The immunomodulatory activity of ArtinM on the course of Cryptococcus gattii infection

As infecções fúngicas invasivas são um problema global em saúde pública que acometem milhões de vidas anualmente. Entre os principais gêneros causadores dessas doenças, a espécie Cryptococcus gattii destaca-se por acometer indivíduos imunocompetentes e imunossuprimidos. Esse patógeno atua no bloqueio da diferenciação das células T helper (Th) 1 (Th1) e 17 (Th17) através da atenuação da resposta pró-inflamatória no tecido pulmonar. Em vista da capacidade imunomoduladora de ArtinM em induzir uma resposta imunológica Th1 e Th17 através da indução da produção de IL-12 por células apresentadoras de antígenos (APCs), e pelo efeito da lectina em células T CD4+, esse efeito imunomodulador de ArtinM promoveu o controle da infecção por P. brasiliensis e C. albicans. Dessa forma, investigamos os efeitos da administração profilática ou terapêutica de ArtinM no controle da infecção experimental por C. gattii. Constatamos que a infecção por C. gattii rapidamente progride no tecido pulmonar, assim como há uma disseminação completa para o coração, fígado, rim, baço, cérebro e sangue após 21 dias de infeção. Na análise do pulmão, o perfil citocínico apresentou baixos níveis de citocinas pró-inflamatórias, e observamos um aumento na expressão relativa dos marcadores de polarização para o perfil M2 (Arginase-1 e YM-1), em todo período analisado. Visto a capacidade reguladora do C. gattii sobre o sistema imunitário do hospedeiro, avaliamos o efeito da administração profilática de ArtinM sobre o curso dessa infecção. Ao longo de 42 dias de infecção, o grupo tratado com ArtinM apresentou uma redução da carga fúngica pulmonar nos dias 28 e 35 pós-infecção, e os níveis de IFN-?, IL-17 e TNF-? aumentaram significativamente no grupo ArtinM. Esse efeito imunomodulador da administração profilática de ArtinM sobre o modelo de infecção experimental por C. gattii levou-nos a avaliar o efeito da administração terapêutica de ArtinM no curso dessa infecção. Essa nova proposta de aplicação de ArtinM resultou na redução da carga fúngica pulmonar de C. gattii após 21 dias de infecção, comparado com o grupo controle. A administração terapêutica de ArtinM foi capaz de elevar o número absoluto de neutrófilos e linfócitos no sangue periférico dos animais infectados, porém os níveis de citocinas não diferiram entre os grupos ArtinM e controle. Além disso, o grupo tratado com ArtinM apresentou menor frequência de células T CD4+ no baço. Diante do efeito imunomodulador a administração profilática ou terapêutica de ArtinM que promoveu um controle parcial da infecção por C. gattii, partimos para a associação desse agente imunomodulador com o antifúngicoR e s u m o | viii Fluconazol. Verificamos que os animais tratados com ArtinM em associação com fluconazol também apresentaram uma redução da carga fúngica pulmonar em comparação ao grupo controle, indicando que ArtinM é passível de ser associada com a terapia convencional antifúngica. Portanto, nossos dados evidenciaram que a atividade imunomodulatória de ArtinM, através da administração profilática ou terapêutica, foi evidenciada no curso da infecção por C. gattii e favoreceu a imunidade do hospedeiro promovendo uma redução parcial da carga fúngica pulmonarInvasive fungal diseases are a global public health problem, that affects millions of individuals each year. Among the species that cause these diseases, Cryptococcus gattii is a major because affects immunosuppressed and immunocompetent individuals. C. gattii regulates the differentiation of helper (Th) T cells 1 (Th1) and 17 (Th17) by attenuating the proinflammatory response in the lungs. We know that the immunomodulatory capacity of ArtinM to induce the Th1 and Th17 immune response through IL-12 production by antigen presenting cells (APCs) and the CD4+ T cells activation by direct effect of ArtinM, these activities of ArtinM promoted the control of P. brasiliensis and C. albicans infection. Thus, we investigated the effects of prophylactic or therapeutic administration of ArtinM in the control of C. gattii infection. We found that C. gattii infection rapidly increase in the lungs and disseminate to the heart, liver, kidney, spleen, brain and blood after 21 days of infection. In the lung analysis, the cytokine profile showed low levels of proinflammatory cytokines and we observed an increase in the relative expression of the polarization markers for the M2 macrophages (Arginase-1 and YM-1) for all period. Given the regulatory ability of C. gattii on the host immune response, we evaluated the effect of prophylactic administration of ArtinM on the course of C. gattii infection. During 42 days of infection, the ArtinM-treated group had a reduction in the pulmonary fungal burden on days 28 and 35 post-infection, and levels of IFN-?, IL-17 and TNF-? were significantly increased in the ArtinM group. This immunomodulatory effect of prophylactic administration of ArtinM on the C. gattii infection led us to evaluate the effect of the therapeutic administration of ArtinM on the course of this infection. This new strategy for the ArtinM administration promoted a reduction in the lung fungal burden of C. gattii after 21 days of infection compared to control group. Therapeutic administration of ArtinM was able to increase the absolute number of neutrophils and lymphocytes in the peripheral blood of infected mice, but the cytokine levels did not differ between the ArtinM and control groups. In addition, the ArtinM-treated group showed decrease of the CD4+ T cell frequency in the spleen. Considering the immunomodulatory effect, the prophylactic or therapeutic administration of ArtinM promoted a partial control of C. gattii infection, and instigate us to investigate the association of ArtinM and Fluconazole. We found that the treatment with ArtinM in combination with fluconazole also promoted a reduction in the pulmonary fungal burden, which suggested that ArtinM works well inA b s t r a c t | xi association with antifungal drugs. Therefore, our data showed that the immunomodulatory activity of ArtinM, via prophylactic or therapeutic administration, was evidenced in the course of C. gattii infection and contributed the host immune response to reduce the lung fungal burde

The immunomodulatory activity of ArtinM on the course of Cryptococcus gattii infection

As infecções fúngicas invasivas são um problema global em saúde pública que acometem milhões de vidas anualmente. Entre os principais gêneros causadores dessas doenças, a espécie Cryptococcus gattii destaca-se por acometer indivíduos imunocompetentes e imunossuprimidos. Esse patógeno atua no bloqueio da diferenciação das células T helper (Th) 1 (Th1) e 17 (Th17) através da atenuação da resposta pró-inflamatória no tecido pulmonar. Em vista da capacidade imunomoduladora de ArtinM em induzir uma resposta imunológica Th1 e Th17 através da indução da produção de IL-12 por células apresentadoras de antígenos (APCs), e pelo efeito da lectina em células T CD4+, esse efeito imunomodulador de ArtinM promoveu o controle da infecção por P. brasiliensis e C. albicans. Dessa forma, investigamos os efeitos da administração profilática ou terapêutica de ArtinM no controle da infecção experimental por C. gattii. Constatamos que a infecção por C. gattii rapidamente progride no tecido pulmonar, assim como há uma disseminação completa para o coração, fígado, rim, baço, cérebro e sangue após 21 dias de infeção. Na análise do pulmão, o perfil citocínico apresentou baixos níveis de citocinas pró-inflamatórias, e observamos um aumento na expressão relativa dos marcadores de polarização para o perfil M2 (Arginase-1 e YM-1), em todo período analisado. Visto a capacidade reguladora do C. gattii sobre o sistema imunitário do hospedeiro, avaliamos o efeito da administração profilática de ArtinM sobre o curso dessa infecção. Ao longo de 42 dias de infecção, o grupo tratado com ArtinM apresentou uma redução da carga fúngica pulmonar nos dias 28 e 35 pós-infecção, e os níveis de IFN-?, IL-17 e TNF-? aumentaram significativamente no grupo ArtinM. Esse efeito imunomodulador da administração profilática de ArtinM sobre o modelo de infecção experimental por C. gattii levou-nos a avaliar o efeito da administração terapêutica de ArtinM no curso dessa infecção. Essa nova proposta de aplicação de ArtinM resultou na redução da carga fúngica pulmonar de C. gattii após 21 dias de infecção, comparado com o grupo controle. A administração terapêutica de ArtinM foi capaz de elevar o número absoluto de neutrófilos e linfócitos no sangue periférico dos animais infectados, porém os níveis de citocinas não diferiram entre os grupos ArtinM e controle. Além disso, o grupo tratado com ArtinM apresentou menor frequência de células T CD4+ no baço. Diante do efeito imunomodulador a administração profilática ou terapêutica de ArtinM que promoveu um controle parcial da infecção por C. gattii, partimos para a associação desse agente imunomodulador com o antifúngicoR e s u m o | viii Fluconazol. Verificamos que os animais tratados com ArtinM em associação com fluconazol também apresentaram uma redução da carga fúngica pulmonar em comparação ao grupo controle, indicando que ArtinM é passível de ser associada com a terapia convencional antifúngica. Portanto, nossos dados evidenciaram que a atividade imunomodulatória de ArtinM, através da administração profilática ou terapêutica, foi evidenciada no curso da infecção por C. gattii e favoreceu a imunidade do hospedeiro promovendo uma redução parcial da carga fúngica pulmonarInvasive fungal diseases are a global public health problem, that affects millions of individuals each year. Among the species that cause these diseases, Cryptococcus gattii is a major because affects immunosuppressed and immunocompetent individuals. C. gattii regulates the differentiation of helper (Th) T cells 1 (Th1) and 17 (Th17) by attenuating the proinflammatory response in the lungs. We know that the immunomodulatory capacity of ArtinM to induce the Th1 and Th17 immune response through IL-12 production by antigen presenting cells (APCs) and the CD4+ T cells activation by direct effect of ArtinM, these activities of ArtinM promoted the control of P. brasiliensis and C. albicans infection. Thus, we investigated the effects of prophylactic or therapeutic administration of ArtinM in the control of C. gattii infection. We found that C. gattii infection rapidly increase in the lungs and disseminate to the heart, liver, kidney, spleen, brain and blood after 21 days of infection. In the lung analysis, the cytokine profile showed low levels of proinflammatory cytokines and we observed an increase in the relative expression of the polarization markers for the M2 macrophages (Arginase-1 and YM-1) for all period. Given the regulatory ability of C. gattii on the host immune response, we evaluated the effect of prophylactic administration of ArtinM on the course of C. gattii infection. During 42 days of infection, the ArtinM-treated group had a reduction in the pulmonary fungal burden on days 28 and 35 post-infection, and levels of IFN-?, IL-17 and TNF-? were significantly increased in the ArtinM group. This immunomodulatory effect of prophylactic administration of ArtinM on the C. gattii infection led us to evaluate the effect of the therapeutic administration of ArtinM on the course of this infection. This new strategy for the ArtinM administration promoted a reduction in the lung fungal burden of C. gattii after 21 days of infection compared to control group. Therapeutic administration of ArtinM was able to increase the absolute number of neutrophils and lymphocytes in the peripheral blood of infected mice, but the cytokine levels did not differ between the ArtinM and control groups. In addition, the ArtinM-treated group showed decrease of the CD4+ T cell frequency in the spleen. Considering the immunomodulatory effect, the prophylactic or therapeutic administration of ArtinM promoted a partial control of C. gattii infection, and instigate us to investigate the association of ArtinM and Fluconazole. We found that the treatment with ArtinM in combination with fluconazole also promoted a reduction in the pulmonary fungal burden, which suggested that ArtinM works well inA b s t r a c t | xi association with antifungal drugs. Therefore, our data showed that the immunomodulatory activity of ArtinM, via prophylactic or therapeutic administration, was evidenced in the course of C. gattii infection and contributed the host immune response to reduce the lung fungal burde

Immunomodulation via carbohydrate ligands in a vaccine strategy helps in the control of Cryptococcus gattii infection

As infecções fúngicas acometem milhões de vidas a cada ano e, nesse caso, a criptococose apresenta importante relevância ao atingir indivíduos imunocomprometidos e indivíduos saudáveis, representando uma das principais causas de morbidade e mortalidade mundial entre os indivíduos HIV+. A baixa eficácia e os efeitos colaterais associados aos agentes antifúngicos têm destacado a importância do desenvolvimento de novas abordagens imunoterapêuticas para o tratamento da infecção por Cryptococcus gattii. Nos estudos que constituem a presente tese, uma estratégia de imunização frente a infeção por C. gattii foi desenvolvida e, para isso, foi utilizado agonistas seletivos do receptor de Dectin-1 ou agonista de TLR2/CD14 que atuaram como adjuvante no protocolo vacinal. Inicialmente, camundongos BALB/c receberam a administração do adjuvante ArtinM e, em seguida, houve a etapa de imunização, e esse protocolo não alterou significativamente os níveis de citocinas e concentração de células CD3+ e CD11b+ no tecido pulmonar dos camundongos tratados com ArtinM, comparado com o grupo imunizado. Esse protocolo de imunização associado ao adjuvante ArtinM não contribuiu na redução da carga fúngica pulmonar e na curva de sobrevivência de camundongos infectados com C. gattii. Isso levou a alteração do protocolo de imunização que modificou para uma alta concentração de levedura inativada de C. gattii, com intervalos maiores entre as imunizações. Em seguida, camundongos C57BL/6 que receberam o adjuvante ArtinM, previamente a imunização, apresentaram níveis elevado de IL-10 e uma expressão relativa de IL-23 aumentada nos pulmões, e foi detectada uma redução na carga de C. gattii no tecido pulmonar. Em paralelo, o protocolo de imunização citado acima foi avaliado na administração de agonista do receptor de dectina-1 como adjuvantes. Para isso, camundongos BALB/c ou C57BL/6 receberam curdlan ou peptídeo β-glucano (BGP), previamente a imunização com C. gattii, e após 14 dias da última imunização foram infectados com C. gattii. Após 14 dias de infecção, os animais foram eutanasiados para avaliação. O adjuvante curdlan restaurou os níveis de TNFα que foram reduzidos no grupo de animais submetidos somente a etapa de imunização com C. gattii. A área média e a frequência relativa das células titãs de C. gattii nos pulmões de camundongos BALB/c tratados com curdlan foram reduzidas. No entanto, isso não reduziu a carga fúngica pulmonar e não diminuiu o infiltrado inflamatório no parênquima pulmonar dos camundongos BALB/c. Por outro lado, o adjuvante curdlan induziu altos níveis IFN-γ e IL-10 e diminuiu a carga de C. gattii nos pulmões de camundongos C57BL/6, o que não foi replicado em camundongos tratados com peptídeos β-glucano. Por fim, a avaliação de macrófagos alveolares de linhagem AMJ2-C11, oriundos de camundongos C57BL/6, que demonstrou uma baixa expressão do receptor de dectina, foram utilizados na incubação com curdlan que resultou na polarização dos macrófagos AMJ2-C11 para o perfil M1, no aumento da expressão de TLR2, e maior atividade fagocítica sobre C. gattii. No entanto, curdlan não induziu uma maior atividade fungicida de AMJ2-C11 sobre C. gattii. A utilização de inibidores de Syk, e de tirosina quinases da família Src demonstrou a especificidade da atuação de curdlan em células AMJ2-C11 via receptor de dectina-1. Assim, o adjuvante ArtinM e o adjuvante curdlan favoreceram o controle da infecção por C. gattii, com destaque, para o modelo de infecção com C. gattii em camundongos C57BL/6. O presente estudo terá importantes implicações no desenvolvimento de novas abordagens imunoterapêuticas para tratar a infecção por C. gattii.Fungal infections affect millions of lives each year and, in this case, cryptococcosis has an important relevance in reaching immunocompromised and healthy individuals, representing one of the main causes of morbidity and mortality worldwide among HIV+ individuals. The low efficacy and side effects associated with antifungal agents have highlighted the importance of developing new immunotherapeutic approaches for the treatment of Cryptococcus gattii infection. In the studies that constitute the present thesis, an immunization strategy against C. gattii infection was developed and, for this, selective agonists of the Dectin-1 receptor or TLR2/CD14 agonist were used, which were as an adjuvant in the vaccine protocol. Initially, BALB/c mice received the administration of adjuvant ArtinM previously the immunization protocol, and this protocol did not significantly alter the cytokine levels and concentration of CD3+ and CD11b+ cells in the lung tissue of the mice treated with ArtinM, compared to with the immunized group. This immunization protocol associated with the adjuvant ArtinM did not contribute to the reduction of pulmonary fungal burden and the survival curve of mice infected with C. gattii. This led to a change in the immunization protocol that modified to a high concentration of inactivated C. gattii yeast, with longer intervals between immunizations. Then, C57BL/6 mice that received the adjuvant ArtinM, prior to immunization, showed elevated levels of IL-10 and an increased relative expression of IL-23 in the lungs, and a reduction in the burden of C. gattii in the lung tissue was detected. In parallel, the above-cited immunization protocol was evaluated in the administration of dectin-1 agonists as adjuvants. For this, BALB/c or C57BL/6 mice received curdlan or peptide β-glucans (BGP), prior to immunization with C. gattii, and 14 days after the last immunization they were infected with C. gattii. After 14 days of infection, the animals were euthanized for evaluation. The adjuvant curdlan restored the levels of TNF-α that were reduced in the group of animals submitted only to the immunization protocol with C. gattii. The mean area and relative frequency of C. gattii titan cells in the lungs of BALB/c mice treated with curdlan were reduced. However, the pulmonary fungal burden was not reduced and the inflammatory infiltrate in the parenchyma of BALB/c mice was not reduced. On the other hand, the adjuvant curdlan induced high levels of IFN-γ and IL-10 and decreased the burden of C. gattii in the lungs of C57BL/6 mice, which was not replicated in mice treated with β-glucan peptides. Finally, the evaluation of AMJ2-C11, a alveolar macrophages cell line from C57BL/6 mice, which demonstrated a low expression of the dectin-1, were used in the incubation with curdlan that resulted in the polarization of AMJ2-C11 macrophages to the M1 profile, increased expression of TLR2, and increased phagocytic activity on C. gattii. However, curdlan did not induce greater fungicidal activity of AMJ2-C11 on C. gattii. The use of Syk inhibitors and tyrosine kinases of the Src family demonstrated the specificity of the action of curdlan in AMJ2-C11 cells via the dectin-1 receptor. Thus, the adjuvant ArtinM and the adjuvant curdlan favored the control of C. gattii infection, especially for the model of infection with C. gattii in C57BL/6 mice. The present study will have important implications for the development of new immunotherapeutic approaches to treat C. gattii infection

The Response of IL-17-Producing B Cells to ArtinM Is Independent of Its Interaction with TLR2 and CD14

ArtinM, a d-mannose-binding lectin from Artocarpus heterophyllus, activates antigen-presenting cells by recognizing Toll-like receptor (TLR)2 and cluster of differentiation (CD)14 N-glycans, induces cytokine production, and promotes type 1 T helper (Th1) immunity, a process that plays an assisting role in the combat against fungal infections. We recently demonstrated that ArtinM stimulates CD4+ T cells to produce interleukin (IL)-17 through direct interaction with CD3. Here, we further investigated the effects of ArtinM on the production of IL-17 by B cell activation. We showed that ArtinM activates murine B cells, increasing IL-17 and IL-12p40 production. The direct effect of ArtinM was sufficient to induce IL-17 production in B cells, and we did not find differences in the levels of IL-17 between the B cells purified from the wild-type (WT) and knockout (KO) mice for TLR2 or CD14 in the presence of ArtinM. Thus, the effects of ArtinM on splenic B cells through carbohydrate recognition may contribute to Th17 immunity; however, the mechanism involved is not associated with the interaction of ArtinM with TLR2 and CD14. The current work represents a pioneering effort in the understanding of the induction of IL-17 by lectins in B cells

scFv against HSP60 of Strongyloides sp. and Its Application in the Evaluation of Parasite Frequency in the Elderly

The present study is aimed at evaluating serological method using scFv anti-Strongyloides sp. and reporting the frequencies of the results with conventional parasitological technique (faeces) in elderly individuals. Among 112 elderly individuals (≥60 years of age), 14.28% were positive for at least one enteroparasite, with one individual positive for S. stercoralis. Sera were evaluated for the presence of anti-Strongyloides sp. antibodies using total or detergent fraction extracts of Strongyloides venezuelensis, which presented positivity rates of 19.64% and 10.71%, respectively. An anti-HSP60 single-chain variable fragment from Strongyloides sp. was used to detect parasite antigens, with 5.36% (6 individuals) of ELISA-positive individuals returning a positive result. While the serological test indicates previous or recent infection and may be limited by antigen purification, the anti-HSP60 method reflects the presence of Strongyloides sp. immune complexes and exhibits greater sensitivity and specificity. Our results demonstrate the variable occurrence of enteroparasites in elderly individuals residing in long-term nursing homes and validate a novel epidemiological tool to describe infection cases by Strongyloides sp

Galectin-3 Inhibits Paracoccidioides brasiliensis Growth and Impacts Paracoccidioidomycosis through Multiple Mechanisms

Paracoccidioidomycosis (PCM) is the most prevalent systemic mycosis in Latin America. Although the immune mechanisms to control PCM are still not fully understood, several events of the host innate and adaptive immunity are crucial to determine the progress of the infection. Mammalian β-galactoside-binding protein galectin-3 (Gal-3) plays significant roles during microbial infections and has been studied for its immunomodulatory roles, but it can also have direct antimicrobial effects. We asked whether this protein plays a role in Paracoccidioides brasiliensis. We report herein that Gal-3 indeed has direct effects on the fungal pathogen, inhibiting fungal growth and reducing extracellular vesicle stability. Our results suggest a direct role for Gal-3 in P. brasiliensis infection, with beneficial effects for the mammalian host.The thermodimorphic pathogenic fungi Paracoccidioides brasiliensis and Paracoccidioides lutzii are the etiologic causes of paracoccidioidomycosis (PCM), the most prevalent systemic mycosis in Latin America. Galectin-3 (Gal-3), an animal β-galactoside-binding protein, modulates important roles during microbial infections, such as triggering a Th2-polarized immune response in PCM. Herein, we demonstrate that Gal-3 also plays other important roles in P. brasiliensis infection. We verified that Gal-3 levels are upregulated in human and mice infections and established that Gal-3 inhibited P. brasiliensis growth by inhibiting budding. Furthermore, Gal-3 affected disruption and internalization of extracellular vesicles (EVs) from P. brasiliensis by macrophages. Our results suggest important protective roles for Gal-3 in P. brasiliensis infection, indicating that increased Gal-3 production during P. brasiliensis infection may affect fungal growth and EV stability, thus promoting beneficial effects that could influence the course of PCM. The finding that Gal-3 has effects against P. brasiliensis together with previously reported effects against Cryptococcus neoformans suggests that molecule has a general antifungal role in innate defenses against fungal pathogens

Adjuvant Curdlan Contributes to Immunization against <i>Cryptococcus gattii</i> Infection in a Mouse Strain-Specific Manner

The low efficacy and side effects associated with antifungal agents have highlighted the importance of developing immunotherapeutic approaches to treat Cryptococcus gattii infection. We developed an immunization strategy that uses selective Dectin-1 agonist as an adjuvant. BALB/c or C57BL/6 mice received curdlan or β-glucan peptide (BGP) before immunization with heat-killed C. gattii, and the mice were infected with viable C. gattii on day 14 post immunization and euthanized 14 days after infection. Adjuvant curdlan restored pulmonary tumor necrosis factor- α (TNF-α) levels, as induced by immunization with heat-killed C. gattii. The average area and relative frequency of C. gattii titan cells in the lungs of curdlan-treated BALB/c mice were reduced. However, this did not reduce the pulmonary fungal burden or decrease the i0,nflammatory infiltrate in the pulmonary parenchyma of BALB/c mice. Conversely, adjuvant curdlan induced high levels of interferon-γ (IFN-γ) and interleukin (IL)-10 and decreased the C. gattii burden in the lungs of C57BL/6 mice, which was not replicated in β-glucan peptide-treated mice. The adjuvant curdlan favors the control of C. gattii infection depending on the immune response profile of the mouse strain. This study will have implications for developing new immunotherapeutic approaches to treat C. gattii infection

Adjuvant Curdlan Contributes to Immunization against Cryptococcus gattii Infection in a Mouse Strain-Specific Manner

The low efficacy and side effects associated with antifungal agents have highlighted the importance of developing immunotherapeutic approaches to treat Cryptococcus gattii infection. We developed an immunization strategy that uses selective Dectin-1 agonist as an adjuvant. BALB/c or C57BL/6 mice received curdlan or &beta;-glucan peptide (BGP) before immunization with heat-killed C. gattii, and the mice were infected with viable C. gattii on day 14 post immunization and euthanized 14 days after infection. Adjuvant curdlan restored pulmonary tumor necrosis factor- &alpha; (TNF-&alpha;) levels, as induced by immunization with heat-killed C. gattii. The average area and relative frequency of C. gattii titan cells in the lungs of curdlan-treated BALB/c mice were reduced. However, this did not reduce the pulmonary fungal burden or decrease the i0,nflammatory infiltrate in the pulmonary parenchyma of BALB/c mice. Conversely, adjuvant curdlan induced high levels of interferon-&gamma; (IFN-&gamma;) and interleukin (IL)-10 and decreased the C. gattii burden in the lungs of C57BL/6 mice, which was not replicated in &beta;-glucan peptide-treated mice. The adjuvant curdlan favors the control of C. gattii infection depending on the immune response profile of the mouse strain. This study will have implications for developing new immunotherapeutic approaches to treat C. gattii infection

Systemic effects in naïve mice injected with immunomodulatory lectin ArtinM

<div><p>Toll-like receptors (TLR) contain N-glycans, which are important glycotargets for plant lectins, to induce immunomodulation. The lectin ArtinM obtained from <i>Artocarpus heterophyllus</i> interacts with TLR2 N-glycans to stimulate IL-12 production by antigen-presenting cells and to drive the immune response toward the Th1 axis, conferring resistance against intracellular pathogens. This immunomodulatory effect was demonstrated by subcutaneously injecting (s.c.) ArtinM (0.5 μg) in infected mice. In this study, we evaluated the systemic implications of ArtinM administration in <i>naïve</i> BALB/c mice. The mice were s.c. injected twice (7 days interval) with ArtinM (0.5, 1.0, 2.5, or 5.0 μg), LPS (positive control), or PBS (negative control) and euthanized after three days. None of the ArtinM-injected mice exhibited change in body weight, whereas the relative mass of the heart and lungs diminished in mice injected with the highest ArtinM dose (5.0 μg). Few and discrete inflammatory foci were detected in the heart, lung, and liver of mice receiving ArtinM at doses ≥2.5 μg. Moreover, the highest dose of ArtinM was associated with increased serum levels of creatine kinase MB isoenzyme (CK-MB) and globulins as well as an augmented presence of neutrophils in the heart and lung. IL-12, IFN-γ, TNF-α, and IL-10 measurements in the liver, kidney, spleen, heart, and lung homogenates revealed decreased IL-10 level in the heart and lung of mice injected with 5.0 μg ArtinM. We also found an augmented frequency of T helper and B cells in the spleen of all ArtinM-injected <i>naïve</i> mice, whereas the relative expressions of T-bet, GATA-3, and ROR-γt were similar to those in PBS-injected animals. Our study demonstrates that s.c. injection of high doses of ArtinM in <i>naïve</i> mice promotes mild inflammatory lesions and that a low immunomodulatory dose is innocuous to <i>naïve</i> mice.</p></div

Blood leukogram of <i>naïve</i> BALB/c mice after ArtinM administration.

<p>Total and differential leukocyte counting was performed in blood samples obtained at days 0 (<b>A</b>), -9 (<b>B</b>), -8 (<b>C</b>), -2 (<b>D</b>), and -1 (<b>E</b>) from animals that received ArtinM at the doses specified in each panel, LPS (positive controls), or PBS (negative controls). Results are expressed as mean ± SEM (<b>A</b>) and mean ± SD (<b>B-E</b>). Differences were considered significant when p < 0.05 (*) compared to the PBS control group.</p