Category Archives: Hormone

Quantitative Analyses Quantitative measurements indicated that the mean cross-sectional profile area of seminiferous tubules in FOR-KO mice was significantly lower compared with wild-type mice at all ages examined (Fig. 9 and Table 1). In addition, both groups showed significant changes in mean profile areas of the tubules as the age of the animals increased (Fig. 9 and Table 1). The mean profile area of tubules in wild-type mice rose by about 5700 ^m2; per tubule from 3 mo and 6 mo and then increased by an additional 14 300 ^m2 per tubule between 6 mo and 12 mo of age (Fig. 9). (more…)

In the mid and apical regions of the seminiferous epithelium of wild-type mice, the heads of elongating spermatids were deeply embedded in niches of Sertoli cell processes (Fig. 7a). The cytoplasm of the latter contained mitochondria, lysosomes, and ER cisternae, all embedded in a finely flocculent ground substance. In addition, bundles of filaments overlaid by ER cisternae, and forming the so-called ectoplasmic specializations, were closely applied to the spermatid heads (Fig. 7a). In the semilunar affected areas of the seminiferous epithelium of FORKO mice, a gross enlargement and dilation of the apical Sertoli cell cytoplasm was evident (Fig. 7b). (more…)

Sertoli and germ cells were closely associated with each other, with very small intercellular spaces positioned between the two. However, in FORKO mice, the seminiferous epithelium showed large dilated spaces surrounding nearby germ cells (Figs. 5b and 6). The basal Sertoli cell cytoplasm contacted the basement membrane and displayed various organelles such as lysosomes, mitochondria, endoplasmic reticulum (ER), and the Golgi apparatus, embedded in a finely flocculent ground substance (Figs. 5b and 6). However, large dilated spaces not connected to basal areas of Sertoli cell cytoplasm, due to the plane of section, were often observed in mid and apical regions of the epithelium (Fig. 5b). (more…)

An intense anti-androgen binding protein (ABP) reaction was noted over the cytoplasm of Sertoli cells at all stages of the cycle of the seminiferous epithelium of wild-type mice; the reaction extended from the base of the epithelium to the lumen (Fig. 4, a, c, and e). No staining was observed over germ cells. In FORKO mice, the staining over Sertoli cells at all stages of the cycle appeared weak and in some areas completely missing (Fig. 4, b, d, and f). A nonspecific staining of Leydig cells was evident in the interstitial spaces inherent to the anti-ABP protein construct (see Materials and Methods), and the fact that Leydig cells express GSTs. Control sections showed no staining over the epithelium (Fig. 4a, inset). (more…)

In the midregion of the epithelium, the dilated spaces often separated chains of round spermatids from each other. In this way, they formed cords or ribbons, which gave the epithelium an anastomotic appearance due to their extensive nature (Fig. 2, a and b). The large dilated spaces of varying sizes also surrounded elongating spermatids, which they often completely enveloped (Figs. 1, b and d, and 2, a and b). The interior of the dilated spaces often contained membranous profiles of varying sizes and spherical particulate material (Fig. 2b). (more…)

Light Microscopic Appearance of the Testis of FORKO Mice At 3 and 6 mo of age, the seminiferous tubules in the testis of FORKO mice showed smaller profile areas compared with wild-type mice (Fig. 1, a and b). In addition, while the testis of wild-type mice displayed a homogenous, compact seminiferous epithelium, where germ and Sertoli cells were closely associated with each other (Fig. 1, a and c), the FORKO mice presented a varying and vacuolated appearance (Figs. 1, b and d, and 2, a and b). Although some tubules at both ages seemed normal in cross-sectional profile (approximately 40%), nearly one half of the circumference of seminiferous epithelium of other tubules in the FORKO mice appeared disrupted, showing large dilated spaces between the epithelial cells (Figs. 1, b and d, and 2, a and b). (more…)

Western Blot Analysis A total of 6 mice at 6 mo of age (wild type, n = 3; FORKO, n = 3) were used for Western blot analysis. One testis of each mouse was extracted and cut into four equal pieces. Each piece was then frozen in liquid nitrogen for subsequent protein extraction. The frozen pieces of testis were homogenized, suspended in lysis buffer containing detergent and a protease inhibitor cocktail, and the solution was centrifuged at 11 000 X g for 15 min at 4°C. The supernatant was removed and quantified for protein using the Bradford Assay (Bio-Rad Laboratories Inc., Richmond, CA). (more…)

The sections were then incubated at 37°C in a humidified chamber for 90 min with 100 |xl of primary antibody diluted in Tris-buffered saline (TBS). Following washes in 0.1% Tween20 in TBS, the slides were incubated with secondary antibody (1:250; 100 |xl) labeled with horseradish peroxidase for 30 min at 37°C in a humidified chamber Reactions were revealed with dia-minobenzidine tetrahydrochloride (DAB). Sections were counterstained with methylene blue, dehydrated in ethanol and Histoclear, and mounted with cover slips using Permount. TBS substitution for primary antibody was used as a negative control. No reactions were observed in these sections. (more…)

LM Immunocytochemistry The following affinity-purified polyclonal antibodies were used at 1: 100 dilution (v/v) for routine peroxidase immunostaining: 1) antiprosa-posin antibody (provided by Dr. C.R. Morales, McGill University, Montreal, Canada; purified and characterized as described previously ) and 2) rabbit anti-ABP antibody prepared against a glutathione sulfo transferase (GST)-ABP fusion protein (provided by Dr. G.L. Hammond, University of Western Ontario, London, Canada). (more…)

Scaled digital images of 5-^m-thick paraffin sections of seminiferous tubules from these animals were captured on a Zeiss Axio-scop 2 equipped with an AxiocamMR camera, and the peripheral outline of selected tubules was traced and the profile areas determined using the appropriate measurement tool available in Version 3.1 of the Axiovision Imaging Software (Carl Zeiss Canada Ltd., Montreal, QC). (more…)