Navegando por Autor "Novaes, Rômulo Dias"
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Item Amlodipine increases the therapeutic potential of ravuconazole upon Trypanosoma cruzi infection.(2020) Machado, Yara Almeida; Bahia, Maria Terezinha; Caldas, Ivo Santana; Mazzeti, Ana Lia; Novaes, Rômulo Dias; Vilas Boas, Breno Raimundo; Santos, Lorena Júnia de Souza; Martins Filho, Olindo Assis; Marques, Marcos José; Diniz, Lívia de FigueiredoMining existing agents that enhance the therapeutic potential of ergosterol biosynthesis inhibitors (EBI) is a promising approach to improve Chagas disease chemotherapy. In this study, we evaluated the effect of ravuconazole, an EBI, combined with amlodipine, a calcium channel blocker, upon Trypanosoma cruzi experimental infection. In vitro assays confirmed the trypanocidal activity of both compounds in monotherapy and demonstrated an additive effect (sum of the fractional inhibitory concentration [FIC] 0.5) of the combined treatment without additional toxicity to host cells. In vivo experiments, using a murine model of the T. cruzi Y strain in a short-term protocol, demonstrated that amlodipine, although lacking trypanocidal activity, dramatically increased the antiparasitic activity of underdosing ravuconazole regimens. Additional analysis using long-term treatment (20 days) showed that parasitemia relapse until 60 days after treatment was significatively lower in mice treated with the combination (4 out of 14 mice) than ravuconazole monotherapy (10 out of 14 mice), even in the presence of immunosuppressant pressure. Furthermore, the combined therapy was well tolerated and protected the mice from mortality. The treatments also impacted on the cellular and humoral immune response of infected animals, inducing a reduction of serum cytokine levels in all ravuconazole-treated mice. Our findings demonstrate that amlodipine is efficacious in enhancing the antiparasitic activity of ravuconazole in an experimental model of T. cruzi infection and indicates a potential strategy to be explored in Chagas disease treatment.Item Basal and β-Adrenergic cardiomyocytes contractility dysfunction induced by dietary protein restriction is associated with downregulation of SERCA2a expression and disturbance of endoplasmic reticulum Ca2+ regulation in rats.(2014) Penitente, Arlete Rita; Novaes, Rômulo Dias; Silva, Marcelo Eustáquio; Silva, Márcia Ferreira da; Quintão Júnior, Judson Fonseca; Guatimosim, Silvia; Cruz, Jader dos Santos; Chianca Júnior, Deoclécio Alves; Natali, Antônio José; Neves, Clóvis AndradeBackground: The mechanisms responsible for the cardiac dysfunction associated with dietary protein restriction (PR) are poorly understood. Thus, this study was designed to evaluate the effects of PR on calcium kinetics, basal and β-adrenergic contractility in murine ventricular cardiomyocytes. Methods: After breastfeeding male Fisher rats were distributed into a control group (CG, n = 20) and a protein-restricted group (PRG, n = 20), receiving isocaloric diets for 35 days containing 15% and 6% protein, respectively. Biometric and hemodynamic variables were measured. After euthanasia left ventricles (LV) were collected for histopathological evaluation, SERCA2a expression, cardiomyocytes contractility and Ca2+ sparks analysis. Results: PRG animals showed reduced general growth, increased heart rate and arterial pressure. These animals presented extracellular matrix expansion and disorganization, cardiomyocytes hypotrophy, reduced amplitudes of shortening and maximum velocity of contraction and relaxation at baseline and after β-adrenergic stimulation. Reduced SERCA2a expression as well as higher frequency and lower amplitude of Ca2+ sparks were observed in PRG cardiomyocytes. Conclusion: The observations reveal that protein restriction induces marked myocardial morphofunctional damage. The pathological changes of cardiomyocyte mechanics suggest the potential involvement of the β-adrenergic system, which is possibly associated with changes in SERCA2a expression and disturbances in Ca2+ intracellular kinetics.Item Concomitant exercise training attenuates the cardioprotective effects of pharmacological therapy in a murine model of acute infectious myocarditis.(2019) Mendonça, Andréa Aparecida dos Santos; Gonçalves, Reggiani Vilela; Silva, Thaiany Goulart de Souza e; Maldonado, Izabel Regina dos Santos Costa; Silva, André Talvani Pedrosa da; Natali, Antônio José; Novaes, Rômulo DiasWhen administered alone, preinfection exercise training and benznidazole-based chemotherapy induce cardioprotection in Chagas disease. However, the effect of concomitant exercise and benznidazole treatment is unknown. We investigated whether exercise and specific chemotherapy could interact to modulate parasitemia, inflammation, redox status and heart damage in a murine model of T. cruzi infection. Wistar rats were randomized into an uninfected control group (CNT) and four groups infected with T. cruzi: sedentary untreated (SUN) and treated (STR), and trained untreated (TUN) and treated (TTR). Running training was administered 5 days/ week for 4 weeks. Treated animals concomitantly received 100 mg/kg/day benznidazole. Heart inflammation and reactive damage were not detected in CNT animals. Compared to SUN, TUN animals presented increased levels of parasitemia, myocarditis, nitric oxide, hydrogen peroxide, protein carbonyl, malondialdehyde, cytokines (IFN-γ, TNF-α, IL-4, IL-6, IL-10 and IL-17), catalase, superoxide dismutase and glutathione reductase activity, as well as reduced heart non-protein antioxidant levels (P < 0.05). TTR animals exhibited higher levels of parasitemia, myocarditis, hydrogen peroxide, malondialdehyde, IFN-γ, TNF-α and IL-6 than STR animals (P < 0.05), which showed the lowest levels of all analyzed parameters compared to the other groups (P < 0.05). Our findings indicate that exercise aggravates acute infection. When concomitantly administered with benznidazole, exercise training impaired parasitic control and chemotherapy-induced cardioprotection in T. cruzi-infected rats. Considering that exercise training and T. cruzi infection constitute independent metabolic challenges, the negative effects of concomitant treatment are potentially related to the overlapping oxidative and immunoinflammatory demands of exercise and the infection itself.Item Could pre-infection exercise training improve the efficacy of specific antiparasitic chemotherapy for Chagas disease?(2019) Santos, Elda Gonçalves dos; Gonçalves, Reggiani Vilela; Silva, Thaiany Goulart de Souza e; Maldonado, Izabel Regina dos Santos Costa; Santos, Eliziária Cardoso dos; Silva, André Talvani Pedrosa da; Natali, Antônio José; Novaes, Rômulo DiasConsidering a potential exercise-drug interaction, we investigated whether exercise training could improve the efficacy of specific antiparasitic chemotherapy in a rodent model of Chagas disease. Wistar rats were randomized into five groups: sedentary and uninfected (CT); sedentary and infected (SI); sedentary, infected and treated (SIT); trained and infected (TI); trained, infected and treated (TIT). After 9-weeks running training, the animals were infected with T. cruzi and followed up for 4 weeks, receiving 100 mg kg−1 day−1 benznidazole. No evidence of myocarditis was observed in CT animals. TI animals exhibited reduced parasitemia, myocarditis, and reactive tissue damage compared to SI animals, in addition to increased IFN-γ, IL-4, IL-10, heart non-protein antioxidant (NPA) levels and glutathione-s transferase activity (P < 0.05). The CT, SIT and TIT groups presented similar reductions in parasitemia, cytokines (IFN-γ, TNF-α, IL-4, IL-10, IL-17 and MCP-1), inflammatory infiltrate, oxidative heart damage and antioxidant enzymes activity compared to SI and TI animals, as well as reduced heart microstructural remodeling (P < 0.05). By modulating heart inflammation and redox metabolism, exercise training exerts a protective effect against T. cruzi infection in rats. However, the antiparasitic and cardioprotective effects of benznidazole chemotherapy are more pronounced, determining similar endpoints in sedentary and trained T. cruzi-infected rats.Item Effects of Trypanosoma cruzi infection on myocardial morphology, single cardiomyocyte contractile function and exercise tolerance in rats.(2011) Novaes, Rômulo Dias; Penitente, Arlete Rita; Gonçalves, Reggiani Vilela; Silva, André Talvani Pedrosa da; Neves, Clóvis Andrade; Maldonado, Izabel Regina dos Santos Costa; Natali, Antônio JoséThe aim of this study was to investigate the effects of Trypanosoma cruzi (T. cruzi) infection on myocardial morphology, single cardiomyocyte contractile function and exercise tolerance in rats. Adult Wistar rats were randomized into control (n = 14) and infected (n = 14) groups. Infected animals were inoculated with T. cruzi Y strain (300,000 trypomastigotes ⁄ 50 g body weight). After 9 weeks, the animals were subjected to a treadmill running protocol. Then, the right atrium (RA) and left ventricle (LV) were removed for morphological and cell contractile evaluation. The infected animals exhibited a significant reduction in distance travelled, total time to fatigue and workload. In addition, these animals had hypertrophy, increased myocardial cellularity, and an increase in the proportion of collagen and blood vessels. RA and LV myocytes from infected animals showed marked contractile dysfunction under basal conditions and a reduced contractile response to b-adrenergic stimulation. The workload of infected animals was correlated closely with the amplitude of cell shortening of RA and LV myocytes. T. cruzi infection influenced the myocardial morphology and the mechanical properties of RA and LV single myocytes negatively and reduced exercise tolerance. Single cardiomyocyte contractile dysfunction could constitute an additional mechanism of cardiac impairment and reduced exercise tolerance in this infection.Item Elemental mapping of cardiac tissue by scanning electron microscopy and energy dispersive X-ray spectroscopy: proof of principle in Chaga’s disease myocarditis model.(2013) Novaes, Rômulo Dias; Maldonado, Izabel Regina dos Santos Costa; Natali, Antônio José; Neves, Clóvis Andrade; Silva, André Talvani Pedrosa daItem In vitro and in silico evaluation of the schistosomicidal activity of eugenol derivatives using biochemical, molecular, and morphological tools.(2022) Souza, Isabella Maria Monteiro de; Novaes, Rômulo Dias; Gonçalves, Reggiani Vilela; Fialho, Felipe Leonardo Bley; Carvalho, Diogo Teixeira; Souza, Thiago Belarmino de; Dias, Danielle Ferreira; Lavorato, Stefânia Neiva; Souza, Raquel Lopes Martins; Marques, Marcos José; Castro, Aline PereiraBackground: Eugenol shows both antibacterial and antiparasitic activities, suggesting that it might be evaluated as an option for the treatment of praziquantel-resistant schistosome. Methods: The in vitro activities of three eugenol derivatives (FB1, FB4 and FB9) on adult worms from Schistosoma mansoni were examined by fluorescence and scanning electron microscopy to analyze effects on the excretory system and integument damage, respectively. Biochemical tests with verapamil (a calcium channel antagonist) and ouabain (a Na+ /K+-ATPase pump inhibitor) were used to characterize eugenol derivative interactions with calcium channels and the Na+/K+-ATPase, while in silico analysis identified potential Na+/K+-ATPase binding sites. Results: The compounds showed effective doses (ED50) of 0.324 mM (FB1), 0.167 mM (FB4), and 0.340 mM (FB9). In addition, FB4 (0.322 mM), which showed the lowest ED50, ED90 and ED100 (p < 0.05), caused the most damage to the excretory system and integument, according to both fluorescence and scanning electron microscopy analysis. The death of adult worms was delayed by ouabain treatment plus FB1 (192 versus 72 hours) and FB9 (192 versus 168 hours), but the response to FB4 was the same in the presence or absence of ouabain. Besides, no changes were noted when all of the eugenol derivatives were combined with verapamil. Moreover, FB1 and FB9 inhibited Na+/K+-ATPase activity according to in silico analysis but FB4 did not show a time- dependent relationship and may act on targets other than the parasite Na+/K+-ATPase. Conclusion: Eugenol derivatives, mainly FB4 when compared to FB1 and FB9, seem to act more effectively on the integument of adult S. mansoni worms.Item In vitro tripanocidal effect of 1,8-dioxooctahydroxanthenes (xanthenodiones) and tetraketones and improvement of cardiac parameters in vivo.(2020) Menezes, Ana Paula de Jesus; Silva, Milene Lopes da; Pereira, Wagner Luiz; Costa, Guilherme de Paula; Horta, Aline Luciano; Mendonça, Andréa Aparecida dos Santos; Carneiro, Ana Cláudia Alvarenga; Souza, Débora Maria Soares de; Novaes, Rômulo Dias; Teixeira, Róbson Ricardo; Silva, André Talvani Pedrosa daObjective: Trypanosoma cruzi infection affects millions of people worldwide, and the drugs available for its treatment have limited efficacy. 1,8-Dioxooctahydroxanthenes and tetraketones are compounds with important biological applications. The aim of this study was to assess the trypanocidal and inflammatory activities of nine 1,8-dioxooctahydroxanthenes (1–9) and three tetraketones (10–12). Methods and results: By in vitro killing assay, three compounds were able to eliminate CL TdTomato expressing strain of T. cruzi, 9 (IC50 = 30.65mM), 10 (IC50 = 14.11mM), and 11 (IC50 = 26.43mM). However, only 9 was not toxic to Vero cells. Next, to evaluate the in vivo antitrypanosomal and immunological efficacy of 9, Swiss mice were infected with the Y and CL strains of T. cruzi and treated for 10 days with 50 mg/kg of 9. This compound reduced the cardiac inflammatory infiltration in animals infected with both strains. Rank's ligand (RankL), CCL2, and interferon (IFN)-g were measured in the cardiac tissue homogenate of the Y-strain-infected animals, and no interference of 9 was observed. However, compound 9 increased the RankL and interleukin (IL)-10 levels in CL-infected mice. No hepatic and renal toxicity was observed. Conclusion: Our findings showed that 1,8-dioxooctahydroxanthene has antiparasitic effect and ameliorates the cardiac inflammatory parameters related to T. cruzi infection.Item Integrative transcriptome analysis of SARS‐CoV‐2 human‐infected cells combined with deep learning algorithms identifes two potential cellular targets for the treatment of coronavirus disease.(2023) Gonçalves, Ricardo Lemes; Souza, Gabriel Augusto Pires de; Terceti, Mateus de Souza; Castro, Renato Fróes Goulart de; Silva, Breno de Mello; Novaes, Rômulo Dias; Malaquias, Luiz Cosme Cotta; Coelho, Luiz Felipe LeomilSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) quickly spread worldwide, leading coronavirus disease 2019 (COVID-19) to hit pandemic level less than 4 months after the frst ofcial cases. Hence, the search for drugs and vaccines that could prevent or treat infections by SARS-CoV-2 began, intending to reduce a possible collapse of health systems. After 2 years, eforts to fnd therapies to treat COVID-19 continue. However, there is still much to be understood about the virus’ pathology. Tools such as transcriptomics have been used to understand the impact of SARS-CoV-2 on dif- ferent cells isolated from various tissues, leaving datasets in the databases that integrate genes and diferentially expressed pathways during SARS-CoV-2 infection. After retrieving transcriptome datasets from diferent human cells infected with SARS-CoV-2 available in the database, we performed an integrative analysis associated with deep learning algorithms to determine diferentially expressed targets mainly after infection. The targets found represented a fructose transporter (GLUT5) and a component of proteasome 26s. These targets were then molecularly modeled, followed by molecular docking that identifed potential inhibitors for both structures. Once the inhibition of structures that have the expression increased by the virus can represent a strategy for reducing the viral replication by selecting infected cells, associating these bioinformatics tools, therefore, can be helpful in the screening of molecules being tested for new uses, saving fnancial resources, time, and making a personalized screening for each infectious disease.Item Modulation of inflammatory and oxidative status by exercise attenuates cardiac morphofunctional remodeling in experimental Chagas cardiomyopathy.(2016) Novaes, Rômulo Dias; Gonçalves, Reggiani Vilela; Penitente, Arlete Rita; Bozi, Luiz Henrique Marchesi; Neves, Clóvis Andrade; Maldonado, Izabel Regina dos Santos Costa; Natali, Antônio José; Silva, André Talvani Pedrosa daAims: The rational basis that explains the benefits of exercise therapy on Chagas cardiomyopathy (ChC) is poorly understood. This study investigated the impact of an exercise programon exercise performance, heart parasitism, immunoinflammatory response, fibrogenesis, oxidative damage, and cardiomyocytes contractility in experimental ChC. Main methods: Wistar rats were subjected to a 9-week treadmill running training and challenged with Trypanosoma cruzi. Control animals remained sedentary. Physical and metabolic performance, cardiac morphology, cytokines, chemokines, nitric oxide, oxidative tissue damage, cardiomyocyte morphology and contractility were analyzed. Key findings: Exercise training was efficient to improve physical performance and anaerobic threshold in trained animals. By increasing cardiac and serum levels of cytokines (TNF-α, IFN-γ, and IL-6), chemokines (MCP-1 and CX3CL1), the myocardial activity catalase and superoxide dismutase, and reducing lipid and protein oxidation in cardiac tissue, exercise training seem to be a beneficial strategy to mitigate the progression and severity of Chagas-associated cardiomyopathy. Significance: The protective adaptations to the host triggered by exercise training contributed to reduce cardiac parasitism, inflammation, fibrosis and cardiomyocytes atrophy. Although exercise training does not affect nitric oxide levels in cardiac tissue from infected animals, this strategy enhanced the efficiency of endogenous antioxidant mechanisms, restricting oxidative tissue damage with positive repercussions to cardiomyocytes biomechanics in rats.Item Naringin accelerates the regression of pre-neoplastic lesions and the colorectal structural reorganization in a murine model of chemical carcinogenesis.(2014) Sequetto, Priscila Lima; Oliveira, Tânia Toledo de; Maldonado, Izabel Regina dos Santos Costa; Augusto, Luís Eugênio Franklin; Mello, Vanessa Joia de; Pizziolo, Virgínia Ramos; Almeida, Márcia Rogéria de; Silva, Marcelo Eustáquio; Novaes, Rômulo DiasThe aim of this study was to investigate the effect of Naringin on pre-neoplastic colorectal lesions induced by chemical carcinogen in rats. Female Wistar rats weighing 130.8 ± 27.1 g received weekly one subcutaneous injection of 1,2-dimethylhydrazine (DMH, 20 mg/kg) for 10 weeks. The animals were divided into 5 groups with 6 animals in each group. Group 1: 0.9% saline; Group 2: DMH + 0.9% saline; Group 3: DMH + Naringin (10 mg/kg); Group 4: DMH + Naringin (100 mg/kg); Group 5: DMH + Naringin (200 mg/kg). G2 and G3 showed a significant increase in ACF number, AgNOR/nucleus and mitosis compared to G1. G4 and G5 presented a significant reduction in these parameters compared to G2. The number of cells producing acidic and neutral mucins, red blood cells and the level of antioxidant minerals, such as copper, magnesium, selenium and zinc, were significantly reduced in G2 and G3, but similar in G4 and G5 compared to G1. Naringin, especially at 200 mg/kg, was effective in reducing the number of pre-neoplastic lesions in rats exposed to DMH. Some of these effects may be due to reduction in cellular proliferation and tissue levels of iron together with the recovery of antioxidant mineral levels induced by this flavonoid.Item Nonsteroidal anti-inflammatory is more effective than anti-oxidant therapy in counteracting oxidative/nitrosative stress and heart disease in T. cruzi-infected mice.(2017) Novaes, Rômulo Dias; Santos, Eliziária Cardoso dos; Fialho, Maria do Carmo Queiroz; Gonçalves, Wagner Gonzaga; Sequetto, Priscila Lima; Silva, André Talvani Pedrosa da; Gonçalves, Reggiani VilelaWe compared the relevance of ibuprofen, vitamins C and E to control oxidative/nitrosative stress and heart disease in mice infected by Trypanosoma cruzi. Swiss mice were randomized into five groups: control, uninfected; infected without treatment; and infected treated with vitamins C, E or ibuprofen. Animals were inoculated with 2000 trypomastigote forms of T. cruzi. After 20 days, infected mice presented reduced vitamin C and E tissue levels, high cytokines (interferon gamma, tumour necrosis factor-α, interleukin 10 and chemokine ligand 2), prostaglandin F2α (PGF2α) and nitric oxide (NO) cardiac production, intense myocarditis and reactive tissue damage, which was directly correlated with the intensity of the inflammatory infiltrate and the degree of pathological cardiac remodelling. Vitamins C and E supplementation were irrelevant to counteract reactive tissue damage and myocarditis in infected animals. Conversely, ibuprofen reduced tissue levels of cytokines, PGF2α and NO, as well as lipid and protein oxidation, antioxidant enzyme activity and the cardiac damage, without interfering with heart parasitism. Our results do not support the applicability of vitamin C and E supplementation in the management of acute Chagas cardiomyopathy. By controlling the inflammatory infiltrate, anti-inflammatory-based therapy proved to be a more rational strategy than a direct antioxidant therapy in attenuating oxidative/nitrosative stress and cardiac damage.Item Parasitaemia and parasitic load are limited targets of the aetiological treatment to control the progression of cardiac fibrosis and chronic cardiomyopathy in Trypanosoma cruzi-infected dogs.(2019) Caldas, Ivo Santana; Menezes, Ana Paula de Jesus; Diniz, Lívia de Figueiredo; Nascimento, Alvaro Fernando da Silva do; Novaes, Rômulo Dias; Caldas, Sérgio; Bahia, Maria TerezinhaIt is still unclear whether the progression of acute to chronic Chagas cardiomyopathy is predominantly associated with the limited efficacy of aetiological chemotherapy, or with the pharmacological resistance profiles and pathogenicity of specific Trypanosoma cruzi strains. Thus, we tested the hypothesis that parasitic load could be a limited target of aetiological chemotherapy to prevent chronic cardiomyopathy in dogs infected by different T. cruzi strains. Animals were infected with benznidazole-susceptible (Berenice-78) and -resistant (VL-10 and AAS) strains of T. cruzi. A quantitative real-time PCR strategy was developed to comparatively quantify the parasite load of the three different strains using a single standard curve. For dogs infected with the VL-10 strain, benznidazole treatment reduced cardiac parasitism during the acute phase of infection. However, similar parasite load and collagen deposition were detected in the myocardium of treated and untreated animals in the chronic phase of the infection. In animals infected with the AAS strain, benznidazole reduced parasite load, myocarditis and type III collagen deposition in the acute phase. However, increased type III collagen deposition was verified in the chronic phase. Dogs infected with the Berenice-78 strain showed a parasitological cure and no evidence of myocardial fibrosis. Parasitic load and cardiac fibrosis presented no correlation in acute or chronic phases of T. cruzi infection. Our findings in a canine model of Chagas disease suggest that parasite burden is a limited predictor for disease progression after treatment and show that benznidazole, although not inducing parasitological cure, is able to prevent total fibrosis in the early stages of infection, as well as complete prevention of cardiac damage when it eliminates parasites at the onset of infection.Item Parasite control and skeletal myositis in Trypanosoma cruzi-infected and exercised rats.(2017) Novaes, Rômulo Dias; Gonçalves, Reggiani Vilela; Penitente, Arlete Rita; Cupertino, Marli do Carmo; Maldonado, Izabel Regina dos Santos Costa; Silva, André Talvani Pedrosa da; Natali, Antônio JoséNon-pharmacological strategies have been rarely described in the treatment of infectious diseases.Although exercise training has been recently incorporated in the clinical management of Chagas disease,the rationale basis that supports this indication is poorly understood. Thus, we investigated the effectof an aerobic exercise on the parasitism, inflammation and oxidative tissue damage in a murine modelof Trypanosoma cruzi-induced skeletal myositis. Wistar rats were randomized into four groups: trainednot infected (TNI) and infected (TI), sedentary not infected (SNI) and infected (SI). A running trainingprogram was administered 5 days/week for 9 weeks. Then, infected animals were inoculated with T. cruziand followed up for another 9 weeks. Exercise training induced beneficial adaptations by increasing timeto fatigue and lactate threshold in TNI and TI animals. SI animals presented higher parasitemia, skele-tal muscle parasitism, cell necrosis, leukocyte infiltration, cytokines levels, reactive oxygen species andnitric oxide production, thiobarbituric acid reactive substances, carbonyl proteins, myosin heavy chain Idepletion, and increased catalase (CAT) and superoxide dismutase (SOD) activities. Beyond attenuationin all these variables, TI animals showed reduced TNF- , CCL-2/MCP-1 and CX3CL1, and increased IL-10muscle levels. Furthermore, these animals presented higher CAT and SOD activities and reduced lipidand protein oxidation. Taken together, our findings indicated that exercise training induced a protectivephenotype in T. cruzi-infected mice, enhancing host defenses against the parasite and attenuating thepathological remodeling associated with skeletal myositis, aspects potentially associated to an improvedimmunological and redox balance in infected animals.Item Protein restriction after weaning modifies the calcium kinetics and induces cardiomyocyte contractile dysfunction in rats.(2013) Penitente, Arlete Rita; Novaes, Rômulo Dias; Chianca Júnior, Deoclécio Alves; Silva, Márcia Ferreira da; Silva, Marcelo Eustáquio; Souza, Aline Maria Arlindo de; Fonseca, Silvia Carolina Guatimosim; Almeida, Pedro William Machado de; Alves, Márcia Netto Magalhães; Felix, Leonardo Bonato; Neves, Clóvis Andrade; Natali, Antônio JoséProtein restriction (PR) is associated with cardiovascular diseases. The purpose of this study was to investigate the effects on single ventricular cardiomyocyte contractile function of a short-term PR after weaning. Male Fischer rats that were 28 days old were randomly divided into a control group (CG, n = 16) and a protein-restricted group (PRG, n = 16). After weaning, CG and PRG animals received isocaloric diets containing 15 and 6% protein, respectively, for 35 days. Biometric parameters were then measured, and the hearts were removed for the analysis of contractile function and calcium transient in isolated cardiomyocytes of the left ventricule (LV), and the quantification of calcium and collagen fibers in LV myocardium. PRG animals had lower body weight (BW) and LV weight (LVW), an increased LVW to BW ratio and a higher proportion of collagen fibers than CG animals. PRG animals exhibited reduced tissue levels of calcium, reduced the length, width and volume of cardiomyocytes and their sarcomere length compared to CG animals. Cardiomyocytes from PRG animals had a lower amplitude of shortening, a slower time to the peak of shortening and a longer time to half-relaxation than those from the CG. Cardiomyocytes from PRG animals also presented a lower peak of calcium transient and a longer calcium transient decay time than CG animals. Taken together, the results indicate that short-term PR after weaning induces a marked structural remodeling of the myocardium parenchyma and stroma that coexists with contractile dysfunctions in single LV cardiomyocytes of rats, which is probably associated with pathological changes of the intracellular calcium kinetics, rather than inadequate available amounts of this mineral in cardiac tissue.Item Pyrrolidine dithiocarbamate reduces alloxan-induced kidney damage by decreasing nox4, inducible nitric oxide synthase, and metalloproteinase-2.(2020) Pereira, Bruna Pinheiro; Valle, Gabriel Tavares do; Salles, Bruno César Côrrea; Costa, Karla Cristinne Mancini; Angelo, Marilene Lopes; Torres, Larissa Helena Lobo; Novaes, Rômulo Dias; Ruginsk, Silvia Graciela; Tirapelli, Carlos Renato; Paula, Fernanda Borges de Araújo; Ceron, Carla SperoniWe examined the effect of the NFκB inhibitor pyrrolidine-1-carbodithioic acid (PDTC) on inducible nitric oxide synthase (iNOS), matrix metalloproteinase-2 (MMP-2) activity, and oxidative and inflammatory kidney damage in alloxan-induced diabetes. Two weeks after diabetes induction (alloxan-130 mg/kg), control and diabetic rats received PDTC (100 mg/kg) or vehicle for 8 weeks. Body weight, glycemia, urea, and creatinine were measured. Kidney changes were measured in hematoxylin/eosin sections and ED1 by immunohistochemistry. Kidney thiobarbituric acid reactive substances (TBARS), superoxide anion (O2−), and nitrate/nitrite (NOx) levels, and catalase and superoxide dismutase (SOD) activities were analyzed. Also, kidney nox4 and iNOS expression, and NFkB nuclear translocation were measured by western blot, and MMP-2 by zymography. Glycemia and urea increased in alloxan rats, which were not modified by PDTC treatment. However, PDTC attenuated kidney structural alterations and macrophage infiltration in diabetic rats. While diabetes increased both TBARS and O2 − levels, PDTC treatment reduced TBARS in diabetic and O2 − in control kidneys. A decrease in NOx levels was found in diabetic kidneys, which was prevented by PDTC. Diabetes reduced catalase activity, and PDTC increased catalase and SOD activities in both control and diabetic kidneys. PDTC treatment reduced MMP-2 activity and iNOS and p65 NFκB nuclear expression found increased in diabetic kidneys. Our results show that the NFκB inhibitor PDTC reduces renal damage through reduction of Nox4, iNOS, macrophages, and MMP-2 in the alloxan-induced diabetic model. These findings suggest that PDTC inhibits alloxan kidney damage via antioxidative and anti-inflammatory mechanisms.Item The role of IL‐10 in immune responses against Pseudomonas aeruginosa during acute lung infection.(2020) Belo, Valéria Aparecida; Pereira, Jéssica Assis; Souza, Sara Franchin Duarte de; Tana, Fernanda de Lima; Pereira, Bruna Pinheiro; Lopes, Débora de Oliveira; Ceron, Carla Speroni; Novaes, Rômulo Dias; Corsetti, Patrícia Paiva; Almeida, Leonardo Augusto dePseudomonas aeruginosa is considered an opportunistic pathogen of great clinical importance. The clearance of this bac- terium occurs through recognition of the pathogen by innate immune system receptors, leading to a lung infammatory response. However, this response must be controlled via immunoregulatory pathways. In this study, we evaluate the role of endogenous murine IL-10 after acute infection with the virulent strain P. aeruginosa PA14. To assess the role of IL-10, intratracheal infection with the PA14 strain was performed in C57BL/6 or IL-10 KO mice. The PA14 strain was recovered in both types of animals, although IL-10 KO mice presented a higher number of viable bacteria in the lung when compared to the C57BL/6 group. Histopathological and stereological analyses showed that IL-10 KO mice had higher tissue damage and infammatory infltrate when compared to control animals. The activity of MMP-9 but not MMP-2, as well as IL-6 and TNF-α expression, were augmented in the lungs of infected animals and was much more evident in IL-10 KO animals when compared to the other analyzed groups. This work indicates that endogenous IL-10 control P. aeruginosa infection, the expression of pro-infammatory genes, MMP-9 activity and histopathological processes of the infectious process in question.Item Trypanosoma cruzi infection alters glucose metabolism at rest and during exercise without modifying the morphology of pancreatic islets in rats.(2012) Novaes, Rômulo Dias; Gonçalves, Reggiani Vilela; Penitente, Arlete Rita; Silva, André Talvani Pedrosa da; Neves, Clóvis Andrade; Natali, Antônio José; Maldonado, Izabel Regina dos Santos CostaThis study investigated the effects of Trypanosoma cruzi infection on pancreatic morphology and glucose metabolism at rest and during exercise. Wistar rats were randomized into control (CG = 10) and infected (IG = 10) groups. The IG animals were inoculated with T. cruzi Y strain (300,000 trypomastigotes/50 g). After 9 weeks, the animals were subjected to glucose (OGTT) and insulin (ITT) tolerance tests and a treadmill running protocol. Blood glucose, lactate and time to fatigue were determined. After euthanasia, the pancreases were removed for morphological and biochemical analyses. The IG presented abnormal glucose kinetics in OGTT and a similar glucose curve in ITT compared to the CG. During the exercise test, the IG showed anticipation of time to fatigue. At the point of fatigue, no difference was found in blood glucose and lactate between the groups. There was a significant correlation between lactate levels and the time to fatigue. The IG presented marked pancreatic inflammation, fibrosis and protein oxidation. The number of _ cells in the IG animals was not reduced. T. cruzi infection impaired pancreas morphology and glucose metabolism at rest and during exercise in rats, which could constitute an additional mechanism in the induction of exercise intolerance in Chagas’ disease.Item Trypanosoma cruzi infection induces morphological reorganization of the myocardium parenchyma and stroma, and modifies the mechanical properties of atrial and ventricular cardiomyocytes in rats.(2013) Novaes, Rômulo Dias; Penitente, Arlete Rita; Gonçalves, Reggiani Vilela; Silva, André Talvani Pedrosa da; Peluzio, Maria do Carmo Gouveia; Neves, Clóvis Andrade; Natali, Antônio José; Maldonado, Izabel Regina dos Santos CostaBackground: This study investigates morphofunctional adaptations of the heart stroma and parenchyma in rats that are chronically infected with Trypanosoma cruzi. Methods: Four-month-old male Wistar rats were randomized into control (n=14) and infected (n=14) groups. Infected animals were inoculated with T. cruzi Y strain. After 9 weeks, the animals were euthanized, and the right atrium (RA) and left ventricle (LV) were removed for biochemical, stereological, and cardiomyocyte mechanical analyses. Results: Infected animals presented cardiomyocyte atrophy and myocardial fibrosis. For these animals, the total volume, length, surface area, and cross-sectional area of cardiomyocytes were significantly reduced, and the total interstitial and collagen volumes were significantly increased in the RA and LV compared to the controls. The total volume and length of blood vessels were significantly increased in the LV, and the total blood vessel surface area was significantly higher in the RA of infected animals. RA and LV cardiomyocytes from infected animals exhibited a significant reduction in cell shortening (43.02% and 24.98%, respectively), prolongation of the time to the peak of contraction (17.09%) and the time to half relaxation (23.68%) compared to non-infected animals. Lipid hydroperoxides, but not mineral concentrations, were significantly increased in the RA and LV from infected animals, showing an inverse correlation with cell shortening. Conclusions: T. cruzi infection induces global structural remodeling of the RA and LV in rats. This remodeling coexists with cardiomyocyte contractility dysfunction, which is possibly related to the abnormal organization of the myocardial stroma and increased cellular lipid peroxidation.Item Uso de fluorescência em um método de disector modificado para estimar o número de miócitos no tecido cardíaco.(2012) Novaes, Rômulo Dias; Penitente, Arlete Rita; Silva, André Talvani Pedrosa da; Natali, Antônio José; Neves, Clóvis Andrade; Maldonado, Izabel Regina dos Santos CostaFundamento: Métodos convencionais de disector atualmente requerem consideráveis custos financeiros, técnicos e operacionais para estimar o número de células, incluindo cardiomiócitos, em uma área de 3D. Objetivo: Usar a microscopia de fluorescência em um método de disector modificado para determinar o número de miócitos no tecido cardíaco em condições normais e patológicas. Métodos: O estudo empregou ratos Wistar machos com quatro meses de idade e peso de 366,25 ± 88,21 g randomizados em grupos controles (GC, n = 8) e infectados (GI, n = 8). Os animais do GI foram inoculados com cepa Y de T. cruzi (300.000 tripomastigotas/50 g). Após oito semanas, os animais foram pesados e sacrificados. Os Ventrículos Esquerdos (VE) foram removidos para análise estereológica da densidade numérica de cardiomiócitos (Nv [c]) e o número total dessas células no VE (N [c]). Esses parâmetros foram estimados usando um disector fluorescente (DF) e comparados com os métodos convencionais de disector óptico (DO) e disector físico (DFi). Resultados: Em ambos os métodos de disector, os animais do GI apresentaram queda significativa de Nv[c] e N[c] em comparação com os animais do GC (p > 0,05). Uma correlação forte, igual ou superior a 96%, foi obtida entre DF, DO e DFi. Conclusão: O método DF parece ser igualmente confiável para determinar Nv[c] e N[c] em condições normais e patológicas, apresentando algumas vantagens em relação aos métodos convencionais de disector: redução de cortes histológicos e imagens na análise estereológica, redução do tempo de análise das imagens, a construção de DF em microscópios simples, utilizando o modo de epifluorescência, distinção de planos de disector em ampliações inferiores.