Glauco Jonas Lemos Santos

Himatanthus drasticus (Apocynaceae) latex reduces oxidative stress and modulates CD4+, CD8+, FoxP3+ and HSP-60+ expressions in Sarcoma 180-bearing mice

ETHNOPHARMACOLOGICAL RELEVANCE In Brazil, latex of Himatanthus drasticus is used to treat inflammation, wound healing and cancer. The present study evaluated the antitumoral potential of H. drasticus latex (HdCL) in Sarcoma 180-bearing mice (S180). MATERIALS AND METHODS HdCL was obtained in Crato-CE, Brazil. Qualitative phytochemicals assays, nuclear magnetic resonance (NMR) and microbiological analyzes were performed. Swiss mice were divided into six groups, according to tumor forms: 1) ascitic model, GI (Control; 0.9% saline), GII (S180asc) and GIII (S180asc/HdCL/14 days); 2) solid model, GIV (Control; 0.9% saline), GV (S180sol) and GVI (S180sol/HdCL/10 days). HdCL and 0.9% saline were administered at 0.2 mL, SID, by gavage, for 10 or 14 days. For ascitic model, 0.5 mL of S180 suspension (4×106 cells/mL) was inoculated intraperitoneally and for solid model, cells were inoculated subcutaneously (25 µL) on the right hind paw of mice. Blood samples were collected for hematological and oxidative stress evaluation. Thickness, volume and weight of paws were measured in solid model. After euthanasia, spleen, liver and kidney were collected in order to assess the relative organ weight. Tissue fragments of paws and popliteal lymph nodes (PLN) were analyzed by H&E and CD4+, CD8+, HSP-60+ and Foxp3+ immunohistochemistry. RESULTS HdCL presented milky aspect and pinkish supernatant. Phenols, flavonols, flavanones, free steroids and cinnamoyl derivatives of lupeol, α-amyrin and β-amyrin were detected at the phytochemistry analysis. HdCL did not alter the relative weight of organs, hematological parameters and volume of ascitic fluid recovered. In solid model, HdCL reduced (P < 0.05) paw volume, but did not altered thickness, paw weight and histological parameters. S180sol induced necrosis, metastasis and destruction of bone, cartilage and muscles. Bleeding, vessel congestion and oncocytes were observed in PLN. In paw, HdCL did not alter FoxP3+ and HSP-60+ expressions but reduced the CD4+ and CD8+ expressions, while at PLN, HdCL reduced the expressions of all markers. HdCL decreased (P < 0.05) serum levels of malondialdehyde in ascitic model. CONCLUSIONS Treatment with HdCL reduced oxidative damage and modulated the expressions of CD4+, CD8+, FoxP3+and HSP-60+ in S180 solid tumor model, which can be associated to the presence of triterpenes, such as α-amyrin, β-amyrin and lupeol cinnamate. Present data emphasizes the importance of immune system in cancer and highlights the evaluation of the pharmacological properties of plants used by population as phytoterapics.

CD4+, CD8+, FoxP3+ and HSP60+ Expressions in Cellular Infiltrate of Canine Mammary Carcinoma in Mixed Tumor

Background: Cancer is a complex process that receive many influences of the tumor microenvironment. The participation of immune system cells and proteins in tumor microenvironment is not yet completely understood. Thus, the aim of this study was to evaluate the infiltrate cellular, subpopulations of T-lymphocytes and HSP60 of canine mammary carcinoma in mixed tumor (CMCMT). Materials, Methods & Results: Female dogs (n = 20) were selected after Canine mammary tumor (CMT) diagnosis and data were achieved throughout clinical-pathological information. Clinical staging was evaluated and tumor biopsies were processed by histology and cellular infiltrate was performed according criteria and grade. Survival curve were generated by Kaplan-Meier and the lymphocytic infiltrate were compared by Log-Rank followed Chi-Square χ². For immunolabeling it was used anti-CD4, anti-CD8, anti-FoxP3 and HSP60 monoclonal antibodies and were attributed scores from 0 to 3. Clinical-pathological relationship was analyzed using Spearman correlation. This study was approved by the Committee for Ethics in Research using Animals (CEUA-UECE), protocol 12247080-2. Our data showed a mean age of 9.3 years-old, the size of tumors presented more than 5 cm (50%), which were located in inguinal mammary glands (70%), and CMTs shows I (70%) and II (30%) grade. The cellular infiltrate was distributed both in peri and intratumoral regions, dispersed multifocally with moderate intensity and lymphocytes were the major populations found into tumors (n = 826 ± 220). In relationship to cellular infiltrate with CMT grade it was observed that lymphocytes (ρ = 0.28) and plasma cells (ρ = 0.22) showed a slight positive correlation, and an opposed negative correlation of neutrophils (ρ = -0.1) and macrophages (ρ = -0.38). CMT presents moderate lymphocytic infiltrate (< 800 lymphocytes), shows higher (P = 0.01) survival rates as compared to intense lymphocytic infiltrate (≥ 800 lymphocytes). FoxP3+ showed lower intensity while CD4+ and CD8+ expression were concentrated surrounding of lymphocytic infiltrate tumor region. HSP60+ was observed in the inflammatory and tumor cells. Discussion: Our data are according to a greater risk to the development of breast tumor in old bitches, not castrated and before or after puberty, as well as the use of contraceptives based on progesterone and estrogen. In relation to size of tumor, these findings reinforce that there is a relationship of tumor size with a higher malignancy grade and with a worse prognosis. The predominant tumor location was in the inguinal breasts that is attributed to the high activity of the mammary glands to hormonal stimuli. CMT with low clinical staging are associated with greater overall survival of affected bitches. In relation to tumor microenvironment, it has been reported that heterogeneous populations of the immune system cells often infiltrate the mammary tumors, whose lymphocytes are the main cells. It is suggested that tumor lymphocytosis may be necessary for malignant behavior of the tumor microenvironment. On the other hand, macrophages and neutrophils play an important role that may favor or inhibit the tumor cells development in the tumor microenvironment. In our work, CD4, CD8 and FoxP3 labeling were distributed in peri and intratumoral regions, and consequently, these markers can be used as prognostic for CMT, as well as being a potential target for anticancer therapies. This is the first work that presents results about the participation of HSP60 in CMT, however this data needs further investigation. HSP60 participates as a potent activator of the immune system through its peptides and other HSP types were studied in mammary carcinomas in bitches and presenting results that indicate the association of these proteins with the carcinogenesis process.

Adesão leucocitária e MDA sérico em cães naturalmente infectados por Leishmania infantum

The present study evaluated the oxidative stress and leukocyte adhesion (LA) in dogs naturally infected by Leishmania infantum. Healthy dogs (HD = 10) and dogs affected by canine visceral leishmaniasis (CVL) in symptomatic form (SD= 10) were previously submitted to indirect immunofluorescence reaction (IIF), enzyme-linked immunosorbent assay (ELISA) and detection of parasites in bone marrow aspirates. Serum was used to assess malondialdeid (MDA) by thiobarbituric acid reactive species (TBARS) assay and AL was determined by nylon column method using whole blood stored in EDTA, citrate and heparin. AL data was expressed as percentage and MDA data as mean ± standard deviation, both submitted to the unpaired student’s T test (p < 0.05). Heparin samples showed higher levels of AL in CS group (55.62 %, p < 0.05) when compared with EDTA and citrate (10.46% and 5.28%). Citrate and EDTA inhibit AL in healthy and sick dogs, while heparin preserved AL in both groups. Neutrophil proportion in heparin samples were lower (85% to 67%, p <0.05) compared with citrate and EDTA, which in turn remained practically unchanged (83% to 80%). MDA levels were higher in SD (0.0117μM ± 0.002) when compared to HD (0.0057μM ± 0.001) (p <0.05). These data support the conclusion that CVL induces oxidative stress enhance and leukocyte adhesion increase, indicating systemic inflammatory response. The choice of anticoagulant is an important decision for implementing LA assays.

Inflammatory response evaluation in naturally infected dogs by Leishmania infantum

Canine visceral leishmaniasis (CVL) is manifested by tissue damage in dogs infected with Leishmania infantum and is presented as asymptomatic or symptomatic disease. Different clinical pictures are associated with the host response, which is triggered by the inflammatory response and specific immune response. This study aimed to evaluate the inflammatory response in L. infantum naturally infected dogs. Dogs (n=12) were divided into two groups: seropositive (n=6) and seronegative (n=6). Blood samples were collected for determination of total leukocyte and neutrophil counts and serum levels of C-reactive protein (CRP), fibrinogen (FIB) and thiobarbituric acid reactive species (TBARS). Data were expressed as mean and standard deviation and to compare the groups Student t test was used (p < 0.05). Seropositive animals showed elevated neutrophils count (p < 0.05), CRP levels above the reference values for the species (p < 0.05) and greater levels of TBARS (p < 0.05) compared to Revista Brasileira de Higiene e Sanidade Animal Brazilian Journal of Hygiene and Animal Sanity ISSN: 1981-2965

Biometric and organs macroscopic evaluation of wildlife southern caracara (Caracara plancus, MILLER, 1777) captured at Fortaleza Metropolitan Region, Ceará

Samuel Salgado Tavares 2 , Mariana Andrioli Pinheiro 3 , Isaac Mourão Xavier 3 , Matheus Silva Feitosa 3 , Pedro Ernesto de Araujo Cunha 3 , Lidia Sampaio Batista 3 , Ligia Santos Oliveira 3 , Rodrigo Fonseca de Medeiros Guedes 4 , Belarmino Eugênio Lopes Neto 5 , Glauco Jonas Lemos Santos 5 , Marcio Gomes Alencar Araripe 5 , Diana Célia Sousa Nunes-Pinheiro 6 _________________________________________________________________________________________________________________