Effects of Growth Hormone on In Situ Culture of Bovine Preantral Follicles are Dose Dependent.

The objective of this study was to evaluate different concentrations of growth hormone (GH) on the development of bovine preantral follicles cultured included in the ovarian tissue (in situ) on the rates of morphologically normal, viable, primordial and developing follicles, as well as the oocyte and follicle diameter and ultrastructural analysis. Ovarian fragments collected from cows with no cross-breeds defined were cultured in situ for 1 and 7 days in minimal essential medium (α-MEM+) supplemented with different concentrations of recombinant human GH (0, 10, 25, 50 ng/ml). The ovarian fragments non-cultured (control) and cultured were processed for classic histology, mechanical isolation and electron transmission microscopy (MET). The parameters underwent anova (Tukey's and Dunnett's tests) and chi-square test (χ(2) ). After 7 days of culture, the treatment with 50 ng/ml GH showed no differences with fresh control (p > 0.05) and had greater effectiveness than in the 0, 10 and 25 ng/ml GH concentrations of the morphologically normal follicles. Regarding the primordial follicles, a reduction was observed in the 50 ng/ml GH concentration concomitant with the significant increase in developing follicles, differing from both the fresh control and the other GH concentrations tested. In addition, 50 ng/ml GH showed a larger follicle and oocyte diameter when compared to the other treatments cultured. Similar structures were ultrastructurally observed in the control group, 50 ng/ml GH. Follicles cultured in 10 ng/ml GH showed nuclear invagination, vacuoles and lesioned basal membrane. Hence, it is concluded that 50 ng/ml GH is the most effective concentration for the development of preantral follicles cultured in situ.

Effects of IGF-1 on In Vitro Culture of Bovine Preantral Follicles are Dose-Dependent.

This study aimed at assessing the effect of different concentrations of the growth factor similar to insulin 1 (IGF-1) in the development, survival and ultrastructure of the bovine preantral follicles cultured in situ. Fragments of bovine ovarian cortical tissue were cultured during 1 and 7 days in 1 ml of α-MEM(+) , supplemented with different concentrations of human recombinant IGF-1 (0, 30, 70 and 100 ng/ml), in an incubator at 37°C and 5% of CO2 in 24-well plates with total replacement of the medium every 2 days. Non-cultured ovarian fragments (control) and ovarian fragments cultured during 1 and 7 days were processed for classic histology, mechanical isolation and electron transmission microscopy (ETM). Parameters such as normality, viability, activation, development, diameter and ultrastructure were evaluated. All statistical analyses were carried out using sas Version 9.2. The results showed that the percentage of follicles morphologically normal in the IGF-1 30 ng/ml treatment was similar to the fresh control (p > 0.05) both on the day 1 and on the day 7 of in vitro culture. In the viability analysis, the cultured treatments maintained the percentage of viable follicles during the entire culture period (p > 0.05). After 7 days of culture, the IGF-1 30 ng/ml treatment showed higher percentages of developing follicles (48.33%) than those of the fresh control (22.22%) and the cultured treatments (p < 0.05). Also, after 7 days of culture, IGF-1 30 ng/ml presented a higher follicular diameter when compared to the control and other concentrations of IGF-1 tested. Ultrastructurally, the non-cultured control and IGF-1 30 ng/ml, after 7 days of culture, showed conserved oocytes, nuclei and organelles. Hence, it is concluded that IGF-1 30 ng/ml was the most efficient concentration for the development of bovine preantral follicles cultured in vitro.

Genetic analysis of Aspergillus nidulans unstable transformants obtained by the biolistic process.

Mitotically unstable Aspergillus nidulans argB+ transformants obtained by the biolistic process were studied in the present work. Hybridization signals from undigested DNA and pulsed-field chromosomal bands of the transformants suggested the introduced plasmid occurred as free concatenated molecules. Fifteen vigorous growth sectors released from the transformants were analysed in order to understand the mechanisms involved in their formation. All sectors showed the integration of exogenous genes into the fungal genome by homologous or heterologous recombinant events.

Transformation of Aspergillus nidulans by microprojectile bombardment on intact conidia.

This paper describes transformation of intact conidia of Aspergillus nidulans, auxotrophic for arginine, by using the biolistic process. The plasmid employed was pFB39, carrying the argB gene. The transformation frequency obtained was 81 transformants/microgram of DNA. Classical genetics and molecular analysis were conducted to analyse transformants and to determine in which chromosome integration took place.

Electrophoretic variation in esterases in 3 wild-type and respective mutant strains of Aspergillus flavus.

Wild-type strains and auxotrophic mutants of Aspergillus flavus, differing regarding aflatoxin production, were tested for esterases isozymes. Esterases variation was found in all strains used, and a possible correlation between the pattern of esterase bands and aflatoxin production is suggested.