The applicability of hematoxylin-eosin staining plus fluorescence or confocal laser scanning microscopy to the study of elastic fibers in cartilages

This study focuses on the use of hemotoxylin-eosin staining plus fluorescence microscopy for the investigation of elastic fibers in some elastic cartilages. We have observed that elastic fibers are consistently imaged by the proposed procedure and the resolution attained is similar to that obtained with the classical Weigert's fuchsin-resorcin. The results also demonstrate that elastin autofluorescence gives little or no contribution to the final fluorescence and that the use of the confocal laser scanning microscope adds to the resolution, permits the use of thicker sections and reveals of minute structural features. We conclude that this is a relevant tool in elastin research.

Fluorescence and confocal laser scanning microscopy imaging of elastic fibers in hematoxylin-eosin stained sections

We have studied the possibility of associating fluorescence microscopy and hematoxylin-eosin staining for the identification of elastic fibers in elastin-rich tissues. Elastic fibers and elastic laminae were consistently identified by the proposed procedure, which revealed itself to be easy and useful for the determination of such structures and their distribution. The fluorescence properties of stained elastic fibers are due to eosin staining as revealed by fluorescence analysis of the dye in solution, with no or only minor contribution by the elastin autofluorescence. The main advantage of this technique resides in the possibility of studying the distribution of elastic fibers in file material without further sectioning and staining. The use of the confocal laser scanning microscope greatly improved the resolution and selectivity of imaging elastic fibers in different tissues. The determination of the three-dimensional distribution and structure of elastic fiber and laminae using the confocal laser scanning microscope was evaluated and also produced excellent results.

Anatomopathological analysis of sentinel and nonsentinel lymph nodes in breast cancer: Hematoxylin-eosin versus immunohistochemistry

The authors compare the detection of metastases in sentinel lymph nodes (SLNs) and nonsentinel lymph nodes (NSLNs) using hematoxvlin-eosin (HE) staining versus immunohistochemistry (IHC). Thirty-six patients with breast carcinoma undergo exeresis of the primary tumor and of 50 SLNs and 491 NSLNs. Sentinel lymph nodes are sectioned into transverse slices of 2- to 3-mm thickness, and a cytologic smear and a frozen section were obtained from each slice. The slices are completely cut into serial sections at 100-mu m intervals. Two consecutive 4-mu m-thick sections are then obtained from each level and were prepared for HE staining and IHC. Nonsentinel lymph nodes are evaluated similarly to SLNs. The authors obtain 4076 SLN sections and 32 012 NSLN sections, fora total of 36 088 sections. A comparison of HE staining versus IHC based on the total number of sections shows a sensitivity of 93.8%, a negative predictive value of 98.9%, and an accuracy of 99.1 %. The values obtained by HE staining are similar to those obtained by IHC.

Micromorphology Of The Dental Pulp Is Highly Preserved In Cancer Patients Who Underwent Head And Neck Radiotherapy.

Teeth are often included in the radiation field during head and neck radiotherapy, and recent clinical evidence suggests that dental pulp is negatively affected by the direct effects of radiation, leading to impaired sensitivity of the dental pulp. Therefore, this study aimed to investigate the direct effects of radiation on the microvasculature, innervation, and extracellular matrix of the dental pulp of patients who have undergone head and neck radiotherapy. Twenty-three samples of dental pulp from patients who finished head and neck radiotherapy were analyzed. Samples were histologically processed and stained with hematoxylin-eosin for morphologic evaluation of the microvasculature, innervation, and extracellular matrix. Subsequently, immunohistochemical analysis of proteins related to vascularization (CD34 and smooth muscle actin), innervation (S-100, NCAM/CD56, and neurofilament), and extracellular matrix (vimentin) of the dental pulp was performed. The morphologic study identified preservation of the microvasculature, nerve bundles, and components of the extracellular matrix in all studied samples. The immunohistochemical analysis confirmed the morphologic findings and showed a normal pattern of expression for the studied proteins in all samples. Direct effects of radiotherapy are not able to generate morphologic changes in the microvasculature, innervation, and extracellular matrix components of the dental pulp in head and neck cancer patients.

Efeito de baixas doses de radiação X em feridas suturadas e não suturadas - estudo experimental em ratos

The present work evaluated the effect of low doses of X-irradiation on the repairing process of sutured and nonsutured skin wounds in rats. For that, rats underwent a surgical proceedure, in which a 20 x 5-millimeter rectangular wound approximately 2-millimeter-deep was made in the dorsal region of each animal, and were divided in four groups: nonirradiated nonsutured; irradiated nonsutured ; nonirradiated sutured and irradiated sutured. The animals under irradiation were protected, during exposure, with a 2-millimeter-thick lead apron in such a way that only the incision was irradiated. Each animal was submitted to 18 seconds of exposure, undergoing a total of 7.4 rads. The evaluation of the effects of X-rays on the repairing process was carried out through microscopic observation by means of hematoxylin-eosin staining for morphological evaluation, and silver impregnation under polarized light for the observation of collagen synthesis. The results have shown that X-irradiation has caused delay in the repairing process, but it did not stop its development. The irradiated nonsutured group was considered to show the greater delay when compared with the other groups.

Effectiveness of different final irrigation protocols in removing debris in flattened root canals

This study evaluated in vitro the capacity of debris removal from the apical third of flattened root canals, using different final irrigation protocols. Thirty human mandibular central incisors with a mesiodistal flattened root were prepared using rotary instrumentation by Endo-Flare 25.12 and Hero 642 30.06, 35.02, 40.02 files, irrigated with 2 mL of 1% NaOCl after each file. The specimens were randomly distributed into 5 groups according to the final irrigation of root canals: Group I: 10 mL of distilled water (control), Group II: 10 mL of 1% NaOCl for 8 min, Group III: 2 mL of 1% NaOCl for 2 min (repeated 4 times), Group IV: 10 mL of 2.5% NaOCl for 8 min, and Group V: 10 mL of 2.5% NaOCl for 2 min (repeated 4 times). The apical thirds of the specimens were subjected to histological processing and 6-μm cross-sections were obtained and stained with hematoxylin-eosin. The specimens were examined under optical microscopy at ×40 magnification and the images were subjected to morphometric analysis using the Scion image-analysis software. The total area of root canal and the area with debris were measured in square millimeters. Analysis of variance showed no statistically significant difference (p>0.05) among the groups GI (2.39 ± 3.59), GII (2.91 ± 2.21), GIII (0.73 ± 1.36), GIV (0.95 ± 0.84) and GV (0.51 ± 0.22). In conclusion, the final irrigation protocols evaluated in this study using the Luer syringe presented similar performance in the removal of debris from the apical third of flattened root canals.

Alterações do trato digestório de cães da raça Golden Retriever afetados pela distrofia muscular

O modelo experimental canino Golden Retriever portador da Distrofia Muscular (GRMD) é o melhor substituto entre os modelos animais para estudar a Distrofia Muscular de Duchenne. Além da musculatura estriada, a doença pode afetar a musculatura estriada cardíaca e a musculatura lisa, e desta forma, o funcionamento do trato digestório, já que o músculo liso é o elemento primário dos órgãos tubulares. Através de estudo morfológico descritivo, o objetivo deste trabalho foi verificar se a distrofia muscular afeta a arquitetura geral do trato digestório e como se dispõe sua estrutura muscular em animais afetados. Foram realizadas avaliações descritivas macro e microscópicas com colorações de Hematoxilina-Eosina, Tricrômio de Masson e Picrosirius. Entre os resultados apresentados, verificou-se que o esôfago e o fígado dos animais afetados encontraram-se alterados, assim como o estômago não ocupava seu lugar habitual. O músculo diafragma apresentava-se atrofiado e diferenças histológicas foram encontradas na camada muscular do sistema gastrointestinal, em geral. Outras estruturas do tubo digestório de GRMDs apresentaram-se de maneira similar a de um animal normal.

Desenvolvimento morfológico dos ovários em fetos eqüinos sem raça definida

O objetivo deste trabalho foi caracterizar histologicamente o desenvolvimento ovariano pré-natal em fetos eqüinos sem raça definida (SRD) nos trópicos, ressaltando surgimento e quantificação de ovogônias e folículos. Foram obtidos 131 fetos em frigorífico, os quais foram mensurados quanto ao comprimento (cm) equivalente à distância cefalococcígea (CR), seguido de corte das gônadas e fixação das amostras em Bouin por 24 horas. As idades foram estimadas por regressão. Foram preparados cortes histológicos ovarianos de 7µm de espessura, os quais foram corados com hematoxilina-eosina. O surgimento de ovogônias, folículos primordiais, em crescimento e antrais foi verificado, respectivamente, em fetos com CR entre 4,1 e 6,5cm (40 a 49 dias de gestação (DG)); 9,1 e 12,0cm (60 a 69 DG); 23,1 e 29,0cm (116 a 135 DG) e 50,1 e 65,0cm (201 a 245 DG). A quantidade máxima de ovogônias, folículos primordiais, em crescimento e antrais foi observada em fetos com CR entre, 4,1 e 6,0cm (40 a 49 DG); 16,1 e 23,0cm (90 a 115 DG); 29,1 e 36,0cm (136 a 155 DG) e 50,1 e 97,0cm (201 a 320 DG), respectivamente. Os resultados sugerem que o desenvolvimento morfológico dos ovários fetais de eqüinos SRD nos trópicos é mais precoce que o observado nos relatos de literatura referentes a estudos de eqüinos de diversas raças em regiões temperadas.

Morphology of the male agouti accessory genital glands (Dasyprocta prymnolopha Wagler, 1831)

The morphology of the accessory genital glands of the male agouti was studied in twenty-three animals that were raised in captivity. Twenty animals had their genital glands dissected in situ for macroscopic description. The samples of each gland were recovered, embedded in paraffin, sliced and stained by Hematoxylin-Eosin technique. It was founded four pairs of glands: the vesicular glands, the coagulating glands, the prostate and the bulbourethral glands. Histological characteristics of the vesicular, coagulating and prostate glands showed similar morphology, within the pseudostratified columnar epithelium. The tubulo-alveolar type of the bulbourethral glands showed a lack of connective tissue among the tubules, a small amount of red stained presented it the cytoplasm, and the presence of vacuoles in the tissue. This study concluded that the agouti showed to have similar morphological aspect described in the others species of rodents.

Effect of GaAlAs Laser Irradiation on the Epiphyseal Cartilage of Rats

Objective: To study the effect of an 830-nm gallium-aluminum-arsenic (GaAlAs) diode laser at two different energy densities (5 and 15 J/cm(2)) on the epiphyseal cartilage of rats by evaluating bone length and the number of chondrocytes and thickness of each zone of the epiphyseal cartilage. Background Data: Few studies have been conducted on the effects of low-level laser therapy on the epiphyseal cartilage at different irradiation doses. Materials and Methods: A total of 30 male Wistar rats with 23 days of age and weighing 90 g on average were randomly divided into 3 groups: control group (CG, no stimulation), G5 group (energy density, 5 J/cm(2)), and G15 group (energy density, 15 J/cm(2)). Laser treatment sessions were administered every other day for a total of 10 sessions. The animals were killed 24 h after the last treatment session. Histological slides of the epiphyseal cartilage were stained with hematoxylin-eosin (HE), photographed with a Zeiss photomicroscope, and subjected to histometric and histological analyses. Statistical analysis was performed using one-way analysis of variance followed by Tukey's post hoc test. All statistical tests were performed at a significance level of 0.05. Results: Histological analysis and x-ray radiographs revealed an increase in thickness of the epiphyseal cartilage and in the number of chondrocytes in the G5 and G15 groups. Conclusion: The 830-nm GaAlAs diode laser, within the parameters used in this study, induced changes in the thickness of the epiphyseal cartilage and increased the number of chondrocytes, but this was not sufficient to induce changes in bone length.

Esophageal Ulcer in Brazilian Patients with HIV: Prevalence and Comparative Analysis Among Diagnostic Methods

Esophageal ulcer (EU) represents an important comorbidity in AIDS. We evaluated the prevalence of EU, the accuracy of the endoscopic and histologic methods used to investigate viral EU in HIV-positive Brazilian patients and the numerical relevance of tissue sampling. A total of 399 HIV-positive patients underwent upper gastrointestinal (UGI) endoscopy. HIV-positive patients with EU determined by UGI endoscopy followed by biopsies were analyzed by the hematoxylin-eosin (HE) and immunohistochemical (IH) methods. EU was detected in 41 patients (mean age, 39.2 years; 23 males), with a prevalence of 10.27%. The median CD4 count was 49 cells/mm(3) (range, 1-361 cells/mm(3)) and the viral load was 58,869 copies per milliliter (range, 50-77,3290 copies per milliliter). UGI endoscopy detected 29 of 41 EU suggestive of cytomegalovirus (CMV) infection and 7 of 41 indicating herpes simplex virus (HSV) infection. HE histology confirmed 4 of 29 ulcers induced by CMV, 2 of 7 induced by HSV, and 1 of 7 induced by HSV plus CMV. IH for CMV and HSV confirmed the HE findings and detected one additional CMV-induced case. UGI endoscopy showed 100% sensitivity and 15% specificity for the diagnosis of EU due to CMV or HSV compared to HE and IH. HE proved to be an adequate method for etiologic evaluation, with 87% sensitivity and 100% specificity compared to IH. The number of samples did not influence the etiologic evaluation. The data support the importance of IH as a complementary method for HE in the diagnosis of EU of viral etiology.

Influence of Laser Photobiomodulation on Collagen IV During Skeletal Muscle Tissue Remodeling After Injury in Rats

Objective: The aim of the present study was to determine the effect of GaAlAs low-level laser therapy (LLLT) on collagen IV remodeling of the tibialis anterior (TA) muscle in rats after cryolesion. Background: Considerable interest exists in skeletal muscle regeneration in situations such as repair after exercise-induced muscle injury, after muscle transplantation, in muscular dystrophy, exercise-induced muscle injury, and the recovery of strength after atrophy due to disuse. A number of studies have demonstrated the potential of LLLT in facilitating the muscle-healing process; however, no consensus is found in the literature regarding the best laser-irradiation parameters. Methods: Adult male Wistar rats (n = 45) were used and randomly divided into three groups: control (n = 5); nontreated cryolesioned group (n = 20), and LLLT-cryolesioned group (n = 20). The cryolesioned groups were analyzed at 1, 7, 14, and 21 days after the injury procedure. Laser irradiation was performed 3 times per week on the injured region by using the GaAlAs laser (660 nm; beam spot of 0.04 cm(2), output power of 20 mW, power density of 500 mW/cm(2), and energy density of 5 J/cm(2), for 10 sec). The muscles were removed, frozen, cryosectioned, and then stained with hematoxylin-eosin for the visualization of general morphology or used for immunohistochemical analysis of collagen IV. Results: It was demonstrated that LLLT promotes an increase in collagen IV immunolabeling in skeletal muscle in the first 7 days after acute trauma caused by cryoinjury, but does not modify the duration of the tissue-repair process. Even with LLLT, the injured muscle tissue needs similar to 21 days to achieve the same state of organization as that in the noninjured muscle. Conclusion: The collagen IV content is modulated in regenerating skeletal muscle under LLLT, which might be associated with better tissue outcome, although the histologic analysis did not detect tissue improvement in the LLLT group.

Lercanidipine reduces matrix metalloproteinase-2 activity and reverses vascular dysfunction in renovascular hypertensive rats

Increased expression/activity of matrix metalloproteinases (MMPs), especially MMP-2, plays a role in the vascular alterations induced by hypertension, and increased oxidative stress is a major factor activating MMPs. Here, we hypothesized that lercanidipine, a calcium channel blocker, could attenuate the increases in oxidative stress and MMP-2 expression/activity in the two-kidney, one-clip (2K-1C) hypertensive rats. Sham-operated or 2K-1C hypertension rats were treated with lercanidipine 2.5 mg/kg/day (or vehicle) starting three weeks after hypertension was induced. Systolic blood pressure was monitored weekly. After five weeks of treatment, aortic rings were isolated to assess endothelium-dependent and independent relaxations. Quantitative morphometry of structural changes in the aortic wall were studied in hematoxylin/eosin sections. Aortic MMP-2 levels were determined by gelatin zymography. Aortic MMP-2/tissue inhibitor of metalloproteinases (TIMP)-2 mRNA levels were determined by quantitative real-time RT-PCR. Plasma thiobarbituric acid reactive substances concentrations were determined using a fluorometric method. Lercanidipine attenuated 2K-1C hypertension (224 12 versus 183 11 mm Hg in 2K-1C rats and 2K-1C + Lercandipine rats, respectively; P < 0.01) and prevented the reduction in endothelium-dependent vasorelaxation found in 2K-1C rats. Increased MMP-2 and Pro-MMP-2 levels were found in the aortas of 2K-1C rats (all P < 0.05). Lercandipine attenuated 2K-1C-induced increases in MMP-2 by more than 60% and blunted 2K-1C-induced increases in oxidative stress (both P < 0.001). While hypertension-induced significant aortic wall hypertrophy and approximately 9-fold increases in the ratio of MMP-2MMP-2 mRNA expression (both P < 0.05), lercandipine did not affect these changes. These results suggest that lercanidipine produces antihypertensive effects and reverses the endothelial dysfunction associated with 2K-1C hypertension, probably through mechanisms involving antioxidant effects leading to lower MMP-2 activation. (C) 2008 Elsevier B.V. All rights reserved.

Antioxidant treatment reduces matrix metalloproteinase-2-induced vascular changes in renovascular hypertension

Mounting evidence indicates that structural and functional vascular changes associated with two-kidney, one-clip (2K-1C) hypertension result, at least in part, from altered activity of matrix metalloproteinases (MMPs). Because MMPs are upregulated by increased formation of reactive oxygen species (ROS), we hypothesized that antioxidant approaches could attenuate the increases in MMP-2 expression/activity and the vascular dysfunction and remodeling associated with 2K-1C hypertension. Sham-operated or 2K-1C hypertensive rats were treated with tempol 18 mg/kg/day or apocyanin 25 mg/kg/day (or vehicle). Systolic blood pressure was monitored weekly. After 8 weeks of treatment, aortic rings were isolated to assess endothelium-dependent and -independent relaxation. Quantitative morphometry of structural changes in the aortic wall was studied in hematoxylin/eosin sections. Aortic and systemic ROS levels were measured using dihydroethidine and thiobarbituric acid-reactive substances, respectively. Aortic MMP-2 levels and activity were determined by gelatin and in situ zymography, fluorimetry, and immunohistochemistry. Tempol and apocyanin attenuated 2K-1C hypertension (181 +/- 20.8 and 192 +/- 17.6 mm Hg, respectively, versus 213 +/- 18 mm Hg in hypertensive controls; both p<0.05) and prevented the reduction in endothelium-dependent vasorelaxation found in 2K-1C rats. Tempol, but not apocyanin (p>0.05), prevented the vascular remodeling found in 2K-1C rats (all p<0.01). Tempol was more effective than apocyanin in attenuating hypertension-induced increases in oxidative stress (both p<0.05), MMP-2 levels, and MMP-2 activity in hypertensive rats (all p<0.05). Our results suggest that antioxidant approaches decrease MMP-2 upregulation and attenuate the vascular dysfunction and remodeling during 2K-1C hypertension. (C) 2009 Elsevier Inc. All rights reserved.

Synchronous adenocarcinoma of the major and minor duodenal papilla

A 50-year-old woman presented with pancreatitis, fluctuant jaundice, weight loss, and abdominal pain. Contrast-enhanced computed tomography and abdominal ultrasound showed slight dilatation of the biliary tree and gallbladder without calculi. Endoscopy demonstrated a tumor protruding from the papilla of Vater. First endoscopically biopsy diagnosed no tumor, and a second biopsy diagnosed as papillary adenocarcinoma. The patient underwent duodenopancreatectomy. The specimen was fixed in formalin (10%). The tissue was processed routinely, and paraffin sections were stained with hematoxylin-eosin and periodic acid Schiff. Gross examination showed two tumors seen as prolapsed nodules growing isolated from the minor and major duodenal papillae measuring 1.5 and 1.0 cm, respectively, both covered by duodenal mucosa and the histologic study of both lesions demonstrated a moderately differentiated tubular adenocarcinoma, which invaded duodenal wall. After surgery, she is alive 24 months without evidence of recurrence.