We have investigated the effect of epidermal growth factor (EGF) on collagen metabolism in clonal MC3T3-E1 cells, an osteoblastic cell line derived from newborn mouse calvaria. EGF significantly increased DNA synthesis, but decreased collagen production. We analyzed the amount of total collagen synthesis and degradation products of collagen together with the level of the enzyme responsible for extracellular collagen degradation, to investigate whether the decreased collagen production was due to a decrease in total collagen synthesis or to an increase in collagen degradation. Total collagen synthesis, determined by total hydroxyproline synthesized, was significantly decreased in cells cultured in medium containing EGF, but the amount of collagen degradation products and the level of animal collagenase activity were not increased. Analysis of the collagen type produced by the cells in the absence of EGF showed that 95% of the collagen recovered was type I and 3% was type III. The decreased level of collagen accumulated by cells cultured in the presence of EGF was explained only by the decreased rate of type I collagen synthesis. These results indicate that EGF selectively inhibits type I collagen synthesis in the clonal osteoblastic cell line, MC3T3-E1.
The effect of epidermal growth factor (EGF) on clone MC3T3-El cells that have osteoblastic activity was examined by phase-contrast microscopy and electron microscopy; hydroxyproline content, collagen synthesis, collagen pattern, and alkaline phosphatase (ALP) activity were also determined. We found that EGF (0.4 ng/ml) transformed the cells from their normal polygonal shape to a spindle-like morphology by 8 h. This hormone also caused dose-related suppression of hydroxyproline content and ALP activity which was detectable 2 days and 1 day, respectively, after EGF addition. Indomethacin did not affect hydroxyproline content and ALP activity, suggesting that the effect of EGF on the cells may not be mediated by prostaglandins. Epidermal growth factor at concentrations of 2 to 50 ng/ml significantly decreased collagen synthesis in the cells, whereas protein synthesis was stimulated. Electron microscopy demonstrated that collagen fiber formation was also reduced by EGF; an immature type of fibril was observed compared with the typical cross-striated one in the controls. Moreover, the hormone treatment also resulted in the appearance of type III collagen in addition to the type I already present in the cells. These suppressive effects of EGF on MC3T3-El cells in vitro suggest that this hormone may be involved in bone remodelling in vivo as well.
The effects of prostaglandins (PGs) on the induction of alkaline phosphatase (ALP) were investigated in osteoblastic clone MC3T3-E1 cells cultured in serum-free medium. Prostaglandin E2 (PGE2) stimulated ALP activity in the cells in a dose-dependent fashion with a maximal effect which was about twice that in the control cells at concentrations of 100-500 ng/ml. Actinomycin D and cycloheximide inhibited the stimulative effect of PGE2 on ALP activity in the cells. PGE2-induced and native ALPs in the cells were of the same type as that in adult mouse calvaria, being heat-labile, L-homoarginine- and levamisole-sensitive, and L-phenylalanine-insensitive. Isobutyl methylxanthine (IBMX), a cAMP phosphodiesterase inhibitor, stimulated the inductive effect of PGE2 on ALP activity at 0.1 mM, at which concentration IBMX alone had little effect on the activity. PGE2 also increased the intracellular cAMP content in a dose-dependent fashion with a maximal effect at 100 ng/ml. PGE1, PGF1 alpha, and PGF2 alpha (primary PGs like PGE2) increased the activity. Our present results suggest that PGs stimulate the differentiation of osteoblasts and are involved in bone formation in vivo, as well as in bone resorption.
Prostate glands of adult guinea pigs were stained for nerve growth factor (NGF) and epidermal growth factor (EGF) by immunohistochemical methods. Both NGF and EGF were localized diffusely in the cytoplasm of the glandular epithelial cells, and also in their secretory products. These findings suggest that NGF and EGF are synthesized, stored, and secreted by the glandular epithelial cells of the prostate.
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