Actin Filament Morphology in Living and Nonliving Cultured Mesangial Cells: Formation and Dissolution

Abstract

Mesangial cells are contractile and are believed to play a role in the regulation of glomerular filtration. Actin filaments constitute an integral part of the cytoskeleton of these cells. To visualize actin filaments in both living and fixed acetone-extracted (nonliving) mesangial cells, we have used the fluorophore nitrobenzoxadiazole-phallacidin. Under the fluorescence microscope, mesangial cells displayed continuous staining. Distinct linear structures running from one end to the other were actin filaments or stress fibers (SF). SF were rather feathery and of slightly curvilinear appearance in living versus fixed, acetone-extracted cells. Formation of SF started a 4 h, and the majority of the cells had well developed SF within 24 h. To find out whether adhesion to a substrate was necessary to SF formation, we plated mesangial cells on variable thickness of poly-HEMA (2-hydroxyethyl methacrylate). This plastic surface at a high concentration (10^-1M) produces poor adherence of mesangial cells. At 10^-1M of poly-HEMA, the cells were rounded even after 48 h and did not develop any SF. In contrast, at a lower concentration (10^-4M), mesangial cells were well spread, and SF were readily observed. These data indicated that spreading as well as formation of SF were directly related to the adhesion properties of the substrate. To evaluate the role of SF in contraction and relaxation of mesangial cells, we have first treated these cells with angiotensin II (5 × 10^-7M) or dibutyryl cyclic adenosine monophosphate (5 × 10^-4M) and then labeled them with nitrobenzoxadiazole phallacidin. Dibutyryl cyclic adenosine monophosphate caused dissolution of SF. These were well preserved or rather brighter in angiotensin II treated cells. It appears that SF played an important role in maintaining basal tonic contractility in cultured mesangial cells.