VALIDATION OF COLLAGENOUS PROTEIN-SYNTHESIS AS AN INDEX FOR ANGIOGENESIS WITH THE USE OF MORPHOLOGICAL METHODS

A method providing a biochemical index for the evaluation of promoters or inhibitors of angiogenesis in the chick chorioallantoic membrane ( CAM) is here described and validated. This method is based on the dete rmination of collagenous protein synthesis which takes place during ne w vessel formation. Validation was done by comparing collagenous prote in synthesis to morphological methods of determining vascular density either by counting the number of vessels intersecting three concentric rings or by computer-assisted image analysis. Five compounds which pr omote or inhibit angiogenesis in the CAM were used for this purpose. T he protein kinase C activator 4-beta-phorbol 12-myristate 13-acetate a nd alpha-thrombin increased collagenous protein synthesis and these re sults correlated with those obtained by using the two morphological me thods mentioned above. Similarly, the inhibitors of angiogenesis, Ro31 8220, tricyclodecan-9-yl xanthate (D609), and 8,9-dihydroxy-7-methyl-b enzo[b]quinolizinium bromide (GPA1734), reduced collagenous protein sy nthesis and vascular density (determined by image analysis or by count ing the number of vessels intersecting three concentric rings) to a co mparable degree. These results indicate that collagenous protein synth esis can be used as a reliable, reproducible, and unbiased index of an gogenesis in the chick chorioallantoic membrane. (C) 1995 Academic Pre ss, Inc.