Ricardo Santana de Lima

Transplanted bone marrow cells repair heart tissue and reduce myocarditis in chronic chagasic mice.

A progressive destruction of the myocardium occurs in approximately 30% of Trypanosoma cruzi-infected individuals, causing chronic chagasic cardiomyopathy, a disease so far without effective treatment. Syngeneic bone marrow cell transplantation has been shown to cause repair and improvement of heart function in a number of studies in patients and animal models of ischemic cardiopathy. The effects of bone marrow transplant in a mouse model of chronic chagasic cardiomyopathy, in the presence of the disease causal agent, ie, the T. cruzi, are described herein. Bone marrow cells injected intravenously into chronic chagasic mice migrated to the heart and caused a significant reduction in the inflammatory infiltrates and in the interstitial fibrosis characteristics of chronic chagasic cardiomyopathy. The beneficial effects were observed up to 6 months after bone marrow cell transplantation. A massive apoptosis of myocardial inflammatory cells was observed after the therapy with bone marrow cells. Transplanted bone marrow cells obtained from chagasic mice and from normal mice had similar effects in terms of mediating chagasic heart repair. These results show that bone marrow cell transplantation is effective for treatment of chronic chagasic myocarditis and indicate that autologous bone marrow transplant may be used as an efficient therapy for patients with chronic chagasic cardiomyopathy.

Modulation of chagasic cardiomyopathy by interleukin-4: dissociation between inflammation and tissue parasitism.

Chronic chagasic cardiomyopathy (CChC) is characterized by an inflammatory reaction which may eventually lead to heart enlargement, arrythmia, and death. As described herein, interleukin-4-deficient mice mount increased specific T helper (Th) 1 immune responses when infected with Trypanosoma cruzi, as compared to wild-type mice. Interestingly, these mice had reduced parasitism and mortality and exacerbated inflammation in their hearts, demonstrating a clear dissociation between inflammation and parasite load. The modulation of these phenomena so as to maximize host and parasite survivals may depend on a fine balance between Th responses, in which a Th1 response will, on one hand, control parasitism and, on the other hand, enhance heart inflammation throughout the course of the infection.

Reversion of gene expression alterations in hearts of mice with chronic chagasic cardiomyopathy after transplantation of bone marrow cells

Chronic chagasic cardiomyopathy is a leading cause of heart failure in Latin American countries, being associated with intense inflammatory response and fibrosis. We have previously shown that bone marrow mononuclear cell (BMC) transplantation improves inflammation, fibrosis, and ventricular diameter in hearts of mice with chronic Chagas disease. Here we investigated the transcriptomic recovery induced by BMC therapy by comparing the heart transcriptomes of control, chagasic, and BMC transplanted mice. Out of the 9390 unique genes quantified in all samples, 1702 had their expression altered in chronic chagasic hearts compared to those of normal mice. Major categories of significantly upregulated genes were related to inflammation, fibrosis and immune responses, while genes involved in mitochondrion function were downregulated. When BMC-treated chagasic hearts were compared to infected mice, 96% of the alterations detected in infected hearts were restored to normal levels, although an additional 109 genes were altered by treatment. Transcriptomic recovery, a new measure that considers both resotrative and side effects of treatment, was remarkably high (84%). Immunofluorescence and morphometric analyses confirmed the effects of BMC therapy in the pattern of inflammatory-immune response and expression of adhesion molecules. In conclusion, by using large-scale gene profiling for unbiased assessment of therapeutic efficacy we demonstrate immunomodulatory effects of BMC therapy in chronic chagasic cardiomyopathy and identify potentially relevant factors involved in the pathogenesis of the disease that may provide new therapeutic targets.

Gene Expression Changes Associated with Myocarditis and Fibrosis in Hearts of Mice with Chronic Chagasic Cardiomyopathy

Chronic chagasic cardiomyopathy is a leading cause of heart failure in Latin American countries. About 30% of Trypanosoma cruzi-infected individuals develop this severe symptomatic form of the disease, characterized by intense inflammatory response accompanied by fibrosis in the heart. We performed an extensive microarray analysis of hearts from a mouse model of this disease and identified significant alterations in expression of approximately 12% of the sampled genes. Extensive up-regulations were associated with immune-inflammatory responses (chemokines, adhesion molecules, cathepsins, and major histocompatibility complex molecules) and fibrosis (extracellular matrix components, lysyl oxidase, and tissue inhibitor of metalloproteinase 1). Our results indicate potentially relevant factors involved in the pathogenesis of the disease that may provide new therapeutic targets in chronic Chagas disease.

The MHC Gene Region of Murine Hosts Influences the Differential Tissue Tropism of Infecting Trypanosoma cruzi Strains

We have previously demonstrated that both parasite genetic variability and host genetic background were important in determining the differential tissue distribution of the Col1.7G2 and JG T. cruzi monoclonal strains after artificial infections in mice. We observed that the JG strain was most prevalent in hearts of mouse lineages with the MHC haplotype H-2 d (BALB/c and DBA2), while Col1.7G2 was predominant in hearts from C57BL/6 mice, which have the H-2 b haplotype. To assess whether the MHC gene region indeed influenced tissue tropism of T. cruzi, we used the same two parasite strains to infect C57BL/6 (H-2 b) and C57BLKS/J (H-2 d) mice; the latter strain results from the introgression of DBA2 MHC region into the C57BL/6 background. We also performed ex vivo infections of cardiac explants from four congenic mice lineages with the H-2 b and H-2 d haplotypes arranged in two different genetic backgrounds: C57BLKS/J (H-2 d) versus C57BL/6 (H-2 b) and BALB/c (H-2 d) versus BALB/B10-H2b (H-2 b). In agreement with our former observations, Col1.7G2 was predominant in hearts from C57BL/6 mice (H-2 b), but we observed a clear predominance of the JG strain in hearts from C57BLKS/J animals (H-2 d). In the ex vivo experiments Col1.7G2 also prevailed in explants from H-2 b animals while no predominance of any of the strains was observed in H-2 d mice explants, regardless of the genetic background. These observations clearly demonstrate that the MHC region influences the differential tissue distribution pattern of infecting T. cruzi strains, which by its turn may be in a human infection the determinant for the clinical forms of the Chagas disease.

Granulocyte colony-stimulating factor treatment in chronic Chagas disease: preservation and improvement of cardiac structure and function

This study investigates the effects of granulocyte colony‐stimulating factor (G‐CSF) therapy in experimental chronic chagasic cardiomyopathy. Chagas disease is one of the leading causes of heart failure in Latin America and remains without an effective treatment other than cardiac transplantation. C57BL/6 mice were infected with 103 trypomastigotes of Trypanosoma cruzi, and chronic chagasic mice were treated with G‐CSF or saline (control). Evaluations following treatment were functional, immunological, and histopathological. Comparing hearts of G‐CSF‐treated mice showed reduced inflammation and fibrosis compared to saline‐treated chagasic mice. G‐CSF treatment did not alter the parasite load but caused an increase in the number of apoptotic inflammatory cells in the heart. Cardiac conductance disturbances in all infected animals improved or remained stable due to the G‐CSF treatment, whereas all of the saline‐treated mice deteriorated. The distance run on a treadmill and the exercise time were significantly greater in G‐CSF‐treated mice when compared to chagasic controls, as well as oxygen consumption (VO2), carbon dioxide production (VCO2), and respiratory exchange ration (RER) during exercise. Administration of G‐CSF in experimental cardiac ischemia had beneficial effects on cardiac structure, which were well correlated with improvements in cardiac function and whole animal performance.—Macambira, S. G., Vasconcelos, J. F., Costa, C. R. S., Klein, W., Lima, R. S., Guimaraes, P., Vidal, D. T. A., Mendez, L. C., Ribeiro‐dos‐Santos, R., Soares, M. B. P. Granulocyte colony‐stimulating factor treatment in chronic Chagas disease: preservation and improvement of cardiac structure and function. FASEB J. 23, 3843–3850 (2009). www.fasebj.org

Antioxidant activity and acute toxicity of Neoglaziovia variegata (Bromeliaceae)

Antioxidant activities of Neoglaziovia variegata were evaluated by using 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging and β-carotene-linoleic acid bleaching and was compared with ascorbic acid, butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT). The total phenolics content of the extracts was determined by the Folin-Ciocalteu method. Total flavonoid was also determined. The most significant total phenolic content was of 543.50 ± 9.38 mg of gallic acid equivalent/g for ethyl acetate extract (AcOEt), which presented the best antioxidant activity (IC 50 5.08 ± 0.20 μg/ml) for DPPH scavenging. The acute toxicity of Nv-EtOH was performed 2.0 g/kg intraperitoneally and 5.0 g/kg orally in mice. No mortality and no toxicity signs were observed, indicating low toxicity of the extract. Blood was removed after 14 days for laboratory analysis of hematological and biochemical parameters. Alterations of aspartate aminotransferase (AST) and creatinine were observed. The data obtained showed that the doses induced microscopic alterations in the liver and kidney. In conclusion, the Nv-EtOH can be considered of low toxicity. Keywords: Antioxidant activity, acute toxicity, Neoglaziovia variegata , Bromeliaceae

Terapia celular na doença de Chagas

A doenca de Chagas, que ocorre no Mexico e nas Americas Central e do Sul, continua representando um grave problema de saude publica. A prevalencia global da infeccao humana pelo Trypanosoma cruzi foi estimada em 16-18 milhoes de casos no ano de 2005, sendo corrigida para aproximadamente 28 milhoes de pessoas no ano de 2007, segundo a Organizacao Mundial de Saude. A cardiopatia chagasica cronica e a forma mais comum de cardiomiopatia nas Americas Central e do Sul e a principal causa de morte por doenca cardiovascular em areas endemicas. Ate o momento nao existe nenhum tratamento eficiente para esta doenca a nao ser o tratamento farmacologico ou o transplante cardiaco nos individuos que desenvolvem um quadro mais grave da doenca. Trabalhos atuais tem mostrado o uso de celulas-tronco de varias origens em modelos animais e humanos de doencas do coracao, como infarto do miocardio, destacando uma melhora em aspectos como neovascularizacao, regeneracao do musculo cardiaco, aumento da fracao de ejecao e melhora na qualidade de vida dos individuos tratados. Estes dados induziram os pesquisadores a investigar os efeitos terapeuticos do transplante de celulas mononucleares de medula ossea em um modelo murino e em individuos chagasicos cronicos. Esta revisao tem por objetivo mostrar os trabalhos realizados usando a terapia celular na cardiopatia chagasica cronica.

Bone marrow cells migrate to the heart and skeletal muscle and participate in tissue repair after Trypanosoma cruzi infection in mice

Infection by Trypanosoma cruzi, the aetiological agent of Chagas disease, causes an intense inflammatory reaction in several tissues, including the myocardium. We have previously shown that transplantation of bone marrow cells (BMC) ameliorates the myocarditis in a mouse model of chronic Chagas disease. We investigated the participation of BMC in lesion repair in the heart and skeletal muscle, caused by T. cruzi infection in mice. Infection with a myotropic T. cruzi strain induced an increase in the percentage of stem cells and monocytes in the peripheral blood, as well as in gene expression of chemokines SDF‐1, MCP1, 2, and 3 in the heart and skeletal muscle. To investigate the fate of BMC within the damaged tissue, chimeric mice were generated by syngeneic transplantation of green fluorescent protein (GFP+) BMC into lethally irradiated mice and infected with Trypanosoma cruzi. Migration of GFP+ BMC to the heart and skeletal muscle was observed during and after the acute phase of infection. GFP+ cardiomyocytes and endothelial cells were present in heart sections of chimeric chagasic mice. GFP+ myofibres were observed in the skeletal muscle of chimeric mice at different time points following infection. In conclusion, BMC migrate and contribute to the formation of new resident cells in the heart and skeletal muscle, which can be detected both during the acute and the chronic phase of infection. These findings reinforce the role of BMC in tissue regeneration.

Creation of a contusion injury method for skeletal muscle in rats with differing impacts

Purpose: To realize a morphological examination of the musculoskeletal tissue, assessing the effect of a contusion method for the production in rat gastrocnemius, comparing the inflammatory responses generated by different impacts. Methods: For the analysis of a contusion method, twelve female Wistar rats were distributed into four groups. The lesion was generated by 324 g of mass that was dropped from different predetermined heights for each group (30, 45, 60 and 70 cm). Results: In the analysis of musculoskeletal tissue, the response to injury varied according to the mass of the height drop onto the muscle. Only the group that was injured from 70 cm responded with uniform and severe inflammation, whereas the groups 30, 45 and 60 cm showed inflammation in some regions of the tissue with mild and moderate infiltrates. Conclusion: The method with the 324-gram mass dropped from a 70-cm height onto the gastrocnemius muscle of rats seems to be the most suitable for the production of muscle injury in these animals after 72 hours, showing an important inflammatory infiltrate.