Bi-functional action of transforming growth factor-beta on DNA synthesis in early passage human fetal fibroblasts.

We investigated the influence of transforming growth factor-beta (TGF-beta) on DNA synthesis in human fetal fibroblasts, as measured by the incorporation of [3H]thymidine and cell replication. In serum-free medium, without additional peptide growth factors, TGF-beta had no action on thymidine incorporation. However, in the presence of 0.1% v/v fetal calf serum, TGF-beta exhibited a bi-functional action on the cells. A dose-dependent stimulation of [3H]thymidine incorporation, and an increase in cell number, occurred with fibroblasts established from fetuses under 50 g body weight, with a maximum stimulation seen at 1.25 ng/ml. For fibroblasts from fetuses of 100 g or greater body weight, TGF-beta caused a dose-related decrease in thymidine uptake with a maximal inhibition at 2.5 ng/ml, and a small decrease in cell number. When DNA synthesis was stimulated by the addition of somatomedin-C/insulin-like growth factor I, epidermal growth factor, or platelet-derived growth factor, their actions were potentiated by the presence of TGF-beta on cells derived from fetuses under 50 g body weight, but inhibited on cells obtained from the larger fetuses weighing more than 100 g. Similar results were found for changes in cell number in response to TGF-beta when stimulated by SM-C/IGF I. The ability of TGF-beta to modulate [3H] thymidine incorporation did not involve a change in the time required for growth-restricted cells to enter the S phase of the replication cycle. These data suggest that TGF-beta may exert either a growth-promoting or growth-inhibiting action on human fetal connective tissues in the presence of other peptide growth factors, which is dependent on fetal age and development.

Bovine retinal angiogenesis factor is a small molecule (molecular mass less than 600).

Two methods were used to extract angiogenic activity from bovine retina. Both methods initially gave rise to nondialyzable (greater than 10,000 Mr) fractions with angiogenic activity. However, after anion exchange chromatography, 20% of the extracts from one of the two methods (method 2) contained a small molecule with angiogenic activity (Mr 300-600). Alcohol treatment of high molecular mass fractions from both methods also released a low molecular mass angiogenic factor (Mr 300-600). No angiogenic activity was left in the nondialyzable residue. The high molecular mass angiogenic fraction obtained by method 1 after DEAE-cellulose chromatography contained a protein immunologically and electrophoretically identical to bovine serum albumin. The low molecular mass retinal angiogenic factor was able to stimulate microvessel endothelial cell proliferation as well as being positive in the chick chorioallantoic membrane test. The presence of a protein carrier system for a small angiogenesis factor is proposed. This would explain discrepancies in the apparent molecular mass of retinal angiogenic factors described previously.

Potentiating effect of heparin in the activation of procollagenase by a low-Mr angiogenesis factor.

A low-Mr freely dialysable endothelial cell-stimulating angiogenesis factor (ESAF) from conditioned medium of a mouse lymphoma cell line has previously been shown to activate latent skin fibroblast procollagenase. Activation comparable with the maximum that can be achieved with trypsin is obtained with chemically undetectable amounts of the factor. We now show that when even smaller amounts of ESAF are used heparin is able to potentiate its action in this system. The relationship between this activity and the mechanism of angiogenesis, which is itself potentiated by heparin, is discussed.

Extraction, isolation and characterization of neutral salt soluble type V collagen from fetal calf skin.

Collagen was extracted with neutral salt solution and examined for the presence of type V collagen. A fraction which was insoluble in both 0.02 M Na2HPO4, pH 9.2 and phosphate-buffered saline (PBS) pH 7.2 at 4 degrees C contained both alpha 1(V)- and alpha 2(V)-chains demonstrated by sodium dodecyl sulfate polyacrylamide gel electrophoresis, diethylaminoethyl cellulose ion exchange chromatography, amino acid analysis and segment long spacing (SLS) crystallites. SLS crystallites showed a globular N-terminal extension peptide attached to the type V collagen monomer. Ion exchange chromatography also demonstrated the presence of a third, minor component, which was identified as the alpha 3(V)-chain. In certain extractions, components corresponding to the partially processed procollagen chains of type V collagen were also observed.