A method is described for the rapid isolation of a plasma membrane fraction containing a high concentration of intact bile canaliculi from the rat liver . Isolated bile canaliculi retain most of the ultrastructural features exhibited in the intact liver cell . The final fraction contains 5'-nucleotidase activity at approximately the same concentration as that in previous preparations of plasma membranes . In the presence of 0 .01 M Mg++, 5'-nucleotidase exhibits a double pH optimum at pH values of 7 .5 and 9 .5. The activities of glucose-6-phosphatase and alkaline phosphatase are present in low amounts . Cytochrome P-450 is not detectable . Na+-K+-activation of ATPase is observed to the extent of 20-36 0/, in about half of the assays. The availability of a method for preparation of intact bile canaliculi should prove useful for studying the biochemical events associated with the transport of bile constituents into canaliculi .

A B S T R A CT Polymorphonuclear neutrophils (PMN) from patients with chronic granulomatous disease of childhood have impaired bactericidal activity and are deficient in diphosphopyridine nucleotide, reduced form of, (NADH) oxidase. Since hydrocortisone had been shown to inhibit NADH oxidation, experiments were undertaken to determine the effect of hydrocortisone on several parameters of human PMN function.The phagocytic and bactericidal capacity of PMN with or without hydrocortisone (2.1 mM) was determined by quantitation of cell-free, cell-associated, and total bacteria. Phagocytosis of Staphylococcus autreus and several gram-negative rods was unimpaired by the presence of hydrocortisone in the media. In contrast, killing of bacteria was markedly impaired by hydrocortisone. After 30 min of incubation, there were 20-400 times as many bacteria surviving in hydrocortisone-treated PMN as in simultaneously run controls without hydrocortisone.The defect of intracellular killing noted in the presence of hydrocortisone was not related to impaired degranulation. Quantitative kinetic studies of degranulation revealed no difference in the release of granule associated acid phosphatase in hydrocortisone-treated and control PMN after phagocytosis. Electron microscopy of PMN also indicated that the presence of hydrocortisone had no effect on the extent of degranulation after phagocytosis. These observations were confirmed by studies using histochemical techniques to detect lysosomal enzymes.After phagocytosis, hydrocortisone-treated P.N demonstrated less NADH oxidase activity, oxygen consumption, and hydrogen peroxide production than postphagocytic control PMN. In addition, Nitro blue tetrazolium dye reduction was diminished in hydrocortisonetreated PMN.

No abstract

No abstract