Olcay Semiz

The effect of growth hormone on ovarian follicular response and oocyte nuclear maturation in young and aged mice.

The aim of this study was to investigate the possible effects of recombinant human growth hormone (rhGH) on the ovarian follicle numbers and the nuclear maturity status of ovulated oocytes. Aged and young Balb-C mice were randomly divided into three groups: (i) control, (ii) stimulated with follicle stimulating hormone (FSH) and (iii) stimulated with FSH+rhGH. Ovaries were collected for histological and morphometric investigations. For the determination of nuclear maturation of ovulated oocytes, additional aged and young mice were randomly divided into two groups: (i) stimulated with FSH and (ii) stimulated with FSH+rhGH. In young mice, developing ovarian follicle numbers were significantly higher with rhGH; while in aged mice, there was no statistically significant difference between groups. In young mice co-stimulated with FSH and rhGH, atretic follicle numbers were significantly higher. In aged mice, atretic follicle numbers significantly decreased with FSH while there was a slight decrease with rhGH and this was not significant. The percentage of metaphase-II oocytes were significantly higher in both young and aged mice groups co-stimulated with FSH and rhGH. In conclusion, co-treatment with FSH and rhGH for ovarian stimulation in older mice appears to cause a slight decrease in the number of atretic follicles and thus improves the collection of high numbers of mature, fertilizable oocytes.

Carbofuran Alters Centrosome and Spindle Organization, and Delays Cell Division in Oocytes and Mitotic Cells

Although many countries banned of its usage, carbofuran (CF) is still one of the most commonly used carbamate derivative insecticides against insects and nematodes in agriculture and household, threatening the human and animal health by contaminating air, water, and food. Our goal was to evaluate the potential toxic effects of CF on mammalian oocytes besides mitotic cells. Caspase-dependent apoptotic pathway was assessed by immunofluorescence and western blot techniques. Alterations in the meiotic spindle formation after CF exposure throughout the in vitro maturation of mice oocyte-cumulus complexes (COCs) were analyzed by using a 3D confocal laser microscope. Maturation efficiency and kinetics were assessed by direct observation of the COCs. Results indicated that the number of TUNEL-positive cells increased in CF-exposed groups, particularly higher doses (>250 µM) in a dose-dependent fashion. The ratio of anticleaved caspase-3 labeled cells in those groups positively correlated with TUNEL-positivity. Western blot analysis confirmed a significant increase in active caspase-3 activity. CF caused a dose-dependent accumulation of oocytes at prometaphase-I (PM-I) of meiosis. Partial loss of spindle microtubules (MTs) was noted, which consequently gave rise to a diamond shape spindle. Aberrant pericentrin foci were noted particularly in PM-I and metaphase-I (M-I) stages. Conclusively, CF (1) induces programmed cell death in a dose-dependent manner, and (2) alters spindle morphology most likely through a mechanism that interacts with MT assembly and/or disorientation of pericentriolar proteins. Overall, data suggest that CF could give rise to aneuploidy or cell death in higher doses, therefore reduce fertilization and implantation rates.

Assessment of resting electrocardiogram, P wave dispersion and duration in different genders applying for registration to the School of Physical Education and Sports – results of a single centre Turkish Trial with 2093 healthy subjects

BACKGROUND The 12-lead electrocardiogram shows a broad range of abnormal patterns in trained athletes. The primary end point of this study was to investigate P wave dispersion, and P wave durations and related factors in different genders applying for registration to the School of Physical Education and Sports. METHODS AND RESULTS From 2006 to 2009, a total of 2093 students - 1674 boys with a mean age of 19.8 plus or minus 1.9 years and 419 girls with a mean age of 19.1 plus or minus 1.8 years - were included in the study. All 12 leads of the resting electrocardiogram were evaluated for P wave dispersion and electrocardiogram abnormalities. Baseline parameters such as age, body weight, body height, and body mass index, as well as electrocardiogram findings such as P wave maximal duration and P wave dispersion, were significantly higher in boys than in girls. Of all the parameters tested with correlation analysis, only gender (p = 0.03) (r = 0.04), body weight (p < 0.001) (r = 0.07), body height (p = 0.004) (r = 0.06), and body mass index (p = 0.01) (p = 0.05) were correlated with P wave dispersion. CONCLUSION The frequencies of all electrocardiogram abnormalities, P wave dispersion, and P wave maximal duration were higher in boys as compared with girls in an unselected student population applying for registration to the School of Physical Education and Sports; in addition, P wave dispersion was correlated with gender, body weight, body height, and body mass index.

Two convenient methods for nuclear labeling in confocal microscopy using visible-lasers

Abstract The objective of this study was to implement a simple and reliable fluorescent staining method specific to DNA for the visualization of cell nuclei using 488-nm and 543-nm visible lasers or conventional mercury lamps. The staining properties of two different reagents, chromomycin A, (C-A3) and 7-aminoactinomycin-D (7-AA-D), were analyzed in several types of mammalian cells and tissues with different fixation and preparation methods. Results were obtained using a scanning laser confocal microscope. Experiments were initially conducted in paraformaldehyde-fixed cell cultures to optimize the staining conditions. C-A3 (100 μg/mL) and 7-AA-D (10 μM) showed consistent and specific binding to nuclear DNA and emitted green (-580nm) and red (-647nm) colors, respectively. No dose change was needed for formaldehyde-fixed cells in the same conditions. In paraffin-embedded or frozen sections, C-A3 exhibited optimum labeling at 140 μglmL, whereas 7-AA-D was found optimum at 10 μM concentration. Both dyes displayed similar staining patterns regarding the specificity to chromatin and chromosomes but not to the RNA molecules. Major RNA-containing regions in cells, such as nucleolus or cytoplasm, were entirely devoid of C-A3 and 7-AA-D staining. All stages of mitotic chromosomes, as well as picnotic and apoptotic cells, were successfully labeled. Penetration of dyes for labeling giant cells (e.g., mouse oocytes) was achieved by a mild detergent extraction procedure. C-A3 and 7-AA-D are highly specific, easily handled fluorescent markers in PFA and FA fixed cultured cells, paraffin-embedded, and frozen sections. They can easily be the first choice for a specific DNA labeling as a primary or a counterstain in any fluorescent microscope, especially in confocal microscopes lacking UV laser. (The J Histotechnol 26:147, 2003) Submitted April 17, 2003; Accepted July 31, 2003