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M K Pratten

Publications and source records attributed to M K Pratten.

52 records · Page 3Linked to original sources
PubMed1981

Effect of molecular size of 125I-labelled poly(vinylpyrrolidone) on its pinocytosis by rat visceral yolk sacs and rat peritoneal macrophages.

Rates of pinocytosis of different molecular-weight distributions of 125I-labelled poly(vinylpyrrolidone) by rat visceral yolk sacs and rat peritoneal macrophages were measured in vitro. Four preparations of mean molecular weights 50 000, 84 000, 700 000 and 7 000 000, were used. Macrophages captured the highest-molecular-weight preparation more rapidly than the other preparations. In contrast, rate of capture by the yolk sac decreased with increasing molecular weight. Incubations with a very-high-molecular-weight fraction derived from the 7 000 000-average-mol. wt. preparation clearly demonstrated that very large polymer molecules are not accumulated by the yolk sac, but are preferentially captured by macrophages. Analysis of the 125I-labelled poly(vinylpyrrolidone) internalized by the two cell types confirmed that low-molecular-weight material is preferred by the yolk sac, whereas the macrophage is less discriminating.

Animals↗
PubMed1981

Serum-dependence of fluid-phase pinocytosis and specificity in adsorptive pinocytosis of simple proteins in rat peritoneal macrophages.

An increase in the concentration of serum resulted in a very marked increase in the rate of fluid-phase pinocytosis in rat peritoneal macrophages, as measured by the uptake of 125I-labelled polyvinylpyrrolidone. In contrast, the rate of uptake of colloidal [198Au]gold, an adsorptive substrate, decreased when the serum concentration was increased. This lessened uptake of colloidal gold must be due to competition by serum components. Studies on the specificity of pinocytosis of simple proteins, using 125I-labelled H4 and M4 forms of porcine lactate dehydrogenase, and formaldehyde-denatured and untreated bovine serum albumin as substrates, suggest that positive charge and hydrophobicity determine adsorptive pinocytosis of simple proteins in peritoneal macrophages. The rate of fluid-phase pinocytosis and the specificity of adsorptive pinocytosis in peritoneal macrophages are very similar to those reported for Kupffer cells.

Adsorption↗
PubMed1981

Uptake by rat peritoneal macrophages of 125I-labelled polyvinylpyrrolidone entrapped within liposomes.

Rat peritoneal macrophages in vitro capture 125I-labelled polyvinylpyrrolidone entrapped within either negatively or positively charged liposomes more rapidly than they do the free macromolecule. The uptake of negatively charged liposomes was linear with time over 10 h, whilst the uptake of positively charged ones, although more rapid, was more transient. Neither type of liposome was taken up in the presence of 2,4-dinitrophenol (100 microgram/ml), and 5 mM calcium chloride increased the uptake of negatively charged liposomes. The enhanced uptake of 125I-labelled polyvinylpyrrolidone when presented in liposomes must have been a consequence of entrapment rather than of a simple interaction between lipid and polyvinylpyrrolidone since the presence of the lipids employed or of empty liposomes had no effect on the uptake of unentrapped 125I-labelled polyvinylpyrrolidone.

Animals↗
PubMed1981

Pinocytic uptake of divinyl ether-maleic anhydride (pyran copolymer) and its failure to stimulate pinocytosis.

The effect of DIVEMA (pyran copolymer) and three DIVEMA derivatives on the pinocytic uptake of 125I-labeled PVP and colloidal 198Au by the rat visceral yolk sac and by rat peritoneal macrophages was studied in vitro. Contrary to expectations from some earlier data, there was no enhancement of pinocytosis and in some cases inhibition was seen. [14C]DIVEMA and 125I-labelled DIVEMA were accumulated rapidly by rat peritoneal macrophages, the results indicating that this is by an adsorptive pinocytic mechanism.

Animals↗
PubMed1981

Endocytosis of simple proteins by rat yolk sacs and by rat peritoneal macrophages incubated in vitro.

The specificity of endocytosis of the 125I-labelled forms of LDH H4 and M4 and BSA was investigated in rat yolk sac and resident rat peritoneal macrophages incubated in vitro. The rate of uptake of the more basic LDH M4 by yolk sac is only slightly higher than that of LDH H4, but in macrophages the isoenzyme M4 is taken up three times more rapidly than the H4 form. Both cell types possess little affinity for BSA, but an increase in the hydrophobicity of this protein by formaldehyde-denaturation resulted in a several-fold increase in uptake by yolk sac cells as well as by macrophages. Apparently, yolk sacs and macrophages both contain binding sites for positively charged and hydrophobic proteins, but, in yolk sac hydrophobicity is relatively more important, whereas in macrophages positive charge is the main characteristic for adsorptive pinocytosis of simple proteins. In this respect, the binding characteristics of peritoneal macrophages resemble those of sinusoidal liver cells. The study also compares the serum-dependence of fluid-phase pinocytosis in both cell types; a decrease in serum concentration gives rise to an increase in the rate of pinosome formation in yolk sac, but has the opposite effect in macrophages.

Animals↗
PubMed1979

Mechanism of polycation stimulation of pinocytosis.

Synthetic polycations cause a stimulation in the rate of tissue accumulation of colloidal 198Au by the rat visceral yolk sac (at 17.5 days of gestation) and rat peritoneal macrophages cultured in vitro. The mechanism of stimulation has been elucidated in these two cell types by using a dual-substrate technique, and by examining the differential effects of poly(D-lysine) and poly(L-lysine) and of metabolic and cytoskeletal inhibitors. Polycations cause aggregation of colloidal 198Au in the culture medium and increase its affinity for the plasma membrane. In the rat peritoneal macrophage this polycation-colloidal gold complex is pinocytosed, thus enhancing the intracellular accumulation of the radio-labelled substrate. In contrast, the rat visceral yolk sac cannot internalize this complex, and so the substrate accumulates extracellularly. This mechanism of polycation modification affords the opportunity for differential uptake of a substrate into distinct cell types.

Animals↗
PubMed1979

Effects of temperature, metabolic inhibitors and some other factors on fluid-phase and adsorptive pinocytosis by rat peritoneal macrophages.

Low temperature, NaF and 2,4-dinitrophenol could each abolish the pinocytic uptake of 125I-labelled poly(vinylpyrrolidone) or colloidal [198Au]gold by rat peritoneal macrophages cultured in vitro. Cytochalasin B caused only partial inhibition, even at 10 microgram/ml, and colchicine (10 or 25 microgram/ml) inhibited uptake of colloidal [198Au]gold much more than that of 125I-labelled poly(vinylpyrrolidone). Dibutyryl cyclic AMP and ouabain were without effect on uptake of 125I-labelled poly(vinylpyrrolidone), and slight stimulation was seen with ATP and theophylline. Uptake of 125I-labelled poly(vinylpyrrolidone) was abolished by EGTA (5mM), but restored by adding CaCl2 (5mM). The results appear not to support the conventional criteria for the division of pinocytic phenomena into macropinocytosis, requiring a metabolic energy supply and cytoskeletal components, and micropinocytosis, requiring neither.

Adsorption↗
PubMed1979

Comparative studies on the nature of purified cytomembranes of the rabbit parotid gland.

Centrifugation procedures have been evolved for isolating purified samples of rough endoplasmic reticulum, Golgi, zymogen granule and plasmalemmal membranes from homogenates of rabbit parotid gland tissue. The purification process was monitored using morphometry and enzyme and chemical marker assays. The membrane preparations were analysed by sodium dodecylsuphate (SDS) polyacrylamide gel electrophoresis, quantitative phospholipid thin layer chromatography and by enrichment studies. The results were used to evaluate various possible general models for the behaviour of membranes during the secretory cycle of parotid acinar cells.

Animals↗
PubMed1977

A quantitative study of pinocytosis and intracellular proteolysis in rat peritoneal macrophages.

A method for the culture of rat peritoneal macrophages in vitro is described, in which pinocytic uptake of colloidal [198 Au]gold, 125I--labelled poly(vinylpyrrolidone) and [14C]sucrose proceeds at contant and fairly reproducible rates for several hours. The rat of uptake of colloidal [198 Au]gold, which wxhibited some inter-batch variation, was approx. 100 times that of the other two substrates. Colloidal gold did not affect the rate of uptake of 125I-labelled poly(vinylpyrrolidone) and therefore its own high rate of uptake could not be attributed to a stimulation of the formation of pinocytic vesicles. It conclude that uptake of collodial gold is highly dependent on adsorption on binding sites on the plasma membrane. Uptake of formaldehyde-treated 125I-labelled bovine serum albumin was followed by the release of [125I]iodo-L-tyrosine into the culture medium and took place at a rate intermediate between those of collodial [198Au]gold and the other two non-digestible substrates, 125I-labelled poly(vinylpyrrolidone) and [14C]sucrose.

Animals↗
PubMed1977

Compartmentation of enzymes in the rabbit parotid salivary gland. A study by enzyme histochemical, tissue fractionation and morphometric techniques.

Rabbit parotid gland was chosen as a suitable model tissue for studying the role of membranes in enzyme secretion by acinar cells. The study was initiated using subcellular fractionation techniques. During these experiments the effects of various tissue disruption conditions such as the medium and the duration and vigour of homogenization were explored and the results assessed by enzyme and chemical assays and both quantitatively and qualitatively by electron microscopy. A series of basic fractions was isolated and marker enzyme or chemical assays selected for each of the relevant membrane types (rough endoplasmic reticulum, Golgi apparatus, zymogen granule, plasmalemma). A parallel study was effected using enzyme histochemical methods applied to frozen sections. Interesting comparisons could then be made between histochemical and biochemical methods of enzyme demonstration. These comparisons are discussed. The basic fractions provide the material from which specimens of purified membranes of the four species can be obtained. The isolation and characterization of such purified membranes is the subject of another report.

Acid Phosphatase↗
PubMed1976

Correlative morphological and biochemical study of the effects of isoprenaline on the organelle and membrane content of the rabbit parotid gland.

A quantitative study has been made on the enzymic, chemical and ultrastructural changes that occur in the parotid glands of rabbits as a result of Isoprenaline-induced secretion. Emphasis has been placed on correlating changes in organelle and membrane content which are evident 2 hr after Isoprenaline administration and which have been measured stereologically with the levels of appropriate enzymic or chemical markers, taking into account the contribution made by both the acinar and duct tissue. Lower protein, alpha-amylase and beta-glycerophosphatase levels correlated with reductions in zymogen granule and lysosome volume with whilst plasmalemmal and Golgi membrane areas and their marker enzyme concentrations remained unchanged. However, declines in alkaline phosphatase and succinate dehydrogenase activity (illustrated histochemically), and p-nitrophenyl phosphatase activity at pH 4-5 in the presence of tartrate occurred without any detectable decrease in membrane area. Conversely, an increase in rough endoplasmic reticulum area was measured stereologically but no increases in chemical markers were detected. The extent of correlation of the data is discussed in the context of the mechanism of secretion and the action of Isoprenaline.

Amylases↗
PubMedDate not supplied

Uptake and digestion of 125I-labelled bovine serum albumin by the rat visceral yolk sac cultured in vitro as a closed vesicle.

A system for culturing the rat visceral yolk sac in vitro as a closed vesicle--the 'giant' yolk sac--has been employed to investigate the vectorial nature of the uptake and digestion of exogenous protein substrates. Uptake of 125I-labelled formaldehyde-denatured bovine serum albumin by such yolk sacs was found to be similar to that observed in yolk sacs removed directly from the mother at 17.5 days' gestation, provided that homologous serum was used as a culture medium. However, unlike the control yolk sacs, giant yolk sacs tended to accumulate substrate within the tissue with increasing culture time. The concentration of digestion products released to the inside of the closed vesicle was found to be greater than that released to the surrounding culture medium at time intervals up to five hours. Giant yolk sacs preloaded with 125I-labelled bovine serum albumin were found to release material to the culture medium or the inside of the vesicle almost entirely in the acid-soluble (digested) form. This system is a useful model for studying the polar nature of epithelia, particularly those involved in the uptake and transport of nutritional and/or informational macromolecules.

Animals↗
PubMedDate not supplied

Differences in binding characteristics of rat and human transferrin by rat visceral yolk sac placenta.

Previous work has shown that human serum supplemented with rat transferrin can support the normal growth of cultured rat conceptuses, but that supplementation with human transferrin has no such effect. Such results strongly suggest an hypothesis of species-specificity. This potential specificity was therefore investigated by comparing uptake, binding, competition and vectorial transport of both rat and human transferrin using two well-established systems, those of 17.5 day rat visceral yolk sacs and anembryonic yolk sacs in culture. The results of these investigations show that human transferrin displays a lower rate of uptake and lower binding affinity for the rat transferrin receptor than does rat transferrin. Human transferrin competes poorly with rat transferrin for receptor occupancy unless present in 20-fold excess. Both molecules are taken up by receptor-mediated endocytosis and are processed in a similar manner. Anembryonic yolk sac experiments show that a greater proportion of intact rat transferrin is transported to the exocoelomic fluid than is intact human transferrin. Binding analyses show a difference in binding affinities of the two molecules for the rat transferrin receptor and also that human transferrin exhibits negative cooperativity in its binding. This evidence strongly supports an hypothesis of species-specificity in the binding of transferrin to the transferrin receptor in the rat visceral yolk sac.

Animals↗
PubMedDate not supplied

Processing of fluorescently labelled insulin and insulin-like growth factor-I by the rat visceral yolk sac.

Insulin and the structurally related insulin-like growth factor I (IGF-I) are mitogenic peptides which have been implicated in the embryonic development of the rat. In addition to factors produced by the developing embryo itself, it is likely that maternally-derived growth factors play an important role also, with their postulated initial site of action being the extraembryonic membranes, which surround the embryo throughout gestation. We have examined the processing of these potential regulatory factors by the visceral yolk sac on the 17th day of gestation, using fluorescently-labelled ligands and fluorescence microscopy. Both insulin and IGF-I are rapidly internalized at the yolk sac surface, and appear in the tissue within discrete vesicular structures. Interestingly, in some cases when both labelled proteins are added simultaneously they do not appear to coexist within vesicles. Instead, insulin appears to remain within vesicles close to the apical surface of the yolk sac whereas IGF-I appears to penetrate the tissue more deeply, being readily transported to the internal face of the epithelium. It appears, therefore, that there is some difference in the sorting mechanisms of these related proteins, although the physiological significance of this observation is not clear.

Animals↗
PubMedDate not supplied

The role of exogenous growth-promoting factors and their receptors in embryogenesis.

During organogenesis, the cells of the embryo may require growth factors that promote a cascade of intracellular events. An absolute requirement for exogenous insulin by presomite 9.5-d rat embryos grown in culture has been demonstrated. The uptake and processing of insulin and insulin-like growth factor-I showed different uptake and localization patterns. When epidermal growth factor (EGF) or "long EGF" is added to media depleted of low molecular weight material, a dose-dependent improvement in growth is observed. Furthermore, the specific EGF receptor signal transduction inhibitor Tyrphostin 47 can inhibit embryonic growth when it is administered in culture. When Tyrphostin 47 was microinjected into embryos on Day 11 and their growth and differentiation evaluated on Day 12 of gestation, a dose-dependent decrease in developmental score was observed. Thus, exogenous growth factors may be essential to normal rat development and these may be synthesized locally in the decidua or placental tissues. Perturbations to ligand-receptor interactions may be a mechanism for dysmorphogenesis.

Animals↗