Astroglial-induced in vitro angiogenesis: requirements for RNA and protein synthesis

Astrocytes are believed to affect microvascular endothelial cell differentiation in brain and retina. Bovine retinal microvessel endothelial cells formed capillary-like structures when cocultured with C6 astroglial cells or in the absence of C6 cells in response to the reconstituted basement membrane protein Matrigel. Using quantitative computer-assisted image analysis, the requirements for RNA and protein synthesis in these two complementary models of in vitro microvessel morphogenesis were examined. Astroglial-dependent capillary-like structure formation was inhibited by up to 87% in a dose-dependent fashion by cycloheximide (0.01-0.1 micrograms/ml), puromycin (0.1-0.25 micrograms/ml), and actinomycin D (0.01-0.025 micrograms/ml). In contrast, the astroglial-independent process in response to Matrigel was not affected by these metabolic inhibitors. These findings suggest that capillary-like structures form in response to astroglial cells in two distinct sequential stages. The first consists of inductive astroglial-endothelial interactions requiring both RNA and protein synthesis. This initiates endogenous endothelial morphogenic events that do not appear to require RNA or protein synthesis, consistent with posttranslational regulatory mechanisms. The first astroglial-dependent step is relevant to the regulation of microvessel formation in brain and retina, whereas the second may represent a morphogenic pathway common to microvessel formation in many tissues.