q000627:therapeutic use*
D048948:14-3-3 Proteins; D000923:Anticestodal Agents; D009574:Nitro Compounds; D013844:Thiazoles; C041747:nitazoxanide