Avaliação de sete protocolos para obtenção de plasma rico em plaquetas na espécie equina

Pereira, Rudiney Soares; Zacarias, Gian Vitor Freitas; Cantarelli, Camila; Junior, Marcos Corrêa; Silva, Gabriele Biavaschi da; Barbosa, Anna Laethicia Trindade; Brass, Karin Érica; Côrte, Flávio Desessards De La

doi:10.1590/s0103-84782013005000052

Cited by 10 publications

(17 citation statements)

References 17 publications

(14 reference statements)

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“…Os valores médios desse fator de crescimento (8.693,8±3.362,58pg/mL) foram relativamente altos, sendo semelhantes aos obtidos por Pereira et al (2013) (7.634±1,218pg/mL) quando o mesmo protocolo de obtenção foi utilizado, ou seja, duas centrifugações, sendo a primeira com 120xg/10 min, e a segunda com 240xg/10 min. Adicionalmente, foram bem mais elevados tanto no PRP como no plasma (2.717,71±900,61pg/mL) do que os obtidos por Giraldo et al (2013) que utilizaram a mesma técnica (ELISA) e os mesmos kits (R&D Systems) para quantificar esse fator de crescimento no PRP em gel (PRG--P) ativado farmacologicamente com gluconato de cálcio a 10%, em equinos das raças crioula argentina e colombiana (2.264,0±1.196,0 pg/mL no PRP; 1.012,0±426,0 pg/ mL no plasma), com idade acima de 10.1 anos.…”

Section: Discussionunclassified

“…Adicionalmente, embora a presente pesquisa não tenha revelado correlação, há relato de associação entre a presença de leucócitos e de TGF-β1 no PRP, onde valores mais elevados das células brancas resultariam em maiores concentrações do fator de crescimento (Pereira et al 2013). Conforme mencionado previamente, esse estudo utilizou um PRP-PL, o que pode, portanto, ter resultado em menor concentração do TGF-β1.…”

Section: Discussionunclassified

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Quantificação de fatores de crescimento na pele de equinos tratada com plasma rico em plaquetas

et al. 2014

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ABSTRACT.-Souza M.V., Pinto J.O., Costa M.M., Santos E.C., Garcia S.L.R. & Oliveira L.L. 2014.[Quantification of growth factors in horse skin treated with platelet-rich plasma.] Quantificação de fatores de crescimento na pele de equinos tratada com plasma rico em plaquetas. Pesquisa Veterinária Brasileira 34(6):599-612. Departamento de Veterinária, Universidade Federal de Viçosa, Campus Universitário s/n, Viçosa, MG 36570-000, Brazil. E-mail: msouza@ufv.br Platelet-rich plasma (PRP) is a product derived from total blood centrifugation, rich in bioactive factors, such as growth factors. Despite largely used in healing processes, there is a controversy whether the therapy is effective in promoting skin healing. The objective of this study was to quantify and compare the concentrations of the factors TGF-β1 and PDGF-BB in PRP, blood plasma and skin, at different phases of the healing process of skin treated or not with PRP. Seven healthy crossbred 16 to 17-year-old geldings (16.14±0.63) were used. Three quadrangular-shaped lesions (6.25cm 2 ) were surgically induced into the right and left gluteal regions of all animals. Twelve hours after induction of the wounds, 0.5mL of the PRP was administered in each of the four edges of the wound in one of the gluteal regions (Treated group = TG) randomly chosen. The contralateral region was used as control (CG). The wounds were submitted to daily cleaning with Milli-Q water, and samples were obtained with a 6mm diameter biopsy Punch. Six skin biopsies were obtained, the first carried out immediately after the production of the wound (T0), and the others 1 (T1), 2 (T2), 7 (T3), and 14 (T4) days after the lesion was induced. The sixth biopsy (T5) was obtained after complete healing of the skin, which occurred at about day 37 (36.85±7.45, CG; 38.85±6.46, TG). EDTA blood samples were also obtained, at all the times mentioned. Quantification of TGF-β1 and PDGF-BB growth factors on the skin, PRP, and blood plasma was carried out by the ELISA technique. Data were statistically analyzed by the t test, Pearson correlation and regression, at a significance level of 5%. No difference was found between the groups in the values of the two growth factors measured on the skin, at the different times. Also, no correlation was found between the amount of growth factors present in the skin and plasma. On the other hand, a positive correlation was observed between PRP and skin in the treated group, for the growth factors TGF-β1 (r=0.31) and PDGF-BB (r=0.38), as well as between both growth factors present in PRP (r=0.81). Considering the growth factor concentrations at T0, the highest skin values (p<0.05) of TGF-β1, in both groups, occurred at T3 and T5. Higher values (p<0.05) of PDGF-BB occurred at T4 (TG) and T5 (CG). No plasma changes occurred at the concentration of these factors in relation to T0, suggest-1 Recebido em 5 de outubro de 2013.Aceito para publicação em 5 de junho de 2014.

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Section: Discussionunclassified

Quantificação de fatores de crescimento na pele de equinos tratada com plasma rico em plaquetas

et al. 2014

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“…In both protocols, a small amount of red blood cells was present in the PRP, with a smaller amount in Protocol I (0.51/µL ± 0.79) in relation to Protocol II (0.65/µL ± 0.44) with a similar outcome for leukocyte counts. It is unknown whether a higher concentration of erythrocytes and leukocytes would affect the usage and outcomes of therapeutic PRP (Marx 2004, Carmona Ramírez & Prades 2006, Vendramin et al 2006), but previous investigations suggested that a small concentration of leukocytes would be desirable in order to maximize benefits arising from the increased platelet and GF concentrations in PRP (Pereira et al 2013).…”

Section: Discussionmentioning

confidence: 99%

“…Various techniques for equine PRP preparation have been suggested (DeRossi et al 2009, Vendruscolo et al 2012) but there is no consensus regarding the gold standard protocol. Differences in centrifugation time, speed, and resting of the sample could potentially influence the concentration of platelets and GFs in the final product, which likely influences the effects of PRP itself (Da Fontoura Pereira et al 2013). Table 1 summarizes information available in the current literature concerning GFs, while Table 2 summarizes published protocols for harvesting PRP.…”

Section: Introductionmentioning

confidence: 99%

Protocols for preparation of platelet rich plasma (PRP) in Quarter Horses

et al. 2019

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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.

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“…PRP was prepared according to the protocol developed by Pereira et al (2013). Whole blood (500 ml) was aseptically drawn from the jugular vein in a blood collection bag containing anticoagulant made of citrate, phosphate, dextrose, and adenine (CPDA).…”

Section: Prp Preparationmentioning

confidence: 99%

Cryopreservation protocol for equine platelet-rich plasma

Kwirant

Côrte

Brass

et al. 2016

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In this preliminary study, a new equine platelet-rich plasma (PRP) cryopreservation protocol was evaluated. PRP was obtained by a double centrifugation technique of whole blood collected from 8 adult healthy ponies. A fresh sample of PRP was analyzed for total platelet count, mean platelet volume (MPV), and platelet morphology. Upon morphological evaluation, 200 platelets were counted using a differential interference contrast microscope with a 40x phase objective and classified as activated (with pseudopodia), inactivated (normal discoid shape), or uncertain state (spherical shape, without pseudopodia). Two other PRP samples, one containing DMSO as a cryoprotectant and the other without DMSO, were stored in a mechanical freezer at -80ºC. After 14 days, the frozen samples were thawed and submitted to the same analysis as described above. The fresh PRP showed a platelet count of 830 (±95.3) x10 3 µL -1 , an MPV of 5.2 (±0.07) fL, and composed of 4% activated platelets. There was no significant difference in platelet count, MPV, and activated platelets between fresh and 6% DMSO frozen PRP samples (617.9±65.5x10 3 µL -1 ; 5.3±0.06fL; 9.5%) (p>0.05). On the other hand, samples frozen without DMSO showed a significantly lower platelet count (519.6±66.1x10 3 µL -1 ), higher MPV (5.7±0.08fL), and more activated platelets (13.9%) than the other groups (p<0.05).The 6% DMSO was able to preserve platelet morphology in PRP stored at -80 o C for 14 days, but studies on platelet function of thawed PRP are still needed. Key words: Platelets. Storage. Equine. Dmso. ResumoNeste estudo preliminar, um novo protocolo de criopreservação de plasma rico em plaquetas (PRP) de equinos foi avaliado. O PRP foi obtido através de dupla centrifugação do sangue total coletado de oito pôneis clinicamente saudáveis. Uma amostra de PRP fresco foi analisada quanto ao número total de plaquetas, volume plaquetário médio (VPM) e morfologia plaquetária. Na avaliação morfológica 200 plaquetas foram contadas utilizando um microscópio com contraste de fase com objetiva de 40x e classificadas como ativadas (com pseudópodes), inativas (forma discóide normal) ou estado incerto (forma esférica, mas sem peseudópodes). Duas outras amostras de PRP foram armazenadas em um freezer mecânico a -80 o C, uma contendo dimetil sulfóxido (DMSO) como crioprotetor e outra sem 3 µL -1 ), maior VPM (5,7±0,08fL) e mais plaquetas ativadas (13,9%) do que os outros grupos (p<0,05). DMSO a 6% foi capaz de preservar a morfologia plaquetária no PRP armazenado a -80 o C durante 14 dias, mas estudos a respeito da função plaquetária ainda não necessários. Palavras-chave: Plaquetas. Armazenamento. Equino. Dmso.

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Avaliação de sete protocolos para obtenção de plasma rico em plaquetas na espécie equina

Cited by 10 publications

References 17 publications

Quantificação de fatores de crescimento na pele de equinos tratada com plasma rico em plaquetas

Quantificação de fatores de crescimento na pele de equinos tratada com plasma rico em plaquetas

Protocols for preparation of platelet rich plasma (PRP) in Quarter Horses

Cryopreservation protocol for equine platelet-rich plasma

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