We used complementary biochemical and immunological techniques to establish that an endothelial cell transmembrane glycoprotein, GP116, is a CD44-like molecule and binds directly both to extracellular matrix components (e.g., hyaluronic acid) and to ankyrin. The specific characteristics of GP116 are as follows: (i) GP116 can be surface labeled with Na125I and contains a wheat germ agglutinin-binding site(s), indicating that it has an extraceHlular domain; (ii) GP116 displays immunological cross-reactivity with a panel of CD44 antibodies, shares some peptide similarity with CD44, and has a similar 52-kDa precursor molecule, indicating that it is a CD44-like molecule; (iii) GP116 displays specific hyaluronic acid-binding properties, indicating that it is a hyaluronic acid receptor; (iv) GP116 can be phosphorylated by endogenous protein kinase C activated by 12-O-tetradecanoylphorbol-13-acetate and by exogenously added protein kinase C; and (v) GP116 and a 20-kDa tryptic polypeptide fragment of GP116 from the intracellular domain are capable of binding the membrane-cytoskeleton linker molecule, ankyrin. Furthermore, phosphorylation of GP116 by protein kinase C significantly enhances GP116 binding to ankyrin. Together, these findings strongly suggest that phosphorylation of the transmembrane glycoprotein GP116 (a CD44-like molecule) by protein kinase C is required for effective GP116-ankyrin interaction during endothelial cell adhesion events.

Although the study was unable to detect any statistically significant differences in outcome between the study groups, this small pilot study shows preliminary findings which may indicate an increase in the short-term eradication of BCC using a PC-based vehicle as compared to conventional petrolatum-based formulations.

A micro method is described for the assay of choline-containing phospholipids in serum and high density lipoproteins (HDL) using an automated microtiter plate reader. The method is adapted from the enzymic method of Takayama, Itoh, Nagasaki, and Tanimuzu (Clin. Chim. Acta 79, 93-98, 1977) using phospholipase D, choline oxidase, and peroxidase coupled with the color generating system phenol and 4-amino-antipyrine. The micro method requires 5 microL of serum or HDL sample, and 42 samples can be assayed in duplicate in one run using a 96-well flat-bottom microtiter plate. The reaction is linear up to 400 mg/dL and the lower limit of detection is 0.25 mg of choline-containing phospholipids per assay. The coefficient of variation within an assay is 0.86-0.79%, and day-to-day variation is 0.9-1.5%. Results obtained by the micro method are in excellent agreement with those obtained by the procedure of Takayama et al. (r = 0.997). The supernatant left after removal of low density lipoproteins and very low density lipoproteins from serum and precipitation with heparin/manganese chloride reagent can thus be conveniently used for the micro assay of choline phospholipids in HDL.

After intravenous injection of liposomes prepared from egg yolk phospholipids into rabbits, the phospholipids were readily assimilated by the lipoproteins, and there were increases in the circulating levels of cholesterol and phospholipids. The increases in cholesterol level were mainly due to increases of free cholesterol. Gradient ultracentrifugation showed that the lipoproteins decreased in density, and gel filtration chromatography showed that they increased in particle size. Upon electrophoresis, they exhibited slower mobility. Liposomes recovered from rabbits 3 hr after the injection contained free cholesterol, apolipoproteins A-I, E and traces of C. The apolipoprotein may target the liposomes for uptake by hepatocytes. Incubation of the liposomes with rabbit red blood cell membranes in vitro caused a decrease in cholesterol content of the membranes. However, the cholesterol/phosphate ratio in red blood cells isolated from the rabbits after the injection of liposomes did not change significantly, suggesting rapid replenishment of red blood cell cholesterol in vivo, possibly by equilibration with lipoprotein cholesterol or tissue cholesterol. These results suggest that the injection of phospholipid liposomes may have an antiatherogenic effect by the removal of tissue cholesterol and enhancing hepatic disposal of cholesterol through the reverse cholesterol transport mechanism.

No abstract