Identification of psoralen loaded PLGA microspheres in rat skin by light microscopy

Drug delivery systems involving the use of polymers are widely studied and discovery of biocompatible polymers has become the focus of research in this area. Psoralen loaded poly(DL-lactide-co-glycolide) (PLGA) microspheres to be used in PUVA therapy (psoralen and UVA irradiation (ultraviolet A, 320-400 nm) of psoriasis were identified in paraffin sections by histological analysis. The psoralen loaded PLGA microspheres were prepared using the solvent evaporation technique. They were spherical and possessed an external smooth surface as observed by scanning electron microscopy (SEM) analysis. This study describes a modification in the routine preparation of microsphere samples for examination by light microscopy. The changes involved fixative agents and/or stains allowing the identification of microspheres containing a non-fluorescent material. The preservation and identification of microspheres in tissues for histological processing in paraffin was greatly improved by these modifications as proven by our results. (c) 2007 Elsevicr Ltd. All rights reserved.

Expression of human leucocyte antigen-G primarily targets affected skin of patients with psoriasis

P>BackgroundThe nonclassical human leucocyte antigen (HLA)-G molecule has been well recognized as a tolerogenic molecule and few studies have evaluated the role of the molecule in inflammatory cutaneous autoimmune diseases.ObjectivesTo evaluate the expression of HLA-G in skin specimens of patients with psoriasis and to analyse its correlation with epidemiological and clinical variables.MethodsThirty untreated patients with psoriasis and 32 healthy individuals were enrolled. Immunohistochemistry was applied to identify HLA-G expression in formalin-fixed paraffin-embedded cutaneous skin biopsies.ResultsSoluble and membrane-bound HLA-G expression was detected in 30 (90%) of the skin specimens from patients presenting clinical and histopathological features of psoriasis. Although infiltrating lymphomononuclear cells of the dermis exhibited HLA-G expression, the epidermis was primarily targeted. HLA-G expression was also observed in 27% (three of 11) of the specimens that exhibited no clinical and histopathological features of psoriasis (nonaffected areas). In contrast, skin specimens obtained from healthy individuals exhibited no HLA-G expression (P < 0 center dot 0001). The intensity of HLA-G expression was not associated with type I/II psoriasis, Psoriasis Area and Severity Index score or clinical forms.ConclusionsAs the HLA-G molecule was consistently expressed in affected and, to a lesser extent, in nonaffected areas of untreated patients with psoriasis, irrespective of the severity of the clinical variants, one may hypothesize that the presence of HLA-G may be responsible, at least in part, for the regulation of autoimmune effector cells.

Galectin-3 Overexpression in Invasive Laryngeal Carcinoma, Assessed by Computer-assisted Analysis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Detection of HPV in mouth floor squamous cell carcinoma and its correlation with clinicopathologic variables, risk factors and survival

The human papillomavirus (HPV) has been historically associated with head and neck cancers, although its role in oral carcinogenesis remains poorly defined. The purpose of this study was to investigate the prevalence of HPV in mouth floor squamous cell carcinoma and correlate it with clinicopathologic variables, risk factors and survival. HPV presence was evaluated by nested polymerase chain reaction (nPCR) in 29 paraffin-embedded specimens of mouth floor squamous cell carcinoma. HPV DNA was detected in 17.2% (5 of 29) of the specimens; the highest prevalence was observed in non-smoking patients over the age of 60 years. All HPV DNA positive specimens were detected in men with clinical stage III and IV lesions, being most of which were moderately differentiated. Despite this correlation there were no statistically significant differences observed among the analyzed variables, including patients' survival. The relatively low incidence of HPV DNA present in these tumors suggests that this virus does not, by itself, have a significant role in the development of mouth floor squamous cell carcinoma. J Oral Pathol Med (2008) 37: 593-598

Chromosomal imbalances are uncommon in chagasic megaesophagus

Background: Chagas' disease is a human tropical parasitic illness and a subset of the chronic patients develop megaesophagus or megacolon. The esophagus dilation is known as chagasic megaesophagus (CM) and one of the severe late consequences of CM is the increased risk for esophageal carcinoma (ESCC). Based on the association between CM and ESCC, we investigated whether genes frequently showing unbalanced copy numbers in ESCC were altered in CM by fluorescence in situ (FISH) technology.Methods: A total of 50 formalin-fixed, paraffin-embedded esophageal mucosa specimens (40 from Chagas megaesophagus-CM, and 10 normal esophageal mucosa-NM) were analyzed. DNA FISH probes were tested for FHIT, TP63, PIK3CA, EGFR, FGFR1, MYC, CDKN2A, YES1 and NCOA3 genes, and centromeric sequences from chromosomes 3, 7 and 9.Results: No differences between superficial and basal layers of the epithelial mucosa were found, except for loss of copy number of EGFR in the esophageal basal layer of CM group. Mean copy number of CDKN2A and CEP9 and frequency of nuclei with loss of PIK3CA were significantly different in the CM group compared with normal mucosa and marginal levels of deletions in TP63, FHIT, PIK3CA, EGFR, CDKN2A, YES and gains at PIK3CA, TP63, FGFR1, MYC, CDNK2A and NCOA3 were detected in few CM cases, mainly with dilation grades III and IV. All changes occurred at very low levels.Conclusions: Genomic imbalances common in esophageal carcinomas are not present in chagasic megaesophagus suggesting that these features will not be effective markers for risk assessment of ESCC in patients with chagasic megaesophagus.

Glycogen distribution in ameloblasts and odontoblasts of newborn rats of sialoadenectomized dams. Histochemical study

In order to get information about the distribution of glycogen in ameloblasts and odontoblasts, studies were made of newborn rats of sialoadenectomized dams and newborn rats of control dams. Rodent offspring were decapitated on the 1st, 3rd, 5th, 7th, and 9th days after birth. Their heads were fixed in 10% neutral formalin solution, decalcified in sodium citrate-formic acid and embedded in paraffin, and frontal sections were prepared. Sections 6 micrometers thick were stained by specific histochemical reactions to detect glycogen. Based on the results obtained, it was concluded that the amount of glycogen was lower in the cytoplasms of ameloblasts and odontoblasts of experimental animals when compared to controls.

Peripheral giant cell granuloma. An immunohistochemical and ultrastructural study.

OBJECTIVE: To study the nature of multinucleated and mononuclear cells from peripheral giant cell granuloma (PGCG). MATERIALS AND METHODS: Formalin-fixed, paraffin-embedded sections of 40 cases of PGCG were immunohistochemically stained for vimentin, alpha I-antichymotrypsin, CD68, S-100 protein, lysozyme, leucocyte common antigen (LCA), factor VIII-related antigen and muscle cell actin. Six cases of PGCG were also studied by transmission electron microscopy. RESULTS: Vimentin, alpha I-antichymotrypsin and CD68 were expressed in both the mononuclear and multinucleated giant cells. Dendritic mononuclear cells, positive for S-100 protein, were noted in 67.5% of the lesions, whereas lysozyme and leucocyte common antigen were detected in occasional mononuclear cells. Ultrastructural examination showed mononuclear cells with signs of phagocytosis and sometimes interdigitations with similar cells. Others presented non-specific characteristics and the third type exhibited cytoplasmic processes and occasional Birbeck granules. Some multinucleated giant cells showed oval nuclei, abundant mitochondria and granular endoplasmic reticulum whereas others presented with irregular nuclei and a great number of cytoplasmic vacuoles. CONCLUSIONS: Immunohistochemical and ultrastructural results suggest that PGCGs of the jaws are composed mainly of cells of the mononuclear phagocyte system and that Langerhans cells are present in two thirds of the lesions.

Reorganizacao do processo condilar da articulacao temporomandibular apos condilectomia unilateral, em camundongos tratados com zidovudina (AZT)

In order to analyze the reorganization of the condylar process after unilateral condylectomy in animals which have received AZT, 30 albino mice, 30 days old were used. The condylectomized animals were divided in two groups, one have received distilled water and the other AZT, orally during 10 days. After 5, 10, 15, 30 and 45 days of the surgery the animals of both groups were sacrificed, their heads removed and fixed in 10% formalin. After decalcification the pieces received histological routine treatment to be included in paraffin. The slices were stained by hematoxylin/eosin method. The analysis of results showed that: 1. the condylar repair is similar on both groups; 2. the results reaffirm those found in the literature, that the articulation after condylectomy is located in an anterior position and that the articular disc is not a determinant factor on condylar reorganization.

Apoptosis in the early developing periodontium of rat molars

Development of the periodontium involves a series of complex steps that result in the formation of root dentine, cementum, bone and fibres of the ligament. These precisely controlled and timed events require the participation of the enamel organ derived epithelial cells of Hertwig's (HRS) and ectomesenchymal cells of the dental follicle. These events involve rapid turnover of the tissues and cells, including disappearance of epithelial cells of HRS. Thus, it seemed likely to us that programmed cell death (apoptosis) may play a role in the development of the periodontium. Fragments of first molars, obtained from 14- and 29-day-old rats, were fixed in glutaraldehyde-formaldehyde and processed for light and electron microscopy. For the TUNEL method for detection of apoptosis, specimens were fixed in 4% formaldehyde and embedded in paraffin. Results confirmed that epithelial cells of HRS maintain a close relationship with the forming dentine root, and that they may become trapped in the dentino-cemental junction. Some of the epithelial cells exhibited ultrastructural features which are consistent with the interpretation that they were undergoing programmed cell death, i.e. apoptosis. Periodontal fibroblast-like cells showed typical images of apoptosis and engulfed apoptotic bodies. TUNEL positive structures were present in all corresponding regions. It seems therefore that apoptosis of epithelial cells of HRS and fibroblast-like cells of the periodontal ligament constitutes an integral part of the developmental process of the tissues of the periodontium. (C) 2000 Wiley-Liss, Inc.

Nuclear phenotypes and morphometry of human secretory prostate cells: A comparative study of benign and malignant lesions in Brazilian patients

Prostatic lesions in Brazilian patients with benign prostatic hyperplasia (BPH, 26 cases) or adenocarcinoma (AC, 25 cases) were compared by qualitative microscopy and morphometric analysis. In 12 cases of BPH, prostate regions with no histological alterations were considered as controls (Ct). Archival material consisted of formalin-fixed, paraffin-embedded specimens obtained from prostatic transurethral resection and radical prostatectomy. Haematoxylin/eosin (HE)-stained sections were used to estimate the nuclear areas, perimeters and form factor values. HE-stained sections from AC specimens were also used for Gleason grading. BPH, AC and Ct could be discriminated by their nuclear areas and nuclear perimeters, but not by the nuclear form factor parameter. No significant differences were found when the AC data were compared using the combined version or the predominant grade version of the Gleason score (p = 0.8380 for nuclear area; p = 0.6076 for nuclear perimeter; p = 0.9202 for nuclear form factor; n = 200 nuclei per patient). This finding indicates that there is extensive heterogeneity in the size and shape of the nucleus in AC cells. These results also show that although the nuclear morphometry served to discriminate BPH and AC from each other and from Ct, it was not sufficient to correlate AC lesions with their respective Gleason scores in the human population analyzed.

Influence of age on combined effects of cyclosporin and nifedipine on rat alveolar bone

Background: There is some evidence showing that cyclosporin A (CsA) and nifedipine (NIF) affect bone metabolism. The purpose of this work was to study the effects of CsA and NIF, given alone or concurrently, on alveolar bone of rats of different ages. Methods: Rats 15, 30, 60, and 90 days old were treated daily with 10 mg/kg body weight of CsA subcutaneously injected and/or 50 mg/kg body weight of NIF/day given orally for 60 days. Alveolar bone of the first lower molars was morphologically and stereologically evaluated in serial 5 μm bucco-lingual paraffin sections, stained with hematoxylin and eosin. Serum calcium and alkaline phosphatase levels were measured in all animals at the end of the experimental period. Results: Rats treated with CsA or NIF alone or CsA and NIF concurrently showed decreased alveolar bone density. CsA was more effective than NIF. A significant decrease in serum calcium was found only in animals treated with CsA or CsA/NIF. The results were similar regardless of age. Conclusions: These results indicate that the decrease in the alveolar bone volume in rats caused by CsA and NIF alone or concurrently is not age dependent. Furthermore, NIF (50 mg/kg) did not further increase the loss of alveolar bone volume induced by CsA (10 mg/kg).

Anatomia foliar com ênfase nos tricomas secretores e análise cromatográfica do óleo essencial de Melissa officinalis L. (Lamiaceae)

Adult leaves of Melissa officinalis L. (Lamiaceae) harvested during the time of budding had been used in the study of the leaf anatomy in the identification of the secretory structures. The material was fixed in FAA 50%, dehydrated in alcoholic series (ethyl), infiltrated in paraffin, sectioned at 13 mm, staining and later analyzed through optic microscopy. Also it was carried through a study in gaseous chromatography for attainment of the rude essential oil. Analyses of transversal sections of the leaf of Melissa officinalis, has identified the presence of two types of trichomes secretory: peltate and capitate, beyond the presence of trichome tector. Trichome capitate, identified in literature as (type I), presents variations in its morphology in relation to the cells number stalk and the head secretory cells number. The chromatographic analysis of the essential oil identified the presence of monoterpenes, in two major components, responsible for more than 87.8% of the relative composition in the rude oil, beyond sesquiterpenes in smaller proportions.

Interação de Paracoccidioides brasiliensis com células endoteliais

Paracoccidioidomycosis has a variety of clinical manifestations and Paracoccidioides brasiliensis, the causative agent, may infect many tissues, most importantly the lungs. Migration of pathogenic yeasts to the endothelial cell layer is considered a prerequisite for multiple organ invasion and dissemination of the fungus. In this study of the adhesion of P. brasiliensis to endothelial cells in vitro, we investigated whether this adhesion could represent a mechanism of dissemination. To this end, as well as using conventional optical microscopy, an alternative in vivo technique was developed, to detect the presence of fungal cells in umbilical cords embedded in paraffin wax. An experiment on the migration of P. brasiliensis through an endothelial cell monolayer was carried out, and the migration of yeast cells was greater, and took less time, in control wells with no cells. The fungus crossed the monolayer, but, compared to control wells, the migration-rate was about 30% lower. This shows that the monolayer only partially blocked migration of the fungus. In these experiments, we had great difficulty finding P. brasiliensis adhered to the cell monolayer, when it was examined at different times, suggesting that migration of the fungus across the endothelial layer is very fast, and cannot normally be observed in cell culture in vitro. Thus, P. brasiliensis can cross the endothelium rapidly and probably invades deeper tissue.

Expression of betacatenin in carcinoma in pleomorphic adenoma, pleomorphic adenoma and normal salivary gland: an immunohistochemical study

OBJECTIVES: Pleomorphic adenomas are the most frequent type of epithelial salivary gland neoplasms, and their malignant counterpart, the carcinoma in pleomorphic adenomas, is much less common. Beta-catenin is a cell adhesion molecule associated with the invasion and metastasis of carcinomas of the head and neck, esophagus. The objective of this study was to detect the expression of beta-catenin in pleomorphic adenomas, carcinomas in pleomorphic adenomas and normal salivary glands to discuss its role in the development of these two lesions. STUDY DESIGN: The expression of beta-catenin (BD Transduction Laboratories) was analyzed by immunohistochemistry in formalin-fixed, paraffin embedded specimens by the avidin-biotin-peroxidase complex method in 16 pleomorphic adenomas (12 from minor salivary glands), 3 carcinomas in pleomorphic adenomas (all from palate) and 10 normal salivary glands as control group (5 from major and 5 from minor salivary glands). RESULTS: All cases of glands, adenomas and carcinomas in pleomorphic adenomas have membranous and cytoplasmic immunostaining. Nuclear beta-catenin immunostaining was not observed. The antibody presented a fine granular arrangement in the cytoplasm and cellular membrane of duct and acinic cells. Higher beta-catenin index rates were seen mainly in salivary gland ducts and in ductal structures in the adenomas and carcinomas in pleomorphic adenomas. There was protein loss in pleomorphic adenomas and cytoplasmic accumulation in carcinoma in pleomorphic adenomas. CONCLUSIONS: The present study showed participation of the loss of beta-catenin adhesion molecule in the development of pleomorphic adenoma, and that the cytoplasmic accumulation of the molecule takes part in the malignant transformation of the pleomorphic adenoma into carcinoma in pleomorphic adenoma.

Effects of tissucol™ and epsilon aminocaproic acid in the healing process following dental extraction in dehydrated rats

A histological study was conducted of the alveolar bone healing process following tooth extraction of dehydrated rats after the implantation of fibrin adhesive (TISSUCOL™) associated to previous irrigation of the wound with a 5% epsilon aminocaproic acid solution (EACA). Seventy two rats were used, divided into three groups receiving different treatments after the surgical procedure. In group I, the gingival mucosa was sutured after extraction of the right upper incisor. In groups II and III, chronic dehydration was produced by water deprivation for 9 days (3 days in the preoperative period and 6 days in the postoperative period). In the animals of Group II, after tooth extraction, the gingival mucosa was sutured in the same way as performed in group I. In group III, after extraction, the dental socket was irrigated with 5% EACA, followed by implantation of the fibrin adhesive (TISSUCOL™). The mucosa was sutured in the same way as performed in the other groups. At 3, 7, 15 and 21 postoperative days, the animals were sacrificed in number of 6 for each group. Specimens containing the dental socket were removed and fixed in 10% formalin and decalcified in an equal part formic acid and sodium citrate solution. After routine processing, the specimens were embedded in paraffin for microtomy. We obtained 6 μm semi-serial slices that were stained with hematoxylin and eosin for histological evaluation. The results showed that the water deprivation in the pre- and postoperative periods caused a delay in the alveolar bone healing process. The use of the fibrin adhesive (TISSUCOL™) produced an improvement in the fibrinolytic picture caused by dehydration.