Carmelita de Lima Bezerra Cavalcanti

Acridinium ester conjugated to lectin as chemiluminescent histochemistry marker

Abstract Cell differentiation/dedifferentiation includes changes in oligosaccharide composition and distribution in the cell surface glycoconjugates. Lectins have been used as auxiliary tools in histopathological diagnosis of mammary, uterus and brain pathologies. Acridinium ester (AE) conjugated to biomolecules has been employed in chemiluminescent analytical applications. This work aimed to use a lectin, concanavalin A (Con A), conjugated to AE as a chemiluminescent histochemistry tool. Biopsies of normal and infiltrating duct carcinoma (IDC) of mammary tissues were treated by a Con A–AE derivative. Photon emission, observed during the breakage of the chemical bound between Con A and AE, was quantified, expressed in relative light units (RLU) and correlated to the labelling of the normal and transformed tissues. The results demonstrated that RLU presented a linear relationship with the labelled tissue area in the range 0.125–1.0 cm2 (r=0.98). Furthermore, RLU was much higher for the IDC (1283.920×103±220.621×103) than the normal tissue (2.565×103±0.247×103), namely, about 500 times higher. The Con A–AE conjugation efficiency, differential staining of normal and IDC tissues, and quantification of results contribute to a decrease in the subjectivity in routine histopathological diagnoses and indicate that acrydinum ester can join other lectin marker to be used in histochemistry.

Chemiluminescent Detection of Carbohydrates in the Tumoral Breast Diseases

Nowadays, there is an increase of investigations into the fibroadenoma, mainly because some studies have shown that the occurrence of fibroadenoma is linked to an increased risk of developing breast carcinoma. Currently, the chemiluminescence biomarkers are applied for validation methods and screening. Here, a lectin chemiluminescence is proposed as new histochemistry method to identify carbohydrates in mammary tumoral tissues. The lectins concanavalin A (Con A) and peanut agglutinin (PNA) conjugated to acridinium ester were used to characterize the glycocode of breast tissues: normal, fibroadenoma, and invasive duct carcinoma (IDC). The lectin chemiluminescence expressed in relative light units (RLU) was higher in fibroadenoma and IDC than in normal tissue for both lectins tested. The relationship RLU emission versus tissue area described a linear and hyperbolic curve for IDC and fibroadenoma, respectively, using Con A whereas hyperbolic curves for both transformed tissues using PNA. RLU was abolished by inhibiting the interaction between tissues and lectins using their specific carbohydrates: methyl-α-d-mannoside (Con A) and galactose (PNA). The intrinsic fluorescence emission did not change with combination of the lectins (Con A/PNA) to the acridinium ester for hydrophobic residues. These results represent the lectin chemiluminescence as an alternative of histochemistry method for tumoral diagnosis in the breast.

Topical Application Effect of the Isolectin Hydrogel (Cramoll 1,4) on Second-Degree Burns: Experimental Model

This study aimed at evaluating the use of hydrogel isolectin in the treatment of second-degree burns. Twenty male rats were randomly divided into two groups (G1 = treatment with hydrogel containing 100 μg/mL Cramoll 1,4 and G2 = Control, hydrogel). After 7, 14, 21, 28, and 35 days, animals were euthanized. On the 7th day, G1 showed intense exudates, necrosis and edema. On the 14th day, G1 showed tissue reepithelialization and moderate autolysis. On the 21st day, G1 showed intense fibroblastic proliferation, presence of dense collagen, and moderate fibrosis. On the 28th day, G1 showed complete tissue epithelialization. On the 35th day, G1 showed modeled dense collagen. The significant wound contraction was initiated from day, 14 in the G1. There were no significant differences in biochemical and hematological parameters analyzed. These results extend the potential of therapeutic applications for Cramoll 1,4 in the treatment of thermal burns.

Lectin staining patterns in human gastric mucosae with and without exposure to Helicobacter pylori

The aim of the present study was to evaluate qualitative changes in the glycoconjugate expression in human gastric tissue of positive and negative patients for Helicobacter pylori, through lectins: Wheat Germ Agglutinin (WGA) and Concanavalin A (Con A). The lectins recognized differently the glycoconjugates in the superficial mucous layer at the gastric tissues. The results suggest a significant change in the carbohydrate moieties present on the surface of the gastric cells during infection.

Immunohistochemiluminescence detection: A quantitative tool in breast cancer HER-2 status evaluation

Her-2 status evaluation in breast cancer has prognostic and treatment response value but its interobserver variation among pathologists is a problem since it is not quantitatively assayed. This study presents an immunohistochemiluminescence method to quantify Her-2 in breast cancer. Anti-Her-2 antibody was conjugated to acridinium ester (AE) and used to evaluate/quantify Her-2 status in breast Invasive Ductal Carcinoma (IDC, n = 50) comparing with traditional immunohistochemistry. Anti-HER-2-AE results were expressed in Relative Lights Units (RLU) and showed to be able to distinguish and quantify the differences between the three groups of Her-2 status. 3+ Her-2 status presented the highest RLU (246,982 × 103 ± 2.061 × 103) compared to 2+ (76,146 × 103 ± 0.290 × 103), negative (27,415 × 103 ± 1.445 × 103) and normal tissues (27,064 × 103 ± 2.060). Status differences were significant between 3+ and 2+ (p = 0.0025); 2+ and negative (p = 0.0003), and +3 and +1 (p = 0.0001) beside this, normal breast control RLU was 27,064 × 103 ± 2,060 × 103, similar to negative cases. Results showed that anti-HER-2-AE conjugate was effective in breast tumors Her-2 status evaluation, allowing its quantitative establishment to consequently decrease the subjectivity in prognostic and predictive information intrinsic to this test.

Carbohydrates Detection in the Hepatic Egg - Granuloma System Using Lectin Histochemistry

El objetivo del estudio fue evaluar el sistema de los huevos de los granulomas en los tejidos hepaticos, utilizando histoquimica de lectinas esquistosomiasis. Ocho ratones suizos experimentales fueron infectados con una cepa local de Schistosoma mansoni y luego a los cuarenta dias fueron sometidos a la perfusion y se prepararon cortes de higado (4 |J,m). Las muestras de los tejidos fueron incubadas con las siguientes peroxidasas lectinas conjugadas: aglutinina de mani (PNA), aglutinina de germenn de trigo (WGA), Concanavalin A (Con A). Todas las lectinas reconocieron las glicoconjugadas en el tegumento del gusano adulto. El tejido hepatico con WGA presento mayor coloracion seguido de PNA y Con A. El PNA presento la mas intensa tincion de los huevos mientras el granuloma del sistema WGA tino las celulas hepaticas sinusoides y las Con A estuvieron siempre presentes en los anillos de la fibrosis y alrededor de los granulomas hepaticos del parenquima. WGA y PNA indicaron la presencia de residuos de N - acetil - glucosamina y galactosa en la superficie de los huevos de Schistosoma mansoni en los granulomas hepaticos de esquistosomiasis.

Evaluating the effectiveness of gel formulation of irradiated seed lectin Cratylia mollis during bone repair in rats

Context: Regeneration corresponds to the replacement of damaged cells with ones that have the same morphology and function. For experimental evaluation of materials that may favor the process of bone healing, defects are created with dimensions that prevent spontaneous regeneration. For the development and use of new drugs, it is necessary to study its effects in vitro, which depends on the formulation, concentration, and rate of irradiation in vivo and the route and frequency of administration; thus, it is possible to characterize the physiological and molecular mechanisms involved in the response and cellular effects. Objective: The objective of this study was to assess the effect of Cramoll-1,4 on the process of bone repair. Materials and Methods: A formulation of biopharmaceutical lectin Cramoll-1,4 at a concentration of 300 mg/100 mL was applied in a single application via gamma radiation and its effect on the process of bone repair in rats was assessed. Results: Histologically, it was observed that the bone defect is coated by loose connective tissue rich in fibroblasts, providing a range similar to the thick bone original and competing with site of new bone formation. This prevented direct contact between the formulation and experimental bone tissue, as, despite its proven effectiveness in experiments on the repair of skin lesions, the formulation used did not promote bone stimulation that would have promoted the tissue repair process. Conclusion: Because of the direct interference of loose tissue repair that prevented direct contact of the implant with the bone interface, the formulation did not promote bone stimulation.

Comparing the Immunoexpression of FUT3 and FUT6 between Prostatic Adenocarcinoma and Benign Prostatic Hyperplasia.

Prostatic Adenocarcinoma (PA) and Benign Prostatic Hyperplasia (BPH) have their etiology not fully understood mainly in glycidic aspects. Glycan changes are associated with cell alterations where glycosylation is carried out by glycosyltransferases, such as fucosyltransferases (FUTs). These enzymes catalyze the insertion of L-fucose residues in a variety of glycan structures often in the final stage of glycosylation. The present study aimed to investigate the expression of FUT3 and FUT6 in PA and BPH as well as to correlate immunostaining of these transferases with PA clinic-histopathologic data. The FUT3 and FUT6 expressions were evaluated by immunohistochemistry in formalin-fixed, paraffin-embedded biopsies of PA (n=40) and BPH (n=40). FUT3 and FUT6 showed a high expression in both prostatic diseases, especially FUT6. FUT6 was more immunoexpressed in PA cases than the FUT3 (p<0.0001) as well as in BPH cases but in a not significant way (p=0.0661). Besides, FUT3 was more expressed in BHP lesion than in PA cases (p<0.0001). Our study presented a new data about FUT3 and FUT6 expression in PA and BPH, revealing high FUT6 expression in both lesions and FUT3 overexpression in BHP in relation to PA, proposing that this enzyme could be a promising biomarker for benign prostate alterations.

Histopathological and Digital Morphometrical Evaluation of Uterine Leiomyoma in Brazilian Women.

The current study aims to evaluate histopathological and digital morphometrical aspects associated with uterine leiomyomas in one hundred and fifty (150) patients diagnosed with leiomyoma. Uterine tissues were subjected to the histopathological and digital morphometric analyses of the interstitial collagen distribution. The analysis of medical records indicates that most of the women diagnosed with uterine leiomyomas (68.7%) are between 37 and 48 years old. As for the anatomic location of the tumors, approximately 61.4% of the patients had intramural and subserosal lesions. In 50% of the studied cases, the patients developed uterine leiomyomatosis (with more than eight tumors). As for the morphometric study, the average size of the interstitial collagen distribution held approximately 28.53% of the capture area, whereas it was of 7.43% in the normal tissue adjacent to the tumor. Another important aspect observed in the current study was the high rate of young women subjected to total hysterectomy, a fact that resulted in early and definitive sterility.