THAISE YUMIE TOMOKANE
Projetos de Pesquisa
Unidades Organizacionais
Departamento de Patologia, Faculdade de Medicina
LIM/50 - Laboratório de Patologia das Moléstias Infecciosas, Hospital das Clínicas, Faculdade de Medicina
LIM/50 - Laboratório de Patologia das Moléstias Infecciosas, Hospital das Clínicas, Faculdade de Medicina
11 resultados
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- In situ study of cellular immune response in human cutaneous lesions caused by Leishmania (Viannia) panamensis in Panama(2021) GONZALEZ, Kadir; CALZADA, Jose Eduardo; TOMOKANE, Thaise Yumie; PACHECO, Carmen Maria Sandoval; FLORES, Gabriela Venicia Araujo; GOMES, Claudia Maria Castro; CORBETT, Carlos Eduardo Pereira; SALDANA, Azael; LAURENTI, Marcia DalastraAims: Leishmaniasis is considered a disease with multiple clinical/immunopathological characteristics, depending on the immunity of the host and the species of the parasite. In Panama, the most prevalent species that causes localized cutaneous leishmaniasis (LCL) is Leishmania (Viannia) panamensis, and its immune response is poorly studied. Therefore, we evaluated by immunohistochemistry, the in situ immune response during this infection. Methods and Results: Biopsies from Panamanian patients with LCL were collected and processed by histological techniques. Infection by L. (V.) panamensis was demonstrated by isolation in culture and molecular characterization by Hsp70-RFLP. The in situ immune response was assessed by immunohistochemistry. The immune response was characterized by predominance of T cells, mainly CD8 cells that showed positive correlation with IFN-gamma and Granzyme B. CD4 cells presented positive correlation with both IFN-gamma and IL-13, pointed by mixed cellular immune response. Regulatory response was characterized by FoxP3 cells, which showed positive correlation to IL-10 but not with TGF-beta. Conclusions: L. (V.) panamensis infection triggers a mixed cellular immune response, characterized by the presence of pro-inflammatory, anti-inflammatory and regulatory elements in the skin lesion of Panamanian patients. These data contribute to a better understanding of the immunopathogenesis of Leishmania Viannia infection in Panama.
- Performance of immunohistochemistry as a useful tool for the diagnosis of cutaneous leishmaniasis in Panama, Central America(2019) GONZALEZ, K.; CALZADA, J. E.; DIAZ, R.; PAZ, H.; GARCIA, V.; MIRANDA, A.; TOMOKANE, T.; PUGA, S.; SALDANA, A.; LAURENTI, M.Cutaneous leishmaniasis (CL) is one of the most frequent parasitic zoonoses in Panama. Currently, conventional, molecular and histopathological tests are performed to diagnose CL. Immunohistochemistry (IHC) has proven to be a valuable tool to facilitate the diagnosis of leishmaniasis and to study the cellular immune response developed during the infection. Therefore, considering the absence of IHC in the diagnostic routine in Panama, the objective of this study is to demonstrate the usefulness of this test as a complementary diagnostic tool for improving the sensitivity of histopathology (HP) and helping to study the cellular immune response of patients. Samples from patients with suspected CL were analysed by intradermal reaction of Montenegro (IDRM), smears, culture, PCR (Viannia, Hsp-70), HP and IHC. According to the diagnostic criteria, 95.8% of patients were positive for Leishmania sp., that was characterized as Leishmania (V.) panarnensis by PCR-HSP7O/RFLP. From positive samples, Leishmania was detected by the tested diagnostic methods in the following degrees: 100% by IDRM, 60% by smears, 93.3% by culture, 100% by kDNA PCR, 78.3% by PCR Hsp-70, 50% by HP and 73.9% by IHC. Although IHC had a poor correlation (k = 0.191) with the diagnostic criteria, the sensitivities of both HP (76.1%) and smears (89.1%) were improved by combining them with IHC. IHC considerably improved the detection of the Leishmania parasites in the histopathological sections, supporting the need to implement this diagnostic tool in Panama. In addition, immunohistochemistry allows evaluation of the patient's immune response and thus provides new guidelines for the treatment and control of CL in Panama.
- Chromosomal segments may explain the antibody response cooperation for canine leishmaniasis pathogenesis(2020) BATISTA, Luis F. S.; TORRECILHA, Rafaela B. P.; SILVA, Rafaela B.; UTSUNOMIYA, Yuri T.; SILVA, Thais B. F.; TOMOKANE, Thaise Y.; PACHECO, Acacio D.; BOSCO, Anelise M.; PAULAN, Silvana C.; ROSSI, Claudio N.; COSTA, Gustavo N. O.; MARCONDES, Mary; CIARLINI, Paulo C.; NUNES, Caris M.; MATTA, Vania L. R.; LAURENTI, Marcia D.Visceral leishmaniasis (VL) is marked by hyperactivation of a humoral response secreting high quantity of immunoglobulins (Igs) that are inaccessible to intracellular parasites. Here we investigated the contributions of the antibody response to the canine leishmaniasis pathogenesis. Using correlation and genome-wide association analysis, we investigated the relationship of anti-Leishmania infantum immunoglobulin classes levels with parasite burden, clinical response, renal/hepatic biochemical, and oxidative stress markers in dogs from endemic areas of VL. Immunoglobulin G (IgG) and IgA were positively correlated with parasite burden on lymph node and blood. Increased IgG, IgA and IgE levels were associated with severe canine leishmaniasis (CanL) whereas IgM was elevated in uninfected exposed dogs. Correlations of IgM, IgG and IgA with creatinine, urea, AST and ALT levels in the serum were suggested an involvement of those Igs with renal and hepatic changes. The correlogram of oxidative radicals and antioxidants revealed a likely relationship of IgM, IgG and IgA with oxidative stress and lipid pemxidation in the blood, suggested as mechanisms mediating tissue damage and CanL worsening. The gene mapping on chromosomal segments associated with the quantitative variation of immunoglobulin classes identified genetic signatures involved with reactive oxygen species generation, phagolysosome maturation and rupture, free iron availability, Thl/Th2 differenciation and, immunoglobulin clearance. The findings demonstrated the roles of the antibody response as resistance or susceptibility markers and mediators of CanL pathogenesis. In addition we pinpointed candidate genes as potential targets for the therapy against the damage caused by exacerbated antibody response and parasitism in VL.
- Reactivity of purified and axenic amastigotes as a source of antigens to be used in serodiagnosis of canine visceral leishmaniasis(2020) SILVA, Thais Bruna Ferreira da; SILVEIRA, Fernando Tobias; TOMOKANE, Thaise Yumie; BATISTA, Luis Fabio da Silva; NUNES, Juliana Barbosa; MATTA, Vania Lucia Ribeiro da; PASSERO, Luiz Felipe Domingues; LAURENTI, Marcia DalastraAlthough there is a great diversity of techniques and antigens used in the serodiagnosis of canine visceral leishmaniasis (CVL), total sensitivity and specificity have not yet been found. Since the use of amastigote forms in the indirect immunofluorescence assay has shown an improvement in the specificity of the test for the diagnosis of CVL, the performance of amastigotes forms of L. (L.) infantum chagasi as antigen source were evaluated in automatized ELISA WA using crude antigen of axenic amastigote and purified amastigote from spleen of hamster chronically infected comparing with ELISA using total antigen produced with promastigote forms of L. (L.) infantum chagasi. One hundred and fifteen sera from dogs with positive parasitological diagnosis by PCR were used. The animals were classified into 2 groups: symptomatic (n = 67) and asymptomatic (n = 48) animals, in accordance with the clinical signs and laboratory tests were. As control, ninety-four sera from dogs with negative parasitological diagnosis were included. No significant difference was found in sensitivity, specificity, predictive values and accuracy between ELISA using whole antigens produced with both axenic and purified amastigotes in comparison with promastigotes forms. Correlation and concordance between the three total antigens tested in ELISA was observed. According to the similar performance among antigens, data pointed out to use antigen from promastigote forms for diagnosing canine leishmaniasis, especially due the easily in the production, lower cost and the abundance of correlative literature.
- Leishmania sp identification by PCR associated with sequencing of target SSU rDNA in paraffin-embedded skin samples stored for more than 30 years(2011) LIMA, Ana Carolina S. de; ZAMPIERI, Ricardo A.; TOMOKANE, Thaise Y.; LAURENTI, Marcia D.; SILVEIRA, Fernando T.; CORBETT, Carlos E. P.; FLOETER-WINTER, Lucile M.; GOMES, Claudia M. C.Paraffin-embedded samples commonly stored at educational and research institutions constitute tissues banks for follow-up or epidemiological studies; however, the paraffin inclusion process involves the use of substances that can cause DNA degradation. In this study, a PCR protocol was applied to identify Leishmania strains in 33 paraffin-embedded skin samples of patients with American cutaneous leishmaniasis. DNA was obtained by the phenol-chloroform protocol following paraffin removal and then used in PCR or nested PCR based on the nucleotide sequence of the small subunit ribosomal RNA (SSU rDNA). The amplicons obtained were cloned and sequenced to determine the single nucleotide polymorphism that distinguishes between different Leishmania species or groups. This assay allowed to distinguish organisms belonging to the subgenus Viannia and identify L. (Leishmania) amazonensis and L. (L.) chagasi of the Leishmania subgenus. Of the 33 samples, PCR and nested PCR identified 91% of samples. After sequencing the PCR product of 26 samples, 16 were identified as L. (L.) amazonensis, the other 10 contain organisms belonging to the L. (Viannia) sub-genus. These results open a huge opportunity to study stored samples and promote relevant contributions to epidemiological studies.
- Comparative evaluation of the DPP (R) CVL rapid test for canine serodiagnosis in area of visceral leishmaniasis(2014) LAURENTI, M. D.; LEANDRO JR., M. V. de Santana; TOMOKANE, T. Y.; LUCCA, H. R. L. De; ASCHAR, M.; SOUZA, C. S. F.; SILVA, R. M.; MARCONDES, M.; MATTA, V. L. R. daWe investigated the performance of the DPP (R) canine visceral leishmaniasis (CVL) rapid test, a novel immunochromatographic assay launched by BioManguinhos (Brazil), which was recently included in the new Brazilian protocol for screening CVL in serological surveys. The present study compared the DPP (R) with the ELISA and IFA produced by BioManguinhos (Brazil) both with L. major-like antigens and with in-house tests using Leishmania infantum chagasi (in-house ELISA and in-house IFA). We analyzed the sera from clinically symptomatic (n = 47) and asymptomatic (n = 38) infected dogs from an endemic area of CVL, as well as from healthy (n = 18) dogs, in addition to the sera of dogs (n = 81) infected with other pathogens. The DPP (R) and the in-house ELISA showed a sensitivity of 90.6% and 94.1%, respectively, and specificity of 95.1% and 97.5%, respectively, and both presented cross-reactivity only with the sera of dogs with babesiosis, 44% for the DPP (R) and 22% for the in-house ELISA. The clinical groups were detected equally by the two assays. The ELISA BioManguinhos, IFA BioManguinhos, and in house-IFA showed a good sensitivity, 90.6%, 96.5% and 89.4%, respectively, but very low specificity, 77.8%, 69.1% and 65.8%, respectively, due to the high cross-reactivity with the sera from the animals harboring other pathogens. The in-house ELISA provided the highest accuracy (95.8%), followed by the DPP (R) (92.7%), ELISA BioManguinhos (84.3%), IFA BioManguinhos (83.1%), and in-house IFA (78.0%). The simultaneous use of the DPP (R) and ELISA BioManguinhos reached a sensitivity of 99.1% and 82.1% when used sequentially. In conclusion, the DPP (R) performed well as serological test for CVL, and detected both asymptomatic and symptomatic dogs in equal proportions. Although its sensitivity is not ideal yet, discarding the IFA and including the DPP (R) improved the accuracy of the new Brazilian CVL diagnostic protocol, particularly of detecting truly infected dogs. Moreover, considering the higher specificity of DPP (R) (95.1% vs 77.8%), positive predictive value (95.1% vs 81.1%) and positive likelihood value (18.3% vs 4.1%) in comparison with the ELISA BioManguinhos, the use of DPP (R) as a confirmatory test instead of a screening test is suggested.
- Evaluation of systemic immunity in atypical cutaneous leishmaniasis caused by Leishmania (L.) infantum chagasi(2022) LAURENTI, Marcia Dalastra; SOSA-OCHOA, Wilfredo; FLORES, Gabriela Venicia Araujo; PACHECO, Carmen Maria Sandoval; TOMOKANE, Thaise Yumie; OLIVEIRA, Luanda Mara da Silva; ZUNIGA, Concepcion; SILVEIRA, Fernando Tobias; CORBETT, Carlos Eduardo PereiraIn some central-American countries, Leishmania (L.) infantum chagasi infection can cause non-ulcerated or atypical cutaneous leishmaniasis (NUCL) in addition to the classic clinical form, visceral leishmaniasis (VL). Little is known about the host-parasite relationship that can contribute to the determination of one or another clinical form. The present study had the objective to evaluate the humoral and cellular immunity in the sera of individuals affected by NUCL to improve the comprehension of this atypical host-parasite interaction. Based on clinical and laboratory diagnosis, serum of 80 individuals was collected to evaluate the cytokines and immunoglobulins profile of NUCL (n = 47), VL patients (n = 5), and negative controls (n = 28). Cytokines were detected using Cytokine Bead Array (CBA) Human Th1/Th2/Th17 kit according to the manufacturer's instructions; class (IgG and IgM), and subclass of (IgG1 and IgG2) immunoglobulins was evaluated by ELISA using specific antigens. The concentration of TNF-alpha, IFN-gamma, IL-2 and IL-4 cytokines in NUCL, VL and control was present below the detection threshold of CBA kit. IL-6, IL-10 and IL-17A cytokines was lower in NUCL compared to LV patients. Regarding to immunoglobulins, NUCL patients produced 4.0 times more IgG than the control, while VL patients produced 6.6 times more; and IgM level was 1.6 times higher in NUCL and 2.6 times in VL patients compared to the control. Concerning the immunoglobulins subclass, only VL patients showed positive reaction for IgG1, and IgG2 did not show positive reaction among the groups. The results showed a weak cellular and humoral systemic immune response in NUCL patients.
- Exposure to Leishmania spp. infection and Lutzomyia spp. in individuals living in an area endemic for visceral leishmaniasis in Brazil(2020) HIRATA, Karina Yukie; OLIVEIRA SOBRINHO, Edenilson Borges de; RIGON, Lais; UTSUNOMIYA, Yuri Tani; TOMOKANE, Thaise Yumie; LAURENTI, Marcia Dalastra; MARCONDES, MaryIntroduction: This study aimed to investigate human exposure to Leishmania spp. infection and sandflies in an area endemic for the disease. Methods: The presence of antibodies specific for Leishmania spp. and saliva of Lutzomyia spp. and that of L. infantum DNA in blood were evaluated. Results: Antibodies against Leishmania spp. and sandfly saliva were observed in 20.8% and 37.7% of individuals, respectively. DNA of Leishmania spp. was amplified from the blood of one patient. Conclusions: The results suggest that Leishmania spp. infection may be underdiagnosed in this area.
- Leishmania (L.) infantum chagasi isolated from skin lesions of patients affected by non-ulcerated cutaneous leishmaniasis lead to visceral lesion in hamsters(2023) FLORES, Gabriela V. Araujo; PACHECO, Carmen M. Sandoval; FERREIRA, Aurea F.; TOMOKANE, Thaise Yumie; NUNES, Juliana B.; COLOMBO, Fabio A.; SOSA-OCHOA, Wilfredo H.; ZUNIGA, Concepcion; SILVEIRA, Fernando T.; CORBETT, Carlos E. P.; LAURENTI, Marcia D.In Central America, Leishmania (L.) infantum chagasi infection causes visceral leishmaniasis (VL) and non -ulcerated cutaneous leishmaniasis (NUCL). The aim of the present study was to evaluate the course of an experimental infection in hamsters caused by L. (L.) infantum chagasi isolated from patients affected by NUCL compared with a strain isolated from a patient with VL. Stationary phase parasites in culture were inoculated through subcutaneous and intraperitoneal routes in hamsters. Following the post-infection times, a histopath-ological study, parasite load and cytokine determination in skin from the cutaneous inoculation site and viscera were performed. Animals subcutaneously infected with the different strains did not develop macroscopic lesions at the inoculation site, and the histopathological changes in the dermis were very slight. Regarding the histo-pathological study of the viscera, we observed the portal mononuclear inflammatory infiltrate, the presence of nodules in the hepatic parenchyma and the proliferation of macrophages in the spleen, which increased over the infection course. Overall, the parasite load in the liver and spleen and in the total IgG titres in the sera of infected hamster showed an increase with the time of infection, regardless of the route of inoculation. Regarding cellular immunity, we did not observe an increase or decrease in pro-and anti-inflammatory cytokines compared to the healthy control, except for IL-10, which was evident in the infected animals. The data showed that strains iso-lated from NUCL cause visceral lesions in the hamsters regardless of the route of inoculation, and they were similar to parasites isolated from VL humans.
- Canine antibody response to Lutzomyia longipalpis saliva in endemic area of visceral leishmaniasis(2016) BATISTA, Luis Fabio da Silva; MATTA, Vania Lucia Ribeiro da; TOMOKANE, Thaise Yumie; PACHECO, Accio Duarte; SILVEIRA, Fernando Tobias; ROSSI, Claudio Nazaretian; MARCONDES, Mary; LAURENTI, Marcia DalastraIntroduction: Canine exposure to Lutzomyia longipalpis bites and the potential of Leishmania infantum transmissibility for the vector were evaluated. Methods: Immunoglobulin G (IgG) anti-Lu longipalpis saliva and-L. infantum, and blood parasite load were determined in dogs from endemic areas of visceral leishmaniasis. Results: Blood parasitism was similar between symptomatic and asymptomatic dogs. IgG anti-L. infantum was higher in symptomatic dogs, but IgG anti-Lu. longipalpis saliva was mostly observed in higher titers in asymptomatic dogs, indicating vector preference for feeding on asymptomatic dogs. Conclusions: Our data suggest a pivotal role of asymptomatic dogs in L. infantum transmission in endemic areas.