Immunohistochemical detection of macrophage migration inhibitory factor in fetal and adult bovine epididymis: release by the apocrine secretion mode?

Originally defined as a lymphokine inhibiting the random migration of macrophages, the macrophage migration inhibitory factor (MIF) is an important mediator of the host response to infection. Beyond its function as a classical cytokine, MIF is currently portrayed as a multifunctional protein with growth-regulating properties present in organ systems beyond immune cells. In previous studies, we detected substantial amounts of MIF in the rat epididymis and epididymal spermatozoa, where it appears to play a role during post-testicular sperm maturation and the acquisition of fertilization ability. To explore its presence in other species not yet examined in this respect, we extended the range of studies to the bull. Using a polyclonal antibody raised against MIF purified from bovine eye lenses, we detected MIF in the epithelium of the adult bovine epididymis with the basal cells representing a prominently stained cell type. A distinct accumulation of MIF at the apical cell pole of the epithelial cells and in membranous vesicles localized in the lumen of the epididymal duct was obvious. In the fetal bovine epididymis, we also detected MIF in the epithelium, whereas MIF accumulation was evident at the apical cell surface and in apical protrusions. By immunoelectron microscopy of the adult bovine epididymis, we localized MIF in apical protrusions of the epithelial cells and in luminal membrane-bound vesicles that were found in close proximity to sperm cells. Although the precise origin of the MIF-containing vesicles remains to be delineated, our morphological observations support the hypothesis that they become detached from the apical surface of the epididymal epithelial cells. Additionally, an association of MIF with the outer dense fibers of luminal spermatozoa was demonstrated. Data obtained in this study suggest MIF release by an apocrine secretion mode in the bovine epididymis. Furthermore, MIF localized in the basal cells of the epithelium and in the connective tissue could be responsible for regulating the migration of macrophages in order to avoid contact of immune cells with spermatozoa that carry a wide range of potent antigens.

Cytoplasmic carbonic anhydrase II of rat coagulating gland is secreted via the apocrine export mode.

Two different pathways for protein secretion are described for epithelial cells of rat coagulating gland and dorsal prostate: the classical merocrine and the alternative apocrine release mode. Apocrine-secreted proteins are synthesized on cytoplasmic polyribosomes and are subsequently exported in protrusions on the apical cell surface (aposomes). In this article we report the identification and purification to homogeneity of a 29-kD protein from the secretion of rat coagulating gland. N-terminal amino acid sequence analyses revealed 100% identity to rat brain carbonic anhydrase II (CAH II). In addition, the 29-kD protein showed CAH enzyme activity. On Western blot analysis, a polyclonal anti-CAH II antibody raised in rabbit reacted specifically with the rat and human but not bovine CAH II isoforms. Immunohistochemical studies on rat coagulating gland showed strong labeling for CAH II protein in aposomes. Immunoelectron microscopy confined CAH II protein to the cytoplasm and aposomes, whereas no staining was visible in the compartments of the classical merocrine route, the endoplasmic reticulum and Golgi apparatus. The resident cytoplasmic protein lactate dehydrogenase, however, was not found in the secretion. Taken together, the morphological and biochemical data clearly indicate that cytoplasmic CAH II from rat coagulating gland is specifically selected and then secreted via the apocrine pathway.

Estrogen-induced morphological and immunohistochemical changes in stroma and epithelium of rat ventral prostate.

Prostatic smooth muscle cells have been regarded to play a major pathogenetic role during the development of benign prostatic hyperplasia (BPH) in elderly men. Altered hormonal signals (increased estrogen) have been made responsible for the "metabolic" transformation of prostatic smooth muscle cells, which were thought to produce increased amounts of connective tissue fibers observed in BPH. In order to find out the role of metabolically "activated" smooth muscle cells, hormone stimulation experiments were performed in male rats. The effects of androgen deprivation and estrogen stimulation were recorded by semiquantitative analysis of intermediate and myofilament proteins in stromal smooth muscle cells. In castrated or estrogen-treated or estrogen-treated and castrated animals, the reduction of the glandular lumen is the most obvious morphological alteration, accompanied by an increase in connective tissue. Regressive changes occurred most rapidly in castrated animals (already within the first week), slower in castrated estrogen-treated animals and still slower in normal estrogen-treated animals. Regression of the epithelium was accompanied by a marked decrease in immunoreactivity for prostatic binding protein (PBP) in castrated animals, while PBP immunoreactivity in estrogenized animals was retained for up to 6 weeks. Smooth muscle cells became atrophic in castrated animals. This effect was attenuated in estrogen-treated animals. There was no indication for enhanced collagen synthesis by smooth muscle cells. Actin and desmin-immunoreactivity were only slightly altered in experimental animals and showed a changed distribution pattern. Prostatic smooth muscle cells respond less markedly to hormonal alterations than do the fibroblasts.

Phenotypic modulation of the canine prostate after long-term treatment with androgens and estrogens.

Fine structural alterations of the canine prostate induced by long-term treatment of castrated adult animals with estrogens and/or androgens and also in combination with antiandrogens and/or antiestrogens for six months have been studied with particular respect to their topographic location within the gland. Three major patterns of structural responses of the epithelium have been distinguished: squamous metaplasia, atrophy, and hypertrophy, while in stroma, regression, hypertrophy, or sclerosis were observed. In addition to cellular alterations of stromal fibrocytes and smooth muscle cells, characteristic changes in the arrangement, distribution, and pattern of the different stromal elements occurred. General squamous metaplasia of the epithelium and regressive alterations of stromal cells were most obvious in animals treated with estradiol plus androstanediol. Atrophy of the epithelium and stromal sclerosis were the salient features of antiandrogen-treated castrated animals, while hypertrophy or hyperplasia of both the epithelium and stroma was a major finding in androstanediol-substituted castrated animals. Combined treatment caused rather heterogeneous structural patterns seemingly dependent on the location within the gland. The results indicate that the prostatic epithelial cells dispose of a broad variety of structural reaction patterns that, in case of combined hormonal treatment, are expressed in a manner typical for their locations within the ductal system of the gland. However, with the exception of combined treatment with estradiol, tamoxifen, and androstanediol of castrated dogs, none of the experimental protocols used induced a morphologic response of the gland comparable to that seen in human benign prostatic hyperplasia. The canine prostate therefore is of rather limited value as a model for human BPH.