Identification of Cryptosporidium spp. oocysts in fecal smears stained with Heidenhain's iron hematoxylin

There is no paucity of methods for diagnosing Cryptosporidium spp. infection. The merits of immunoassays notwithstanding, microscopic identification of Cryptosporidium spp. oocysts in fecal samples remains an important diagnostic procedure. It owes the persistence of its use to such characteristics as dispensing with expensive equipment and kits, requiring only basic laboratory facilities, and having a low probability of false positive results when permanent slides are prepared, which can be re-examined in case of doubt. Cryptosporidium spp. oocysts can be readily identified in fecal smears prepared according to a regressive iron hematoxylin staining technique. The number of steps and their duration, as well as costs, were reduced to a minimum without loss of image quality and permanence of the preparations.

Staining of intestinal protozoa with heidenhain's iron Hematoxylin

Due to its unique properties, iron hematoxylin has been traditionally recommended for staining intestinal protozoa. This process can be simplified by reducing the number of steps and periods of permanence of the slides in some of the liquids used, without detriment to the quality of the results. Thus iron hematoxylin becomes adequate for routine use. Hematoxylin is a natural dye extracted from Haematoxylon campechianum, of the family Leguminosae. It must first be 'ripened', i.e. oxidized to hematein, which reacts with ferric ammonium sulphate to produce the ferric lake (iron hematoxylin), a basic dye. Iron hematoxylin most frequently stains regressively, i.e. the slides are first overstained and then differentiated.

Detection of Dientamoeba fragilis in patients with HIV/AIDS by using a simplified iron hematoxylin technique

INTRODUCTION: Studies strongly indicate Dientamoeba fragilis as one of the causes of diarrhea in human immunodeficiency virus (HIV) patients. METHODS: The objective of the present study was to evaluate the prevalence of D. fragilis associated with the causes of diarrhea in 82 HIV/ AIDS patients hospitalized at the Instituto de Infectologia Emílio Ribas from September 2006 to November 2008. RESULTS: In total, 105 samples were collected from 82 patients. Unprotected sex was the most frequent cause of HIV infection (46.3%), followed by the use of injectable or non-injectable drugs (14.6%). Patients presented with viral loads of 49-750,000 copies/ mL (average: 73,849 ± 124,850 copies/mL) and CD4 counts ranging of 2-1,306 cells/mm³ (average: 159 ± 250 cells/mm³). On an average, the odds of obtaining a positive result by using the other techniques (Hoffman, Pons and Janer or Lutz; Ritchie) were 2.7 times higher than the chance of obtaining a positive result by using the simplified iron hematoxylin method. Significant differences were found between the methods (p = 0.003). CONCLUSIONS: The other techniques can detect a significantly greater amount of parasites than the simplified iron hematoxylin method, especially with respect to Isospora belli, Cryptosporidium sp., Schistosoma mansoni, and Strongyloides stercoralis, which were not detected using hematoxylin. Endolimax nana and D. fragilis were detected more frequently on using hematoxylin, and the only parasite not found by the other methods was D. fragilis.

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.

Testosterone Therapy Differently Regulates The Anti- And Pro-inflammatory Cytokines In The Plasma And Prostate Of Rats Submitted To Chronic Ethanol Consumption (uchb).

Ethanol consumption damages the prostate, and testosterone is known by anti-inflammatory role. The cytokines were investigated in the plasma and ventral prostate of UChB rats submitted or not to testosterone therapy by ELISA and Western blot, respectively. Additionally, inflammatory foci and mast cells were identified in the ventral prostate slides stained by hematoxylin and eosin and toluidine blue, respectively. Inflammatory foci were found in the ethanol-treated animals and absent after testosterone therapy. Plasma levels of IL-6 and IL-10 were not changed while TNFα and TFG-β1 were increased in the animals submitted testosterone therapy. Regarding to ventral prostate, IL-6 did not alter, while IL-10, TNFα, and TFG-β1 were increased after testosterone therapy. Ethanol increases NFR2 in addition to high number of intact and degranulated mast cell which were reduced after testosterone therapy. So, ethanol and testosterone differentially modulates the cytokines in the plasma and prostate.

Desquamation Is A Novel Phenomenon For Collective Prostate Epithelial Cell Deletion After Castration.

The mechanism underlying castration-induced prostate regression, which is a classical physiological concept translated into the therapeutic treatment of advanced prostate cancer, involves epithelial cell apoptosis. In searching for events and mechanisms contributing to prostate regression in response to androgen modulation, we have frequently observed the collective deletion of epithelial cells. This work was undertaken to characterize this phenomenon hereafter named desquamation and to verify its presence after 17β-estradiol (E2) administration. Electron microscopy revealed that the desquamating cells had preserved cell-cell junctions and collapsed nuclear contents. The TUNEL reaction was negative for these cells, which were also negative for cleaved caspases-8, -9, -3 and nuclear apoptosis-inducing factor. Detailed analyses revealed that the condensed chromatin was first affected detaching from the nuclear lamina, which was observable after lamin A immunohistochemistry, suggesting the lack of lamin A degradation. A search in animals treated with supraphysiological E2 employed as an alternative anti-androgen treatment revealed no desquamation. The combined treatment (Cas + E2 group) caused changes particular to each treatment, including desquamation. In conclusion, desquamation appeared as a novel phenomenon contributing to collective prostate epithelial cell deletion, distinct from the classical castration-induced apoptosis and particular to the androgen deprivation resulting from surgical castration, and should be considered as part of the mechanisms promoting organ regression.

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.

Effect Of Particles Of Ashes Produced From Sugarcane Burning On The Respiratory System Of Rats.

The practice of burning sugarcane obtained by non-mechanized harvesting exposes workers and the people of neighboring towns to high concentrations of particulate matter (PM) that is harmful to health, and may trigger a series of cardiorespiratory diseases. The aim of this study was to analyze the chemical composition of the micro-particles coming from sugarcane burning residues and to verify the effects of this micro-particulate matter on lung and tracheal tissues. Micro-particulate matter (PM10) was obtained by dissolving filter paper containing burnt residues in NaCl solution. This material was instilled into the Wistar rats' nostrils. Histological analyses (hematoxylin and eosin - HE) of cardiac, lung and tracheal tissues were performed. Inflammatory mediators were measured in lung tissues by using ELISA. The chemical composition of the particulate material revealed a large quantity of the phthalic acid ester, high concentrations of phenolic compounds, anthracene and polycyclic aromatic hydrocarbons (PAH). Histological analysis showed a reduction in subjacent conjunctive tissue in the trachea, lung inflammation with inflammatory infiltrate formation and reduction of alveolar spaces and a significant increase (p<0.05) in the release of IL-1α, IL-1β, IL-6, and INF-γ in the group treated with PM10 when compared to the control group. We concluded that the burning sugarcane residues release many particles, which have toxic chemical compounds. The micro-particulate matter can induce alterations in the respiratory system.

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.

Periapical repair after root canal filling with different root canal sealers

The aim of this study was to evaluate periapical repair after root canal filling with different endodontic sealers. Sixty-four root canals from dog´s teeth were filled, divided into 4 groups (n=16). Root canals were instrumented with K-type files and irrigated with 1% sodium hypochlorite solution. Root canals were filled in the same session by active lateral condensation of the cones and sealers: Intrafill, AH Plus, Roeko Seal and Resilon/Epiphany System. After 90 days, the animals were euthanized and the tissues to be evaluated were processed and stained with hematoxylin and eosin. For histopathological analysis, the following parameters were evaluated: inflammatory process, mineralized tissue resorption, and apical mineralized tissue deposition. Histopathological analysis demonstrated that Intrafill had less favorable results in terms of apical and periapical repair, compared to the other sealers (p<0.05). AH Plus, Roeko Seal, and Epiphany sealers had similar and satisfactory results (p>0.05). In conclusion, AH Plus and the materials Roeko Seal and Epiphany are good options for clinical use in Endodontics.

Histopathological evaluation of different methods of experimental induction of periapical periodontitis

This study evaluated histopathologically different methods of experimental induction of periapical periodontitis. The radiographic and microbiological evaluations have been performed in a previous investigation. Fifty-seven root canals from dogs' teeth were assigned to 4 groups. In GI (n=14) and GII (n=14), the root canals were exposed to oral environment for 180 days; in GIII (n=14) and GIV (n=15) the root canals were exposed for 7 days and then the access cavities were restored and remained sealed for 53 days. The root apices of GI and GIII were perforated, whilst those of GII and GIV remained intact. After induction of periapical periodontitis, the dogs were euthanized. Serial sections were obtained and stained with hematoxylin and eosin. Data of the histopathological evaluation were submitted to Kruskal-Wallis and Dunn's tests at 5% significance level. The inflammatory periapical reaction and resorption of mineralized tissues were less intense in GII than in the other groups (p<0.05). There was no histopathological difference among the experimentally induced periapical lesions in the teeth with coronal sealing. On the other hand, when coronal sealing was not performed, greater intensity of induced periapical periodontitis was observed in the teeth with apical perforation.

Bone quality associated with daily intake of coffee: a biochemical, radiographic and histometric study

Caffeine induces loss of calcium and influences the normal development of bone. This study investigated the effects of coffee on bone metabolism in rats by biochemical measurement of calcium, bone densitometry and histometry. Male rats, born of female treated daily with coffee and with coffee intake since born, were anesthetized, subjected to extraction of the upper right incisor, and sacrificed 7, 21 and 42 days after surgery. Blood and urine samples were taken, and their maxilla radiographed and processed to obtain 5-µm-thick semi-serial sections stained with hematoxylin and eosin. The volume and bone quality were estimated using an image-analysis software. The results showed significantly greater amount of calcium in the plasma (9.40 ± 1.73 versus 9.80 ± 2.05 mg%) and urine (1.00 ± 0.50 versus 1.25 ± 0.70 mg/24 h) and significantly less amount in bone (90.0 ± 1.94 versus 86.0 ± 2.12 mg/mg bone), reduced bone mineral density (1.05 ± 0.11 versus 0.65 ± 0.15 mmAL), and lower amount of bone (76.19 ± 1.6 versus 53.41 ± 2.1 %) (ANOVA; p≤0.01) in animals treated with coffee sacrificed after 42 days. It may be concluded that coffee/caffeine intake caused serious adverse effects on calcium metabolism in rats, including increased levels of calcium in the urine and plasma, decreased bone mineral density and lower volume of bone, thus delaying the bone repair process.

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.