Gene expression of human lung cancer cell line CL1-5 in response to a direct current electric field

Abstract

Background: Electrotaxis is the movement of adherent living cells in response to a direct current (dc) electric field (EF) of physiological strength. Highly metastatic human lung cancer cells, CL1-5, exhibit directional migration and orientation under dcEFs. To understand the transcriptional response of CL1-5 cells to a dcEF, microarray analysis was performed in this study.

Methodology/principal findings: A large electric-field chip (LEFC) was designed, fabricated, and used in this study. CL1-5 cells were treated with the EF strength of 0 mV/mm (the control group) and 300 mV/mm (the EF-treated group) for two hours. Signaling pathways involving the genes that expressed differently between the two groups were revealed. It was shown that the EF-regulated genes highly correlated to adherens junction, telomerase RNA component gene regulation, and tight junction. Some up-regulated genes such as ACVR1B and CTTN, and some down-regulated genes such as PTEN, are known to be positively and negatively correlated to cell migration, respectively. The protein-protein interactions of adherens junction-associated EF-regulated genes suggested that platelet-derived growth factor (PDGF) receptors and ephrin receptors may participate in sensing extracellular electrical stimuli. We further observed a high percentage of significantly regulated genes which encode cell membrane proteins, suggesting that dcEF may directly influence the activity of cell membrane proteins in signal transduction.

Conclusions/significance: In this study, some of the EF-regulated genes have been reported to be essential whereas others are novel for electrotaxis. Our result confirms that the regulation of gene expression is involved in the mechanism of electrotactic response.

Figure 1. Assembly drawing of the large electric-field chip (LEFC).

The LEFC had connecting holes for the medium inlet/outlet and the agar salt bridges. Cells were cultured in the micro-chamber (the cell culture region). The width, length, and thickness of the micro-chamber were 24mm, 75mm, and 70 µm, respectively.

Figure 2. Lateral view of the electrotaxis system.

A LEFC was integrated with a transparent ITO heater chip, two Ag/AgCl electrodes with phosphate-buffered saline (PBS) as electrolyte, two agar salt bridges (1.5% agar in PBS), a syringe pump, a DC power supply, an ampere-meter, and an inverted microscope.

Figure 3. Simulated EF in the cell culture region of the LEFC.

The EF strength along the dotted line (between the two arrows) was shown. More than 85% culture area was exposed to the EF strength of 300+/−15mV/mm.

Figure 4. Biological processes correlated with the EF-regulated genes.

The significantly regulated genes listed in Table 2 (up-regulated) and Table 3 (down-regulated) were categorized with their biological function according to the Gene Ontology annotation.

Figure 5. Protein-protein interaction between the products of the EF-regulated genes associated with adherens junction.

The diagram showed that the EF up-regulated Fyn, ACP1, and c-Src might directly interact with two kinds of membrane receptors, PDGF receptors and ephrin receptors. Green arrow: positive effect; Red arrow: negative effect; gray arrow: unspecified effect.