Isabel Rodrigues Rosado

Comparison of the osteogenic potential of mesenchymal stem cells from the bone marrow and adipose tissue of young dogs

BackgroundThe aim of the present study was to compare the osteogenic potential of mesenchymal stem cells extracted from the bone marrow (BM-MSCs) and adipose tissue (AD-MSCs) of young dogs. The following parameters were assessed: dimethyl thiazolyl diphenyl tetrazolium (MTT) conversion, alkaline phosphatase (ALP) activity, collagen and mineralised matrix synthesis, and the expressions of osterix, bone sialoprotein (BSP), and osteocalcin (OC).ResultsMTT conversion was greater in BM-MSCs compared to AD-MSCs after 14 and 21 days of differentiation; ALP activity was greater in differentiated AD-MSCs on day 7; collagen synthesis was greater in BM-MSCs on days 14 and 21; the percentage of mineralized area per field was greater in BM-MSCs compared to AD-MSCs; osterix expression was greater in BM-MSCs in days 14 and 21, and BSP and OC expression levels were greater in BM-MSCs at all the investigation time-points.ConclusionsIt was concluded that the osteogenic potential was greater in BM-MSCs than AD-MSCs when extracted from young dogs.

Orthopedic implant of a polyhydroxybutyrate (PHB) and hydroxyapatite composite in cats

The purpose of this study was to evaluate the tissue response to a 70% polyhydroxybutyrate and 30% hydroxyapatite composite in the form of a bone implant, placed intracortically in the distal metaphyseal of the right femur, and subcutaneous implants in cats. Samples of the composite were implanted subcutaneously in the dorsolumbar region and the distal metaphyseal region of the right femur of the animals. The study used 12 neutered adult mixed breed cats, weighing an average of 3.5 kg. The cats were randomly divided into three groups: GI, GII and GIII, according to the length of the assessment period. The assessments of their subcutaneous and bone tissues were performed at 15, 30 and 45 days and at 30, 60 and 90 days, respectively. The subcutaneous and bone reactions to the composites were characterized by granulomatous inflammation with a predominance of macrophages and giant cells. The results showed that the composites triggered a chronic local inflammatory response, despite their clinical acceptance.

Emprego experimental da placa de compósito poli-hidroxibutirado/hidroxiapatita na fixação femoral em gatos

The composite of polyhydroxybutyrate (PHB) 70% and hydroxyapatite (HA) 30% was evaluated as plate for bone fixation in cats. The employed composite plates presented six orifices and measured 60 x 10mm, length and width, respectively, with thickness ranging from 3 to 5mm according to the region. The composite plate was used in the fixation of femoral osteotomy in four cats, in a total of six interventions. There were ruptures in five plates (83.3%) until day 4 and in one plate (16.7%) on the day 21, when it was possible to observe an exuberant osseous callus. The result of the plate deployment in the cat showed that the composite does not have sufficient strength to be used as plate of femoral fixation in cats.

Head trauma as a possible cause of central diabetes insipidus in a cat

A 13-month-old female domestic shorthair cat presented with a 10-month history of polyuria and polydipsia that began after having been hit by a car. Neurological examination revealed visual deficits and an absent bilateral menace response. Hematological and serum biochemical analyses were within reference values, but hyposthenuria was identified. Failure to concentrate urine during the water deprivation test followed by an increase in urine specific gravity after administration of synthetic antidiuretic hormone (ADH) suggested a diagnosis of central diabetes insipidus. Subcutaneous or oral administration of synthetic ADH was effective in central diabetes insipidus treatment during the 19-month follow-up.

Association of riluzole and dantrolene improves significant recovery after acute spinal cord injury in rats

BACKGROUND CONTEXT Damage to the spinal cord can result in irreversible impairment or complete loss of motor, sensory, and autonomic functions. Riluzole and dantrolene have been shown to provide neuroprotection by reducing neuronal apoptosis after brain and spinal cord injury (SCI) in several animal models of neurologic disorders. As these drugs protect the injured spinal cord through different mechanisms, we investigated the cumulative effects of riluzole and dantrolene. PURPOSE This study aimed to investigate the neuroprotective efficacy of the combined administration of riluzole and dantrolene in experimental thoracic SCI. STUDY DESIGN Twenty-nine Wistar rats were laminectomized at T12 and divided in five groups. Rats in GI (n=6) underwent laminectomy alone and were treated with placebo. Rats in GII (n=6) underwent laminectomy followed by SCI and were treated with placebo. Rats in GIII (n=5) underwent laminectomy followed by SCI and were treated with riluzole and placebo 15 minutes and 1 hour after laminectomy, respectively. Rats in GIV (n=6) underwent laminectomy followed by SCI and were treated with placebo and dantrolene 15 minutes and 1 hour after laminectomy, respectively. Rats in GV (n=6) underwent laminectomy followed by SCI and were treated with riluzole and dantrolene 15 minutes and 1 hour after laminectomy, respectively. A compressive trauma was performed to induce SCI. METHODS Behavioral testing of hind limb function was performed using the Basso Beattie Bresnahan locomotor rating scale, which revealed significant recovery in the group treated with the association of riluzole and dantrolene compared with other groups. After euthanasia, the spinal cord was evaluated using light microscopy and immunochemistry with anti-NeuN and transferase dUTP nick-end-labeling (TUNEL) staining. RESULTS Animals treated with the association of riluzole and dantrolene showed a larger number of NeuN-positive neurons adjacent to the epicenter of injury (p≤.05). Furthermore, the TUNEL staining was similar between animals treated with riluzole and dantrolene and those that did not receive spinal cord trauma (p>.05). CONCLUSIONS These results showed that riluzole and dantrolene have a synergistic effect in neuroprotection after traumatic SCI by decreasing apoptotic cell death.

Efeito do produto iônico do biovidro 60S na diferenciação osteogênica de células-tronco mesenquimais do tecido adiposo de cães

The aim was to evaluate the ionic product of 60S bioglass (BV60S) in osteogenic differentiation of mesenchymal stem cells from adipose tissue (ADMSCs) in dogs. ADMSCs were differentiated without the ionic product (OST) and with the ionic product (PI-OST) for 7, 14 and 21 days. We evaluated the MTT, alkaline phosphatase (ALP), collagen mineralization and expressions of osterix (OSX), bone sialoprotein (BSP), osteonectin (ON) and osteocalcin (OC). The PI-OST group had a lower MTT conversion to 7days and higher conversion at 21 days. The ALP activity was higher in the OST group at 14 and 21 days. Collagen synthesis was higher in the OST group at 7 and 21 days. A higher mineralized area in the PI-OST group was observed at all times. There were no differences in expressions of OSX and OC between groups. We observed increased expression of BSP in the PI-OST group at 14 and 21 days. The expression of ON was higher in the OST group at 14 days. It was concluded that the ionic product of BV60S promotes in vitro osteogenesis of MSC-AD from dogs.

Systemic effects induced by intralesional injection of ω-conotoxin MVIIC after spinal cord injury in rats

BackgroundCalcium channel blockers such as conotoxins have shown a great potential to reduce brain and spinal cord injury. MVIIC neuroprotective effects analyzed in in vitro models of brain and spinal cord ischemia suggest a potential role of this toxin in preventing injury after spinal cord trauma. However, previous clinical studies with MVIIC demonstrated that clinical side effects might limit the usefulness of this drug and there is no research on its systemic effects. Therefore, the present study aimed to investigate the potential toxic effects of MVIIC on organs and to evaluate clinical and blood profiles of rats submitted to spinal cord injury and treated with this marine toxin. Rats were treated with placebo or MVIIC (at doses of 15, 30, 60 or 120 pmol) intralesionally following spinal cord injury. Seven days after the toxin administration, kidney, brain, lung, heart, liver, adrenal, muscles, pancreas, spleen, stomach, and intestine were histopathologically investigated. In addition, blood samples collected from the rats were tested for any hematologic or biochemical changes.ResultsThe clinical, hematologic and biochemical evaluation revealed no significant abnormalities in all groups, even in high doses. There was no significant alteration in organs, except for degenerative changes in kidneys at a dose of 120 pmol.ConclusionsThese findings suggest that MVIIC at 15, 30 and 60 pmol are safe for intralesional administration after spinal cord injury and could be further investigated in relation to its neuroprotective effects. However, 120 pmol doses of MVIIC may provoke adverse effects on kidney tissue.

Conotoxin MVIIA improves cell viability and antioxidant system after spinal cord injury in rats

This study evaluates whether intrathecal MVIIA injection after spinal cord injury (SCI) elicits neuroprotective effects. The test rats were randomly distributed into six groups— sham, placebo, MVIIA 2.5 μM, MVIIA 5 μM, MVIIA 10 μM, and MVIIA 20 μM—and were administered the treatment four hours after SCI. After the optimal MVIIA dose (MVIIA 10 μM) was defined, the best time for application, one or four hours, was analyzed. Locomotor hind limb function and side effects were assessed. Forty-eight hours after the injury and immediately after euthanasia, spinal cord segments were removed from the test rats. Cell viability, reactive oxygen species, lipid peroxidation, and glutamate release were investigated. To examine the MVIIA mechanism of action, the gene expressions of pro-apoptotic (Bax, nNOS, and caspase-3, -8, -9, -12) and anti-apoptotic (Bcl-xl) factors in the spinal cord tissue samples were determined by real-time PCR, and the activities of antioxidant enzymes were also investigated. Application of intrathecal MVIIA 10 μM four hours after SCI prompted a neuroprotective effect: neuronal death decreased (22.46%), oxidative stress diminished, pro-apoptotic factors (Bax, nNOS, and caspase-3, -8) were expressed to a lesser extent, and mitochondrial viability as well as anti-apoptotic factor (Bcl-xl) expression increased. These results suggested that MVIIA provided neuroprotection through antioxidant effects. Indeed, superoxide dismutase (188.41%), and glutathione peroxidase (199.96%), reductase (193.86%), and transferase (175.93%) expressions increased. Therefore, intrathecal MVIIA (MVIIA 10 μM, 4 h) application has neuroprotective potential, and the possible mechanisms are related to antioxidant agent modulation and to intrinsic and extrinsic apoptotic pathways.

Luxação patelar medial grau iv em gato: Relato de caso

Este trabalho tem como objetivo relatar um caso de luxacao patelar medial grau IV em gato, abordando aspectos clinicos e cirurgico. A luxacao de patela caracteriza-se por sua saida do sulco troclear. Comumente essa patologia e congenita e se nao tratada precocemente pode gerar graves alteracoes osteomusculares. A luxacao patelar nao e comum em gatos especialmente o grau IV. Foi atendido um gato com historico de claudicacao, apoio parcial e atrofia do membro pelvico esquerdo. No exame fisico foi observada a patela luxada medialmente de forma irredutivel, incapacidade de extensao completa do membro e desvio medial da tuberosidade tibial. O exame radiografico mostrou a patela luxada medialmente e desvios angulares de femur distal e tibia proximal. No tratamento cirurgico foram realizados trocleoplastia, desmotomia medial da capsula articular, liberacao parcial do reto femoral e vasto medial, transposicao da tuberosidade tibial, sutura antirotacional, imbricacao lateral de capsula articular e reforco do retinaculo lateral com fascia lata. No pos-operatorio o animal permaneceu com bandagem de Robert Jones por dez dias e recebeu como medicacao meloxicam, cefalexina e condroton. O tratamento foi eficiente no realinhando do mecanismo extensor e na manutencao da patela no sulco troclear. O animal voltou a utilizar o membro aos 10 dias de pos-operatorio. Conclui-se que a associacao de tecnicas cirurgicas realizadas promove o alinhamento do das estruturas do mecanismo extensor e a estabilidade da patela no sulco troclear, sendo eficiente no tratamento da luxacao patelar medial em gatos.

Estudo comparativo da diferenciação osteogênica das células tronco mesenquimais da medula óssea e do tecido adiposo de cães adultos

O objetivo deste estudo foi comparar o potencial osteogenico das celulas tronco mesenquimais extraidas da medula ossea (CTM-MO) com as do tecido adiposo (CTM-AD) de caes adultos. As celulas foram caracterizadas fenotipicamente quanto a expressao de CD29, CD90, CD34 e CD45 e submetidas a diferenciacao adipogenica e condrogenica por 21 dias e osteogenica por 7, 14 e 21 dias. Foram constituidos quatro grupos: 1) CTM-MO em meio osteogenico, 2) CTM-MO em meio basal, 3) CTM-AD em meio osteogenico e 4) CTM-AD em meio basal. Aos 7, 14 e 21 dias de diferenciacao osteogenica as culturas foram submetidas as avaliacoes da conversao de MTT em formazan, da atividade da fosfatase alcalina (FA), da sintese de colageno e de matriz mineralizada, avaliacao do numero de celulas por campo e foram quantificados os transcritos genicos para osterix, sialoproteina ossea (BSP), osteonectina (ON) e osteocalcina (OC). Tanto as celulas extraidas da medula ossea quanto do tecido adiposo mostraram elevada expressao de marcadores para celulas tronco e baixa expressao de marcadores de celulas hematopoieticas (menor que 2%). Alem disso, foram capazes de se diferenciar em osteoblastos, condrocitos e adipocitos. As CTM-AD submetidas a diferenciacao osteogenica mostraram maior conversao do MTT em formazan que as CTM-MO, sob mesmas condicoes aos 7 e 21 dias. O numero de celulas por campo, a atividade da FA, a sintese de colageno e de matriz mineralizada foram superior nas CTM-AD em diferenciacao, em relacao as CTM-MO sob as mesmas condicoes, em todos os tempos estudados. As expressoes de osterix, BSP e OC foram predominantemente superiores nas CTM-MO diferenciadas, mas a expressao de ON foi superior nas CTM-AD diferenciadas aos 7, 14 e 21 dias. Conclui-se que as CTM-AD apresentam maior potencial osteogenico que as CTM-MO quando extraidas de caes adultos.