Evaluation of pancreatic cancer cell migration with multiple parameters in vitro by using an optical real-time cell mobility assay device

Abstract

BackgroundMigration of cancer cell correlates with distant metastasis and local invasion, which are good targets for cancer treatment. An optically accessible device “TAXIScan” was developed, which provides considerably more information regarding the cellular dynamics and less quantity of samples than do the existing methods. Here, we report the establishment of a system to analyze the nature of pancreatic cancer cells using TAXIScan and we evaluated lysophosphatidic acid (LPA)-elicited pancreatic cell migration.MethodsPancreatic cancer cell lines, BxPC3, PANC-1, AsPC1, and MIAPaCa-2, were analyzed for adhesion as well as migration towards LPA by TAXIScan using parameters such as velocity and directionality or for the number of migrated cells by the Boyden chamber methods. To confirm that the migration was initiated by LPA, the expression of LPA receptors and activation of intracellular signal transductions were examined by quantitative reverse transcriptase polymerase reaction and western blotting.ResultsScaffold coating was necessary for the adhesion of pancreatic cancer cells, and collagen I and Matrigel were found to be good scaffolds. BxPC3 and PANC-1 cells clearly migrated towards the concentration gradient formed by injecting 1 μL LPA, which was abrogated by pre-treatment with LPA inhibitor, Ki16425 (IC50 for the directionality ≈ 1.86 μM). The LPA dependent migration was further confirmed by mRNA and protein expression of LPA receptors as well as phosphorylation of signaling molecules. LPA1 mRNA was highest among the 6 receptors, and LPA1, LPA2 and LPA3 proteins were detected in BxPC3 and PANC-1 cells. Phosphorylation of Akt (Thr308 and Ser473) and p42/44MAPK in BxPC3 and PANC-1 cells was observed after LPA stimulation, which was clearly inhibited by pre-treatment with a compound Ki16425.ConclusionsWe established a novel pancreatic cancer cell migration assay system using TAXIScan. This assay device provides multiple information on migrating cells simultaneously, such as their morphology, directionality, and velocity, with a small volume of sample and can be a powerful tool for analyzing the nature of cancer cells and for identifying new factors that affect cell functions.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-017-3218-4) contains supplementary material, which is available to authorized users.

Reduction of Real-Time Imaging of M1 Macrophage Chemotaxis toward Damaged Muscle Cells is PI3K-Dependent

“…TAXIScan is an assay device for studying cell dynamics in vitro, which functions by providing two-dimensional images of cell migration. Accordingly, this device can provide markedly more information, including morphology as well as quantitative analysis, compared to existing methods such as the Boyden chamber method [ 13 , 24 , 25 , 26 ]. Interestingly, the directionality and velocity of chemotactic cells can be provided by analyzing the cell images.…”

Newly developed anti‐S100A8/A9 monoclonal antibody efficiently prevents lung tropic cancer metastasis

“…To precisely assess the 5 antibodies in vitro , we used a TAXIScan device (Fig. e ) that provides two‐dimensional images of cell migration over time for the cell tracing period and also provides additional information about velocity as well as directionality of a ligand in a quantitative analysis . Using this advanced approach, we found that Ab45 had the highest activity for the inhibition of S100A8/A9‐stimulated chemotaxis of B16‐BL6 cells, which consistently appeared in both velocity and directionality (Fig.…”

“…A cell migration assay was performed using the EZ‐TAXIScan device (ECI, Kawasaki, Japan) as described previously . Briefly, cancer cells were maintained with appropriate media and were detached on the day of the experiment with Trypsin/EDTA.…”

“…For analysis of the time‐lapse images, “TAXIScan analyzer 2” software that was included with the device was used. The velocity and the directionality of each migrating cell were calculated using the traced data as described previously . Statistical analysis for the velocity and directionality was done by Student's t‐test.…”

Metastasis in Pancreatic Ductal Adenocarcinoma: Current Standing and Methodologies

“…Despite providing several advantages over in vivo methods (including the capacity to fine-tune experimental parameters, and being relatively inexpensive and easy to use), these methods represent endpoint studies, are limited in their capacity to study multicellular interactions, and do not provide information beyond the number of migrated cells [160,162]. Optical mobility assay devices such as the TaxiScan, on the other hand, can be used to obtain additional information on migrating cells, including morphology, directionality and velocity [163]. Using these methods, several studies have shown that only a small fraction of pancreatic cancer cells are capable of invading the ECM [164][165][166][167][168][169].…”