Inclusões de quitosana no subcutâneo de rato: avaliação clínica, histológica e morfométrica

FUNDAMENTOS: A quitosana é polímero derivado da quitina, com vários tipos de aplicação na área médica. OBJETIVO: Avaliar a biocompatibilidade de membranas de quitosana no subcutâneo de ratos. MÉTODOS: Foram utilizados 20 ratos Wistar machos, nos quais foram implantadas membranas de quitosana, na região mediana dorsal. Os animais foram sacrificados: sete, 15, 30 e 60 dias após a cirurgia, tendo sido avaliados clinicamente durante o período experimental e com fotodocumentação no momento do sacrifício. Após o sacrifício, as membranas e tecidos adjacentes foram removidos e preparados para exame histológico e morfométrico. RESULTADOS: Nenhum animal apresentou efeitos adversos que pudessem ser atribuídos à implantação das membranas. O exame histológico mostrou que as inclusões são lisas e homogêneas e não são colonizadas por células do hospedeiro, sendo circundadas por pseudocápsula composta por fibroblastos e células inflamatórias. A morfometria da pseudocápsula revelou espessura semelhante durante todo o período experimental (P>0,05). CONCLUSÃO: A quitosana pode ser opção para uso como implante não integrado. Novos estudos devem ser realizados para comprovar a biocompatibilidade a longo prazo.

Agaricus blazei (Himematsutake) does not alter the development of rat diethylnitrosamine-initiated hepatic preneoplastic foci

The modifying potential of crude extracts of the mushroom Agaricus blazei Murrill (Himematsutake) on the development and growth of glutathione S-transferase placental form (GST-P)-positive liver foci (liver preneoplastic lesion) was investigated in adult male Wistar rats. Six groups of animals were used. Groups 2 to 5 were given a single i.p. injection of 200 mg/kg b.w. of diethylnitrosamine (DEN) and groups 1 and 6 were treated with saline at the beginning of the experiment. After 2 weeks, animals of groups 3 to 6 were orally treated with three dose levels of aqueous extracts of the mushroom A. blazei (1.2, 5.6, 11.5, and 11.5 mg/ml of dry weight of solids) for 6 weeks. All animals were subjected to two-thirds partial hepatectomy at week 3 and sacrificed at week 8. Two hours before sacrifice, ten animals of each group were administered a single i.p injection of 100 mg/kg of bromodeoxyuridine (BrdU). Apoptotic bodies and BrdU-positive hepatocyte nuclei were quantified in liver sections stained for hematoxylin and eosin (H&E) (eosinophilic foci) and simultaneously stained for GST-P expression (GST-P-positive foci), respectively. The 6-week treatment with A. blazei did not alter the development (number and size) of GST-P-positive foci and did not affect the growth kinetics of liver normal parenchyma or foci in DEN-initiated animals. Our results indicate that the treatment with aqueous extracts of the mushroom A. blazei during the post-initiation stage of rat liver carcinogenesis does not exert any protective effect against the development of GST-P-positive foci induced by DEN. (Cancer Sci 2003; 94: 188-192).

Biological differences between reflux stimulated proliferative stomal lesions and N-methyl-N '-nitro-N-nitrosoguanidine induced carcinomas in Wistar rats.

The morphology and evolution of epithelial lesions that developed at a gastrojejunal stoma due to reflux of duodenal contents were compared with MNNG-induced carcinomas in the pyloric mucosa of rats in a long term experiment. Random bred male Wistar rats were given MNNG in drinking water (100 mg/l) for 12 weeks and then one group was submitted to a gastrojejunal anastomosis at the greater curvature in the oxyntic mucosa, Untreated rats underwent either gastrojejunostomy or gastrotomy. The animals were killed at the 24th and 66th weeks of the experiment. The lesions obtained in the pyloric mucosa and in the mucosa of the gastrojejunal stoma were analyzed histologically using hematoxylin and eosin staining and immunohistochemistry for pepsinogen isoenzyme 1. Duodenal reflux induced proliferative lesions at the gastrojejunal junction that increased in incidence and size with time. Histologically they consisted of benign epithelial proliferation of gastric type. No evidence of malignant transformation within the gastric components of the proliferative lesions at the gastrojejunal stoma was observed even at the 66th week, Adenocarcinomas induced by MNNG in the pyloric mucosa increased in size during the experiment and were morphologically and histochemically distinct from the proliferative lesions at the gastrojejunal junction. In conclusion, proliferative lesions at the gastrojejunal stoma stimulated by duodenal reflux are biologically distinct from adenocarcinomas induced by MNNG in the pyloric mucosa. They do not seem to be precursor lesions of gastric carcinogenesis, as they do not undergo malignant transformation even after long-term, up to 66 weeks, follow-up. (C) 1999 Elsevier B.V. Ireland Ltd. All rights reserved.

Absence of chemopreventive influence of propolis on the rat liver altered foci development

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Imuno-histoquímica para identificação de células neoplásicas no infiltrado ativo de melanomas finos

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Canine corpus luteum regression: Apoptosis and caspase-3 activity

The present study evaluated the occurrence of apoptosis and caspase-3 activity in the canine corpus luteum during the period of luteal regression in eight pregnant and nine nonpregnant diestrus bitches. Intact luteal cells were obtained from corpora lutea in both peripartum pregnant bitches and nonpregnant diestrus bitches at approximately 65 d (range 63-68) after estrus, but not at days 75 and 85 in nonpregnant bitches. In all bitches, apoptotic cells were rarely detected and when present, those cells were more easily detected using the hematoxylin and eosin technique than using the critical electrolyte concentration technique. The luteal structures at 75 and 85 d of diestrus had histological characteristics similar to a corpus albicans. Caspase-3 activity was detected in morphologically normal corpora lutea from both pregnant and diestrus bitches around day 65, and also in the later structures considered corpus albicans tissue. These results suggested that apoptosis may not be the major mechanism involved in canine functional luteal regression, and that caspase-3 participated in both functional and morphological luteolysis and in the tissue reorganization involved in corpus albicans formation. (c) 2006 Published by Elsevier B.V.

Effects of cyclosporin, phenytoin, and nifedipine on the synthesis and degradation of gingival collagen in tufted capuchin monkeys (Cebus apella): Histochemical and MMP-1 and -2 and collagen I gene expression analyses

Background: The purpose of this experimental study was to evaluate the collagen fiber distribution histologically after phenytoin, cyclosporin, or nifedipine therapy and to correlate it with collagen I and matrix metalloproteinase (MMP)-1 and -2 gene expression levels.Methods: Gingival samples from the canine area were obtained from 12 male monkeys (Cebus apella). The mesial part of each sample was assessed by reverse transcription-polymerase chain reaction, whereas the distal part was processed histologically for picrosirius red and hematoxylin and eosin stainings, as well as for collagen IV immunostaining. One week after the first biopsy, the animals were assigned to three groups that received daily oral dosages of cyclosporin, phenytoin, or nifedipine for 120 days. Additional gingival samples were obtained on days 52 and 120 of treatment from two animals from each group on the opposite sides from the first biopsies.Results: Picrosirius red staining showed a predominance of mature collagen fibers in the control group. Conversely, there was an enlargement of areas occupied by immature collagen fibers in all groups at days 52 and 120, which was not uniform over each section. There was a general trend to lower levels of MMP-1 gene expression on day 52 and increased levels on day 120. Phenytoin led to increased levels of MMP-2 and collagen I gene expression on day 120, whereas the opposite was observed in the nifedipine group.Conclusion: Cyclosporin, phenytoin, and nifedipine led to phased and drug-related gene expression patterns, resulting in impaired collagen metabolism, despite the lack of prominent clinical signs.

Effect of alveolex on the bone defects repair stimulated by rhBMP-2: Histomorphometric study

OBJECTIVE: The aim of this study was to evaluate histomorphometrically the effect of alveolex (Propolis 10%) on the repair of bone cavities in the calvaria of rats. MATERIALS and METHODS: A 5 mm diameter bone defect was made in the calvaria of male Wistar rats using the drill-type trephine. The defects were filled with rhBMP-21Alveolex, rhBMP-2, Alveolex, or coagulum. Twenty-eight animals with seven subjects on each were sacrificed 30 days after surgery and samples were fixed and embedded in paraffin. Histological sections stained by HE (hematoxylin and eosin) were obtained from the calvaria bone defect and analyzed by a differential point-counting method. RESULTS: Group I and II, rhBMP-21Alveolex and rhBMP-2, respectively, presented higher levels of newly formed bone than other groups (P < 0.001). There were not significant differences between groups I and II (P > 0.05). In addition, there was not significant difference between groups III and IV, Control-Coagulum and Alveolex, respectively (P > 0.05). CONCLUSION: Alveolex has increased the bone repair in calvaria defects of rats when associated to rhBMP-2, however without significant differences for rhBMP-2 isolated group; Alveolex isolated group showed the lowest levels of newly formed bone with no significant differences to coagulum group (control). Microsc. Res. Tech. 75: 36-41, 2012. (C) 2011 Wiley Periodicals, Inc.

Histomorphometric analysis and immunolocalization of RANKL and OPG during the alveolar healing process in female ovariectomized rats treated with oestrogen or raloxifene

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Periodontal tissue response to coverage of root cavities restored with resin materials: A histomorphometric study in dogs

Background: the purpose of this study was to histomorphometrically evaluate the response of periodontal tissues covering Class V resin restorations in dogs.Methods: After raising a mucoperiosteal flap, bony defects measuring 5 x 5 mm were created on the buccal aspect of the canines of five dogs followed by cavity preparations on the root surface measuring 3 x 3 x 1 mm. Before repositioning the flap to cover the bone defect, the cavities were restored with composite resin (CR) or resin-modified glass ionomer cement (RMGIC) or were left unrestored as control (C). The dogs were euthanized 90 days after surgery. Specimens comprising the tooth and periodontal tissues were removed, processed routinely, cut into longitudinal serial sections in the bucco-lingual direction, and stained with hematoxylin and eosin (H&E) or Masson's trichrome. The most central sections were selected for histomorphometric analysis.Results: Histomorphometric analysis revealed apical migration of epithelial tissue onto the restorative materials (RMGIC and CR). The C group presented significantly longer connective tissue attachment (P < 0.05) than the RMGIC and CR groups and significantly higher bone regeneration (P < 0.05) compared to the RMGIC group. Histologically, the cervical third (CT) of all groups had the most marked chronic inflammatory infiltrate.Conclusions: Within the limits of this study, it can be concluded that the restorative materials used exhibit biocompatibility; however, both materials interfered with the development of new bone and the connective tissue attachment process.

Influence of enamel matrix derivative (Emdogain((R))) and sodium fluoride on the healing process in delayed tooth replantation: histologic and histometric analysis in rats

Although it has already been shown that enamel matrix derivative (Emdogain((R))) promotes periodontal regeneration in the treatment of intrabony periodontal defects, there is little information concerning its regenerative capacity in cases of delayed tooth replantation. To evaluate the alterations in the periodontal healing of replanted teeth after use of Emdogain((R)), the central incisors of 24 Wistar rats (Rattus norvegicus albinus) were extracted and left on the bench for 6 h. Thereafter, the dental papilla and the enamel organ of each tooth were sectioned for pulp removal by a retrograde way and the canal was irrigated with 1% sodium hypochlorite. The teeth were assigned to two groups:in group I, root surface was treated with 1% sodium hypochlorite for 10 min (changing the solution every 5 min), rinsed with saline for 10 min and immersed in 2% acidulated-phosphate sodium fluoride for 10 min; in group II, root surfaces were treated in the same way as described above, except for the application of Emdogain((R)) instead of sodium fluoride. The teeth were filled with calcium hydroxide (in group II right before Emdogain((R)) was applied) and replanted. All animals received antibiotic therapy. The rats were killed by anesthetic overdose 10 and 60 days after replantation. The pieces containing the replanted teeth were removed, fixated, decalcified and paraffin-embedded. Semi-serial 6-mu m-thick sections were obtained and stained with hematoxylin and eosin for histologic and histometric analyses. The use of 2% acidulated-phosphate sodium fluoride provided more areas of replacement resorption. The use of Emdogain((R)) resulted in more areas of ankylosis and was therefore not able to avoid dentoalveolar ankylosis. It may be concluded that neither 2% acidulated-phosphate sodium fluoride nor Emdogain((R)) were able to prevent root resorption in delayed tooth replantation in rats.

Healing process of incisor teeth of diabetic rats replanted after storage in milk

Several local factors that influence the healing process of replanted teeth have been investigated. However, it remains unclear how systemic alterations, such as diabetes mellitus, affect the prognosis of these cases. The purpose of this study was to evaluate the healing process of incisors of non-controlled diabetic rats replanted after storage in bovine long shelf-life (UHT) whole milk. Thirty-two rats were randomly assigned to receive an endovenous injection of either citrate buffer solution (group I - control; n = 16) or streptozotocin dissolved in citrate buffer solution to induce diabetes (group II; n = 16). After confirmation of the diabetic status by analysis of the glycemic levels, the maxillary right incisor of each animal was extracted and immersed in milk for 60 min. The root canals of teeth were then instrumented, and were filled with a calcium hydroxide-based dressing and replanted into their sockets. All animals received systemic antibiotic and were killed by anesthetic overdose 10 and 60 days after replantation. The specimens containing the replanted teeth were removed, fixed, decalcified, and embedded in paraffin. Semi-serial 6-mu m-thick sections were obtained and stained with hematoxylin and eosin for histologic and histometric analyses. The results showed that the connective tissue adjacent to the root surface was less organized in the diabetic animals than in the control animals in both periods; the root dentin was less severely affected by root resorption in the diabetic rats; there were no significant differences between the control and diabetic groups regarding the occurrence of replacement resorption and inflammatory resorption.

Evaluation of alveolar socket response to Angelus MTA and experimental light-cure MTA

Objective. The aim of this study was to evaluate the rat alveolar socket response to implanted polyethylene tubes filled with a light-cured MTA or Angelus MTA.Study design. Fifty-seven Wistar rats received an implant filled with one of the materials in the dental socket of the right central incisor. After 30, 60, and 90 days, animals were killed, and the right hemimaxillas were removed and processed to be analyzed on light microscopy. After embedding in paraffin, 6-mu m-thick sections were obtained and stained with hematoxylin and eosin. Qualitative and quantitative analyses of the reaction were performed.Results. It was observed that the light-cured MTA presented a similar response compared with Angelus MTA, characterized by the reduction of the chronic inflammatory response and organization of the alveolar bone close to the materials, especially at the 90th day of observation.Conclusions. It was concluded that both materials were well accepted by the alveolar tissue of rats, with the formation of mineralized tissue close to the materials. (Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2010;110:e93-e97)

Repair process of surgical defects filled with autogenous bone grafts in tibiae of diabetic rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Delayed replantation of rat teeth after use of reconstituted powdered milk as a storage medium

Minimal extraoral dry storage period and moist storage for the avulsed tooth are identified as key steps for the treatment protocol of tooth replantation. Among the possible moist storage media, bovine milk has stood out because of its capacity of preserving the integrity of the periodontal ligament (PDL) fibers. This condition has attracted the attention to investigate the use of powdered milk, which is one of the presentation forms of bovine milk, as a feasible storage medium in cases of delayed tooth replantation. The aim of this study was to evaluate the healing process after delayed replantation of rat teeth stored in reconstituted powdered milk and long shelf-life (ultra high temperature) whole milk. Forty maxillary right rat incisors were assigned to four groups (n = 10): group I - the teeth were extracted and immediately replanted into theirs sockets; group II - the teeth were stored for 60 min in 200 ml of freshly reconstituted powdered milk; group III - the teeth were stored for 60 min in 200 ml of long shelf-life whole milk; group IV - the teeth were kept dry for the same time. All procedures were performed at room temperature. Next, the root canals of teeth in groups II, III, and IV were instrumented, filled with a calcium hydroxide-based paste, and replanted into their sockets. All animals received systemic antibiotic therapy and were killed by anesthetic overdose 60 days after replantation. The pieces containing the replanted teeth were removed, fixed, decalcified, and paraffin-embedded. Semi-serial 6-mu m-thick sections were obtained and stained with hematoxylin and eosin for histomorphological analysis. There was statistically significant difference (P < 0.05) between groups I and IV regarding the presence of replacement resorption and PDL remnants on root surface. The powdered milk and long shelf-life whole milk presented similar results to each other and may be indicated as storage media for avulsed teeth.