The high prevalence of exercise-induced pulmonary hemorrhage (EIPH) in athletic horses constitutes to be a challenge to the racing industry and a source of major concern to animal welfare. Both experimental and clinical evidence indicate that the use of autologous platelet-rich plasma (PRP) is a promising effector of repair in a variety of pulmonary conditions. The present study evaluated the effect of intrabronchial instillation of PRP on EIPH endoscopic scores from 37 Thoroughbred racehorses. Inclusion criteria were for animals to be EIPH-positive in, at least, two consecutive post-exercise endoscopic exams and to receive 250mg of furosemide IV four hours before racing. Animals were randomly assigned into 3 groups: placebo, control, and PRP instillation. All 37 Thoroughbred racehorses included had EIPH endoscopic scores pre- and post- treatment compared by statistical analysis. The bleeding score from the group receiving PRP was significantly lower than in the control and placebo groups. No adverse effects were observed in any animal during or after the experiment. It was possible to conclude that the intrabronchial instillation of autologous PRP was effective in reducing EIPH scores in racehorses receiving furosemide and that this bioproduct can be considered as a promising coadjuvant in controlling EIPH in athletic horses.
RESUMO.-[Protocolos para o preparo de plasma rico em plaquetas (PRP) em cavalos Quarto de Milha.] Este estudo comparou dois protocolos de preparo de plasma rico em plaquetas (PRP) e avaliou a associação entre dois métodos de contagem plaquetária -um manual e o outro automático através de um estudo prospectivo. Sangue venoso de oito equinos da raça Quarto de Milha foi coletado e em seguida foi centrifugado duas vezes utilizando-se dois protocolos distintos: um com descanso antes da primeira centrifugação e outro após a segunda centrifugação. A contagem plaquetária ao início, no meio e ao final dos protocolos foi realizada manualmente e pelo método automatizado, seguida de comparação entre os dois métodos. Para investigar a degranulação plaquetária ocorrida durante o preparo do PRP, o fator de crescimento vascular endotelial (VEGF) foi mensurado em cada estágio dos protocolos. O método utilizando o descanso da amostra antes da primeira centrifugação proporcionou a obtenção de um PRP mais concentrado, além de o estudo verificar que ambos os métodos de contagem plaquetária (manual e automatizado) ABSTRACT.-Miranda S., Mello Costa M.F., Rebouças N., Ramos M.T., Lessa D.A.B. & Alencar N.X. 2019. Protocols for preparation of platelet rich plasma (PRP) in Quarter Horses. Pesquisa Veterinária Brasileira 39 (8):614-621. This study compared two protocols for preparation of platelet rich plasma (PRP) and evaluated the association between manual and automated methods for platelet count using a prospective study design. Eight clinically healthy Quarter Horses had venous blood samples collected at rest. After collection, blood samples were centrifuged twice, using two different protocols including a period of sample resting, either at the start or at the end of the protocol. Platelet counting at the start of the protocol, during, and after obtaining PRP was conducted manually or with an automated counter, followed by comparison of the two methods. In order to investigate platelet degranulation during the protocol, vascular endothelial growth factor (VEGF) was measured at each preparation stage. The protocol with sample resting before centrifugation yielded a more concentrated PRP, and the study verified that both manual and automated methods are comparable and can be used interchangeably for platelet counting. VEGF concentration did not differ significantly between protocols, or among protocol stages.The results indicate that choice of protocol for PRP preparation will affect the quantity of platelets in the final product, although platelet degranulation was not observed as evidenced by the stable VEGF concentrations measured. A larger yield of non-degranulated platelets in PRP is desirable since more α-granules will be present, therefore Protocol II is recommended. Both manual and automated counts reliably allow clinicians to obtain platelet counts and the choice of utilizing a manual or automated method is unlikely to interfere with evaluation of the final PRP product.
Background: Summer Pasture Associated Obstructive Pulmonary Disease (SPAOPD), or Equine Pasture Asthma (EPA), as termed by Ferrari et al. [17], has been described as an environmentally-induced respiratory disease that occurs during the warmer and more humid months, leading to reversible airway obstruction, persistent and non-specific airway hyper-responsiveness, and chronic neutrophilic airway inflammation. Exacerbation of clinical signs vary according to warm seasons, and range from mild to severe episodes of wheezing, coughing, and laboured breathing in a chronic state that is debilitating for the equine [4]. This report describes two cases of Equine Pasture Asthma that show clinical and environmental similarities with Summer Pasture Associated Obstructive Pulmonary Disease.Case: The patients were crossbreed geldings that have never been stabled and were used for cattle management in a farm in southeastern Brazil. They presented poor performance and a persistent cough for over 3 years. Initially, the respiratory signs were only observed after exercise but, over the years, it gradually progressed to being observed when the horses were at rest. Both animals had a history of regular deworming and were previously treated by other veterinarians with antibiotics, clenbuterol, and mucokinetics. Little improvement was noticed by the owner and the signs returned over time as treatment was often discontinued. Clinical findings were compatible with the grade 3 mucus classification of Gerber et al. [18] as well as with score 2 for Severe Asthma of Davis and Sheats [13]. BALF cytology was done following the technique described by Couetil et al. [10]. Animal 1 presented slides with free yeast; macrophages and mucus with Curschmann’s spiral and counting of 29,7% of neutrophils (NE), 43,7% of lymphocytes (LP), 25,3% of macrophages (MC) and 1,3% of eosinophils (EO). Animal 2 presented slides with phagocytized yeast, mucus and counting of 27% of NE, 38,5% of LP, 33% of MC and 1,5% of EO.Discussion: Diagnostic findings fit the clinical score 2 (from 0 to 3) for Severe Asthma [13], where the animal presented frequent cough with periods of no coughing, nostrils flares in inspiration and exhalation, obvious abdominal flattening and “heave line”, pulmonary auscultation with crackles, and scarce mucous nasal discharge. The cytological findings of our reported cases also falls within the Severe Asthma classification [13], when the specific counting of 300 cells is equivalent to ≥20% of neutrophils on BALF analysis and the animal present increase in respiratory rate/effort at rest. Similar counts were found by Costa et al. [9] counting 200 cells, Rossi et al. [28] also counting 300 cells, and Couetil and Thompson [11] counting 5 fields (of at least 100 cells) on a cytocentrifuge smear. The present report took place in a region of the Rio de Janeiro State, southeastern Brazil, which shows predisposing climatic characteristics similar to those described in previous SPAOPD reports. Yet, this very climate remains somewhat constant throughout the year, as seasonality in the Rio de Janeiro State is not as marked as in the Northern Hemisphere. Given this contrasting aspect, we believe that the term Equine Pasture Asthma, instead of SPAOPD, is more appropriate to describe the cases presented here. Also, to the best of the authors’ knowledge, this is the first documented description in Brazil of Equine Pasture Asthma in animals that were never stabled or fed with hay. This documented evidence of a chronic respiratory condition consistent with Equine Pasture Asthma but little related to seasonal changes presents as a warning to other possible cases that might be unnoticed in equine herds in Brazil and in similar climates.
Alveolar macrophages (AMs) are an essential part of defense mechanisms within the lungs and their phagocytic activity is important for organ homeostasis. The phagocytic ability of AMs obtained from bronchoalveolar lavage from 17 mature mixed-breed pleasure horses (8 healthy and 9 diagnosed with mild equine asthma) was studied through assays with Leishmania (Viannia) braziliensis promastigotes, which enabled the calculation of a phagocytic index (PI) and a survival index (SI). Results indicate that phagocytic activity of AMs in asthma affected horses is similar to healthy horses, while leishmanicidal activity is significantly increased in horses with asthma.
Summary The objectives were: 1. to investigate the occurrence of equine asthma syndrome (EAS) in a group of Campolina and Mangalarga Marchador horses through post‐exercise respiratory endoscopy and cytology; 2. compare the efficiency of different diagnostic criteria; and 3. compare the incidence of EAS by breed, age group and sex. A standardised exercise test was conducted in the field, followed by respiratory endoscopy and bronchoalveolar lavage and fluid collection 30 min post‐exercise. Clinical evaluation pre‐ and post‐exercise, respiratory endoscopy post‐exercise and bronchoalveolar lavage, also post‐exercise, were conducted in 30 horses (15 Campolina and 15 Mangalarga Marchador) aged 3–18 years, without a history of respiratory illness and without clinical evidence of active respiratory disease. All horses were in active training and were considered fit to participate in equestrian competition characteristic of these breeds. Post‐collection, the horses were subdivided into healthy or EAS‐positive, depending on several diagnostic criteria. Statistical comparisons between breeds, age brackets (up to 7 years of age and above 7 years of age) and sex were conducted using appropriate statistical tests. According to bronchoalveolar lavage fluid (BALF) cytology, 56.7% of the horses showed at least one inclusion criteria that contributed to classification as EAS‐positive. Of those, only one was classified as moderate EAS, while the others were classified as mild. Total cell count in the BALF above 530 cells/µL was the single most common diagnostic criteria (100%), followed by increased tracheal mucus (53%) at endoscopy, eosinophilia (47%) and neutrophilia (35%) in BALF. Significant differences were found between macrophage and lymphocyte numbers according to breed, but no other differences between breeds, age brackets, sex or status (healthy vs. EAS‐positive). A high occurrence of EAS was found in competition‐ready Mangalarga and Campolina pacer horses in this study.
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