ABSTRACT
BACKGROUND: Polymorphonuclear neutrophils (PMNs), along with macrophages, are the first leukocytes recruited to the site of infection in dermatophytoses and are responsible for the in fine elimination of the fungus. It has been demonstrated that feline PMNs produce pro-inflammatory cytokines after stimulation with Microsporum canis. The activation of these cells results from the recognition of specific PAMPs (pathogen associated molecular patterns) from M. canis by PRRs (pattern recognition receptors) of PMNs. The C-type lectin receptors (CLRs) and toll-like receptors (TLRs) are the two main PRRs in phagocytic cells that recognize fungal components. HYPOTHESIS/OBJECTIVE: The aim of this study was to evaluate the expression of TLR-2, TLR-4 and dectin-1 mRNA in feline PMNs exposed to different components from M. canis. METHODS: Feline PMNs were stimulated for 2 h or 4 h with either live arthroconidia, heat-killed arthroconidia or secreted components from M. canis. The levels of TLR-2, TLR-4 and dectin-1 mRNA were assessed by RT-qPCR. RESULTS: Results showed an increase of TLR-2 and TLR-4 mRNA levels in feline PMNs stimulated with live and heat-killed arthroconidia, but not in those stimulated with the secreted components from M. canis. No significant variation in dectin-1 mRNA expression was observed in PMNs stimulated with the different fungal components. CONCLUSIONS AND CLINICAL IMPORTANCE: The overexpression of TLR-2 and TLR-4 mRNAs in stimulated feline PMNs suggests that these receptors are involved in the host immune response through the recognition of M. canis PAMPs.
Subject(s)
Cats/metabolism , Gene Expression Regulation/immunology , Microsporum , Neutrophils/physiology , Toll-Like Receptor 2/metabolism , Toll-Like Receptor 4/metabolism , Animals , Cats/immunology , Cells, Cultured , Female , Lectins, C-Type/genetics , Lectins, C-Type/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Spores, Fungal , Toll-Like Receptor 2/genetics , Toll-Like Receptor 4/geneticsABSTRACT
In order to identify protective immunogens against Microsporum canis infection, a purified recombinant keratinolytic metalloprotease (r-MEP3) was tested as a subunit vaccine in experimentally infected guinea pigs. Both humoral and cellular specific immune responses developing towards r-MEP3 were evaluated, by enzyme-linked immunosorbent assay and by in vitro lymphocyte transformation tests respectively. Vaccination induced a strong antibody response, and a significant but transient lymphoproliferative response against the protein. However, the protocol failed to prevent fungal invasion or development of dermatophytic lesions. These results show that under the present experimental conditions, r-MEP3 specific antibodies are not protective against a challenge exposure. They also suggest that in the same model, the induction of cell-mediated immunity towards r-MEP3 is not sufficient, indicating the need for further research in the field of specific immune mechanisms involved in M. canis dermatophytosis.
Subject(s)
Antibodies, Fungal/blood , Antigens, Fungal/immunology , Dermatomycoses/prevention & control , Fungal Vaccines , Microsporum/immunology , Animals , Disease Models, Animal , Enzyme-Linked Immunosorbent Assay , Female , Fungal Vaccines/immunology , Guinea Pigs , Immunity, Cellular , Metalloproteases/immunology , Microsporum/enzymology , Recombinant Proteins/immunology , Treatment Outcome , Vaccination , Vaccines, Subunit/immunologyABSTRACT
A Microsporum canis recombinant 31.5 kDa keratinase and a M. canis crude exo-antigen were tested as vaccines in an experimental infection model in guinea pigs. Animals were vaccinated subcutaneously three times at two-week intervals with either the keratinase, the exo-antigen or the adjuvant alone. Cutaneous challenge was performed blindly. Both humoral and cellular-specific immune responses to M. canis antigens were evaluated every 14 days, while a blind evaluation of clinical lesion development and fungal persistency in skin were monitored weekly. Vaccination induced very high and significant (P < 0.01) antibody responses towards both antigens. High cell-mediated immune responses to both immunogens were also induced by vaccination. After challenge, however, scores reflecting the severity of dermatophytic lesions did not differ significantly between vaccinated and control groups at any time after challenge. These results suggest that, in the guinea pig, the induction of specific immune responses against the M. canis-secreted antigens used in this study are not protective against challenge exposure.