Despite its very potent vasodilating action in vivo, acetylcholine (ACh) does not always produce relaxation of isolated preparations of blood vessels in vitro. For example, in the helical strip of the rabbit descending thoracic aorta, the only reported response to ACh has been graded contractions, occurring at concentrations above 0.1 muM and mediated by muscarinic receptors. Recently, we observed that in a ring preparation from the rabbit thoracic aorta, ACh produced marked relaxation at concentrations lower than those required to produce contraction (confirming an earlier report by Jelliffe). In investigating this apparent discrepancy, we discovered that the loss of relaxation of ACh in the case of the strip was the result of unintentional rubbing of its intimal surface against foreign surfaces during its preparation. If care was taken to avoid rubbing of the intimal surface during preparation, the tissue, whether ring, transverse strip or helical strip, always exhibited relaxation to ACh, and the possibility was considered that rubbing of the intimal surface had removed endothelial cells. We demonstrate here that relaxation of isolated preparations of rabbit thoracic aorta and other blood vessels by ACh requires the presence of endothelial cells, and that ACh, acting on muscarinic receptors of these cells, stimulates release of a substance(s) that causes relaxation of the vascular smooth muscle. We propose that this may be one of the principal mechanisms for ACh-induced vasodilation in vivo. Preliminary reports on some aspects of the work have been reported elsewhere.

The obligatory role of endothelial cells in the relaxation of arterial smooth muscle by acetylcholine

Atherosclerosis, the principal cause of heart attack, stroke and gangrene of the extremities, is responsible for 50% of all mortality in the USA, Europe and Japan. The lesions result from an excessive, inflammatory-fibroproliferative response to various forms of insult to the endothelium and smooth muscle of the artery wall. A large number of growth factors, cytokines and vasoregulatory molecules participate in this process. Our ability to control the expression of genes encoding these molecules and to target specific cell types provides opportunities to develop new diagnostic and therapeutic agents to induce the regression of the lesions and, possibly, to prevent their formation.

The pathogenesis of atherosclerosis: a perspective for the 1990s

Endothelial cells were isolated from freshly obtained human umbilical cords by collagenase digestion of the interior of the umbilical vein. The cells were grown in tissue culture as a homogeneous population for periods up to 5 mo and some lines were subcultured for 10 serial passages. During the logarithmic phase of cell growth, cell-doubling time was 92 h. Light, phase contrast, and scanning electron microscopy demonstrated that cultured human endothelial cells grew as monolayers of closely opposed, polygonal large cells whereas both cultured human fibroblasts and human smooth muscle cells grew as overlapping layers of parallel arrays of slender, spindle-shaped cells. By transmission electron microscopy, cultured endothelial cells were seen to contain cytoplasmic inclusions (Weibel-Palade bodies) characteristic of in situ endothelial cells. These inclusions were also found in endothelial cells lining umbilical veins but were not seen in smooth muscle cells or fibroblasts in culture or in situ. Cultured endothelial cells contained abundant quantities of smooth muscle actomyosin. Cultured endothelial cells also contained ABH antigens appropriate to the tissue donor's blood type; these antigens were not detectable on cultured smooth muscle cells or fibroblasts. These studies demonstrate that it is possible to culture morphologically and immunologically identifiable human endothelial cells for periods up to 5 mo.

/pdf/culture-of-human-endothelial-cells-derived-from-umbilical-3vt4ihjkgj.pdf

Culture of Human Endothelial Cells Derived from Umbilical Veins. IDENTIFICATION BY MORPHOLOGIC AND IMMUNOLOGIC CRITERIA

Ralph L. Nachman

Papers

Culture of Human Endothelial Cells Derived from Umbilical Veins. IDENTIFICATION BY MORPHOLOGIC AND IMMUNOLOGIC CRITERIA

A clinical study of the lupus anticoagulant

Binding of plasminogen to cultured human endothelial cells.

Human Bone Marrow Microvascular Endothelial Cells Support Long-Term Proliferation and Differentiation of Myeloid and Megakaryocytic Progenitors

Isolation and characterization of human bone marrow microvascular endothelial cells: hematopoietic progenitor cell adhesion