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Renal cytosolic extracts from rats of different ages and mononephrectomized rats were incubated with gamma-[32P]ATP and analysed by high resolution two-dimensional electrophoresis and autoradiography. Extracts from new-born and young rats showed a great number of phosphorylated proteins migrating between the origin and Mr 52,000. Among these proteins, the group co-migrating with phosphorylase b (Mr 97,000) was particularly evident in new-born and days-old rats. In extracts from mature rats, other proteins of lower molecular weight, particularly those migrating between Mr 60,000 and 44,000, became intensely phosphorylated. The number and intensity of phosphorylated proteins from extracts of normal and nephrectomized rats, however, did not vary. Activity of cAMP-dependent protein kinase and [3H]cAMP binding was also modified during neonatal development but not in compensatory renal growth. Since cAMP-PK and protein phosphorylation are known to be regulated in response to hormonal stimulations, these results may provide good indications for the understanding of hormonal involvement in kidney growth.

Aqueous extracts from rabbit colon, kidney, testis and small intestinal mucosa were prepared by homogenization and centrifugation at 105,000 g. After precipitation with ammonium sulphate. the 0–50 fraction (F1) and the supernatant (F2) were collected, dialysed against a phosphate buffer and tested on mice in vivo. 1 hr after a single injection of F1 (15 mg content) from colon, the uptake of tritiated thymidine was decreased in jejunal and colonic DNA in mice. This effect, maximal after 3 hr and totally reversible after 7 hr, was found in neither the kidney nor the testis. the F1 fractions of non-digestive organs (kidney, testis) were also found to exert a significant inhibition on thymidine incorporation into intestinal DNA in vivo. F1 fractions of intestinal contents, prepared under the same conditions, exerted no significant effects on DNA synthesis in mouse intestine. Conversely, the colon F2 fraction did not inhibit the synthesis of jejunal and colonic DNA in vivo. A slowing of cellular migration was also noticed in the jejunum and colon of mice injected with colon or small intestine F1, as ascertained radioautographically by determining the position of the leading edge of the labelled cells in jejunal or colonic F1-injected mice. Our results suggest that the F1 fraction of the aqueous extract of rabbit colon contains one or more substances, which may act either on intestinal DNA synthesis or on the G1-S transition of the cellular cycle in the mouse intestine. This reversible and tissue-specific intestinal action appears to inhibit cell proliferation and presents several of the characteristics defining a chalone, as does the action of small intestinal F1 previously reported (Sassier & Bergeron, 1977). However, because of a relative lack of origin specificity of this effect, the physiological significance of our data remains to be ascertained.

An aqueous extract was prepared from the mucosa of rabbit small intestine by homogenization and centrifugation at 105,000 g. After precipitation with ammonium sulfate, the 0–50 fraction (F1) and the supernatant (F2) were collected, dialysed against a phosphate buffer and tested on rats in vitro and mice in vivo. The F1 fraction was found to inhibit thymidine incorporation into rat intestinal DNA in vitro, but this effect was not found to be tissue specific (liver, kidney). Two hours after a single injection of F1 (10 mg protein content), the uptake of tritiated thymidine was decreased in jejunal and colonic DNA in mice. This effect was maximal between 2 and 4 hr and totally reversible after 7 hr; this effect was found in neither the kidney nor the testis. A slowing of cellular migration was also noticed in the jejunum and the colon. Conversely, the F2 fraction did not inhibit the synthesis of jejunal and colonic DNA either in vitro or in vivo. Our results suggest that the F1 fraction of the aqueous extract of rabbit small intestine contains one or more substances which may act either on intestinal DNA synthesis or on the G1–S transition of the cellular cycle in the mouse intestine. This reversible and specific intestinal action appears to inhibit cell proliferation and presents several of the characteristics defining a chalone.

Aqueous extracts from rabbit organs were prepared by homogenization and centrifugation at 105,000 g. After precipitation with ammonium sulphate, the 0-50 fraction was separated by ultrafiltration through Amicon XM 100 and XM 300 membranes yielding two filtrate fractions (U1 and U2) and one retentate fraction (U3). Only U1 and U3 inhibited thymidine incorporation into DNA. After a single injection of U1 from rabbit small intestine, the uptake of tritiated thymidine was decreased in mouse jejunal and colonic DNA. This effect, totally reversible after 7 hr, was found in neither the kidney nor the testis. The U1 fractions of colon and non-digestive organs (kidney, testis) were found not to exert a significant inhibition on thymidine incorporation into intestinal DNA in vivo. The U3 fraction from rabbit small intestine also decreased the uptake of tritiated thymidine in mouse jejunal and colonic DNA in vivo. However, this inhibition was irreversible and not tissue-specific. Slowing of cell migration was also noticed in the jejunum of mice injected with U1 or U3, as ascertained radioautographically by determining the position of the leading edge of the labelled cells in U1- or U3-injected mice compared with controls. A decrease of mitotic activity in U1- and U3-injected mice was recorded 8.5 hr after a single injection of small intestinal fractions. Our results suggest that U1 and U3 from rabbit small intestine contain one or more substances which may act on the G1-S transition of the cell cycle in the mouse intestine. However, only the effect of U1 is reversible and tissue specific. Our data suggest the existence of a factor, having a low molecular weight, which regulates intestinal cell proliferation.