Effects of growth hormone on female reproductive organs

During the last decade many experiments have been performed to study the effects of growth hormone (GH, somatotropin) on reproductive functions. Most of the studies found only slight or no effects of GH treatment, both on the oestrous cycle and on gonadotropin, progesterone. or oestrogen serum levels. In GH-treated animals, elevated levels of insulin-like growth factor I and GH in the serum could be correlated with an increased number of small (< 5 mm in diameter) ovarian follicles, possibly as a consequence of a reduction of apoptosis and follicular atresia. There is still controversy over the effects of GH on in vivo and in vitro embryo production and on the gestation period. Recent studies produced some evidence that GH-receptor is expressed in ovarian tissue, implying a direct role for GH in the ovary.     

Expression of cytokeratin 18 during pre- and post-natal porcine lung development

The expression pattern of the intermediate filament protein cytokeratin 18 (CK 18) is described during pre- and post-natal development of the porcine lung using a monoclonal antibody against human CK 18. Lungs from 16 foetuses in pseudoglandular, canalicular, saccular and alveolar stages of lung development and lungs from 12 pigs ranging in age from birth to 49 days after birth were studied by immunohistochemistry.In the early pseudoglandular stage of development (day 70 of gestation) all the columnar epithelial cells lining the tubular endbuds strongly expressed CK 18 predominantly in the apical cell compartment. A modest staining was found in the more cuboidal cells of the canalicular stage (day 80 of gestation) where the labelling occurred as a distinct positive rim at the apical cell membrane in most of the cells lining the canaliculi. In 96- and 100-day-old foetuses, parts of the gas exchanging area were formed as terminal sacs by extreme attenuation of the epithelium. In this stage, CK 18 was clearly detectable in the flat type I as well as in the cuboidal type II alveolar epithelial cells. A marked change of the CK 18 expression pattern occurred during formation of the alveoli by septal outgrowth and maturation of the epithelium in 105- and 111-day-old foetuses. Differentiated type I cells no longer expressed CK 18, whereas type II cells were still labelled. Moreover, a specific change in the subcellular distribution pattern from the luminal periphery in immature porcine type II cells to a cytoplasmic localization in differentiated type II cells could be observed. Our investigation additionally demonstrated that the epithelium of bronchi, bronchioli and terminal bronchioli expressed CK 18 in all pre- and post-natal developmental stages. From the 96 days of gestation onwards the epithelial cells of developing bronchial glands were also labelled. Our results clearly show that during porcine lung development profound changes in the cellular expression pattern of CK 18 occur and that CK 18 can be regarded as a selective marker for differentiated porcine alveolar type II cells from the 105th day of gestation onwards. We also assume that the intermediate filament CK 18 could be of significance in the maturation process of the type II alveolar cells.     

Immunohistochemical Localization of the Spermadhesin AWN-1 in the Equine Male Genital Tract

Spermadhesins are proteins with various functions in sperm capacitation and zona pellucida binding. In this study the cellular localization of the spermadhesin AWN-1 has been examined in the equine male genital tract. Results obtained by immu-nohistochemical methods reveal that in the horse AWN-1 is synthesized in spermatogonia, in the rete testis, the ductus epi-didymidis and the seminal vesicles. These findings indicate that the cellular origin of spermadhesins is species-specific.     

Subpopulations of Stromal Cells from Long-term Human Bone Marrow Cultures: Ontogeny of Progenitor Cells and Expression of Growth Hormone Receptors *

Long-term culture of bone marrow derived stromal colony forming cells (S-CFC) in matrix and nutrient defined agar medium resulted in stromal cell colonies that pass sequentially through three distinct morphological stages: firstly, aggregated loose syncytium of round to ovoid cells (stage I), a second developmental stage of large branching colonies in which the cells become enlarged, elongated with cytoplasmic projections forming a loosely anastomized network with adjacent cells (stage II), and finally cells become dissociated, loosing their long, thin cytoplasmic filaments and breaking their contacts with one another, but remain large and retain a bi-polar nature (stage III). Cells were also grown in liquid medium in a culture microenvironment closely resembling conditions of haemopoiesis in vitro. Using a panel of well defined monoclonal antibodies reactive against the rat, rabbit and human growth hormone receptors, this study found immunochemical evidence of the presence and localization of binding sites of growth hormone (GH) in the cell membrane and extra-nuclear Golgi area of long-term bone marrow derived human stromal cells in liquid and semi-solid nutrient agar mediums. GH-receptor immunoreactivity was present in small proliferating progenitor cells, myofibroblast-like cells, large reticular fibroblast cells, adipocytes and endothelial cells. Only MAb known to be reactive against human tissue resulted in strong immunoreactivity. The expression of GH-receptors not only on small proliferating, but also on the well differentiated cells, indicates a role for growth hormone on non-progenitor cells. GH-receptor immunoreactivity on differentiating and/or differentiated cells suggests that GH is also necessary for, or has a trophic function in differentiation. We propose that direct GH action is necessary not only for differentiation of progenitor cells as implied by the dual effector hypothesis, but also their subsequent clonal expansion, differentiation and maintenance.     

On the Species Specificity of Sperm Binding and Sperm Penetration of the Zona Pellucida

Sperm binding and sperm penetration of the zona pellucida (zp) are regarded as species‐specific. In this investigation, the interactions between bovine oocytes and porcine, respectively, equine spermatozoa have been studied under in vitro conditions and compared with the normal in vitro fertilization of bovine oocytes by bovine sperm. Surprisingly, many of the heterologous spermatozoa adhered firmly to the bovine oocytes and could not be removed by intense washing. On average, more than 100 boar or equine spermatozoa were bound to the zp of bovine oocytes. Electron microscopic studies clearly demonstrated that porcine sperm attached to the zona and underwent the acrosome reaction. Equine spermatozoa displayed a similar binding affinity, but unlike the porcine spermatozoa even penetrated the zp and were taken up into the oocyte after a longer period of co‐incubation. Considering these new results the dogma of a strict species specificity of sperm zona interactions under in vitro conditions has to be reconsidered.     

Sexual dimorphism of the kidney in the NMRI mouse as shown by Dolichos biflorus agglutinin labelling

The histological affinity pattern of Dolichos biflorus agglutinin (DBA) in kidneys from mice (NMRI, Balb/c, CBA) and rats (Wistar) fixed by perfusion with formalin, Bouin, or HgCl2 was investigated with a horseradish peroxidase conjugate. The animals were examined from fetal stage to adulthood. Adult female NMRI mice exhibited constant DBA labelling, with DBA binding to cells of the proximal and collecting tubules. Moreover the vascular endothelium of the renal papilla was found to be DBA-positive in 50% of adult female animals. In contrast, there was only very little DBA binding in the kidneys of male adult NMRI mice. There was no sexual dimorphism in lectin labelling in kidneys from other strains of mice or from rats.     

Einfluß wiederholter Hodenbiopsien auf die postnatale Entwicklung des Rinderhodens

Inhalt In der vorliegenden Arbeit wird über die Veränderungen am Rinderhoden während der postnatalen Entwicklung nach wiederholter Biopsie berichtet. Bei Jungtieren wurden im Abstand von 3 Monaten mehrmals Hohlnadelpunktionsbiopsien vorgenommen und gleichzeitig die Gewichtszunahme, die Hodengröβe, die Testosteron- und Epitestosteronkonzentration im Blut sowie die üblichen Samenparameter ermittelt. Die Hodenentwicklung und Körpergewichtszunahme der Versuchstiere war im Vergleich zu den Kontrolltieren nicht signifikant unterschiedlich. Spermatologisch lagen jedoch gravierende Unterschiede vor. Die histoiogischen und ultrastrukturellen Untersuchungen zu Versuchsende zeigten, daβ eine groβe Zahl von Tubuli schwere morphologische Veränderungen aufwies. Das Keimepithel war auf Sertolizellen und vereinzelte Spermatogonien reduziert. Bei jenen Tubuli, bei denen noch eine Spermatogenese ablief, entsprachen die Spermatogonien weitgehend dem normalen Bild, während bei den Spermatozyten erster Ordnung und den Spermatiden feinstrukturelle Abweichungen häufig festgestellt werden konnten. Das Hodeninterstitium wurde durch die wiederholten Biopsien offensichtlich nur wenig beeinfluβt. Die Ausbildung und die Zahl der Leydigzellen entsprachen jenen der Kontrolltiere. Die Testosteron- und Epitestosteronkonzentration im Blut der untersuchten Stiere glich den in der Literatur beschriebenen Werten. Contents: The Influence of repeated Biopsies of Cattle Testicles on their postnatal Development This is a report on the pathological changes of cattle testicles during postnatal development after repeated biopsies. In intervals of 3 months repeated cannula puncture biopsies were made with 3 young bulls and at the same time weight increase, size of testicles, the concentration of testosterone and epitestosterone in the blood as well as the usual parameters of semen were examzned. The development of the testicles and increase of body weight of testing animals were not significantly different compared with the control animals. There were, however, dastic spermatological differences. The histological and ultrastructural examinations at the end of the tests showed that with a great number of tubuli significant morphological changes could be identified. The germinal epithelium was reduced to Sertoli cells and isolated spermatogonia. With those tubuli which underwent a spermatogenesis, the spermatogonia largely corresponded to the normal picture, while with the spermatocytes of the first oder and the spermatids there repeatedly showed subtile structural differences. Obviously the testicle interstice was influenced only very little. Formation and number of the Leydig cells corresponded to those of the control animals. The concentration rates of testosterone and epitestosterone in the blood of the tested bulls were analogous to those in bibliography.     

Original Papers

In the present study we report on the histotopographical distribution of lectin binding sites in the trophoblasts of day 18 to day 40 bovine embryos, using the FITC-labeled lectins BPA, Con A, DBA, GS I, GS II, MPA, PNA, SBA, UEA I and WGA. Lectin binding sites localized in giant binucleate cells differ from those localized in uninucleate cells, indicating changes in the biochemical structure of cell surfaces taking place during differentiation. In the trophoblast of the day 40 embryo, a distinct staining of uninucleate cells was seen after incubation with GS I, Con A and MPA, demonstrating N-acetylgalactosamine (GS I), Mannose (Con A) and Galactose (MPA) moieties, whereas giant binucleate cells showed intense reactions after incubation with DBA and WGA, indicating presence of N-acetylgalactosamine (DBA) and N-acetylglucosamine (WGA). GS II (specific for N-acetylglucosamine), SBA (specific for N-acetylgalactosamine) and UEA I (specific for L-Fucose) showed no affinity toward any of the examined tissues. We assume, that carbohydrate moieties in trophoblast cells play an important role in fetomaternal cell-cell adhesion and cell migration during implantation and placentation period.     

Prenatal differentiation of bovine oviductal epithelium: an electron microscopic study

In this study, we investigated the ultrastructural changes during the prenatal differentiation of oviductal epithelium in 16 bovine embryos and fetuses from CRL of 18.0 cm to a CRL of 94.0 cm. Ciliated and secretory cells of bovine uterine tube, a derivative of the Müllerian duct, differentiate to distinct development stages in the prenatal period. The typical cellular pattern, which is generally characteristic for the adult bovine oviduct, is also obtained during fetal life. In the early stages (CRL 18.0/20.4 cm), the bovine oviductal epithelium appears mostly undifferentiated. The epithelial cells show only a few mitochondria, some cisternae of rough endoplasmic reticulum (rER) and a small Golgi-complex. Most of the cytoplasm is filled with a large amount of glycogen, which decreases during later development. Interspersed between the undifferentiated epithelial cells, a few cells undergoing ciliogenesis can be observed. Ciliogenesis increased significantly during the later prenatal developmental stages. At a CRL of 55.0 cm, ciliated cells appear fully differentiated with mature cells covering their luminal surface. Formation of cilia usually use the acentriolar pathway. Fibrous granules occurred initially in association with the Golgi-apparatus and r(ER) in the supranuclear cytoplasm. Fibrous granules later fuse with deuterosomes and give rise to procentrioles, which are translocated to the luminal plasma membrane. There they become arranged in a line just beneath the apical cell membrane and further differentiate to basal bodies from which the formation of cilia and striated rootlets take place. Clear signs of differentiation of secretory cells were first seen in our material in fetuses with a CRL of 51.0 cm and 64.0 cm. These cells contain a well developed rER and Golgi-apparatus with dilated cisterns. In the supranuclear cytoplasm, the number of secretory granules continuously increases during later development and the cells adapt to the morphology of mature secretory cells at the CRL 94.0 cm.