S. Van Cruchten

Apoptosis in the canine endometrium during the estrous cycle.

Apoptotic cell death in the endometria of 58 female dogs in different stages of the estrous cycle was assessed (in formalin-fixed, paraffin-embedded sections) with both the terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate nick end labeling (TUNEL) assay and immunohistochemical detection of caspase-3 activity. For both techniques, the apoptotic index was determined in the surface epithelium, stroma, crypts, and basal glands by counting the percentage of stained cells in a total of 500 cells in each category. In the surface epithelium and stroma, TUNEL- and caspase-3-positive cells were rare (apoptotic index<1) throughout the estrous cycle. However, caspase-3 detection showed a significant increase in the apoptotic index in the stroma during anestrus as well as an increase in the index in both the stroma and surface epithelium in late metestrus. The apoptotic index increased during late metestrus and anestrus in the crypts and basal glands; in the crypts, this increase was significant only when caspase-3 detection was used, whereas in basal glands, significant differences were found for both techniques. In conclusion, apoptosis was present in canine endometrial cells during the estrous cycle, but caspase-3 detection showed more significant differences than the TUNEL assay. Furthermore, a high apoptotic index (suggestive of endometrial desquamation) was not detected in the surface epithelium and there was no significant correlation between the apoptotic index in any cell group and serum progesterone concentrations.

Endogenous occurrence of some anabolic steroids in swine matrices.

Following findings of 17β-19-nortestosterone (150–200 µg kg−1) in pigs of unspecified gender imported into the European Union, a study to determine steroid and hormone levels in swine from six age/gender categories (uncastrated ‘old’ boars, cryptorchids, one intersex, barrows, gilts and sows) was initiated. Indeed, for some hormones there has been a discussion about their being endo- or exogenous. Tissue and urine samples from swine from each of the six categories were obtained in Belgium, France, the Netherlands and the USA. Samples were analysed in three laboratories. Quantitation was obtained for norandrostenedione, 19-nortestosterone and boldenone. The results give a well-documented overview of the status of the presence of these hormones in swine. The data illustrate that uncastrated ‘old’ boars produce the highest percentage of ‘positive’ matrices, followed by the cryptorchids. Concentrations in the matrices of the barrows and the gilts are lower. Also, sow matrices contain low amounts of nor-steroids. Furthermore, urine samples from an intersex pig contains a higher concentration of nortestosterone than sows and can therefore be suspected for illegal use of these hormones. Veterinarians taking samples in pig farms for the analysis of hormones need to be aware of the presence and concentrations of these substances in the different categories.

Cyclic Changes of the Canine Endometrial Surface: An Electron-Microscopic Study

The endometrial surface morphology of 38 dogs during different stages of the estrous cycle was investigated with scanning electron microscopy. The cell surface altered from convex in proestrus and estrus to very variable in early metestrus, flattened in late metestrus and became completely plane in anestrus. Microvilli were numerous and long in proestrus and in estrus, became short and variable in number in early metestrus, decreased further in length in late metestrus and became very short and rare in anestrus. The variable appearance in early metestrus was not influenced by changing the osmolarity of the fixative and might be a physiological process. The number of glandular openings showed little variability throughout the estrous cycle. Ciliated cells were rare but present in all cycle stages except in late metestrus. However, in the latter cycle stage and in anestrus rare single strands were noted. Transmission electron microscopy was used to determine the inner structure of these strands. Microtubuli were detected in transversal and longitudinal sections but without the 9 + 2 arrangement which is characteristic for cilia. The nature and function of these structures remain unclear.

Biological and Chemical Approaches for the Detection and Identification of Illegal Estrogens in Water-based Solutions

The continuous introduction of new products used as growth promoters in animal husbandry, for sports doping and as products for body-building requires residue laboratories to initiate research on developing a strategy for the identification of ’unknown’ components. In this study, a strategy is presented for elucidating the identity, the structure and the possible effects of illegal estrogenic compounds in an unidentified water-based solution. To obtain complete information on the composition and activity of the unidentified product, a multidisciplinary approach was needed. A case-study is described with a ’solution X’ found during a raid. First, in vivo techniques (animal trials with mice, anatomical and histological research) were combined with in vitro techniques (the yeast estrogenic screen (YES)). In a later stage of the investigation, HPLC-fractionation, liquid chromatography–multiple mass spectrometry (LC-MS n and gas chromatography-multiple mass spectrometry (GC-MS n were used. Finally, the identity of ’solution X’ was confirmed in a very low concentration range (10 ng/L estrone and 400 ng/l ethinyloestradiol).