"Lysis and Concentration Technique" improves the parasitological diagnosis of Trypanosoma vivax.

The "Lysis and Concentration Technique" (LCTe) involves lysing red blood cells and concentrating parasites to increase the chances of visualization in low parasitemia and in scenarios of evaluators with less knowledge. The lysis of red blood cells reduced the time of diagnosis by 21 s, showing that the effect produced by the treatment is comparable to the effect of the experience in parasitological examination. In addition, the concentration of parasites was 39.18% higher for slides with high parasitemia and 131.03% for slides with low parasitemia in relation to the standard slide. LCTe proved to be inexpensive, with a total cost of approximately US$0.07 per slide made, which allows it to be easily implemented in most laboratories.

Omega-3 Fatty Acids Alter Systemic Inflammatory Mediators Caused by Apical Periodontitis.

INTRODUCTION: This study investigated the effects of omega-3 polyunsaturated fatty acids (ω-3 PUFAs) on serum inflammatory mediators of rats with pulp exposure-induced apical periodontitis. METHODS: Forty male Wistar rats were divided into the following groups: control, untreated rats (C); control rats treated with ω-3 PUFAs (C-O); rats with pulp exposure-induced apical periodontitis (AP); and rats with pulp exposure-induced apical periodontitis treated with ω-3 PUFAs (AP-O). ω-3 PUFAs were administered orally once a day for 15 days before pulp exposure and continued for 30 days after pulp exposure. The rats were sacrificed, and then blood and jaw samples were collected. Blood analysis was conducted to determine the total number of leukocytes including neutrophils, monocytes, and lymphocytes. Proinflammatory cytokines tumor necrosis factor alpha, interleukin (IL) 6, and IL-17 were quantified by enzyme-linked immunosorbent assay. Histologic analysis was performed to confirm the development of apical periodontitis. The data were statistically evaluated using analysis of variance and the Tukey posttest. The significance level was set at 5%. RESULTS: The development of apical periodontitis was confirmed in all infected groups. Bone destruction was larger in the AP group compared with the AP-O group (P < .05). Blood analysis showed that the AP and AP-O groups showed higher numbers of lymphocytes, leukocytes, monocytes, eosinophils, and expressions of tumor necrosis factor alpha and IL-6 compared with the C and C-O groups (P < .05). In contrast, the presence of leukocytes, lymphocytes, and the expression of IL-6 decreased in the AP-O group compared with the AP group (P < .05). CONCLUSIONS: ω-3 PUFA supplementation influences the systemic effects caused by apical periodontitis, decreasing the number of leukocytes, lymphocytes, and IL-6 in rat blood.

Chronic consumption of alcohol increases alveolar bone loss.

This study evaluated the effects of the chronic consumption of different concentrations of alcohol on the experimental periodontitis (EP). 160 rats were divided into 4 groups: (EP-NT) rats with EP and no alcohol exposure; (EP-A14) rats with EP exposed to 14% alcohol; (EP-A25) rats with EP exposed to 25% alcohol; (EP-A36) rats with EP exposed to 36% alcohol. The animals from the EP-A14, EP-A25 and EP-A36 groups were subjected to different concentrations of alcohol 30 days before EP induction. The histological characteristics, percentage of bone in the furcation (PBF) and bone metabolism in the furcation region were evaluated. The PBF and tartrate-resistant acid phosphatase (TRAP) data were subjected to statistical analysis. The EP-A14, EP-A25 and EP-A36 groups had lower PBFs compared with the EP-NT group. A more severe inflammatory process and a greater number of TRAP+ cells were also observed. In the EP-A14, EP-A25 and EP-A36 groups, the inflammatory process became more severe as the ingested alcoholic concentration increased. An increase in RANKL immunolabeling and a significantly higher number of TRAP+ cells were also observed. We conclude that chronic alcohol consumption increases the severity of experimental periodontitis in a dose-dependent manner by increasing the magnitude of local inflammatory responses and stimulating alveolar bone resorption.

Omega 3 Fatty Acids Reduce the Triglyceride Levels in Rats with Apical Periodontitis.

The aim of this study was to evaluate the influence of the prophylactic and therapeutic supplementation with omega 3 polyunsaturated fatty acids (w-3 PUFAs) on the lipid profile and periapical bone resorption in rats with apical periodontitis. Forty male rats were divided into groups: control rats (C), rats treated with w-3 PUFAs (C+O), rats with pulp exposure-induced apical periodontitis (AP), and rats with AP treated with w-3 PUFAs (AP+O). The administration of w-3 PUFAs was carried out orally once a day for 15 days before pulp exposure and, subsequently, for an additional 30 days after pulp exposure. AP was induced by exposing pulpal tissues to the oral environment. The samples were collected after 30 days. Triglycerides and cholesterol levels were enzymatically measured using the Trinder method. The jaws were collected and submitted for histological analysis. Two-way analysis of variance and Kruskal-Wallis tests were used for statistical analysis, and the significance was set at p<0.05. The triglyceride levels of the AP group were significantly higher than those of the C, C+O and AP+O groups (p<0.05). However, the difference in the cholesterol levels among the groups was not significant (p>0.05). Rats with AP showed larger areas of bone resorption as well as higher inflammatory intensity compared with rats with AP supplemented with w-3 PUFAs. It may be concluded that the presence of multiple AP foci increased the triglyceride levels. In addition, omega 3 supplementation might reduce these levels in rats with AP, as well as the bone resorption areas of periapical tissues.

Endodontic infections increase leukocyte and lymphocyte levels in the blood.

OBJECTIVES: The aim of this study was to determine whether the presence of single or multiple apical periodontitis (AP) alters blood cell counts and cytokine production. MATERIAL AND METHODS: Thirty rats were divided into three groups: a control group comprising rats without AP, a group called 1AP comprising rats with AP in one tooth, and a group called 4AP comprising rats with AP in four teeth. Endodontic infection was induced by pulp exposure of the first right maxillary molar in the 1AP group or by exposing the first and second right maxillary and mandibular molars in the 4AP group. A blood count and cytokine levels were obtained 30 days after infection by collecting blood by cardiac puncture. The maxillae were dissected and stained with hematoxylin and eosin to evaluate the inflammatory infiltrate. The data were tabulated and subjected to statistical analysis (P < 0.05). RESULTS: Histological analysis showed a predominance of mononuclear inflammatory cells. In blood, significant increase of leukocytes, lymphocytes, and tumor necrosis factor alpha (TNF-α) in 4AP compared with the control and 1AP groups (P < 0.05) was observed. In addition, significant decrease of interleukin-4 (IL-4) in 1AP and 4AP groups compared with the control was observed (P < 0.05). CONCLUSIONS: In the rat model, the presence of multiple AP can affect health by increasing lymphocyte and TNF-α levels in the blood. CLINICAL RELEVANCE: The presence of endodontic infections can interfere with the blood profile, altering systemic health.

Evaluation of the progression and treatment of experimental periodontitis in rats subjected to chemotherapy with 5-fluorouracil.

AIM: This study aimed to evaluate the progression and treatment of experimental periodontitis (EP) in rats treated with 5-fluorouracil (5-FU). MATERIALS AND METHODS: One-hundred fifty rats were divided into groups: C--control, EP--EP induction, EP/scaling and root planing (SRP)--EP induction and SRP, 5-FU--systemic treatment with 5-FU, 5-FU/EP--systemic treatment with 5-FU and EP induction, and 5-FU/EP/SRP--systemic treatment with 5-FU, EP induction and SRP. Systemic treatment consisted of the administration of 80 and 40 mg/kg of 5-FU. The animals were euthanised at 7, 15 and 30 days. Histological and immunohistochemical analyses for the detection of the receptor activator of nuclear factor-kappa B ligand (RANKL), osteoprotegerin (OPG) and tartrate-resistant acid phosphatase (TRAP) were performed at the furcation region. A histometric analysis was performed to evaluate the percentage of bone in the furcation region (PBF). RESULTS: In the 5-FU/EP and 5-FU/EP/SRP groups, there was less PBF, greater breakdown of periodontal tissues and increased immunostaining for RANKL compared with the EP and EP/SRP groups. CONCLUSION: It can be concluded that 5-FU aggravates PE progression, impairs the host response to SRP mechanical therapy and does not seem to cause changes in the healthy periodontium.

Morphological changes and parasite load of the adrenal from dogs with visceral leishmaniasis.

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.

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.

Periapical lesions decrease insulin signal and cause insulin resistance.

INTRODUCTION: Inflammatory cytokines are associated with decreased insulin signal transduction. Moreover, local oral inflammation, such as that accompanying periodontal disease, is associated with insulin resistance and type 2 diabetes mellitus. The aim of this study was to evaluate the effect of periapical lesions (PLs) on insulin signaling and insulin sensitivity in rats. We hypothesized that PLs alter systemic insulin signaling and insulin sensitivity via elevated plasmatic tumor necrosis factor α (TNF-α). METHODS: Wistar rats were divided into control (CN) and PL groups. PLs were induced by exposing pulpal tissue to the oral environment. After 30 days, insulin sensitivity was measured using the insulin tolerance test. After euthanization, maxillae were processed for histopathology. Plasmatic concentrations of tumor necrosis factor α (TNF-α) were determined via the enzyme-linked immunosorbent assay. Insulin signal transduction was evaluated using insulin receptor substrate tyrosine phosphorylation status and serine phosphorylation status in periepididymal white adipose tissue via Western blotting. For insulin signaling and insulin tolerance tests, the analyses performed were analysis of variance followed by the Tukey post hoc test. For TNF-α analysis, the Student's t test was used. In all tests, P < .05 was considered significant. RESULTS: The rats with PLs showed higher plasmatic TNF-α, lower constant rate for glucose disappearance values, and reduced pp185 tyrosine phosphorylation status but no change in serine phosphorylation status in white adipose tissue after insulin stimulation. CONCLUSIONS: PLs can cause alterations to both insulin signaling and insulin sensitivity, probably because of elevation of plasmatic TNF-α. The results from this study emphasize the importance of the prevention of local inflammatory diseases, such as PLs, with regard to the prevention of insulin resistance.

Pulpal and periodontal diseases increase triglyceride levels in diabetic rats.

OBJECTIVES: The aim of this study was to evaluate triglyceride and cholesterol levels in diabetic rats and their relationship with pulpal and periodontal diseases. METHODS: Eighty male rats (Rattus norvegicus albinus, Wistar) were divided into the following eight groups comprising ten animals each: normal rats (G1), rats with pulpal diseases (G2), rats with periodontal diseases (G3), rats with both pulpal and periodontal diseases (G4), diabetic rats (G5), diabetic rats with pulpal diseases (G6), diabetic rats with periodontal diseases (G7), and diabetic rats with both periodontal and pulpal diseases (G8). Diabetes was induced by injecting streptozotocin, periapical lesions were induced by exposing pulpal tissue to the oral environment, and periodontal diseases were induced by periodontal ligature. The animals were killed after 30 days, and lipid profile was enzymatically measured using Trinder's method. The total assessed values were statistically analyzed by analysis of variance and Tukey test (p < 0.05). RESULTS: The triglyceride levels of diabetic rats with periodontal disease and of diabetic rats with both periodontal and pulpal diseases were significantly higher than those of normal rats and nondiabetic group rats, respectively. The differences in the cholesterol levels among the groups were not significant. CONCLUSIONS: We found that the association of pulpal and periodontal diseases with diabetes increased triglyceride levels in rats. CLINICAL SIGNIFICANCE: Changes in lipid profile may be related to the presence of oral infections and diabetes.

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.