Investigating contactless high frequency ultrasound microbeam stimulation for determination of invasion potential of breast cancer cells

Abstract

In this article, we investigate the application of contactless high frequency ultrasound microbeam stimulation (HFUMS) for determining the invasion potential of breast cancer cells. In breast cancer patients, the finding of tumor metastasis significantly worsens the clinical prognosis. Thus, early determination of the potential of a tumor for invasion and metastasis would significantly impact decisions about aggressiveness of cancer treatment. Recent work suggests that invasive breast cancer cells (MDA-MB-231), but not weakly invasive breast cancer cells (MCF-7, SKBR3, and BT-474), display a number of neuronal characteristics, including expression of voltage-gated sodium channels. Since sodium channels are often co-expressed with calcium channels, this prompted us to test whether single-cell stimulation by a highly focused ultrasound microbeam would trigger Ca2+ elevation, especially in highly invasive breast cancer cells. To calibrate the diameter of the microbeam ultrasound produced by a 200-MHz single element LiNbO3 transducer, we focused the beam on a wire target and performed a pulse-echo test. The width of the beam was ~17 μm, appropriate for single cell stimulation. Membrane-permeant fluorescent Ca2+ indicators were utilized to monitor Ca2+ changes in the cells due to HFUMS. The cell response index (CRI), which is a composite parameter reflecting both Ca2+ elevation and the fraction of responding cells elicited by HFUMS, was much greater in highly invasive breast cancer cells than in the weakly invasive breast cancer cells. The CRI of MDA-MB-231 cells depended on peak-to-peak amplitude of the voltage driving the transducer. These results suggest that HFUMS may serve as a novel tool to determine the invasion potential of breast cancer cells, and with further refinement may offer a rapid test for invasiveness of tumor biopsies in situ.

Non-contact High-Frequency Ultrasound Microbeam Stimulation for Studying Mechanotransduction in Human Umbilical Vein Endothelial Cells

“…Furthermore, the low-frequency ultrasound affected nuclear processes of the cells including the expression of early response gens c-Fos, c-Jun, and c-Myc while the magnitude of effect depended on the ultrasound frequency (Louw et al 2013). In contrast, we recently demonstrated that high-frequency ultrasound microbeam was better suited for more localized single-cell stimulation (Hwang et al 2012; Hwang et al 2013) than the low-frequency ultrasound. Particularly, high-frequency ultrasound microbeam stimulation (HFUMS) allowed identifying functional characteristics of cells and also discriminating cell types (Hwang et al 2012; Hwang et al 2013).…”

“…In contrast, we recently demonstrated that high-frequency ultrasound microbeam was better suited for more localized single-cell stimulation (Hwang et al 2012; Hwang et al 2013) than the low-frequency ultrasound. Particularly, high-frequency ultrasound microbeam stimulation (HFUMS) allowed identifying functional characteristics of cells and also discriminating cell types (Hwang et al 2012; Hwang et al 2013). In these studies, it was found that HFUMS resulted in more reduction of mitochondrial membrane potentials in breast cancer cells than in normal cells, thus showing its capability to discriminate between the functional characteristics of the cells (Hwang et al 2012; Hwang et al 2013).…”

“…Particularly, high-frequency ultrasound microbeam stimulation (HFUMS) allowed identifying functional characteristics of cells and also discriminating cell types (Hwang et al 2012; Hwang et al 2013). In these studies, it was found that HFUMS resulted in more reduction of mitochondrial membrane potentials in breast cancer cells than in normal cells, thus showing its capability to discriminate between the functional characteristics of the cells (Hwang et al 2012; Hwang et al 2013). In addition, it was observed that the HFUMS elicited different levels of cytoplasmic calcium (Ca 2+ ) elevations in highly and weakly invasive breast cancer cells (Hwang et al 2012; Hwang et al 2013).…”

Acoustic tweezers for studying intracellular calcium signaling in SKBR-3 human breast cancer cells

“…In those studies, the calcium response of the cells to high-frequency ultrasonic stimulation was dependent on the applied acoustic pressure. Therefore, it was shown that both umbilical vein endothelial cells and highly-invasive cancer responded less actively at the lower acoustic excitation (input voltage = ~8 V pp and ~9.5 V pp ) [24]. Similarly, the calcium response of SKBR-3 cells is here little affected by non-FNT-coated microbead attachment under acoustic excitation at 6.3 V pp (Fig.…”

“…3, it is shown that attachment of the acoustically trapped FNT-coated microbead to the SKBR-3 cells clearly induces significant intracellular calcium elevations compared to attachment of the trapped non-FNT microbead to the cells. Our previous studies have shown that 200 MHz ultrasound microbeams themselves were able to promote intracellular calcium elevations in highly-invasive breast cancer cells (MDA-MB-231), but not in weakly-invasive breast cancer cells such as SKBR-3 cells [24]. In addition, such a high-frequency ultrasonic stimulation resulted in considerable cytoplasmic calcium elevations in human umbilical vein endothelial cells [25].…”