Effect of paclitaxel treatment on cellular mechanics and morphology of human oesophageal squamous cell carcinoma in 2D and 3D environments

Abstract

During chemotherapy, structural and mechanical changes in malignant cells have been observed in several cancers, including leukaemia, pancreatic and prostate cancer. Such cellular changes may act as physical biomarkers for chemoresistance and cancer recurrence. This study aimed to determine how exposure to paclitaxel affects the intracellular stiffness of human oesophageal cancer of South African origin in vitro. A human oesophageal squamous cell carcinoma cell line WHCO1 was cultured on glass substrates (2D) and in collagen gels (3D) and exposed to paclitaxel for up to 48 hours. Cellular morphology and stiffness were assessed with confocal microscopy, visually aided morpho-phenotyping image recognition, and mitochondrial particle tracking microrheology at 24 and 48 hours. In the 2D environment, the intracellular stiffness was higher for the paclitaxel-treated than for untreated cells at 24 and 48 hours. In the 3D environment, the paclitaxel-treated cells were stiffer than the untreated cells at 24 hours, but no statistically significant differences in stiffness were observed at 48 hours. In 2D, paclitaxel-treated cells were significantly larger at 24 and 48 hours and more circular at 24 but not at 48 hours than the untreated controls. In 3D, there were no significant morphological differences between treated and untreated cells. The shape heterogeneity was lower for paclitaxel-treated than untreated cells in 3D and similar for the treated and untreated groups in 2D. Future studies with patient-derived primary cancer cells and prolonged drug exposure will help identify physical cellular biomarkers to detect chemoresistance onset and assess therapy effectiveness in oesophageal cancer patients.InsightMechanical changes in cancer cells brought on by chemotherapeutic drug exposure offer possible physical biomarkers to detect chemoresistance and cancer recurrence. This first study on human oesophageal squamous cell carcinoma of South African origin indicated that in vitro paclitaxel treatment induced stiffening and enlarging of the malignant cells in two-dimensional environments at 24 and 48 hours. In physiologically more relevant three-dimensional collagen matrices, however, the paclitaxel treatment led to cellular stiffening at 24 hours but softening after that, without significant changes in cellular size at any time. The in vitro environment’s influence on experimental outcomes needs to be considered in further studies with prolonged drug exposure and patient-derived primary cancer cells.

Advances in mechanical biomarkers

“…Interestingly, cell mechanics is also affected by many of the cancer drugs in the market, specifically targeting elements of the cytoskeleton and membrane of cells. Examples of these drugs are Doxorubicin (membrane peroxidation), 81,148 Taxol (tubulin depolymerization suppression), 149,150 Dexamethasone (actin filaments reorganization), 151,152 and Vincristine or Vinblastine (microtubules polymerization suppression) 83,153 . However, they present important secondary effects like cardiotoxicity, thus novel drugs targeting the cytoskeletal structures are being studied for clinical use.…”

“…Studies demonstrated that filamentous structures can change their mechanical properties in fast biochemical processes like phosphorylation, ion concentration, and associated protein interactions. 41,42,46 depolymerization suppression), 149,150 Dexamethasone (actin filaments reorganization), 151,152 and Vincristine or Vinblastine (microtubules polymerization suppression). 83,153 However, they present important secondary effects like cardiotoxicity, thus novel drugs targeting the cytoskeletal structures are being studied for clinical use.…”

Stem cell mechanoadaptation - Part A - Effect of microtubule stabilization and volume changing stresses on cytoskeletal remodeling

“…The use of small molecules targeting cytoskeleton de-/polymerization has proven useful for elucidating the role of the cytoskeleton in the regulation of cell shape and mechanical properties underpinning cell lineage specification 1 . Paclitaxel (PAX) inhibits tubulin depolymerization, thus stabilizing microtubules, increasing microtubule bundling, and leading to an increase in cell stiffness 6 . F-actin and tubulin act in concert and exhibit compensatory effects in the cell; hence, inhibition of microtubule depolymerization with PAX is accompanied by significant changes in Factin, resulting in dose-dependent decreases in MSC migratory capacity and differentiation potential 7 .…”