Characterizing the Motility of Chemotherapeutics-Treated Acute Lymphoblastic Leukemia Cells by Time-Lapse Imaging

Abstract

Drug resistance is an obstacle in the therapy of acute lymphoblastic leukemia (ALL). Whether the physical properties such as the motility of the cells contribute to the survival of ALL cells after drug treatment has recently been of increasing interest, as they could potentially allow the metastasis of solid tumor cells and the migration of leukemia cells. We hypothesized that chemotherapeutic treatment may alter these physical cellular properties. To investigate the motility of chemotherapeutics-treated B-cell ALL (B-ALL) cells, patient-derived B-ALL cells were treated with chemotherapy for 7 days and left for 12 h without chemotherapeutic treatment. Two parameters of motility were studied, velocity and migration distance, using a time-lapse imaging system. The study revealed that compared to non-chemotherapeutically treated B-ALL cells, B-ALL cells that survived chemotherapy treatment after 7 days showed reduced motility. We had previously shown that Tysabri and P5G10, antibodies against the adhesion molecules integrins α4 and α6, respectively, may overcome drug resistance mediated through leukemia cell adhesion to bone marrow stromal cells. Therefore, we tested the effect of integrin α4 or α6 blockade on the motility of chemotherapeutics-treated ALL cells. Only integrin α4 blockade decreased the motility and velocity of two chemotherapeutics-treated ALL cell lines. Interestingly, integrin α6 blockade did not affect the velocity of chemoresistant ALL cells. This study explores the physical properties of the movements of chemoresistant B-ALL cells and highlights a potential link to integrins. Further studies to investigate the underlying mechanism are warranted.

Keywords: acute lymphoblastic leukemia; cell motility; chemotherapeutics-treated; integrins; time-lapse imaging.

Figure 1

The schematic of the whole system framework for time-lapse cinematography is shown. The system consists of a gas/air route indicated with blue arrows, a temperature feedback route indicated with brown arrows, a temperature control route indicated with green arrows, and a light route indicated with red arrows. An 8-well chamber plated with leukemia cells and human stromal cells was used to monitor the motility of primary ALL cells.

Figure 2

An example of LAX7R co-cultured with HS27a human stromal cells monitored by time-lapse microscopy to display the motility tracks of viable primary B-ALL cells in control medium and treated with chemotherapy. (a,b) illustrate a case of a LAX7R cell migration pattern (white lines) in medium control and with VDL chemotherapeutical treatment for 7 days. The time-lapse image reveals that the migration pattern is tangled at the start point of the migration and displays a weak motility as the cells were treated with VDL (red-dashed circles). The scale bars in (a,b) are 50 nm. (c) A proposed vector plot provides a visualization to simultaneously observe cell motility and migration patterns in both medium and VDL. The arc (red arrows) and radial (blue arrow) indicate a cell’s migration steps and travel distance from its start point. In the study, the 48 steps (12 h recording) were considered in both groups. The travel distance to 90 indicates 26.1 µm as the actual distance. (d) The viability of the medium control and VDL-treated cells on Day 7 was measured by 7-AAD and Annexin V-PE staining using flow cytometry. *** p < 0.001 compared with the medium group, unpaired t-test. The letter M stands for medium.

Figure 3

Effect of chemotherapeutic treatment of primary ALL cells cocultured with human stromal cells on velocity and migratory distance. Velocities of (a) LAX7R, (c) LAX56, and (e) ICN24 cells treated with medium or VDL. Cells were co-cultured with HS27a human stromal cells (+HS27a) or not (-HS27a). The migration distance from the origins of the (b) LAX7R, (d) LAX56, and (f) ICN24 cells that were or were not cocultured with HS27a cells was also measured. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 when compared with their corresponding medium controls (unpaired t-test). NS, not significant.

Figure 4

The effects of integrin blockade on the velocity and migration distance of primary B-ALL cells. B-ALL cells LAX7R (a–c), LAX56 (d–f), and ICN24 (g–i) were treated with VDL (VDL) or without VDL (Medium) along with P5G10 or Tysabri for 7 days. After a washing and medium incubation period of 12 h, ALL cells were time-lapse imaged for 12 h (a,d,g). Cell viability was measured by 7-AAD/Annexin V staining using flow cytometry (b,e,h). The velocities and (c,f,i) migratory distances of the three ALL cells are depicted. The differences were analyzed by one-way ANOVA with Tukey’s test. * p < 0.05 and *** p < 0.001. NS, not significant. The pooled data of three experiments are shown.