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The present study examined the tissue-expression of MRP8 in human placenta using a biotinylated DNA-probe for in situ hybridization. During the first and second trimesters high level and synchronous expression of MRP8 was detected in cytotrophoblasts (Langhans' cells), placental-tissue macrophages (Hofbauer cells), fibroblast-like cells, endothelial cells and monocytic lineages in the foetal capillaries. The highest expression was seen in large and oval-shaped cytotrophoblasts and stromal-cell populations at around 8-11 weeks. At term placentas had low level MRP8 expression chiefly in the myelomonocytic lineages in foetal blood vessels. The peripheral monocytes in the maternal space also expressed MRP8 at high levels during the first and second trimesters, which subsequently decreased at term. We suggest three hypotheses based on these results; (1) The initial expression of MRP8 may occur in two cell lineages of extra-embryonic and intra-embryonic origin in the first two trimesters; (2) the cytotrophoblasts, placental-tissue macrophages and fibroblasts may play important roles in the production of placental hormones and the immuno-regulation of foetal acceptance; and (3) MRP8-expression may be synchronously inhibited once the trophoblasts and stromal cell-constituents have differentiated in the chorionic villi.

Immunohistochemistry revealed initial expression of the stage-specific glycoprotein, GP68, in various mesenchymal tissue substructures of mouse embryos. During the 11-15th days of gestation, GP68 was localized in the primitive meninges, chondroblasts and perichondrium of pre-cartilaginous vertebral bodies and ribs, connective tissue cells of the dermis, the epicardium and endocardium of the heart, the epimysium and perimysium of skeleton musclature, and the basement membranes of splanchnic organs. Double staining for laminin expression indicated coincidental expression in identical tissue substructures. However, laminin was expressed in days 10-18 embryos and the neonate. Therefore, GP68 is coincidentally expressed with laminin in mesenchymal tissues between the 11th and 15th day of gestation, and may play a role as a laminin-associated protein. In the light of these results, a hypothesis concerning the relationship between these two proteins and the mechanisms of non-integrin laminin-associated proteins during normal embryogenesis is discussed further.

The protein A-gold immunocytochemical technique was used to localize the secretory pathway of oestradiol-induced vitellogenin in hepatic parenchymal cells of the cockerel. Liver was removed from experimental birds on the 1st, 4th and 8th day following oestradiol-treatment, and embedded in Lowicryl K4M resin cured at -20 degrees C. In selected electron micrographs the fractional surface area of each of the intracellular compartments was measured by the computer-assisted digitization technique. Labelling was detected over the cisternae of the rough endoplasmic reticulum (RER), the Golgi apparatus, the immature secretory vacuoles (ISV) including condensing vacuoles and the mature secretory vacuoles (MSV). Counts of the gold particles demonstrated an increasing concentration which progressed in the order RER less than Golgi less than ISV less than MSV and identified the secretory pathway of the protein. The highest density of labelling was obtained on the 4th day, when vitellogenin reaches its peak activity. Autophagic activity (or crinophagy) was also found in lysosomes and its labelling intensity increased daily. A hypothesis concerning the secretory pathway of non-stored proteins by the liver is discussed further.

Vitellogenin has been localized at the electron microscopical level in the liver of the cockerel using a colloidal gold technique. White leghorn cockerels were treated with 17 beta-oestradiol to induce vitellogenesis. Pieces of liver were removed from control and experimental birds on the 4th and 8th days following hormone treatment, and embedded in Lowicryl K4M. Vitellogenin was isolated from the plasma of oestradiol-treated cockerels, and the antibody to it elicited in rabbits and made vitellogenin-specific by affinity chromatography on lipovitellin-Sepharose columns. At the light microscopical level, the intensity of immunohistochemical staining was considerably above background levels in oestradiol-treated cockerels. At the electron microscopical level, gold particles indicating antigenic sites of vitellogenin were largely confined to the rough endoplasmic reticulum, Golgi apparatus, immature and lysosomes and phagosomes within hepatocytes and sinusoidal cells respectively. These observations strongly suggest that the intracellular pathway of vitellogenin secretion in chicken hepatocytes under the experimental conditions studied involves external stimuli and secretory vacuoles. The labelling of lysosomes may reflect catabolic turnover (crinophagy).

In laying hen oviduct tissues fixed with buffered-formaldehydes, the definitive localization of endogenous biotin-binding protein and immunogenic avidin was demonstrated in the cytoplasm of tubular gland cells and PAS-positive seromucous cells of the epithelium of the lower third magnum and the isthmus. Smaller amounts of the protein were also associated with tubular gland cells and deep-lining cells as crypt-constituents, excluding epithelial goblet-like cells, of the middle magnum. In biotin-affinity histochemistry, suppression by pre-incubation with biotin was evaluated to be the most satisfactory procedure for positive control stainings. The staining intensity and pattern achieved with biotinylated-horseradish peroxidase was more prominent than that achieved with biotinylated-alkaline phosphatase techniques. Positively stained profiles of tubular gland cells varied considerably with different cellular constituents, having a mosaic-like appearance. In addition, every biotin-binding site was not always stained by immunoperoxidase techniques. This paper discusses the reasons for these heterogeneous localizations resulting from these two staining procedures.

The localization of avidin in the oviduct of the laying hen was investigated using ultrastructural immunoperoxidase techniques. Endogenous avidin was localized in secretory granules of both tubular gland cells and non-ciliated single epithelial cells in the magnum mucosa. These immunospecific granules were electron-dense and heterogeneous with a patchy core and dense peripheral region, especially in acinar cells. The size varied from small to large in the gland cells (500-2200 nm in diameter) and remained small in the epithelial cells (180-720 nm). Columnar epithelial cells containing avidin granules strongly resembled the protodifferentiated tubular gland cells appearing in the magnum mucosa of chicks artificially pretreated with ovarian hormones. On the other hand, no avidin was observed in either epithelial goblet cells or ciliated cells in adult hens, although both cell types were shown to produce avidin in young chicks when synchronized by the administration of progesterone. The present results parallel those obtained with biotinylated enzyme affinity methods in our previous cytochemical study. Therefore, avidin is one of the proteins produced and stored in the secretory granules of the tubular gland cells and protodifferentiated acinar cells present in the epithelial layer of the laying hen oviduct. It is not present in goblet cells. Although the initiation of a synthesis may be triggered by progesterone, it is still not clear whether different hormone dependent proteins are localized in the same granules in both the adult hen and the immature chick.

The localization of lysozyme (LZM) in the human nasal mucoua was ob-served with the use of an unlabelled antibody peroxidase-antiperoxidase (PAP) method. LZM was distributed in serous cells of the nasal submucosal glands. Weak staining occurred in some mucous parts of them which seemed to be so-called seromucous cells classified by Shackleford and Wilborn, which exhibit intermediate characteristics between mucous and serous cells. In the present study, however, it was not clarified whether LZM produced by the serous cells is from blood plasma. No specific staining for LZM was detectable in the epithelial cells. Some tissue blood element cells such as neutrophils and macro-phages also contained antigenic LZM intracellularly in parallel with those of human blood huffy coats.