Lung epithelial cells are subjected to large cyclic forces from breathing. However, their response to dynamic stresses is poorly defined. We measured the complex shear modulus (G(*)(omega)) of human alveolar (A549) and bronchial (BEAS-2B) epithelial cells over three frequency decades (0.1-100 Hz) and at different loading forces (0.1-0.9 nN) with atomic force microscopy. G(*)(omega) was computed by correcting force-indentation oscillatory data for the tip-cell contact geometry and for the hydrodynamic viscous drag. Both cell types displayed similar viscoelastic properties. The storage modulus G'(omega) increased with frequency following a power law with exponent approximately 0.2. The loss modulus G"(omega) was approximately 2/3 lower and increased similarly to G'(omega) up to approximately 10 Hz, but exhibited a steeper rise at higher frequencies. The cells showed a weak force dependence of G'(omega) and G"(omega). G(*)(omega) conformed to the power-law model with a structural damping coefficient of approximately 0.3, indicating a coupling of elastic and dissipative processes within the cell. Power-law behavior implies a continuum distribution of stress relaxation time constants. This complex dynamics is consistent with the rheology of soft glassy materials close to a glass transition, thereby suggesting that structural disorder and metastability may be fundamental features of cell architecture.

Alveolar epithelial cells undergo stretching during breathing and mechanical ventilation. Stretch can modify cell viscoelastic properties, which may compromise the balance of forces in the alveolar epithelium. We studied the viscoelasticity of alveolar epithelial cells (A549) subjected to equibiaxial distention with a novel experimental approach. Cells were cultured on flexible substrates and subjected to stepwise deformations of up to 17% with a device built on an inverted microscope. Simultaneously, cell storage (G') and loss (G'') moduli were measured (0.1-100 Hz) with optical magnetic twisting cytometry. G' and G'' increased with strain up to 64 and 30%, respectively, resulting in a decrease in G''/G' (15%). This stretch-induced response was inhibited by disruption of the actin cytoskeleton with latrunculin A. G' increased with frequency following a power law with exponent alpha = 0.197. G'' increased proportionally to G' but exhibited a more marked frequency dependence at high frequencies. Stretching (14%) caused a fall in alpha (13%). At high stretching amplitudes, actual cell strain (14.4%) was lower than the applied substrate strain (17.3%), which could indicate a partial cell detachment. These data suggest that cytoskeletal prestress modulates the elastic and frictional properties of alveolar epithelial cells in a coupled manner, according to soft glassy rheology. Stretch-induced cell stiffening could compromise the balance of forces at the cell-cell and cell-matrix adhesions.

. Measurement of cell microrheology by magnetic twisting cytometry with frequency domain demodulation. J Appl Physiol 91: [1152][1153][1154][1155][1156][1157][1158][1159] 2001.-Magnetic twisting cytometry (MTC) (Wang N, Butler JP, and Ingber DE, Science 260: 1124-1127, 1993) is a useful technique for probing cell micromechanics. The technique is based on twisting ligand-coated magnetic microbeads bound to membrane receptors and measuring the resulting bead rotation with a magnetometer. Owing to the low signal-to-noise ratio, however, the magnetic signal must be modulated, which is accomplished by spinning the sample at ϳ10 Hz. Present demodulation approaches limit the MTC range to frequencies Ͻ0.5 Hz. We propose a novel demodulation algorithm to expand the frequency range of MTC measurements to higher frequencies. The algorithm is based on coherent demodulation in the frequency domain, and its frequency range is limited only by the dynamic response of the magnetometer. Using the new algorithm, we measured the complex modulus of elasticity (G*) of cultured human bronchial epithelial cells (BEAS-2B) from 0.03 to 16 Hz. Cells were cultured in supplemented RPMI medium, and ferromagnetic beads (ϳ5 m) coated with an RGD peptide were bound to the cell membrane. Both the storage (GЈ, real part of G*) and loss (GЉ, imaginary part of G*) moduli increased with frequency as ␣ (2 ϫ frequency) with ␣ Ϸ 1 ⁄4. The ratio GЉ/GЈ was ϳ0.5 and varied little with frequency. Thus the cells exhibited a predominantly elastic behavior with a weak power law of frequency and a nearly constant proportion of elastic vs. frictional stresses, implying that the mechanical behavior conformed to the so-called structural damping (or constant-phase) law (Maksym GN, Fabry B, Butler JP, Navajas D, Tschumperlin DJ, LaPorte JD, and Fredberg JJ, J Appl Physiol 89: [1619][1620][1621][1622][1623][1624][1625][1626][1627][1628][1629][1630][1631][1632] 2000). We conclude that frequency domain demodulation dramatically increases the frequency range that can be probed with MTC and reveals that the mechanics of these cells conforms to constant-phase behavior over a range of frequencies approaching three decades. cell mechanics; cell viscoelasticity; complex elastic modulus; power law rheology; structural damping; magnetic tweezers MECHANICAL PROPERTIES OF THE CELL play an important role in essential cellular functions such as mechanotransduction, shape stability, motility, apoptosis and DNA synthesis (12-14, 19, 23). Techniques for studying cell mechanics include cell poking (6), atomic force microscopy (18), optical tweezers (28), laser tracking microrheology (27), magnetic bead microrheometry (2), and magnetic twisting cytometry (MTC) (23). MTC, in particular, has proved to be a useful tool for exploring force transmission across the cell membrane and for assessing cell stiffness and its changes (11,15,23,25,26). This technique was first introduced by Crick (4) and Crick and Hughes (5) and was further refined by Valberg (21), Wang et al. in 1993 (23), and, m...