Culture of ciliated and nonciliated cells from rat ductuli efferentes.

The isolation and culture of ciliated and nonciliated cells from rat ductuli efferentes is described. Fragments of epithelium obtained after two collagenase digestions attached to plastic and to extracellular matrix and could be maintained in culture for at least 2 weeks. Ciliary beating in cells grown on epididymal extracellular matrix-coated plastic could be observed for up to 7 days in culture. Although cells maintained on this substrate retained organelles characteristic of cells in vivo, they assumed a flattened, squamous appearance. In contrast, cells growing on the surface of permeable supports impregnated with extracellular matrix were polarized and exhibited a cuboidal/columnar appearance. Androgen binding protein conjugated to colloidal gold was taken up by these cells via coated pits and was found sequentially in uncoated endosomes, multivesicular bodies and lysosomes.

Internalization of colloidal gold-labelled insulin complexes into cultivated human smooth muscle cells.

The binding and internalization of colloidal gold-labelled insulin complexes into cultivated human arterial smooth muscle cells was studied. We could demonstrate that the colloidal gold-labelled insulin complex was bound to specific receptors on the cell surface of smooth muscle cells and internalized after 30 min at 20 degrees C. The gold-labelled insulin complexes entered the cell via vesicles and escaped from them in the cytoplasm. The biological activity of the complexes was determined by measuring their effect on DNA synthesis in smooth muscle cells and incorporation of (14C)-glucose into isolated fat cells. The binding characteristics of the complexes were examined by competitive inhibition of 125I-insulin binding to isolated fat cells in the presence of increasing concentrations of gold-labelled insulin complexes or unlabelled insulin. The gold-labelled insulin complex showed a growth promoting and metabolic effect and was able to compete with 125I-insulin for its binding sites.

Ca2+-independent, protein-mediated fusion of chromaffin granule ghosts with liposomes.

We have investigated the interaction between isolated membrane vesicles from chromaffin granules and large unilamellar phospholipid vesicles (liposomes). Mixing of membrane lipids has been monitored continuously, utilizing the fluorescence resonance energy transfer assay described by Struck et al. ((1982) Biochemistry 20, 4093-4099). To demonstrate coalescence of the internal vesicle volumes the transfer of colloidal gold from the liposomes to the interior of the granule membrane vesicles has been examined. Efficient fusion of the liposomes with the granule membranes was observed. Significant fusion occurred in the absence of Ca2+, although the extent of interaction was enhanced in its presence. The sensitivity of the interaction to pretreatment of the granule membranes with trypsin showed the fusion reaction to be a protein-mediated process.

Reversible pinocytosis in polymorphonuclear leukocytes.

Since pinocytosis has only been recently recognized in polymorphonuclear leukocytes (PMNs), little is known about the fate of pinosomes. Here we report that pinosomes can fuse with the cytoplasmic granules of PMNs. We also find that at least for a short period of time after formation, pinosomes can fuse with the plasma membrane and release their contents to the outside. We present a morphological description and biochemical data on the kinetic parameters of a steady state pool of reversible pinosomes in PMNs. In addition, we have developed conditions under which pinosomes continue to form and fuse with the plasma membrane but fail to fuse with the cytoplasmic granules, i.e., only "reversible" pinocytosis occurs. This inhibition of fusion with the granules is not due to an inability of the pinosomes to move from the surface since under these conditions pinosomes labeled with an electron-dense marker can be seen in the cell interior.

Transferrin endocytosis in reticulocytes: an electron microscope study using colloidal gold.

The endocytosis of transferrin by rabbit reticulocytes was investigated by electron microscopy using transferrin labelled with colloidal gold. This complex was shown to bind and donate its iron to the cells in a manner comparable to native iron-transferrin. After incubation at 37 degrees C approximately 70% of the transferrin-gold particles were located within endocytotic vesicles. Treatment of the cells with pronase, EDTA, metabolic inhibitors and heating to 46 degrees C inhibited the endocytosis. The uptake of colloidal gold complexes of albumin and concanavalin A were compared with that of transferrin. Little endocytosis of these proteins was observed. It is concluded that colloidal gold is a suitable label for the investigation of transferrin endocytosis and that the endocytosis is specific for transferrin and sensitive to the action of reagents which damage membrane receptors, alter membrane function or impair cell metabolism.

Histochemical evidence for the differential surface labeling, uptake, and intracellular transport of a colloidal gold-labeled insulin complex by normal human blood cells.

A colloidal gold-labeled insulin-bovine serum albumin (GIA) reagent has been developed for the ultrastructural visualization of insulin binding sites on the cell surface and for tracing the pathway of intracellular insulin translocation. When applied to normal human blood cells, it was demonstrated by both visual inspection and quantitative analysis that the extent of surface labeling, as well as the rate and degree of internalization of the insulin complex, was directly related to cell type. Further, the pathway of insulin (GIA) transport via round vesicles and by tubulo-vesicles and saccules and its subsequent fate in the hemic cells was also related to cell variety. Monocytes followed by neutrophils bound the greatest amount of labeled insulin. The majority of lymphocytes bound and internalized little GIA, however, between 5-10% of the lymphocytes were found to bind considerable quantities of GIA. Erythrocytes rarely bound the labeled insulin complex, while platelets were noted to sequester large quantities of the GIA within their extracellular canalicular system. GIA uptake by the various types of leukocytic cells appeared to occur primarily by micropinocytosis and by the direct opening of cytoplasmic tubulo-vesicles and saccules onto the cell surface in regions directly underlying surface-bound GIA. Control procedures, viz., competitive inhibition of GIA labeling using an excess of unlabeled insulin in the incubation medium, preincubation of the GIA reagent with an antibody directed toward porcine insulin, and the incorporation of 125I-insulin into the GIA reagent, indicated the specificity and selectivity of the GIA histochemical procedure for the localization of insulin binding sites.

[Distribution, absorption and excretion of 198Au-colloid and Na251CrO4 in the chicken (author's transl)].

In order to clarify the suitability of 198Au-colloid and Na251CrO4 as a marker of transit of the gastrointestinal content of the chicken, the distribution, absorption and excretion of 198Au-colloid and Na251CrO4 in the chicken were examined. Orally administered 198Au-colloid was not absorbed by the gastrointestinal tract and excreted into feces. Intravenously injected 198Au-colloid was not excreted into the gastrointestinal tract through the gastrointestinal wall. On the other hand, orally administered Na251CrO4 was relatively well absorbed by the gastrointestinal tract and distributed over a whole body. Intravenously injected Na251CrO4 was excreted into the gastrointestinal tract through the gastrointestinal wall and bile ducts. It seemed that 198Au-colloid was not adhered to the gastrointestinal mucosa and consequently, the transport of 198Au-colloid in the gastrointestinal tract was not delayed. It is concluded that 198Au-colloid is highly suitable as a marker of gastrointestinal transit of the chicken but Na251CrO4 is unsuitable.

Distribution of 169Yb microspheres and colloidal 198Au following injection into the rectal submucosa in dogs.

A comparison between two tracers, the one colloidal suspension of 198Au, the other 169Yb-labelled microspheres of 8 to 10 micron has been made concerning lymphatic and hematogenous spread from the submucosa in the rectum in dogs. Both tracers have, in principal, the same distribution. It was therefore considered that a colloidal suspension of 198Au is a useful tracer for functional anatomic investigations of lymph drainage.

[Interpretations of uptake defects in hepatic scintiscanning]. / Interpretazione dei difetti di captazione nella scintigrafia epatica.

The absence of precise parameters for the interpretation of defective uptake means that personal experience is of great assistance in the examination of scintiscans. Classification of uptake defects in 208 cases was carried out by two experts in accordance with the following parameters: 1) irregular uptake or distinct defects; 2) central or peripheral defects; 3) increased size of the liver picture; 4) uptake by the spleen. Irregular uptake or central or peripheral uptake defects (with or without spleen uptake in each case) were observed as categories. The series as a whole showed that peripheral defects are associated with a greater frequency of false positives, and diagnosis must therefore be checked, spleen uptake is an accurate pointer to diffuse chronic hepatopathy, and central defects are indicative of substitutive pathology.

Effect of Corynebacterium parvum, methanol-extraction residue of BCG, and levamisole on macrophage random migration, chemotaxis, and pinocytosis.

Three parameters of macrophage function: random migration, chemotaxis, and pinocytosis, were studied in the guinea pig after administration of Corynebacterium parvum, methanol-extraction residue of BCG, and levamisole (LMS), a synthetic anthelmintic. Macrophage migration studies were performed with a modified Boyden chamber. Pinocytosis was assessed by the uptake of colloidal 198Au. After ip administration, each of the three immunostimulators induced an increase in macrophage chemotactic responsiveness and, to a lesser extent and duration, in random motility. Kinetic, dose-response, and time course data for the effect of each agent on macrophage movement were explored. LMS was the most effective stimulator of macrophage activation, which occurred earlier and persisted longer than it did with the other agents. Macrophages from animals receiving each of the agents showed enhanced pinocytosis. Measurement of macrophage random migration, chemotaxis, and pinocytosis appeared to provide a rapid and quantitative assessment of several parameters of macrophage function and, when studied with other immunologic parameters, may provide useful tools for the evaluation of potential immunoadjuvants.

[Pulmonary uptake of 198Au during liver scanning in two patients with amyloidosis (author's transl)]. / Lungenmitspeicherung im 198Au-Leberszintigramm bei zwei Patienten mit Amyloidose

Lung uptake of 198Au-colloid, injected for a liver scan, is demonstrated in two patients with systemic amyloidosis, The incidence, importance and mechanism of this phenomenon is discussed.

[Radiation dose to the patient after administration of radioactive labelled colloids (author's transl)]. / Die Strahlenbelastung des Patienten durch Applikation radioaktiv markierter Kolloide

The radiation dose to the patient after injection of radioactive colloids is calculated for three substances: colloidal radiogold, 99mTc sulfur colloid and 113mIn colloid. For the calculation normal and pathological conditions were assumed.