Effects of Cyclic Nucleotides on Deoxyribonucleic Acid Synthesis in Hypophysectomized Rat Cartilage: Stimulation of Thymidine Incorporation and Potentiation of the Action of Somatomedin by Analogs of Adenosine 3′,5′-Monophosphate or a Cyclic Nucleotide Phosphodiesterase Inhibitor*

Abstract

The actions of cyclic nucleotides on basal and somatomedin-stimulated thymidine incorporation into DNA by costal cartilage from hypophysectomized rats were investigated. Three analogs of cAMP (dibutyryl, 8-bromo, and 8-dimethylamino derivatives, which are alternate activators of cAMP-dependent protein kinase and resistant to degradation by cAMP phosphodiesterase but represent a wide difference in potency as phosphodiesterase inhibitors) in range of concentrations from about 10(-5) to 3 X 10(-4) M enhanced basal and somatomedin-stimulated thymidine incorporation. Each cAMP analog at optimal concentration produced combined effects with a suboptimal concentration of somatomedin which were additive or greater. cAMP itself, 5'-AMP, adenosine, 8-Br-5'-AMP, 8-Br-AMPT, and cGMP at concentrations from 10(-7)--10(-3) M or dibutyryl cGMP at concentrations from 10(-10)--10(-3) M did not reproduce the effects of the cAMP analogs. A phosphodiesterase inhibitor (1-methyl-3-isobutylxanthine) at concentrations of 100 or 500 microM also potentiated the effects of somatomedin. At 100- or 500-microM concentrations, the phosphodiesterase inhibitor increased cartilage levels of cAMP and cGMP. These results suggest a role for cAMP in DNA synthesis in rat cartilage. However, they fail to support the hypothesis that all effects of somatomedin on that process are mediated by cAMP, since stimulation of thymidine incorporation by the hormone can be demonstrated in cartilage maximally stimulated by analogs of cAMP.

Fibroblast growth factor stimulates colony formation of differentiated chondrocytes in soft agar

“…Specificity of growth factor stimulation of chondrocyte colony formation Previous studies have shown that in addition to FGF, other agents, including EGF, insulin, IGF, and dibutyryl cyclic-AMP, stimulate the growth of chondrocytes in monolayer and organ cultures (Bomboy and Salmon, 1980;Prins et al, 1982;Kato et al, 1983). Table 1 shows the effect of these and other mitogens on growth in soft agar of chick embryo chondrocytes.…”

Potent mitogenic effects of parathyroid hormone (PTH) on embryonic chick and rabbit chondrocytes. Differential effects of age on growth, proteoglycan, and cyclic AMP responses of chondrocytes to PTH.

“…This finding indicates that PTH stimulation of chondrocyte phenotypic expression is not linked to its growth stimulation, and that the mitogenic action of PTH is not mediated by cAMP. The effects of cAMP analogue mimicked those of PTH in chondrocytes (5,8,9,29,30), but dibutyryl cAMP enhanced chondrocyte growth (8,9,30) (32). Furthermore, PTH is found in fetal blood of various animals, and its level (10-'1-10-'0 M) is similar to the maternal level (monkey [33]; miniature pig [34]; rabbit [Kato, Y., and T. Koike, unpublished observations]).…”

Effect of multiplication-stimulating activity (MSA) on the cyclic AMP level and proteoglycan synthesis in cultured chondrocytes

“…These findings suggest that with respect to cyclic AMP metabolism, the re¬ sponse of chondrocytes to MSA differs from that of fibroblasts. Recently, Bomboy & Salmon (1980) showed that analogues of cyclic AMP and cyclic nucleotide phosphodiesterase inhibitor markedly potentiate the effect of somatomedin on DNA synthesis in cultured cartilage from hypophysectomized rats. Analogues of cyclic AMP stimulate DNA synthesis in chondrocytes in organ culture (Bomboy & Sal¬ mon 1980), but they suppress DNA synthesis in monolayer cultures (Miller et al 1979;Takigawa et al 1981).…”

“…These findings suggest that the de¬ crease in cyclic AMP level may be related to the action of somatomedin on macromolecule synthe¬ sis. Cyclic AMP analogues have been shown to be potent stimulators of proteoglycan synthesis (Miller et al 1979;Takigawa et al 1981), RNA synthesis (Drezner & Uebovitz 1979) and DNA synthesis (Bomboy & Salmon 1980) in chondrocytes. Taki¬ gawa et al (1981) showed that parathyroid hor¬ mone (PTH) increases ornithine decarboxylase activity and proteoglycan through increase of cyclic AMP.…”