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Cellular traction forces were measured by using a microfabricated substrate, particularly exploring how cytoskeletal structures such as actin filaments and microtubules contribute to traction forces. Smooth muscle cells isolated from bovine aortas were cultured and transfected with fluorescence proteins to visualize cell microstructures and then plated on a micropatterned elastomer substrate with arrays of micropillars. Cell spreading on the substrates produced deflection of micropillars which was used for estimation of cellular traction forces, and was closely associated with organization of stress fibers of actin filaments. Traction forces varied considerably among cells, showing the order of several 10 nN. After disruption of microtubules with nocodazole, traction forces significantly increased and there was no detectable change in formation of stress fibers. To inhibit the ROCK pathway, a signaling pathway of myosin light chain phosphorylation, possibly being induced by disruption of microtubules, significantly depressed the increase in traction forces after the disruption of microtubules. This result indicates that microtubules disassembly may regulate the actomyosin-based contractile system mainly through the ROCK pathway. The present study suggests that formation of stress fibers are mainly involved in cellular traction forces and a contribution of microtubules should include not only a force balance but also rather a modulator of the actomyosin contractile system in actin stress fibers.

Biomechanical Contribution of Cytoskeletal Structures to Traction Forces in Cultured Smooth Muscle Cells