Rosa Cabral

Ringo, a Golden Retriever Muscular Dystrophy (GRMD) dog with absent dystrophin but normal strength

The closest model to human Duchenne Muscular Dystrophy (DMD) is the Golden Retriever Muscular Dystrophy (GRMD) dog, which carries a point mutation in the splice acceptor site in intron 6 of the orthologe X-linked dystrophin gene, leading to the absence of protein in the muscles. These dogs present clinical signs within the first weeks of life involving the limbs as well as masticatory muscles. Diaphragmatic and intercostal muscles impairment leads to progressive respiratory failure. Death occurs from bronchopneumonia and cardiac arrest, usually before 2 years of age. Here, we report the case of Ringo, an exceptional GRMD dog showing an unusually mild course. Currently, at age 4 years and 10 months he is able to run, jump and open doors while standing on his rear paws. He was also able to breed naturally, which apparently has never been reported before. Ringo is descendant of Beth, a GRMD female carrier donated by Dr. Joe Kornegay (University of North Carolina, USA), and all affected descendants carry the same original mutation. The diagnosis in all dogs was established right after birth through DNA genotyping and elevated serum creatine kinase (CK). At birth, Ringo’s serum CK level was increased 10-fold as compared to his three normal sibs. One affected brother had a comparable serum CK while the other had a 20-fold increase. At 15 days serum CK in the affected dogs was 4to 5-fold higher than in normal siblings. Pedigree analysis (Fig. 1) revealed that Ringo had two affected brothers from the same litter. One of them died at 2 weeks of age. The other one is still alive but with a

Collection and evaluation of epididymal sperm in captive agoutis (Dasyprocta aguti)

The objective was to establish a protocol for the collection and evaluation of epididymal sperm in agoutis. Eight males (1-2 y old) underwent left orchidectomy and epididymal sperma were collected by retrograde flush. Average values were flush volume 32 μL, pH 6.9, sperm concentration 748 x 10(6) sperm/mL, with motility 86.5% and vigor 4.6. Viable sperm were present in all flush samples; 66% of sperm were alive, and 41.9% of sperm responded positively to the hypoosmotic test (using distilled water). There were 21.1% morphologically abnormal sperm, of which 2.0 and 19.1% were primary and secondary defects, respectively. The acrosome was intact in 99.5% of sperm. The sperm head was 4.89 ± 0.41 μm long and 3.13 ± 0.35 μm wide, with an area of 13.01 ± 2.01 μm(2). Midpieces were 5.33 ± 0.44 μm long and 0.98 ± 0.13 wide, sperm tails were 29.91 ± 2.29 μm, and overall sperm length was 40.12 ± 2.44 μm. In conclusion, epididymal sperm collection from agoutis was satisfactory; the collected sperm has the potential to be stored, facilitating development of other reproductive biotechnologies for this species.

Myocardial homing after intrapericardial infusion of Bone Marrow Mononuclear Cells

Erika Branco1, Emerson Ticona Fioretto2, Rosa Cabral3, Carlos Alberto Sarmento Palmera4, Guilherme Buzon Gregores4, Angelo Joao Stopiglia4, Paulo Cesar Maiorka4, Pedro Alves Lemos5, Carlos Campos5, Celso Takimura5, Jose Antonio Franchini Ramires5, Maria Angelica Miglino4 Faculdade de Medicina Veterinaria da Universidade Federal Rural da Amazonia – UFRA1, Belem, PA; Universidade Federal do Sergipe – UFS2, Aracaju, SE; Universidade Feral do Piaui – UFPI3, Teresina, PI; Faculdade de Medicina Veterinaria e Zootecnia FMVZ/USP4, Sao Paulo, SP; Instituto do Coracao – Incor5, Sao Paulo, SP, Brasil

Cell Therapy for Fibrotic Interstitial Pulmonary Disease: Experimental Study

Parte superior do formulário Digite um texto ou endereço de um site ou traduza um documento. The aim of this study is to evaluate the histological changes in lung parenchyma of pigs affected by interstitial lung disease induced after the infusion of bone marrow mononuclear cells (BMMCs). Ten female swines were submitted to pulmonary fibrosis induced by a single dose of intratracheal bleomicine sulfate. Animals were arranged into two groups: Group 1: induced‐disease control and Group 2: cell therapy using BMMCs. Both groups were clinically evaluated for 180 days. High‐resolution computed tomography (HRCT) was performed at 90 and 180 days. BMMC sampling was performed in cell therapy group at 90 days. Euthanasia was performed, and samples were collected for histology and immunohistochemistry. The 90‐days HRCT demonstrated typical interstitial lesions in pulmonary parenchyma similarly to human disease. The 180‐days HRCT in Group 1 demonstrated advanced stages of the disease when compared with Group 2. Immunohistochemistry analysis suggests the presence of pre‐existent vessels and neoformed vessels as well as predominant young cells in the injured parenchyma of Group 2. Immunohistochemistry analysis suggests that cell therapy would promote a reconstructive response. Histology and HRCT analysis suggest a positive application of swine as a model for a bleomicine inducing of fibrotic interstitial pulmonary disease. Microsc. Res. Tech., 2011.

Kidney injury and cell therapy: Preclinical study

The aim of this study is to show histological and immunofluorescence analysis of renal parenchyma of agoutis affected by gentamicin‐induced renal disease after the infusion of bone marrow mononuclear cells (BMMC) stained with Hoechst®. Nine agoutis males were divided into three groups: Test group (TG): renal disease by gentamicin induced (n = 3), cell therapy group (CTG): renal disease by gentamicin induced and BMMC infusion (n = 3), and control group (CG): nonrenal disease and BMMC infusion (n = 3). TG and CTG were submitted to the protocol of renal disease induction using weekly application of gentamicin sulfate for 4 months. CG and CTG received a 1 × 108 BMMC stained with Hoechst and were euthanized for kidney examination 21 days after BMMC injection and samples were collected for histology and immunofluorescence analysis. Histological analysis demonstrated typical interstitial lesions in kidney similarly to human disease, as tubular necrosis, glomerular destruction, atrophy tubular, fibrotic areas, and collagen deposition. We conclude that histological analysis suggest a positive application of agoutis as a model for a gentamicin inducing of kidney disease, beyond the immunofluorescence analysis suggest a significant migration of BMMC to sites of renal injury in CTG. Microsc. Res. Tech., 2012.

Anatomy and morphometric aspects of the trachea of Saimiri sciureus Linnaeus, 1758: knowledge for emergency procedures

The aim of this work is to study the morphological characteristics of the trachea of Saimiri sciureus through quantification and measurement of the cartilaginous rings, providing information to facilitate the election of more appropriate endotracheal tube, laryngeal mask or tracheostomy tube for anesthetic and emergency procedures, as it is a species of Neotropical primates most commonly used as biological models, and little is known about their morphology. Nine animals were investigated, being 4 adults and 5 young acquired from the Centro Nacional de Primatas (National Primate Center - CENP) - Ananindeua - PA, which died from natural causes and then fixed in aqueous buffered formalin 10%. Saimiri sciureus trachea comprises an average of 32.8 incomplete rings and an average length of 3.74 cm in young animals, while in adults it demonstrated an average of 30.25 rings and average length of 3.67 cm. The shape of the light and its proportion varied along the trachea. Endotracheal tube with a diameter the 2.0 - 2.5 mm, laryngeal mask number 1.0 or tracheostomy tube neonatal Shiley number 3.0, can be placed in animals weighing 600 g - 1.2 Kg. Given the great importance of the species studied, which is widely used as a biological model, the detailing on the morphology and morphometry of tracheal animal studies provides new approaches needed in respiratory emergency, as well as, facilitates the development of future anesthetic protocols.

Bone marrow cells of swine: Collection and separation

Bone marrow is a source of stem cells for greater and easier access, which is widely studied as a provider of hematopoietic and mesenchymal cells for various purposes, mainly therapeutic by the advances in research involving cell therapy. The swine is an animal species commonly used in the pursuit of development of experimental models. Thus, this study aimed to standardize protocol for collection and separation of bone marrow in swines, since this species is widely used as experimental models for various diseases. Twelve animals were used, which underwent bone marrow puncture with access from the iliac crest and cell separation by density gradient followed by a viability test with an average of 98% of viable cells. Given our results, we can ensure the swine as an excellent model for obtaining and isolation of mononuclear cells from bone marrow, stimulating several studies addressing the field of cell therapy. Microsc. Res. Tech., 2012.

Padrão de distribuição de células mononucleares de medula óssea em tecido cardíaco sadio por diferentes vias de infusão

INTRODUCAO: A medicina regenerativa tem ganho grande importância nos ultimos anos em decorrencia da possibilidade de certas celulas se diferenciarem em linhagens celulares distintas e, assim, reconstruirem o tecido lesado. As celulas-tronco tem despontado como forma alternativa de tratamento para doencas pela sua capacidade de diferenciacao nos mais de 100 tipos de tecido. A medula ossea contem celulas-tronco adultas, hematopoeticas e mesenquimais, que auxiliam na limitacao do remodelamento cardiaco. METODO: Foram utilizados 9 caes com peso entre 25 kg e 30 kg, divididos em tres grupos: intracoronaria, intramiocardica-transendocardica e retrograda venosa. Celulas mononucleares da medula ossea foram coletadas por densidade Ficoll, marcadas com fluorocromo Hoechst e infundidas nas diferentes vias citadas anteriormente. RESULTADOS: Observou-se que, na via intracoronaria, a distribuicao das celulas foi homogenea do epicardio ao endocardio, em atrios e ventriculos direito e esquerdo; na via intramiocardica-transendocardica, as celulas ficaram mais restritas ao local da infusao, com consequente maior numero em ventriculo esquerdo; e na via retrograda venosa, as celulas encontravam-se com padrao de distribuicao semelhante ao da via intramiocardica, contudo sem que as mesmas chegassem a regiao endocardica. Com relacao a quantidade de celulas distribuidas pelo tecido, foi observado que havia diferenca significativa (P < 0,01), quando analisada a interacao via de infusao vs. regiao. CONCLUSAO: Podemos concluir que existe a possibilidade de empregar tecnicas distintas de entrega de celulas para diferentes doencas cardiacas.

Chorion biopsy in mongrel dogs

With the great development of the gestational studies in all of the species, we noticed the necessity of adaptations of these techniques for prenatal diagnosis in dogs. Based on this, we studied the feasibility of chorion biopsy guided by ultrasound. Our results demonstrated accuracy on the sex determination being 2 males and 12 females, as well as it would be possible to identify chromosome alteration due to the quality of samplings. Sex determination was accomplished with the identification of Y gene chromosomes in PCR technique. After the collection, fragments were prepared for light microscopy studies and revealed fetal chorion tissue, blood colloid and erythrocyte. In the whole material we found hemosiderin impregnations due to the hemolysis and to the residue of blood of the placental marginal hematomes. The submitted female dogs to this technique demonstrated normal puppy births without death.

Morphological aspects of buffaloes (Bubalus bubalis) umbilical cord

Bufalo e uma importante fonte de recurso nos rebanhos animais, apresentando uma grande participacao na agropecuaria, provendo leite, carne e forca de trabalho. O Cordao umbilical e responsavel pela troca de nutrientes materno-fetais durante a gestacao, e suas alteracoes podem comprometer o desenvolvimento fetal. Nos investigamos dez uteros gravidicos de bufalos de racas cruzadas em fases diferentes de gestacao. O periodo de gestacao e a idade fetal foram determinados pelo comprimento apice sacral, aplicando formulas previamente estabelecidas. Posteriormente mediu-se o comprimento do cordao umbilical. A rede vascular do cordao umbilical e anastomoses foram observadas por injecao ou latex de neoprene. O cordao umbilical foi estudado a partir de cortes histologicos, corados por HE, picrossirius, azul de Toluidina, orceina e reacao histoquimica de PAS. O cordao umbilical de bufalos e formado por duas arterias centrais, ducto alantoide e duas veias perifericas e apresentam forma de ampulheta. A parede da arteria umbilical e composta por grande quantidade de fibras colagenas e elasticas, fibroblastos e um grande numero de vasa vasorum. O ducto alantoide fica alocado entre as arterias e apresenta um grande numero de pequenos vasos nutritivos. Os vasos nutritivos devem ser cuidadosamente identificados para evitar-se confundi-los com vasa vasorum. O comprimento medio do cabo de cordao umbilical dos bufalos era 11.8cm (minimo de 6.8cm e maximo de 17.4cm).