Hypergravity-induced changes in actin response of breast cancer cells to natural killer cells

Abstract

Although immunotherapy holds promising cytotoxic activity against lymphoma or leukemia, the immunosuppressive mechanisms of solid tumors remain challenging. In this study, we developed and applied a hypergravity exposure system as a novel strategy to improve the responsiveness of breast cancer cells to natural killer (NK) cells for efficient immunotherapy. Following exposure to hypergravity, either in the presence or absence of NK cells, we investigated for changes in the cell cytoskeletal structure, which is related to the F-actin mediated immune evasion mechanism (referred to as “actin response”) of cancer cells. Breast cancer cell line MDA-MB-231 cells were exposed thrice to a 20 min hypergravitational condition (10 × g), with a 20 min rest period between each exposure. The applied hypergravity induces changes in the intracellular cytoskeleton structure without decreasing the cell viability but increasing the cytotoxicity of MDA-MB-231 from 4 to 18% (4.5-fold) at a 3:1 ratio (NK-to-target). Analyses related to F-actin further demonstrate that the applied hypergravity results in rearrangement of the cytoskeleton, leading to inhibition of the actin response of MDA-MB-231. Taken together, our results suggest that the mechanical load increases through application of hypergravity, which potentially improves efficiency of cell-based immunotherapies by sensitizing tumors to immune cell-mediated cytotoxicity.

Increased H3K9me3 and F-Actin Reorganization in the Rapid Adaptive Response to Hypergravity in Human T Lymphocytes

“…This is consistent with the downregulation of the actin-binding gene LOC643980 in hypergravity samples [21] and in the studies showing heightened sensitivity to immune cell-mediated cytotoxicity after F-actin inhibition [63]. A separate study found similar results in cells exposed to hypergravity, where a decrease in the F-actin to G-actin ratio was observed [63]. Additionally, another study found a concentrated distribution of G-actin around the nucleus in rat cells following a parabolic flight [64], possibly contributing to the mitigation of nuclear actin and the structural maintenance of the nucleus [65].…”

“…Furthermore, we observed a rapid decrease in F-actin fluorescence within 20 s of hypergravity exposure, indicating filament depolymerization and cytoskeletal restructuring. This is consistent with the downregulation of the actin-binding gene LOC643980 in hypergravity samples [21] and in the studies showing heightened sensitivity to immune cell-mediated cytotoxicity after F-actin inhibition [63]. A separate study found similar results in cells exposed to hypergravity, where a decrease in the F-actin to G-actin ratio was observed [63].…”

Enhanced cytotoxic activity of natural killer cells from increased calcium influx induced by electrical stimulation

“…For the lactate dehydrogenase (LDH) cytotoxicity assay, the medium was collected from each well after KHYG-1 cell treatment at an effector-to-target (E:T) ratio of 3:1 (24 h) and centrifuged at 600 g for 5 min. NK cell-mediated cytotoxicity was measured according to previously described protocols [ 20 ] and calculated according to the manufacturer’s instructions ( Fig 1B ).…”