We combine a micro-fluidic electric-field cell-culture (MEC) chip with structured-illumination nano-profilometry (SINAP) to quantitatively study the variations of cancer cell filopodia under external direct-current electric field (dcEF) stimulations. Because the lateral resolution of SINAP is better than 150 nm in bright-field image modality, filopodia with diameters smaller than 200 nm can be observed clearly without fluorescent labeling. In the MEC chip, a homogeneous EF is generated inside the culture area that simulates the endogenous EF environment. With this MEC chip-SINAP system, we directly observe and quantify the biased growth of filopodia of lung cancer cells toward the cathode. The epidermal growth factor receptors around the cell edges are also redistributed to the cathodal side. These results suggest that cancer-cell filopodia respond to the changes in EFs in the microenvironment.

We employed direct-current electric fields (dcEFs) to modulate the chemotaxis of lung cancer cells in a microfluidic cell culture device that incorporates both stable concentration gradients and dcEFs. We found that the chemotaxis induced by a 0.5 lM/mm concentration gradient of epidermal growth factor can be nearly compensated by a 360 mV/mm dcEF. When the effect of chemical stimulation was balanced by the electrical drive, the cells migrated randomly, and the path lengths were largely reduced. We also demonstrated electrically modulated chemotaxis of two types of lung cancer cells with opposite directions of electrotaxis in this device. V C 2014 AIP Publishing LLC. [http://dx

Microenvironment stiffening plays a crucial role in tumorigenesis. While filopodia are generally thought to be one of the cellular mechanosensors for probing environmental stiffness, the effects of environmental stiffness on filopodial activities of cancer cells remain unclear. In this work, we investigated the filopodial activities of human lung adenocarcinoma cells CL1-5 cultured on substrates of tunable stiffness using a novel platform. The platform consists of an optical system called structured illumination nano-profilometry, which allows time-lapsed visualization of filopodial activities without fluorescence labeling. The culturing substrates were composed of polyvinyl chloride mixed with an environmentally friendly plasticizer to yield Young's modulus ranging from 20 to 60 kPa. Cell viability studies showed that the viability of cells cultured on the substrates was similar to those cultured on commonly used elastomers such as polydimethylsiloxane. Time-lapsed live cell images were acquired and the filopodial activities in response to substrates with varying degrees of stiffness were analyzed. Statistical analyses revealed that lung cancer cells cultured on softer substrates appeared to have longer filopodia, higher filopodial densities with respect to the cellular perimeter, and slower filopodial retraction rates. Nonetheless, the temporal analysis of filopodial activities revealed that whether a filopodium decides to extend or retract is purely a stochastic process without dependency on substrate stiffness. The discrepancy of the filopodial activities between lung cancer cells cultured on substrates with different degrees of stiffness vanished when the myosin II activities were inhibited by treating the cells with blebbistatin, which suggests that the filopodial activities are closely modulated by the adhesion strength of the cells. Our data quantitatively relate filopodial activities of lung cancer cells with environmental stiffness and should shed light on the understanding and treatment of cancer progression and metastasis.