Properties of cultured endothelium from adult human vessels.

Abstract

Endothelium was isolated from samples of aorta and vena cava obtained from cadaver donors at the time kidneys were harvested for transplantation. Digestion with collagenase and gentle swabbing were used to free the cells from the Intlmal surface. Low density seeding permitted isolation of individual colonies with typical endothellal morphology. Modified Medium 199 supplemented with 10%-20% human plasma-derived serum and an extract from the bovine hypothalamus (500 ^g/ml) enabled subcultured colonies to grow to confluency when culture surfaces were coated with fibronectin (1 /xg/cm 2 ). The presence of Factor VIII antigen was demonstrated using an indirect immunofluorescence technique. A monoclonal antibody to cultured umbilical vein endothelium, specific for endothelium, reacted with the subcultured cells from the aorta and vena cava. Type IV procollagen, flbronectln, and thrombospondln were identified as labeled proteins secreted by cultures of adult endothelium that had been Incubated with 3 H-prollne and 3 H-glycine. When the cultured endothelium was used In a sodlum-m-periodate stimulated T lymphocyte mltogenic culture system, the endothelium exhibited accessory cell function. Prostacyclln production stimulated by incubation with arachldonlc acid and PGH 2 was variable from vessel to vessel. However, average values were lower than normally seen with cultured primary umbilical vein endothelium. (Arteriosclerosis 4:4-13, January/February 1984) M any discoveries concerning endothelial function have been made in recent years following the development of methods for culturing endothelium. Umbilical vein endothelium has been used by many groups while others have studied endothelium from the bovine aorta, pulmonary artery, or vein. Both qualitative and quantitative differences have been observed in functional assays with endothelial cells from these different sources. We have examined adult endothelium under similar conditions to ascertain its functional characteristics in culture. Received March 19, 1982; revision accepted July 26, 1983. Early attempts to culture endothelium from adult aorta and vena cava were unsuccessful. It was necessary to change the culture conditions that we normally used for primary cultures of umbilical vein endothelium. We previously reported 12 that by modifying the culture conditions we have been able to isolate colonies with endothelial morphology. We have grown sufficient numbers of adult aorta and vena cava endothelium to compare with umbilical vein endothelial cultures. Primary cultures of endothelium from the vena cava and aorta of the same donor, subcultured endothelium from aorta and vena cava samples, primary umbilical vein endothelium and umbilical vein endothelium subcultured with the modification used for adult endothelium have been examined for functional ability by a variety of different assays.

Von Willebrand Factor in Cultured Human Vascular Endothelial Cells from Adult and Umbilical Cord Arteries and Veins

“…In cultures of endothelial cells, the prostacyclin production in response to stimuli is somewhat variable (Thble 3). This confirms previous observations (5). The reason for the low prostacyclin production, sometimes seen in adult endothelial cells, could be a real property of these cells, i.e.…”

“…A. van (3)(4)(5) have reported on the properties of cultured human endothelium from adult vessels, including the production of collagens and prostaglandins. However, the synthesis, storage and secretion of the von Wllebrand factor by endothelial cells from other origins than umbilical cord were not studied in much detail.…”

Induction of prostacyclin biosynthesis is closely associated with increased guanosine 3',5'-cyclic monophosphate accumulation in cultured human endothelium.

“…Lastly, we have found (31) that early passage (1st through 3rd) of primary cultures of human umbilical vein endothelium results in a progressive, parallel loss of the capacity of these cells to produce PGI2 or to synthesize cyclic GMP in response to extracellular stimuli. As these passaged cells retain all other characteristic morphologic and physiologic properties of primary endothelial cultures (32), it is clear that the co-availability of physiologically responsive (i.e., with respect to PGI2 and cyclic GMP biosynthesis) and progressively nonresponsive populations of the same cell type provides a unique opportunity to examine the factors involved in the expression of stimulus-response coupling mechanisms as well as the regulatory role of cyclic GMP.…”

Evidence for a discrete UTP receptor in cardiac endothelial cells

“…The endothelial origin of cultured cells was routinely confirmed by positive staining in immunofluorescent assays using FITC-labelled antibodies directed against clotting Factor VIII (Fry et al, 1984). Cultured uterine smooth muscle cells and lung fibroblasts served as negative controls.…”