COMPARTMENTALIZED COCULTURE OF PORCINE ARTERIAL ENDOTHELIAL AND SMOOTH MUSCLE CELLS ON A MICROPOROUS MEMBRANE

摘要

Endothelial and smooth muscle cells were harvested from porcine pulmonary arteries and grown to two passages from primary culture in serum-containing medium. Thereafter, the cells were plated on the opposite sides of a microporous poly(ethylene terephthalate) membrane and cultivated in a chemically defined, serum-free medium. The membrane with pores of 1 mu m diameter allowed the passage of molecules and the extension of cell processes, while maintaining separate homogeneous cell populations. Pores of 3 mu m diameter permitted the crossing of smooth muscle cells through the membrane, The coating of the polymer with constituents of the extracellular matrix optimized cell adhesion, Morphological analysis of the model showed typical cobblestone pattern and ultrastructure of endothelial cells, which lost rapidly the expression of von Willebrand factor hut kept that of angiotensin-converting enzyme. Smooth muscle cells were spindle shaped and specific alpha-actin was revealed by immunochemistry and quantitated by enzyme-linked immunosorbent assay (ELISA). Their ultrastructure featured an intermediate contractile-synthetic phenotype. Permeability studies to different molecules showed a marked reduction of the albumin clearance. Finally, in coculture in the presence of endothelial cells, the smooth muscle cells proliferation was increased, whereas it was not the case in autologous cocultures. In conclusion, such a coculture model may help to a better understanding of the interactions between endothelial and smooth muscle cells that may be important in the pathogenesis of vascular diseases.