* P<0.01 and ** P<0.001 as determined by Student t-test. Cell viability was determined by Celltiter Fluor assay. (E) Patient samples were cultured at 37☌ at atm and 310 mmHg above atm for three days, and then treated with 250 nM daunorubicin for three days at the same pressure levels. Cell viability was determined by Celltiter Fluor assay and IC 50 were calculated. HL60 and TEX cells were cultured at 37☌ at atm and 310 mmHg above atm for three days, and then treated with increasing concentrations of idarubicin (D) and mitoxantrone (E) at the same pressures for another three days.
* P<0.05 and ** P<0.0001 as determined by one-way ANOVA with Bonferroni post-test.
HL60 and TEX cells were cultured at 37☌ at atm and 77 mmHg and 310 mmHg above atm for three days, and then treated with increasing concentrations of daunorubicin at indicated pressures for another three days. Cell viability measured by trypan blue exclusion assay of HL60 and TEX leukemia cells cultured over 4 days at 37☌ at atmospheric pressure (atm) and 310 mmHg above atm. Acute myeloid leukemia (AML) cells at increased pressure display chemoresistance to danorubicin. Furthermore, the effects of increased pressure on chemosensitivity were reversible.įigure 1. 4 3 HL60 and TEX cells grown at increased static pressure were more resistant to daunorubicin, ( Figure 1B), but not idarubicin ( Figure 1C) or mitoxantrone ( Figure 1D and Online Supplementary Figure S2). Next, we explored the impact of increased pressure on the sensitivity of these cells to three anthracyclines used in the treatment of AML: daunorubicin, idarubicin, and mitoxantrone. 2 1 We found that the growth and viability of these cell lines did not change at high pressure ( Figure 1A). A pressure of 310 mmHg above atm corresponds to less than 0.5% of the maximum pressure acting on the head of the femur of an upright 160 lb person, and therefore, would be physiologically relevant as erosion of the trabeculae from leukemic infiltration increases sensitivity to external forces. Two AML cell lines, HL60 and TEX, were cultured at pressures up to 310 mmHg above atmospheric pressure (atm). We designed and manufactured pressure chambers to deliver different levels of physiological pressure to AML cells while maintaining continuous gas exchange ( Online Supplementary Figure S1). Therefore, biomechanical stimulus such as pressure is another environmental factor that can influence chemosensitivity. Using AML cell lines and primary patient samples, we show that high pressure promotes a transition to a gel-like plasma membrane that reduces the intracellular accumulation of daunorubicin in AML cells. Therefore, we sought to establish whether the pressure in the marrow is another environmental factor that can influence chemosensitivity. The pressure in the marrow of AML patients can be 10–20 fold higher compared to patients with solid tumors or non-malignant conditions, 1 possibly due to increased cellularity and fibrosis.
Although environmental factors such as cytokines and stroma are recognized to influence the growth and chemosensitivity of acute myeloid leukemia (AML) cells, little is known about the effect of physical forces such as pressure.
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