Anti-cyclin Antibody Research Articles

Bases for a treatment of cutaneous leishmaniasis with moderate hyperthermia.

Regional hyperthermia at between 38 and 39.5 °C has been used to treat inflammatory processes and, occasionally, skin infections. In areas where leishmaniasis is endemic, hot compresses are applied as anti-parasitic treatment. To identify the bases of leishmaniasis thermal treatment in order to properly regulate it. In vitro-cultured Leishmania mexicana parasites were incubated for variable periods at 37 and then at 25 °C. The parasites were then stained with Annexin V-FITC to detect apoptosis induction and with propidium iodide for viability. Post-treatment growth curves and cell cycle identification with anti-cyclin antibodies were performed. After 30 minutes of exposure to a temperature of 37 °C, a variable proportion of parasites lost their characteristic oval shape and became spherical, without refringence and with condensed nuclei, with these changes suggesting apoptosis, which was confirmed by Annexin V-FITC staining. The number of parasites that underwent apoptosis was proportional to exposure time. Parasites in which apoptosis was observed were stained with anti-cyclin antibodies. Constant, regulated and physiological elevation of temperature for more than 30 minutes induces apoptosis of in vitro-cultured L. mexicana parasites when they are in an active phase of the cell cycle.

Antibody delivery into viable epimastigotes of <i>Trypanosoma cruzi</i> as a tool to study the parasite biology

American trypanosomiasis is a zoonosis of worldwide medical importance and currently there is no effective treatment in chronic patients, hence the importance of the study of protein function of the parasite with the objective of finding new drug targets and to know better the biology of the agent causal (Trypano-soma cruzi). T. cruzi is an RNAi-negative parasite, therefore the silencing genes strategies by RNAi is not possible; for that reason, antibodies may be taken as a tool for studying the parasite proteins function by blocking these molecules with specific antibodies. The aim of this work was to establish a methodology for antibody delivery (antibody transfection) into viable parasites. We used anti-cyclin-A antibody (human origin) in western blot assay with epimastigote of T. cruzi proteins and this recognized a ~55 kDa polypeptide. Several methods for antibody transfection (electroporation, saponin permeabilization and a lipid-based formulation) were tested. The first two methods were unsuccessful. In electroporation was impossible to visualize the antibody inside parasites and with saponin permeabilization, antibodies were successfully introduced, but with loss of parasites viability. The lipid-based formulation method forms noncovalent complexes with antibodies. These complexes are internalized by cells and antibodies are released into the cytoplasm. With this method, a successful antibody delivery was achieved. Anti-cyclin antibodies were visualized in the cytoplasm from fixed transfected parasites (immunofluorescence assays). At 24 h post-transfection, parasites maintained their viability (90%) and were able to arrest the cell cycle in G0/G1-phase of cultured epimastigotes (cell population increased in G0/G1-phase from 50.5% to 66.2% and decreased in S-phase from 47.2% to 26%). It was also observed that anti-cyclin-A antibodies inhibit the parasite population doubling (p T. cruzi, with a simple and cheap technique, which will allows carrying out further studies of this protozoan.

Prognostic value of nuclear DNA quantification and cyclin A in epithelial ovarian cancer

5106 Backgrounds: To investigate the relationship between DNA ploidy, S phase fraction(SPF), cyclin A and clinical prognostic factors such as stage, grade and CA 125, and overall survival in epithelial ovarian cancer. Correlations between DNA ploidy, SPF and cyclin A were also assessed. Methods: Study group consisted of 31 cases of epithelial ovarian cancer, 10 of borderline ovarian tumor and 5 of benign ovarian tumor diagnosed from Feb 2000 to Jan 2003. Patients' sera were sampled after 6th postoperative chemotherapy and the level of CA 125 was assessed with cut off value of 35U/ml. DNA ploidy and SPF were evaluated by flowcytometry of fresh ovarian tissue obtained at the operation. The expression of cyclin A was evaluated by conjugating anticyclin antibody, anti-mouse and rabbit-Ig and streptovidin-HRP to paraffin- blocks of ovarian tissue. Expressivity of 5% was considered as positive expression. Two-sample t-test, chi-square test, and Kaplan-Meier survival curve were used for statistics. Results: DNA ...

Reversible Large-Body Formation from Nuclear Bodies upon Amino Acid(s) Starvation in T24 Cells

AP435 dot, a nuclear dot-like structure that is recognized by a monoclonal antibody AP435 MAb and that seems to correlate with perinuclear intermediate filaments, was identified as a nuclear body by double immunofluorescent staining with AP435 MAb and the nuclear-body-specific antibody αSp100 or mAb 5E10. In T24 cells, nuclear bodies usually appear as small entities with an apparent diameter ranging from 0.2 to 0.7 μm, and several to 20 or more of them are present per nucleus. After long culture without a change in the medium, however, nuclear bodies disappeared while one or more large doughnut-shaped bodies appeared, which had apparent outer diameters of 0.7–1.8 μm. When the medium was changed or medium components were added, large bodies disappeared and many nuclear bodies of normal size reappeared within several hours. Large-body formation was not related to the arrest of DNA synthesis, as revealed by double labeling with AP435 MAb and anti-cyclin antibody. Among the medium components, only an amino acid mixture induced the change from large bodies to nuclear bodies. Large-body formation was also observed in long-cultured HeLa cells. These results suggest that nuclear bodies reversibly aggregate or reorganized to form large bodies upon amino acid(s) starvation.