Morphological changes and parasite load of the adrenal from dogs with visceral leishmaniasis / Alterações morfológicas e carga parasitária da adrenal de cães com leishmaniose visceral

The objective of this study was to analyze morphological changes and parasite loads in the adrenal gland from 45 dogs with visceral leishmaniasis (VL). The animals were from the Zoonosis Control Center of Araçatuba, state of São Paulo, which is an endemic region for the disease. These animals were euthanized due to positive diagnoses of VL. The dogs were classified into asymptomatic, oligosymptomatic and symptomatic groups. The parasite load was determined by immunohistochemistry, using VL-positive dog hyperimmune serum. Nine dogs showed an inflammatory infiltrate composed, predominantly, of plasma cells and macrophages. However, only eight dogs showed macrophages with amastigote forms of the parasite, immunolabeled in the cytoplasm. The medullary and reticular layers were the most affected areas, possibly due to a favorable microenvironment created by hormones in these regions. The density of parasites in the glandular tissue was not associated with clinical signs of VL (P > 0.05). However, the presence of the parasite was always associated with the presence of a granulomatous inflammatory infiltrate. This gland may not be an ideal place for the parasite's multiplication, but the presence of injuries to the glandular tissue could influence the dog's immune system, thus favoring the parasite's survival in the host's different organs.

O objetivo deste estudo foi analisar as alterações morfológicas e a carga parasitária da glândula adrenal de 45 cães com leishmaniose visceral (LV). Os animais eram provenientes do Centro de Controle de Zoonoses (CCZ) de Araçatuba (SP), região endêmica para a doença. Esses animais são submetidos à eutanásia, devido ao diagnóstico positivo para LV. Os cães foram classificados nos grupos assintomático, oligossintomático e sintomático. A determinação da carga parasitária foi feita por imuno-histoquímica, com utilização de soro hiperimune de cão positivo para LV. Em nove cães, verificou-se um infiltrado inflamatório, composto predominantemente por plasmócitos e macrófagos. Entretanto, apenas oito cães apresentaram macrófagos com formas amastigotas do parasito imunomarcadas em seu citoplasma. As camadas medular e reticulada foram as mais afetadas, possivelmente por um microambiente favorável criado pelos hormônios nestas regiões. A densidade de parasitos no tecido glandular não foi relacionada com os sinais clínicos de LV (P > 0,05). No entanto, a presença do parasito sempre esteve associada à presença de infiltrado inflamatório granulomatoso. Possivelmente, essa glândula não é um sítio ideal para a multiplicação do protozoário, mas a presença de injúrias no tecido glandular poderia influenciar o sistema imune do cão, favorecendo a sobrevivência do parasito nos diferentes órgãos do hospedeiro.

A proporção plasma rico em plaquetas/enxerto ósseo autógeno pode influenciar o reparo ósseo de defeitos de tamanho crítico: um estudo radiográfico em calvária de ratos / The proportion platelet-rich plasma/particulate autogenous bone graft may influencebone healing in criticalsize defects: a radiographic study in rat calvaria

Introdução: Tem sido sugerido que enxertos ósseos associados ao Plasma Rico em Plaquetas (PRP) cicatrizam com maior densidade que os enxertos sem PRP. Contudo, esse resultado parece depender da proporção PRP/enxerto ósseo utilizada. Objetivo: O propósito deste estudo foi avaliar, radiograficamente, a influência da proporção PRP/enxerto de Osso Autógeno (OA) no reparo ósseo de defeitos de tamanho crítico (DTCs) criados cirurgicamente em calvárias de ratos. Material e método: Cinquenta ratos foram divididos em cinco grupos: C, OA, OA/PRP-50, OA/PRP-100 e OA/PRP-150. Um DTC de 5 mm de diâmetro foi criado na calvária de cada animal. No Grupo C, o defeito cirúrgico foi preenchido somente com coágulo sanguíneo. No Grupo OA, o defeito foi preenchido com enxerto de OA particulado. Nos grupos OA/PRP-50, OA/PRP-100 e OA/PRP-150, o defeito foi preenchido com enxerto de OA particulado associado a 50, 100 ou 150 µL de PRP, respectivamente. Os animais foram eutanasiados aos 30 dias pós-operatórios. Imagens radiográficas padronizadas das calvárias foram obtidas com o sistema Digora (Soredex, Finlândia). As imagens foram avaliadas por três examinadores por meio de escores, que indicaram o percentual de radiopacidade dos defeitos. Os escores obtidos foram submetidos ao Teste de Kruskal-Wallis (p < 0,05). Resultado: O Grupo C apresentou radiopacidade significativamente menor que os grupos OA, OA/PRP-50, OA/PRP-100 e OA/PRP-150 (p < 0,05). O Grupo OA/PRP-50 apresentou radiopacidade significativamente maior que os grupos OA/PRP-100 e OA (p < 0,05). Conclusão: Dentro dos limites deste estudo, pode-se concluir que a proporção PRP/enxerto de OA influencia a cicatrização de DTCs criados em calvárias de ratos.

Introduction: It has been suggested that bone grafts combined with platelet rich plasma (PRP) present greater density than bone grafts without PRP after healing process. However, this result may depend on proportion PRP/autogenous bone (AB) graft used. Objective: The purpose of this study was to analyze radiographically the influence of the proportion PRP/particulate autogenous bone (AB) graft on bone healing in surgically created criticalsize defects (CSD) in rat calvaria. Material and method: 50 rats were divided into 5 groups: C, AB, AB/PRP-50, AB/PRP-100 and AB/PRP-150. A 5 mm diameter CSD was created in the calvarium of each animal. In Group C the defect was filled by blood clot only. In Group AB the defect was filled with 0.01 mL of AB graft. In groups AB/ RP-50, AB/PRP-100 and AB/PRP-150 the defects were filled with 0.01 mL of AB graft combined with 50, 100 and 150 µL of PRP, respectively. All animals were euthanized at 30 days post-operative. Standardized radiographic images of the rat calvaria were obtained using Digora System (Soredex, Finland). The images were evaluated by three examiners using scores that indicated the percentage of radiopacity of the defect. The obtained data were subjected to statistical analysis (Kruskal-Wallis test, p <0.05). Result: Group C presented radiopacity significantly lower than groups AB, AB/PRP-50, AB/PRP-100 and AB/PRP-150. Group AB/PRP-50 showed radiopacity significantly greater than groups AB/PRP-100 and AB. Conclusion: It can be concluded that the proportion PRP/AB graft influences bone healing in CSD in rat calvaria.

A standardized research protocol for platelet-rich plasma (PRP) preparation in rats

Introduction: The urgent need for studies using standardized protocols to evaluate the real biological effects of PRP has been emphasized by several authors. Objective: The purpose of this study was to standardize a methodology for autologous Platelet-Rich Plasma (PRP) preparation in rats. Material and methods: Twenty-four, 5 to 6-month-old, male rats, weighing 450 to 500 g were used. After general anesthesia, 3.15 ml of blood was collected from each animal, via cannulation of the jugular vein. A standardized technique of double centrifugation was used to prepare PRP. PRP samples and peripheral blood platelets were then manually counted using a Neubauer chamber. Student’s t-test was used to compare the differences between the number of platelets in peripheral blood and PRP samples (p < 0.05). In addition, PRP and peripheral blood smears were stained to see platelets’ morphology. Results: All surgical procedures were well tolerated by the animals and they were healthy during the entire experimental period. PRP samples showed higher significantly platelet concentrations than peripheral blood samples (2,677,583 and 683,680 respectively). Conclusion: Within the limits of this study, it can be concluded that the method used produced autologous PRP with appropriated platelet quantity and quality, in rats.