Viability of Equine Chondrocytes After Exposure to Mepivacaine and Ropivacaine In Vitro.

Chondrocyte health is altered when exposed to local anesthetics, raising concerns as to the long-term effects of local anesthetics intra-articularly for diagnosis and analgesia. To investigate the drug with the lowest toxic potential, the effect of ropivacaine and mepivacaine on chondrocytes was evaluated. Articular cartilage from normal metacarpophalangeal joints of five equine cadaver specimens was used to establish chondrocyte cultures. Following seven days, chondrocytes were exposed to standard culture medium (DMEM), ropivacaine 7.5 mg/ml (ROP7.5), ropivacaine 10 mg/ml (ROP10), mepivacaine 20 mg/ml (MEP20), mepivacaine 30 mg/ml (MEP 30), and 0.9% saline solution (SAL). Chondrocyte viability was evaluated by trypan blue exclusion, MTT, and flow cytometry via cellular staining with propidium iodide. No differences were observed between treatments following trypan blue exclusion assay. A difference was observed between DMEM and all other treatment groups (P < .0001) with a significant viability drop using the MTT assay. Mepivacaine 20 mg/ml and MEP30 exposure between showed greatest decrease in cellular viability compared to SAL, ROP7.5, and ROP10 (P < .0001). Cellular viability decreased as measured by flow cytometry in all groups compared to DMEM and ROP7.5 (P < .02). Interestingly, the trypan blue, MTT, and flow cytometry assays yielded different results. Although there was no difference using trypan blue, MTT demonstrated that ropivacaine-treated cells had lower viability than DMEM, and cytometry found that ROP7.5 did not differ from DMEM. Results in vitro suggest that short-term exposure to ropivacaine may result in less chondrotoxicity than mepivacaine. In vivo studies are warranted investigating long-term effects of local anesthetics on equine articular cartilage.

Evaluation of Three Methods of Platelet-Rich Plasma for Treatment of Equine Distal Limb Skin Wounds.

Wounds localized on the distal limbs in horses are difficulty healing. No studies have been conducted to evaluate homologous platelet-rich plasma (PRP) in horses. The aim of this study was to evaluate the effect upon applying autologous PRP, homologous PRP, and autologous PRP gel on the wound healing process of wounds on the distal limb of horses. To study a possible correlation between the platelet count and wound healing time, four skin wounds of 4 cm2 were surgically created on the dorsolateral aspect of the third metacarpus in eight healthy adult horses. Each skin wound was randomly treated according to the established treatment groups (G): GI, treated with autologous PRP injection on the wound edges; GII treated topically with autologous PRP gel; and GIII homologous PRP injection on the wound edges and control group treated with saline solution injected on the wound edges. The animals were monitored every fifteen days throughout the wound healing process. The wound size was determined using a tape measure. Skin biopsies were obtained on the 15th and 30th day after the first PRP treatment. Autologous PRP gel treatment reduced the healing time in 15 days compared to the wounds of the control group. Histologically, GII presented a higher frequency of mild inflammation and mild to moderate neovascularization of biopsies 1 and 2. Platelet-rich plasma in gel form showed the best result on wound healing of the distal limb of horses in both clinical and histopathological evaluations. The homologous PRP is recommended for the recovery of animals with compromised hemodynamic conditions.

Objective evaluation versus subjective evaluation of flexion tests in the pelvic limb of horses / Avaliação objetiva versus avaliação subjetiva dos testes de flexão no membro pélvico de cavalos

The agreement between subjective and objective evaluation methods was studied to identify claudication in the pelvic limbs of horses before and after flexion tests were performed. Twenty-nine horses were equipped with a wireless system of inertial sensors and evaluated during seven times while trotting. Videos were recorded to be evaluated by three veterinarians, with different levels of experience, to evaluate the agreement between the two different methods and between the evaluators. The evaluators and the objective evaluation had a low rate of agreement, with the exception of moderate agreement between the objective evaluation and evaluator 1 to identify lameness after the left total flexion and moderate agreement in evaluating the response to the tests, between objective evaluation and evaluator 2, after right distal flexion. This shows that there was a low agreement among the evaluators and between them and the objective evaluation for identifying lameness, measuring the degree of lameness and the response to the flexion tests.(AU)

A concordância entre métodos de avaliação subjetiva e objetiva foi estudada para identificar claudicação nos membros pélvicos antes e após a realização dos testes de flexão. Vinte e nove cavalos foram equipados com um sistema de sensores inerciais sem fio e avaliados durante sete momentos enquanto troteavam. Vídeos foram gravados para serem avaliados por três veterinários com diferentes níveis de experiência, a fim de avaliar a concordância entre os dois métodos diferentes e entre os avaliadores. Os avaliadores e a avaliação objetiva tiveram uma baixa taxa de concordância, com exceção de uma moderada concordância entre avaliação objetiva e avaliador 1 para identificar claudicação após o teste de flexão total esquerdo e uma moderada concordância na avaliação da resposta aos testes, entre avaliação objetiva e avaliador 2, após o teste de flexão distal direito. Isso demonstra que houve uma baixa concordância entre os avaliadores e entre estes e a avaliação objetiva para identificação da claudicação, mensuração do grau de claudicação e resposta aos testes de flexão.(AU)

Comparison of the effects of triamcinolone acetonide or platelet-rich plasma on expression of extracellular matrix-related genes in equine healthy chondrocytes in vitro / Comparação dos efeitos do acetonido de triancinolona ou plasma rico em plaquetas na expressão de genes relacionados à matriz extracelular de condrócitos in vitro de equinos adultos

ABSTRACT: In healthy cartilage, chondrocytes maintain an expression of collagens and proteoglycans and are sensitive to growth factors and cytokines that either enhance or reduce type II collagen synthesis. In osteoarthritis, pro-inflammatory cytokines, such as IL-6, induce overexpression of metalloproteinases (MMP) and decreasing synthesis of aggrecan. Use of chondroprotectors agents, such as Platelet-Rich Plasma (PRP) and triamcinolone (TA) are alternatives to reduce the progression of joint damage. In this study, we used chondrocytes extracted from metacarpophalangeal joints of healthy horses as the experimental model. Cells were treated in vitro with PRP or TA. No differences were observed between these treatments in comparison to the control group when the expressions of MMP9, MMP13, IL-6 and ACAN genes were evaluated (P<0.05). With these results, we can suggest that the treatments were not deleterious to equine cultured chondrocyte, once they did not stimulate MMPs and IL-6 synthesis or caused changes in ACAN.

RESUMO: Na cartilagem saudável, os condrócitos mantêm a expressão de colágenos e proteoglicanos, sendo sensíveis a fatores de crescimento e citocinas que aumentam ou reduzem a síntese de colágeno tipo II. Na osteoartrite, citocinas pró-inflamatórias, como a IL-6, estimulam a expressão de metaloproteinases (MMP) e reduzem a síntese de agrecano. O uso de condroprotetores, como o Plasma Rico em Plaquetas (PRP) e triancinolona (TA) é uma alternativa para se reduzir a progressão do dano articular. Neste estudo foram usados condrócitos extraídos das articulações metacarpofalangeanas de equinos saudáveis. As células foram tratadas in vitro com TA ou PRP. Não foram observadas diferenças entre os tratamentos comparando-se com o grupo controle quanto à expressão genética de MMP-9, MMP-13, IL-6 e ACAN (p<0,05). Assim, pode-se sugerir que os tratamentos não foram deletérios ao cultivo de condrócitos, uma vez que não estimularam a síntese de MMP e IL-6 e nem causaram alterações no ACAN.

Can Triamcinolone acetonide, platelet-rich plasma, and pentosan polysulfate sodium induce oxidative stress in cultured equine chondrocytes? / Acetonido de triancinolona, plasma rico em plaquetas e pentosano polissulfato sódico podem ocasionar estresse oxidativo em cultivo de condrócitos de equinos?

ABSTRACT: Progressive deterioration and loss of articular cartilage are the final degenerative events common to osteoarthritis (OA). Reactive oxygen species (ROS) play an important role in this chondrocyte catabolic activity, leading to cell death and matrix components breakdown. Intra-articular corticosteroid injections such as triamcinolone acetonide have been used to control pain and inflammation associated with OA. New treatments for OA, platelet-rich plasma and pentosan polysulphate sodium have also been used and further investigations are necessary to determine their safety in joint cells. In this in vitro study, the use of these three substances (triamcinolone acetonide, platelet-rich plasma, and pentosan polysulphate sodium) in healthy chondrocytes did not alter the antioxidant status when compared to control groups, indicating that they could be considered safe in healthy conditions.

RESUMO: A deterioração progressiva e perda da cartilagem articular são os eventos finais da osteoartrite (OA). Espécies reativas de oxigênio (ROS) têm papel importante na atividade catabólica de condrócitos, levando a morte celular e quebra dos componentes da matriz. Injeções intra-articulares de corticosteroides, como com o acetonido de triancinolona, são usadas para controle da dor e inflamação associadas à OA. Novos tratamentos para a OA, como o plasma rico em plaquetas e o pentosano polissulfato sódico, também tem sido utilizados e necessitam de maiores investigações para determinar sua segurança para as células articulares de equinos. Neste estudo in vitro, o uso destas três substâncias (acetonido de triancinolona, pentosan polissulfato de sódio de plasma rico em plaquetas) em condrócitos saudáveis de equinos não alterou o status antioxidante quando comparado aos grupos controle, indicando que puderam ser considerados seguros em condições saudáveis.

Prevalência de alterações ósseas no tarso de potros Crioulos de até vinte e seis meses de idade / Bone changes prevalence in the tarsus of Crioulo yearlings up to 26 months of age

Com o objetivo de definir a prevalência de alterações ósseas no tarso, foram examinados radiograficamente 77 potros da raça Crioula, com até 26 meses de idade e a relação das lesões com parâmetros biométricos e de manejo. Os animais foram divididos em dois grupos por idade: G1, incluindo potros de nove a 18 meses (34/77); e G2, com animais de 19 a 26 meses (43/77). Os animais também foram divididos pelo tempo de preparo para exposição morfológica em GC, grupo controle (29/77), os animais sem preparo para exposição; e GI, grupo incentivo (48/77), animais com preparo médio de 4,55 meses, e foram separados por sexo em machos (46/77) e fêmeas (31/77). Os dados foram coletados em 24 propriedades de criação ou centros de treinamento/preparo do estado do Rio Grande do Sul. Lesões radiograficamente visíveis foram observadas em 79,2% (61/77) dos animais, 80,4% dos machos, 77,4% das fêmeas, 77,3% dos animais do GC, 79,2% dos animais do GI, 86,7% dos animais do G1 e 76,7% dos animais do G2. O peso médio dos potros de 10 a 12 meses de idade foi 293,2kg, representando 71,3% do peso adulto; o peso médio dos animais com idade de 19 a 26 meses foi de 360,5kg (87,6% do peso adulto); do GC foi de 288,9kg (70,2% do peso adulto); e do GI 341,9kg (83,1% do peso adulto), apesar da idade média dos dois últimos grupos ter sido de apenas 16,8 e 19,0 meses, respectivamente. Observou-se relação positiva entre o aumento do escore corporal dos animais (P=0,01; r=0,3), do tempo de preparo dos animais para exposição morfológica (P=0,0001; r=0,7) e do escore de deposição de gordura na crista do pescoço (P=0,02; r=0,3) com as lesões radiográficas observadas no tarso dos potros. Vários fatores podem estar envolvidos na osteoartrite társica distal, entretanto, em cavalos Crioulos, o fator mais importante presente nas criações foi o sobrepeso.

Aiming to evaluate the prevalence of bone changes, radiographs were taken from both tarsal joints of 77 Crioulo yearlings, up to 26 months of age. The degree of changes was correlated with biometric parameters and management practices. The animals were separated into six groups, according to age: G1, including nine to 18 months old foals (34/77) and G2, with animals between 19 to 26 months of age (43/77); preparation time to halter class shows: GC, including animals who were not submitted to any preparation (29/77), and GI, consisting of young horses prepared to participate (average of 4.55 months of preparation time) in halter shows (48/77); and gender: colts (46/77) and fillies (31/77). Data were obtained from 24 breeding farms or training centers in Rio Grande do Sul state. Results showed that 79.22% (61/77) of the animals presented some degree of radiographically visible lesions, being 80.4% of males, 77.4% of females, 77.3% in GC, 79.2% in GI, 86.7% in G1 and 76.7% in G2. The mean body weight of the 10 to 12 month old yearlings (G1) was 293.2kg, representing 71.3% of the breed's mature weight and that of the 19 to 26 month old (G2) animals was 360.5kg (87.6% of the mature weight). The mean body weight of the GC was 288.9kg (70.2% of the mature weight) and of GI was 341.9kg (83.1% of the mature weight), although the average age of both groups was 16.8 and 19.0 months, respectively. A significant relationship was found between the animal's body condition score (P=0.01; r=0.3), time of preparation to halter class shows (P=0.0001; r=0.7), and neck crest score (P=0.02; r=0.3), and degree of the radiographic lesions found in the tarsi. Several factors can be involved in the development of distal tarsal juvenile osteoarthritis; however, in Crioulo breeding farms, the most important factor present was the horse's overweight.

Oxidative stress parameters in blood, liver, and kidney of diabetic rats treated with curcumin and/or insulin.

This study evaluated the effects of curcumin and/or insulin on antioxidant enzyme activity in blood, liver, and kidney, as well as on lipid peroxidation and delta aminolevulinic dehydratase (δ-ALA-D) activity, and a histopathological analysis of streptozotocin-induced diabetic rats. The animals were divided into six groups (n = 6): control/saline (C); control/curcumin (CCur); diabetic/saline (D); diabetic/insulin (DIns); diabetic/curcumin (DCur); and diabetic/insulin/curcumin (DInsCur). After 30 days of treatment with curcumin and/or insulin, the animals were sacrificed and the liver, kidney, and serum were used for experimental determinations. Results of histopathological analysis showed that the treatment with insulin ameliorate renal and hepatic lesions from both DIns and DInsCur groups. TBARS levels were significantly increased in serum, liver, and kidney in D group and the administration of curcumin and insulin prevented this increase in DIns and DCur groups. The activities of catalase (CAT), superoxide dismutase, and δ-ALA-D presented a significant decrease in the liver and kidney D group when compared to C group (P < 0.05). The animals treated with curcumin and insulin presented an increase of CAT activity, revealing a positive interaction between both substances. The treatments with curcumin or insulin prevented oxidative stress in blood, through modulation of enzymatic antioxidant defenses. These findings contributed to the comprehension that antioxidants from medicinal plants could be used as adjuvant in the treatment of this endocrinopathy and not as single therapy.

Occurrence of Amblyomma longirostre in Ramphastos dicolorus in Southern Brazil / Ocorrência de Amblyomma longirostre em Ramphastos dicolorus no Sul do Brasil

The species Amblyomma longirostre Koch, 1844 is poorly known in the state of Rio Grande do Sul. Usually the adult stage could be found on Sphigurus spp. and the immatures on birds (Passeriformes). Although A. longirostre is distributed in the Neotropical region, from Panama to Uruguay, it also occurs in Central America and the United States. The aim of this study was to report that Ramphastos dicolorus Linnaeus 1766 is a new host record for this tick species.

A espécie Amblyomma longirostre Koch, 1844 é pouco conhecida no Estado do Rio Grande do Sul. Normalmente a fase adulta pode ser encontrada em porquinhos Sphigurus spp., e as fases imaturas podem ser encontradas em aves (Passeriformes). A. longirostre distribui-se na região neotropical, do Panamá ao Uruguai. Ela também ocorre na América Central e nos Estados Unidos. Este estudo tem como objetivo relatar que Ramphastos dicolorus Linnaeus 1766 é um novo hospedeiro dessa espécie de carrapato.

Leptospirose e tritricomonose: isolamento em propriedade com problemas reprodutivos no sul do Brasil / Leptospirosis and tritrichomonosis: property in isolation with reproductive problems in southern Brazil / Leptospirosis y tritricomonosis: aislamiento en propiedad con problemas reproductivos en el sur de Brasil

O objetivo do estudo foi pesquisar quais os agentes etiológicos presentes em touros de duas propriedades rurais com elevados índices de repetição de cio. Da propriedade A foram analisados 23 touros, dos 58 integrados no rebanho, e na propriedade B foram analisados 15 reprodutores. Destes animais, foi colhido sangue para exame de leptospirose e realizado lavado prepucial para cultura de tritricomonose. Na propriedade A, em nenhuma das amostras avaliadas isolou-se trofozoítas de Tritrichomonas foetus. No soro de 15 touros (65,2%) testados para Leptospira foi verificada reação para um ou mais sorovares de L. hardjo, L. wolffi, L. icterohaemorrhagiae e L. butembo, em titulações que oscilaram entre 1/100 e 1/400. Na propriedade B não foi verificada titulação para leptospirose, mas em um dos animais isolaram e reproduziram trofozoìtas de T. foetus em laboratório. Do restante dos animais, cinco foram considerados suspeitos em microscópico óptico, não sendo confirmado o parasitismo em lâmina corada com Giemsa. Todos os bovinos da propriedade A foram submetidos ao tratamento com sulfato de estreptomicina em dose única de 25mg kg-1. O animal positivo para T. foetus foi descartado e os demais estão sendo monitorados mensalmente. Tanto Leptospirose como tritricomonose são enfermidades capazes de acarretar problemas reprodutivos em bovinos no sul do Brasil.

The aim of this study was to research the etiological agents present in bulls from a property with high estrus repetition rate. Twenty three out of 58 bulls in the property A and 15 of the property B were analyzed. Blood samples and preputial washes for leptospirosis and tritrichomonosis research were collected, respectively. In the property A, none of the evaluated animals showed the presence of Tritrichomonas foetus trophozoites, although 15 bulls (65,2%) tested for Leptospira reacted with one or more sorovars of L. hardjo, L. wolffi, L. icterohaemorrhagiae and L. butembo with titles between 100 and 400. Animals from property B were negative for leptospirosis and the isolation and reproduction of T. foetus in laboratory was possible only in one animal. Five other animals were considered suspect in the direct smear technique, although they were negative in the Giemsa staining method. All animals from property A received a single dose of streptomycin sulfate (25mg kg-1) which drastically reduced the estrus repetition rate. The animal with tritrichomonosis was taken out of the herd and the others have been monitored monthly. Even leptospirosis as tritrichomonosis are diseases directly linked with reproductive problems in southern Brazil.

El objetivo de este estudio fue investigar cuales son los agentes etiológicos presentes en toros de dos propiedades rurales con elevados índices repetitivos de celo. De la propiedad “A” fueron analizados 23 toros, de los 58 que integran el rebaño, en la propiedad “B” fueron analizados 15 reproductores. De estos animales, se extrajo muestras de sangre para examen de leptospirosis y se realizó lavado prepucial para cultivo de tritricomonosis. En la propiedad “A”, en ninguna de las muestras evaluadas se aisló trofozoítos de Tritrichomonas foetus, ya en el suero de 15 toros (65,2%) testeados para Leptospira fue verificada reacción para uno o más sorovares de L. hardjo, L. wolffi, L. icterohaemorrhagiae y L. butembo en titulaciones que oscilaron entre 1/100 y 1/400. En la propiedad “B” no fue verificada titulación para leptospirosis, pero en uno de los animales se aisló y se reprodujeron trofozoítos de T. foetus en laboratorio. En los demás animales, cinco fueron considerados sospechosos en microscopio óptico, no siendo confirmado el parasitismo en lámina coloreada con Giemsa. Todos los bovinos de la propiedad A fueron sometidos al tratamiento con sulfato de estreptomicina en dosis única de 25mg kg-1. El animal positivo para T. foetus fue descartado y los demás están siendo monitoreados mensualmente. Tanto Leptospirosis como tritricomonosis son enfermedades capaces de provocar problemas reproductivos en bovinos en el sur de Brasil.