Differentiation of single lymphoma primary cells and normal B-cells based on their adhesion to mesenchymal stromal cells in optical tweezers

Abstract

We have adapted a non-invasive method based on optical tweezers technology to differentiate between the normal B-cells and the B-cell non-Hodgkin lymphoma (B-NHL) cells derived from clinical samples. Our approach bases on the nascent adhesion between an individual B-cell and a mesenchymal stromal cell. In this study, a single B-cell was trapped and optically seeded on a mesenchymal stromal cell and kept in a direct contact with it until a stable connection between the cells was formed in time scale. This approach allowed us to avoid the introduction of any exogenous beads or chemicals into the experimental setup which would have affected the cell-to-cell adhesion. Here, we have provided new evidence that aberrant adhesive properties found in transformed B-cells are related to malignant neoplasia. We have demonstrated that the mean time required for establishing adhesive interactions between an individual normal B-cell and a mesenchymal stromal cell was 26.7 ± 16.6 s, while for lymphoma cell it was 208.8 ± 102.3 s, p < 0.001. The contact time for adhesion to occur ranged from 5 to 90 s and from 60 to 480 s for normal B-cells and lymphoma cells, respectively. This method for optically controlled cell-to-cell adhesion in time scale is beneficial to the successful differentiation of pathological cells from normal B-cells within the fine needle aspiration biopsy of a clinical sample. Additionally, variations in time-dependent adhesion among subtypes of B-NHL, established here by the optical trapping, confirm earlier results pertaining to cell heterogeneity.

Development and Characterization of 3D Hybrid Spheroids for the Investigation of the Crosstalk Between B-Cell Non-Hodgkin Lymphomas and Mesenchymal Stromal Cells

“… 56 Several reports showed that stromal cells chemoprotect tumor cells through direct cell–cell contact; thus, there is an urgent need to develop models which allow studying such direct interactions. 19 Importantly, our group previously investigated the direct interactions between B-cell lymphoma and stromal cells in optical tweezers using a 2D culture, 57 , 58 whereas the 3D organization and cellular microenvironment emerged as critical determinants of lymphoma pathogenesis and drug resistance.…”

Formation of Lymphoma Hybrid Spheroids and Drug Testing in Real Time with the Use of Fluorescence Optical Tweezers

“…This limitation was overcome by the transition to infrared lasers, which enabled longer manipulation times of living cells, resulting in the broader use of optical tweezers in cell research [43]. In our previous works, we investigated the effect of a low-intensity optical laser on living lymphoma cells [7,8,22]. We proved that laser operation at 1064 nm poses a low risk of optical damage to lymphoma cells.…”

“…The precise measurements of the adhesion force, as well as the influence of the laser beam used on living cell viability, have been described in our earlier papers [7,8]. All experiments were carried out at room temperature, i.e., 25 • C. The spheroids formed in the OT were then transferred to an incubator and cultured in a hanging drop in RPMI-1640 (Gibco, Paisley, UK) with 10% bovine serum (Gibco, Paisley, UK) and antibiotics (Sigma-Aldrich Chemie GmbH) under standard conditions.…”

“…Furthermore, the complicated molecular biology of MSCs and their correspondingly complex dynamics remain primarily unknown. In our previous work, we studied via optical tweezers (OT) the formation of nascent adhesion between MSCs and B-NHL lymphomas using a broad range of lymphoma cell lines [7], as well as patient-derived primary cells [8]. We found that the adhesive properties are cell-line specific, and it is possible to distinguish lymphoma from normal B-cells based on the adhesion to the stromal cell.…”

“…A single B-NHL lymphoma cell was trapped, optically seeded on a 3D stromal spheroid, and kept in direct contact until a stable connection was formed. The attachment time of a single cell was estimated as previously described [7,8,21]. The procedure was repeated until the entire surface of the stromal spheroid was covered with lymphoma cells.…”