Remanent Cell Traction Force in Renal Vascular Smooth Muscle Cells Induced by Integrin-Mediated Mechanotransduction

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myogenic response, paramagnetic bead, cell traction force microscopy, calcium signaling

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It was previously demonstrated in isolated renal vascular smooth muscle cells (VSMCs) that integrin-mediated mechanotransduction triggers intracellular Ca2+ mobilization, which is the hallmark of myogenic response in VSMCs. To test directly whether integrin-mediated mechanotransduction results in the myogenic response-like behavior in renal VSMCs, cell traction force microscopy was used to monitor cell traction force when the cells were pulled with fibronectin-coated or low density lipoprotein (LDL)-coated paramagnetic beads. LDL-coated beads were used as a control for nonintegrin-mediated mechanotransduction. Pulling with LDL-coated beads increased the cell traction force by 61 ± 12% (9 cells), which returned to the prepull level after the pulling process was terminated. Pulling with noncoated beads had a minimal increase in the cell traction force (12 ± 9%, 8 cells). Pulling with fibronectin-coated beads increased the cell traction force by 56 ± 20% (7 cells). However, the cell traction force was still elevated by 23 ± 14% after the pulling process was terminated. This behavior is analogous to the changes of vascular resistance in pressure-induced myogenic response, in which vascular resistance remains elevated after myogenic constriction. Fibronectin is a native ligand for α5β1-integrins in VSMCs. Similar remanent cell traction force was found when cells were pulled with beads coated with β1-integrin antibody (Ha2/5). Activation of β1-integrin with soluble antibody also triggered variations of cell traction force and Ca2+ mobilization, which were abolished by the Src inhibitor. In conclusion, mechanical force transduced by α5β1-integrins triggered a myogenic response-like behavior in isolated renal VSMCs.

myogenic vasoconstriction is an autoregulatory mechanism for adjusting local vascular resistance to the increase in arterial pressure, which fluctuates over a wide range under different physiological and pathophysiological conditions (27, 43). In the renal circulation, pressure-induced myogenic constriction in the afferent arteriole is an important protective mechanism to prevent the transmission of elevated systemic pressure to the glomerular capillaries, a critical determinant in the progression of glomerulosclerosis in diabetes, hypertension, and end-stage renal disease. However, the mechanisms that transduce the changes in perfusion pressure into vascular smooth muscle cells (VSMCs) to initiate myogenic constriction are still elusive (7, 15, 17, 28). Many studies on mechanotransduction of myogenic response have focused on the mechanosensitive gating of stretch-sensitive channels in triggering membrane depolarization and Ca2+ influx. However, stretch-sensitive ion channels cannot provide a sustained error signal to maintain the myogenic constriction when the stretch vanishes as the vessel contracts (9, 10, 42). In contrast, integrins are known to transduce extracellular mechanical force into intracellular biochemical signals to affect VSMC contractility (1, 20, 21, 32). We have previously shown that application of mechanical force to renal VSMCs with paramagnetic beads coated with integrin ligands activates subcellular Ca2+ release from ryanodine receptors in the form of Ca2+ sparks (1). The coupling of Ca2+ sparks to the Ca2+-activated Cl− channel may account for the membrane depolarization in VSMCs required for contraction (22). However, it has not been determined whether the integrin-mediated mechanotransduction is sufficient to elicit VSMC contraction for the myogenic response. Cells are anchored to the substratum via focal adhesions, which are composed of cytoskeleton, integrins ,and extracellular matrix. VSMCs exert cell traction force through focal adhesions (34). In the present study, cell traction-force microscopy was used to monitor the changes in cell traction exerted by renal VSMCs on a flexible substrate to examine integrin-mediated contraction at the single cell level.

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American Journal of Physiology-Cell Physiology, v. 304, issue 4, p. C382-C391