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Present concepts of the roles of collagen and elastin in lung elastic behavior and maintenance of lung structure have been largely inferred from anatomical observations or from studies of isolated fibers in vitro. Based on the intimate association of elastin and collagen it has been postulated that elastin contributes little to elastic behavior and that collagen is the major determinant of lung structure. Using clostridial collagenase, pancreatic elastase, and papain we have selectively degraded these fibers and studied the resulting changes in elastic behavior and structure of rat lungs in vitro.Pressure-volume curves were recorded during continuous slow air inflation and deflation (10.5 ml/min) before and after the intratracheal instillation of 0.5 ml of control or enzyme solution. Surface tension-lowering activity of lavaged material was studied. All lungs were fixed inflated at 25 cm H(2)O pressure and whole lung sections were stained for elastin, collagen, and reticulin. Collagenase produced a marked susceptibility to pleural rupture but did not alter elastic behavior or lung structure. Elastase and papain produced segments of lung with increased compliance; this change was not due to alteration in surface forces but was associated with decreased tissue elastic recoil. Histologically, altered tissue recoil correlated well with evidence of damaged elastin fibers. In contrast to previous concepts these results suggest that elastin is the major connective tissue determinant of lung structure and elastic behavior.

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.

Neurite outgrowth in cultured neuroblastoma cells is inhibited in cells grown on a collagen substratum as compared to those grown on glass. Rapid, synchronized initiation of neurite outgrowth occurs after hydrolysis of the underlying collagen with collagenase. Axonated cultures exhibit an increased RNA and protein content as compared to cultures grown on collagen.