Increased hyaluronate and collagen biosynthesis and fibroblast estrogen receptors in macaque sex skin.

Sequential steroid administration of estradiol (E2) and progesterone (P) in spayed pigtailed macaques was used to precisely control the time course of sex skin swelling. After removal of the P implant, the sex skin swelled considerably and the water content of the sex skin increased manyfold over that of back skin. During the swelling phase, hyaluronate biosynthesis in sex skin increased dramatically compared with back skin of the same animals. Collagen synthesis also increased but to a lesser extent. Estrogen receptor levels were undetectable in back skin and very low in spayed animals that had been treated with both E2 and P. After removal of the P implant, both the level of estrogen receptor and the rate of hyaluronate biosynthesis increased. Immunocytochemistry with monoclonal antibodies against the estrogen receptor showed that the dermal fibroblast was the only cell type to stain positively for estrogen receptor. We conclude that the sex skin swelling that follows P withdrawal in pigtailed macaques bearing E2 implants is mediated by estrogen receptors in dermal fibroblasts and is a result of increased hyaluronic acid synthesis by these cells.

Biosynthesis of aortic collagen and glycosaminoglycan following immunological injury.

The effect of immunological injury upon the early in vivo changes in aortic connective tissue metabolism was studied. It was found that a single antigen (bovine serum albumin), when administered in multiple doses, activated collagen synthesis and increased the rate of lgycosaminoglycan synthesis in the arterial wall. The degree of stimulation of both connective tissue components was higher in animals receiving a higher dose of antigen.

The effect of lysosomal enzymes on the proliferation of aortic smooth muscle cells and fibroblasts.

The rate of 3H-thymidine incorporation and cell multiplication was investigated in aortic smooth muscle cells and WI-38 fibroblasts exposed to lysosomal enzymes isolated from liver or alveolar macrophages. It was found that cells grown in the presence of lysosomal enzymes reached higher final densities than untreated cells and the rate of 3H-thymidine incorporation was markedly increased in the enzyme-exposed cultures. The stimulation of 3H-thymidine uptake was detected both in growing and quiescent cultures of smooth muscle cells and WI-38 fibroblasts. The precursor taken up by the lysosomal enzyme treated cells was incorporated into a DN-ase susceptible material. These experiments illustrate that lysosomal enzymes, when present in low concentration in the cellular environment, can act as mitogens and may play a role in the pathogenesis of atherosclerosis.

Effect of calcium on cell proliferation and extracellular matrix synthesis in arterial smooth muscle cells and dermal fibroblasts.

The effect of calcium on cell proliferation and connective tissue formation was studied in cultured vascular smooth muscle cells (SMC) and dermal fibroblasts. Calcium deficiency caused a modest decrease in proliferation of smooth muscle cells but this effect was small compared to that previously observed with fibroblasts. Synthesis of connective tissue components was affected differently in the two cell types. Biosynthesis of proteoglycans was assessed by metabolic labeling of their glycosaminoglycan side chains. Different levels of extracellular calcium did not affect proteoglycan production by fibroblasts, but it was significantly reduced in smooth muscle cells incubated in calcium-deficient medium. Both smooth muscle cells and fibroblasts were able to produce appreciable amounts of collagen in the complete absence of calcium and in both cell types collagen synthesis was increased when calcium was present. Fibroblasts, however, showed a much smaller response to calcium than did smooth muscle cells. In fibroblasts the maximum rate of collagen synthesis was achieved in a narrow range of calcium concentration which was slightly below that found commonly in the tissue culture medium. By contrast, in smooth muscle cells the rate of collagen synthesis increased greatly when calcium was present and this elevated rate persisted even when the cells were exposed to high levels of extracellular calcium. We conclude that these findings may be of significance to the development of atherosclerotic lesions.

The metabolic heterogeneity of rabbit ear cartilage chondroitin sulphate.

The only glycosaminoglycans that can be isolated from the ear cartilage of 2-month-old rabbits are chondroitin 4-sulphate and chondroitin 6-sulphate. These chondroitin sulphates exhibit molecular-weight polydispersity when isolated from tissue by papain digestion. The chondroitin sulphate is metabolically heterogeneous in that radioactive precursors [(14)C]glucose or [(35)S]sulphate are preferentially incorporated into the higher-molecular-weight polymers both in vivo and in vitro. No transfer of radioactivity from the high-molecular-weight chondroitin sulphate to the low-molecular-weight chondroitin sulphate was seen during 15 days in vivo. It is suggested that there are at least two pools of proteoglycan in the tissue. One of these pools is metabolically active whereas the other is not.

Arterial cholesterol content and collagen metabolism in cynomolgus monkeys.

The activity of prolyl hydroxylase and the rate of collagen synthesis were examined in two anatomically different arterial segments of cholesterol-fold cynomolgus monkeys. In the aortic arch vessels the activity of prolyl hydroxylase was increased after six months on a high fat and high cholesterol diet. In this tissue the enzyme activity showed positive correlation with the levels of plasma and tissue cholesterol content. By contrast in the abdominal aortas of the same animals no significant increase of arterial tissue cholesterol or prolyl hydroxylase activity was seen. Despite the elevation of prolyl hydroxylase activity in the arch vessels of hyper cholesterolemic animals, no concomitant increase of collagen synthesis was seen. Neither cholesterolemia nor arterial cholesterol showed a positive correlation with the rate of arterial collagen synthesis. No significant changes of the rate of collagen metabolism were detected at the end of an 18 months intervention phase when the cholesterol content both in plasma and in the arterial tissue was substantially reduced.

Composition and metabolism of symptomatic distal aortic plaque.

This study examines the composition of advanced human atherosclerotic lesions removed by endarterectomy and compares the collagen synthesis in the lesion with that of the intima-media layer of uninvolved vessel wall. The most striking feature of the lesions is an enormous accumulation of calcium that was raised more than tenfold, from a level of about 3% of dry tissue weight in the normal tissue to values approaching 50% in the pathologic specimens. The composition of the organic matrix was also altered. Elastin and glycosaminoglycan content decreased, whereas collagen content more than doubled, thus becoming the major component of the organic matrix. The cholesterol content increased severalfold above normal but even then accounted for less than 10% of the organic matrix. The lesion specimens synthesized collagen in vitro at a greater rate than normal tissue. The endarterectomy specimens with the highest calcium content exhibited the highest rate of collagen synthesis.