A permanent human cell line, EA hy 926, has been established that expresses at least one highly differentiated function of vascular endothelium, factor VII-related antigen. This line was derived by fusing human umbilical vein endothelial cells with the permanent human cell, line A549. Hybrid cells that survived in selective medium had more chromosomes than either progenitor cell type and included, a marker. chromosome from the A549 line. Factor VIII-related antigen can be identified intracellularly in the hybrids by immunofluorescence and accumulates in the culture fluid. Expression of factor VIH-related antigen by these hybrid cells has been maintained for more than 100 cumulative population doublings, including more than 50 passages and three cloning steps. This is evidence that EA-hy 926 represents a permanent line.Differentiated functions of endothelium are critical for the vascularization process in normal and neoplastic tissue, for maintaining the blood-brain barrier, and for hemostasis. Factor VIIIrelated antigen (VIIIR:Ag) is an endothelial cell product (1) involved in the aggregation of platelets, and megakaryocytes are the only other cell type known to express this antigen (2).VIIIR:Ag is present in normal human plasma at about 10 pug/ ml, and decreased levels are found in classical von Willebrand disease, an autosomal dominantly inherited bleeding disorder in humans. VIIIR:Ag circulates as a large molecular complex and its platelet-aggregating activity is directly related to the size distribution of the complex (3). Factor VIII coagulant activity (antihemophilic factor) is also associated with the VIIIR:Ag complex in plasma. We have previously described interspecies hybrids between human endothelial cells and a number of rodent cell lines, in which VIIR:Ag was not expressed (7). There has also been a preliminary report by others (8) of human-rodent hybrids that may express VIIIR:Ag, but the intracellular distribution of antigenicity that they detect is not similar to that of VIIIR:Ag in endothelial cells.Here we report a human intraspecies hybrid, EA-hy 926, that expresses VIIIR:Ag with the same morphological distribution as in primary endothelial cells. MATERIALS AND METHODSCell Culture. Cells were cultured on plastic ware at 370C in a humid atmosphere containing 7% CO2 in air. Culture medium was exchanged every 3-5 days. At confluence, 0.01% trypsin (TRL Worthington, no. LSOO 044 52) in 8 mM phosphate-buffered saline (pH 7.4) with 0.54 mM EDTA and 5.5 mM glucose was used to detach the cells, which were then subcultured at lower cell density.Human umbilical vein endothelial cells (HUV-EC) were isolated as described in detail by Gimbrone (9). The vein of an umbilical cord kept at 40C for 4 hr postpartum was irrigated with phosphate-buffered saline. The endothelial cells were dissociated from the vessel wall with 0.1% collagenase (Sigma, no. C2139) in phosphate-buffered saline at 370C for 20 min. The cells were separated from the collagenase solution by centrifugation and were distributed over...
We have previously described three individuals with a syndrome characterized by hypomagnesemia, hypokalemia, alkalosis, and impaired renal conservation of potassium and magnesium.' This disorder is not associated with an excess of known mineralocorticoids. The urinary excretions of aldosterone, 1 l-hydroxycorticoids, and 17-ketosteroids are not elevated. Aldosterone secretion rates measured in two individuals are high normal but appropriate to their sodium intakes and clinical status. Furthermore, the plasma renin activity is not suppressed but is clearly elevated in all. The basic features of their disorder will be reviewed, emphasizing those aspects that relate to magnesium deficiency in man.Two of the patients are sisters and both are presently well except for complaints related to a chronic, nonspecific dermatitis of many years duration. The skin is thickened with a purple-red hue that is reminiscent of the erythema commonly associated with experimental magnesium depletion in the rat. Both sisters have experienced transient episodes of muscle weakness but otherwise have noted n o symptoms directly attributable to their chemical abnormalities.The occurrence of an unusual syndrome in two siblings prompted an evaluation of their family. The family pedigree is displayed in FIGURE 1. It is noteworthy that their parents are distantly related through a common male ancestor; however, we were unable to demonstrate any similar abnormalities in the other members of the family. The concentration of potassium and magnesium in plasma was normal in the individuals marked by asterisks. Moreover, the patients' parents, their unaffected sibling, and one of their children responded normally to magnesium and potassium restricted diets.The third patient with this syndrome is an unrelated female of similar appearance. No consanguinity was established between her parents, and no similar chemical abnormalities were observed in her immediate family.All three individuals demonstrate an impaired renal conservation of potassium. A representative study is displayed in FIGURE 2. Normal individuals respond to moderate potassium restriction by promptly reducing their urinary excretion of potassium to a level equal to their oral intake. In contrast, the urinary excretion of potassium was significantly greater than the dietary intake in these affected individuals despite their associated hypokalemia.The renal conservation of magnesium was evaluated in a similar fashion, employing a diet that contained approximately 1 mEq of magnesium per 24 hours. These results are tabulated in FIGURE 3. The response of four normal volunteers has *
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.