During follicular development, granulosa cells undergo functional and structural changes affecting their steroidogenic activity. Oestrogen synthesis mainly occurs in the endoplasmic reticulum and relies on aromatase activity to convert androgens that arise from theca cells. In the present study, indicators of mitochondria-related steroidogenic capacity, as steroidogenic acute regulatory (StAR) protein expression and mitochondrial membrane potential (MMP), have been evaluated in bovine granulosa cells (GCs) and related to follicle growth and atresia. Atresia was estimated by morphological examination of follicle walls and cumulus–oocyte complexes (COC) and assessed by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay for apoptosis detection. Bovine ovarian follicles were macroscopically classified according to their atresia grade and grouped into small, medium or large follicles. After follicle opening, the COCs were morphologically classified for follicle atresia and the GCs were collected. Granulosa cells were fixed for immunofluorescence (IF) and TUNEL assay, frozen for western blotting (WB) or freshly maintained for MMP analyses. StAR protein expression was assessed using both IF and WB analyses. The follicle atresia grade could be efficiently discriminated based on either follicle wall or COC morphological evaluations. Granulosa cells collected from small non-atretic follicles showed a higher (P <0.01) MMP and WB-based StAR protein expression than small atretic follicles. For IF analysis, StAR protein expression in large atretic follicles was higher (P <0.05) than that in large non-atretic follicles. These results suggest a role played by mitochondria in GC steroidogenic activity, which declines in healthy follicles along with their growth. In large follicles, steroidogenic activity increases with atresia and is possibly associated with progesterone production.

Exposure to lead may cause changes in the male reproductive system. We evaluated the effect of lead chloride (PbCl2) in vitro on semen quality from 31 individuals. Samples were incubated at room temperature for two exposure times (4 h and 8 h) and with two concentrations of PbCl2 (15 μg/ml or 30 μg/ml). Results showed that PbCl2 significantly inhibited rapid progressive motility and caused an increase in the percentage of tail anomalies in both times and concentrations assessed, as well as a decrease in vitality in the group exposed to 30 μg/ml PbCl2. A significant increase in immotile sperm was also observed between the group control and the groups submitted to lead. Total motility and DNA fragmentation also showed a significant decrease and increase, respectively, after 4 h of incubation in the group exposed to 30 μg/ml and in both groups after 8 h of incubation. In conclusion, PbCl2 affected sperm parameters and DNA integrity, which are essential for male fertility.

Severe malnutrition is widely distributed throughout the world, showing a high prevalence in developing countries. Experimental animal models have been useful to study the effects of malnutrition at different levels and ages. Apoptosis is a well recognised process of cell death occurring under several physiological and pathological conditions. It represents the principal mechanism involved in cell selection in the thymus. Thymocyte apoptosis induction by dexamethasone is one of the best characterised experimental models of programmed cell death. The aim of the present study was to determine whether severe malnutrition increased spontaneous and/or dexamethasone-induced apoptosis in vivo in thymocytes of experimentally malnourished rats during lactation. Thymocytes were obtained from malnourished rats at weaning (21d of age). Apoptosis frequency was estimated by the terminal transferase-mediated dUTP nick end labelling assay. Spontaneous apoptosis was 1·9 (SD 1·0) % IN WELL NOURISHED RATS IN CONTRAST TO 13·3 (sd 3·8) % in malnourished animals; this is seven times greater (P<0·001). Interestingly, the frequency of dexamethasone-induced apoptosis was similar in both groups of animals (47·9 (sd 10·1) % in well nourished rats and 53·8 (sd 8·0) % in malnourished rats). The results obtained in the present study indicate that malnutrition is associated with a significant increase of spontaneously apoptotic cells. In addition, the data showed that the fraction of thymocytes susceptible to dexamethasone-induced apoptosis was similar in well nourished and malnourished animals. The greater levels of spontaneously apoptotic cells associated with malnutrition could be related to alterations of the microenvironment of the thymus and/or to an obstruction of early thymocyte maturation.