Periurethral muscle-derived mononuclear cell injection improves urethral sphincter restoration in rats.

AIMS: Investigate the effect of a novel cell-based therapy with skeletal muscle-derived mononuclear cells (SMDMCs) in a rat model of stress urinary incontinence. METHODS: Male Wistar-Kyoto rats' hind limb muscles were enzymatically dissociated, and SMDMCs were isolated without needing expansion. The cell population was characterized. Twenty female rats underwent urethrolysis. One week later, 10 rats received periurethral injection of 106 cells (SMDMC group), and 10 rats received saline injections (Saline group). Ten rats underwent sham surgery (Sham group). Four weeks after injection, animals were euthanized and the urethra was removed. The incorporation of SMDMCs in the female urethra was evaluated with fluorescence in situ hybridization for the detection of Y-chromosomes. Hematoxylin and eosin, Masson's trichrome staining, and immunohistochemistry for actin and myosin were performed. The muscle/connective tissue, actin and myosin ratios were calculated. Morphological evaluation of the urethral diameters and fractional areas of the lumen, mucosa, and muscular layer was performed. RESULTS: SMDMCs population was consistent with the presence of muscle cells, muscle satellite cells, perivascular cells, muscle progenitor cells, and endothelial cells. SMDMCs were incorporated into the urethra. A significant decrease in the muscle/connective tissue ratio was observed in the Saline group compared with the SMDMC and Sham groups. The proportions of actin and myosin were significantly decreased in the Saline group. No differences were observed in the morphometric parameters. CONCLUSIONS: SDMSC were incorporated into the rat urethra and promoted histological recovery of the damaged urethral sphincter, resulting in decreased connective tissue deposition and increased muscle content.

Endothelial differentiation of canine yolk sac cells transduced with VEGF.

Yolk sac (YS) is the site of blood-cell production where primitive erythroid cells originate and complete their maturation. YS is a source of precursor cells, however its differentiation potential and suitability for cell therapies are not well described. YS can be a cell source when neovascularization is required. This study characterized YS canine cells, transduced with VEGF, to analyze then using Immunocytochemistry, flow cytometry and real time PCR. Immunocytochemistry: positive expression for CD105, PCNA, VEGF and vWF, flow cytometry for CD105, VEGF, PCNA, OCT-4 and RT-qPCR for VEGF, CD31, CD105, PCNA and FLT - 1, indicating that these cells have characteristics of endothelial progenitor and pluripotency. After transduction, the YS cells changed their morphology and showed endothelial-like cells. We suggest, because of their cell surface phenotype as well as their capacity to differentiate into endothelial-like cells, that canine YS represents a source of cells for neovascularization therapies.

Organogenesis of the Musculoskeletal System in Horse Embryos and Early Fetuses.

Musculoskeletal system development involves heterotypical inductive interactions between tendons, muscles, and cartilage and knowledge on organogenesis is required for clarification of its function. The aim of this study was to describe the organogenesis of horse musculoskeletal system between 21 and 105 days of gestation, using detailed macroscopic and histological analyses focusing on essential developmental steps. At day 21 of gestation the skin was translucid, but epithelial condensation and fibrocartilaginous tissues were observed on day 25 of pregnancy. Smooth muscle was seen in lymphatic and blood vessel walls and the beginning of cartilaginous chondrocranium was detected at day 30 of gestation. At day 45, typical chondroblasts and chondrocytes were observed and at day 55, mandibular processes expanded toward the ventral midline of the pharynx. At day 75, muscles became thicker and muscle fibers were seen developing in carpal and metacarpal joints with the beginning of the ossification process. At day 105, major muscle groups, similar to those seen in an adult equine, were observed. The caudal area of the nasal capsule and trabecular cartilages increased in size and became ossified, developing into the ethmoid bone. The presence of nasal, frontal, parietal, and occipital bones was observed. In conclusion, novel features of equine musculoskeletal system development have been described here and each process was linked with an early musculoskeletal event. Data presented herein will facilitate a better understanding of the equine muscular system organogenesis and aid in the detection of congenital deformities. Anat Rec, 299:722-729, 2016. © 2016 Wiley Periodicals, Inc.

Aquapuncture Using Stem Cell Therapy to Treat Mdx Mice.

Duchenne muscular dystrophy (DMD) occurs due to genetic mutations that lead to absence or decrease of dystrophin protein generating progressive muscle degeneration. Cell therapy using mesenchymal stem cell (MSC) has been described as a treatment to DMD. In this work, MSC derived from deciduous teeth, called stem cells from human exfoliated deciduous teeth (SHED), were injected in acupoint as an alternative therapy to minimize muscle degeneration in twenty-two mdx mice. The treatment occurred three times with intervals of 21 days, and animals were analyzed four times: seven days prior treatment (T-7); 10 days after first treatment (T10); 10 days after second treatment (T31); and 10 days after third treatment (T52). Animals were evaluated by wire test for estimate strength and blood was collected to perform a creatinine phosphokinase analysis. After euthanasia, cranial tibial muscles were collected and submitted to histological and immunohistochemistry analyses. Treated groups presented improvement of strength and reduced creatinine phosphokinase levels. Also, a slight dystrophin increase was observed in tibial cranial muscle when aquapuncture was associated SHED. All therapies have minimized muscle degeneration, but the association of aquapuncture with SHED appears to have better effect, reducing muscle damage, suggesting a therapeutic value.

Pilot study: bone marrow stem cells as a treatment for dogs with chronic spinal cord injury.

BACKGROUND: Chronic Spinal Cord injury is a common, severe, and medically untreatable disease. Since the functional outcomes of acute and experimental chronic spinal cord injury have been shown to improve with stem cell therapy, a case study was conducted to test if the application of stem cell also regenerates chronic SCI dysfunction. Transplantation of foetal bone marrow stem cells was applied in seven dogs with chronic spinal cord injury. Magnetic resonance images and assessments of symptoms according to the Olby scale were used to diagnose the severity of injury. RESULT: All dogs improved locomotor and sensory function when examined 90 days after surgery, and showed increased movement of the hind limbs, and were able to stand upright, as well as to take small steps. Tail tone was observed in seven dogs, pain reflexes and defecation return were observed in five dogs. CONCLUSION: The transplantation of bone marrow stem may be a promising, reliable and safe treatment for chronic spinal cord injury.

Morphological and ultrastructural evaluation of the golden retriever muscular dystrophy trachea, lungs, and diaphragm muscle.

Duchenne muscular dystrophy (DMD) is a genetic disease, characterized by atrophy and muscle weakness. The respiratory failure is a common cause of early death in patients with DMD. Golden retriever muscular dystrophy (GRMD) is a canine model which has been extensively used for many advances in therapeutics applications. As the patients with DMD, the GRMD frequently died from cardiac and respiratory failure. Observing the respiratory failure in DMD is one of the major causes of mortality we aimed to describe the morphological and ultrastructural data of trachea, lungs (conductive and respiratory portion of the system), and diaphragm muscle using histological and ultrastructural analysis. The diaphragm muscle showed discontinuous fibers architecture, with different diameter; a robust perimysium inflammatory infiltrate and some muscle cells displayed central nuclei. GRMD trachea and lungs presented collagen fibers and in addition, the GRMD lungs showed higher of levels collagen fibers that could limit the alveolar ducts and alveoli distension. Therefore, the most features observed were the collagen areas and fibrosis. We suggested in this study that the collagen remodeling in the trachea, lungs, and diaphragm muscle may increase fibrosis and affect the trachea, lungs, and diaphragm muscle function that can be a major cause of respiratory failure that occur in patients with DMD.

Microscopy of the umbilical cord of rock cavies--Kerodon rupestris Wied, 1820 (Rodenta, Caviidae).

The rock cavies (Kerodon rupestris) are rodents belonging to family Caviidae, with habitat restricted to northeastern Brazil. Current studies have shown that blood cord has stem and hematopoietic cells both with a high regenerative potential, microscopic studies about the description in rodents and other mammals are scarce. The aim of this study was to describe the structural components of the umbilical cord of rock cavies. Rock cavies embryos at different stages of pregnancy were used, three in the first third, five in middle third, and six in the final third of gestation. They were obtained at the Centre for Wild Animals Multiplication of Federal Rural University of Semi-Arid. In rock cavies was observed that the number of vessels involved in the transport of maternal substances was represented by two major arteries, veins and an allantois duct. The umbilical cord was delimited by amniotic epithelium, formed by a layer of squamous cells, connective tissue with characteristic of the mucosal tissue, corresponding to the Wharton jelly (fetal mesenchyme), where elastic fibers were observed and intercellular spaces were arranged in concentric blades. The morphology observed in rock cavies is similar to that described in the literature for other mammals, including man. The rock cavies are well adapted to captivity and are docile, which makes this animal an important model for study within the field of regenerative medicine. The knowledge of the umbilical cord morphology represents a base point for using this animal as a model for culture and cell therapy.

Diaphragm morphology of guinea pig (Cavia porcellus).

The diaphragm is the main respiratory muscle. Along with other respiratory muscles, the diaphragm is responsible for the muscular contraction that generates the respiratory cycle and, as a consequence, the gaseous interchanges in the lungs. Guinea pigs (Cavia porcellus Linnaeus 1758) are largely used as experimental animals in many biology applications due to their easy management, low cost, and docile behavior. As the diaphragm exerts important effects on lung physiology and function, this study aimed at investigating the morphological characteristics of the muscle, through macroscopic, microscopic, and scanning electron microscopy to add reference data for future studies. We observed a "U"-shaped tendineous center and its morphology was similar to other mammals. These results cooperate with the descriptive and comparative anatomy of mammals, besides can be used as control data for areas of surgery and stem cells.

Characteristics of the equine embryo and fetus from days 15 to 107 of pregnancy.

In spite of numerous, substantial advances in equine reproduction, many stages of embryonic and fetal morphological development are poorly understood, with no apparent single source of comprehensive information. Hence, the objective of the present study was to provide a complete macroscopic and microscopic description of the equine embryo/fetus at various gestational ages. Thirty-four embryos/fetuses were aged based on their crown rump length (CRL), and submitted to macroscopic description, biometry, light and scanning microscopy, as well as the alizarin technique. All observed developmental changes were chronologically ordered and described. As examples of the main observed features, an accentuated cervical curvature was observed upon macroscopic examination in all specimens. In the nervous system, the encephalic fourth ventricle and the encephalic vesicles forebrain, midbrain, and hindbrain, were visualized from Day 19 (ovulation = Day 0). The thoracic and pelvic limbs were also visualized; their extremities gave rise to the hoof during development from Day 27. Development of other structures such as pigmented optical vesicle, liver, tail, cardiac area, lungs, and dermal vascularization started on Days 25, 25, 19, 19, 34, and 35, respectively. Light and scanning microscopy facilitated detailed examinations of several organs, e.g., heart, kidneys, lungs, and intestine, whereas the alizarin technique enabled visualization of ossification. Observations in this study contributed to the knowledge regarding equine embryogenesis, and included much detailed data from many specimens collected over a long developmental interval.

Arquitetura e estrutura dos rins de Gracilinanus microtarsus (Didelphimorphia: Didelphidae) (Wagner, 1842) / Architecture and structure of the kidneys of Gracilinanus microtarsus (Didelphimorphia: Didelphidae) (Wagner, 1842)

A cuíca Gracilinanus microtarsus é um marsupial sul-americano, pertencente à ordem Didelphimorphia e família Didelphidae. Foi estudada a anatomia macroscópica e microscópica dos rins de G. microtarsus. A espécie possui um par de rins localizados na região sublombar da cavidade abdominal adjacente à coluna vertebral, sendo o rim direito mais cranial que o esquerdo. Os rins possuem formato de feijão e ao corte longitudinal possuem áreas bem de nidas como córtex e medula renal, papila, cálice e pelve renal. Em G. microtarsus, o rim é unilobado, apresentando uma superfície lisa e estrutura microscópica similar à de cães e gatos.

Gracilinanus microtarsus is a South American marsupial that belongs to Didelphimorphia order and Didelphidae family. e gross and microscopic anatomy of the kidneys of G. microtarsus was studied. e species has a pair of kidneys located in the sublumbar region of the abdominal cavity, close to the spine. e right kidney is cranial to the le kidney, and both have a bean shape. In a longitudinal cut, the kidney has de ned areas, such as the renal cortex and medulla, as well as the renal papilla, calices and renal pelvis. e kidney of Gracilinanus microtarsus is classi ed as unilobed with a smooth surface and showing microscopic aspect similar to the kidneys of dogs and cats.

Correlação entre o peso, comprimento e anexos fetais de mocós (Kerodon rupestris Wied, 1820) / Correlation between weight, length and fetal membranes of rock cavies (Kerodon rupestris Wied, 1820)

Foi avaliada a existência de correlação entre diferentes anexos fetais de mocós e determinada a relação entre peso do feto e peso da placenta, a relação entre o peso do feto e comprimento do cordão umbilical e a relação entre o peso do feto e comprimento do feto. Foram utilizados anexos fetais e fetos de três, cinco e seis fêmeas, respectivamente, no terço inicial, médio e final de gestação, obtidas no Centro de Multiplicação de Animais Silvestres (CEMAS-UFERSA). Os dados foram expressos em média ± desvio-padrão, bem como valores mínimos e máximos, avaliados pelo programa estatístico GraphPad Prism Versão 6.0. Após a análise dos pressupostos paramétricos, os dados foram submetidos aos Testes de Kruskal-Wallis e Mann-Whitney, e regressão linear. A relação entre peso do feto e peso da placenta demonstrou expressão y = 33,73 + 16,38x com cálculo do coeficiente de correlação entre o peso do feto e o peso da placenta elevado e positivo, evidenciando dependência entre as variáveis (R2 = 0,7251). A relação entre o peso do feto e comprimento do cordão umbilical demonstrou expressão y = 35,64 + 25,64x e a análise de correlação entre as variáveis, positiva (R2 = 0,7201) indicando elevada dependência entre as variáveis. Quanto à relação entre o peso do feto e comprimento do feto, identificou-se uma relação do tipo y = a + bx, cuja expressão definida foi y = 1,26 + 0,41x, com análise de correlação das variáveis apresentando elevada correlação considerando-se o valor de R2 = 0,7890. As variáveis analisadas demonstram uma influência direta no desenvolvimento embrionário e fetal em mocós confirmando a correlação destes com o tamanho do feto.

We evaluated the correlation between different fetal membranes of rock cavies and determined the relationship between fetal weight and placental weight, the relationship between fetal weight and length of the umbilical cord and the relationship between fetal weight and length of the fetus. The fetuses and fetal membranes of three, five and six females respectively in the first third, middle and late gestation were used, obtained from Multiplication Center for Wild Animals of the Federal University Rural of Semiarid (CEMAS-UFERSA). The data were expressed as mean ± standard deviation, minimum and maximum values, evaluated by the statistical program GraphPad Prism version 6.0. After analysis of parametric assumptions, the data were subjected to Kruskal-Wallis’ test and Mann-Whitney’s test and linear regression. The relationship between fetal weight and placental weight showed the expression y = 33.73 + 16.38x with an estimate of the coefficient of correlation between fetal weight and placental weight high and positive, showing dependence between variables (R2 = 0.7251). The relationship between fetal weight and length of the umbilical cord showed expression y = 35.64 + 25.64x and the correlation analysis between variables, positive (R2 = 0.7201) indicating high dependence between variables. Regarding the relationship between fetal weight and length of the fetus, we identified a relationship of the type y = a + bx, whose expression was defined y = 1.26 + 0.41x, with correlation analysis of the variables showing high correlation considering the value of R2 = 0.7890. The variables analyzed demonstrate a direct influence on embryonic andfetal development in rock cavies confirming the correlation of these with the size of the fetus.

Morfologia e topografia do fígado e pâncreas de emas Rhea americana / Morphology and topography of the liver and pancreas of Rhea americana

As emas são aves ratitas nativas do continente sul americano, são consideradas aves primitivas do ponto de vista filogenético que constituem um grupo altamente especializado. Este estudo buscou caracterizar macro e microscopicamente o fígado e pâncreas de emas. O material foi coletado no Centro de Multiplicação de Animais Silvestres (CEMAS), na cidade de Mossoró-RN, Brasil, (Registro IBAMA n° 14.78912). Utilizaram-se 20 animais jovens com idade entre dois e seis meses independente do sexo. Em emas, o fígado se relacionava cranialmente com o ápice do coração, dorsalmente com os pulmões, esôfago e o proventrículo gástrico, caudalmente, com o ventrículo gástrico, o baço, o duodeno e parte do jejuno. Apresentava coloração vermelha escura e possuía apenas dois lobos, sendo o direito ligeiramente menor que o esquerdo. Histologicamente, era revestido por uma cápsula de tecido conjuntivo delgada e cada lóbulo hepático pôde ser identificado pela presença evidente de veias centrais, com muitos sinusoides comunicando-se com elas. O pâncreas, ventralmente, apresentava-se como uma fita fina, formado por um lobo dorsal e um lobo ventral. Longitudinalmente o pâncreas em emas localiza-se no mesentério dorsal desde o fígado até a flexura cranial do duodeno, mantendo-se preso às alças duodenais por ligamentos. Histologicamente, era composto por uma cápsula delgada de tecido conjuntivo denso, com discretos lóbulos separados por tecido conjuntivo capsular, compostos por estruturas tubuloalveolares e ductos. O fígado e pâncreas de emas apresentam padrão morfológico similar ao descrito para aves domésticas.

The rhea are ratites birds native of the South American continent, which are considered primitive birds from the standpoint of the phylogenetic group that is highly specialized .This study aimed to characterize gross and microscopically the liver and rhea pancreas. The material was collected in a Wild Animals Multiplication Center, Mossoró-RN, Brazil, (Registration n° 14.78912 IBAMA). It was used 20 young animals aged between two and six months independent of sex. In the rhea the liver is related to the cranial apex of the heart, dorsally with the lungs, esophagus and gastric proventriculus, caudally with the gastric ventricle, the spleen, the duodenum and part of the jejunum. It was dark red color and had only two lobes, being the right lobe slightly smaller than the left. Histologically was lined by a thin connective tissue capsule and each lobe liver could be clearly identified by the presence of central veins, with sinusoids communicating with the veins. The pancreas, ventrally, showed a thin tape formed by a dorsal lobe and a ventral lobe. Longitudinally the pancreas in rhea is located in the dorsal mesentery since the liver until the cranial duodenal flexure, remaining connected the duodenal handles by ligaments. Histologically was composed of a thin capsule of dense connective tissue, with discrete lobules separated by capsular connective tissue, composed of tubuloalveolares structures and pipelines. The liver and pancreas of rhea have morphology similar to that described for domestic poultry.

Origem e distribuição dos nervos isquiáticos do preá / Origin and distribuition of sciatic nerves of the Galea

O preá do semiárido nordestino (Galea spixii Wagler, 1831) é um roedor pertencente à família Caviidae. Pouca literatura é encontrada sobre essa espécie em relação a sua morfologia e seu comportamento ambiental e reprodutivo. Com o objetivo de entender a morfologia geral, em foco, na inervação do membro pélvico dessa espécie, neste trabalho, foi explorado o nervo isquiático, o qual é o maior de todos os nervos do organismo. Foram utilizados 10 preás (cinco machos e cinco fêmeas) que vieram a óbito por causas naturais, oriundos do Centro de Multiplicação de Animais Silvestres da Universidade Federal Rural do Semiárido (CEMAS/UFERSA). Os animais foram fixados após o óbito em solução aquosa de formaldeído 10 por cento e, após 48 horas de imersão nessa solução, foram dissecados para exposição do nervo isquiático. Dessa forma, os dados obtidos foram compilados em tabelas e expressos em desenhos esquemáticos e fotografias. Os pares de nervos isquiáticos originaram-se de raízes ventrais de L6L7S1 (70 por cento) e de L7S1S2 (30 por cento) e distribuíram-se pelos músculos glúteo profundo, bíceps femural, semitendinoso e semimembranoso.

The Galea of the semi-arid northeast (Galea spixii Wagler, 1831) is a rodent belonging to the family Caviidae. Little literature is found on this species in relation to morphology, environmental and reproductive behavior. With a view to understanding the general morphology, particularly, in the innervation of the pelvic limb of this species, ponder the sciatic nerve, which is the largest of all the nerves of the body. 10 cavies were used (five males, five females) that they had died of natural causes, originating from breeders scientific, legally licensed by IBAMA, the UFRSA, Mossoró, RN. The animals were fixed after the death in an aqueous solution of formaldehyde 10 percent and after 48 hours of immersion in it, were dissected to expose the sciatic nerve. Thus, the data were compiled in tables and expressed in drawings and photographs. The pairs of sciatic nerves were derived from ventral roots L6L7S1 (70 percent) and L7S1S2 (30 percent) and were distributed by the deep gluteal muscles, biceps femural, semitendinosus and semimembranosus muscles.