Continuous wave irradiation at 0.1 terahertz facilitates transmembrane transport of small molecules

Abstract

The bioeffects of terahertz (THz) radiation received growing attention because of its influence on the interactions between biomolecules. Our work aimed to investigate the effects of THz irradiation on cell membrane, especially cell membrane permeability. We found that 0.1 THz irradiation promoted the endocytosis of FM4-64-labeled cells and the inhibition of dynamin attenuated but did not fully abolish the THz promoted endocytosis. Moreover, 0.1 THz irradiation also promoted the transmembrane of the rhodamine, as well as the chemical compounds GDC0941 and H89, evidenced by the confocal microscope observation and the western blotting analysis, respectively. These findings demonstrated 0.1 THz irradiation facilitated the transmembrane transport of small molecules by promoting both the cellular endocytosis and the diffusion process. Our study provided direct evidence that THz could affect the cell membrane permeability, broadened the THz affected cellular physiological processes, and implied its potential application in regulating the cell membrane functions.

Keywords: Membranes; Radiation; Transport phenomena.

Graphical abstract

Figure 1

Experimental setup of THz exposure THz radiation was emitted and directly reached the irradiated samples in an incubator at 37°C. Cells were cultured in a transparent plastic plate and were exposed to the radiation in the period of experiments.

Figure 2

THz radiation enhanced endocytosis in neuronal cells HT22 cells were stained with 10 μM FM4-64 and immediately exposed to 0.1 THz radiation at 33 mW/cm² for 5 min. The cells were then imaged using a laser scanning confocal microscope. Scale bar, 20 μm. The results are shown as Mean ± SD Statistical analyses were performed using two-tailed Student’|'s t test (∗∗∗p < 0.001).

Figure 3

Irradiation-induced endocytosis was independent of dynamin HT22 cells were treated with 10 μM Dynasore or left untreated for 30 min, and subsequently stained with 10 μM FM4-64 and immediately exposed to 0.1 THz irradiation at 33 mW/cm² for another 5 min. The cells were then imaged using a confocal laser scanning microscope. Scale bar, 20 μm. The results are shown as means ± SD Statistical analyses were performed using ANOVA followed by Tukey’|'s post hoc test (∗∗p < 0.01, ∗∗∗p < 0.001 vs. Ctrl group; ##p < 0.01 vs. Dynasore alone group).

Figure 4

THz irradiation increased the diffusion of rhodamine into cells (A) Exposure to THz irradiation increased the diffusion of rhodamine (Rh) B into the cells. HT22 cells were treated with 5 μM RhB and immediately exposed to 0.1 THz irradiation at 33 mW/cm² for 5 min. The cells were then imaged using a laser scanning confocal microscope. (B) Exposure to THz irradiation increased the diffusion of the rhodamine derivative Rh123 into the cells. HT22 cells were treated with 5 μM Rh123 and immediately exposed to 0.1 THz irradiation at 33 mW/cm² for 5 min. The cells were then imaged using a confocal laser scanning microscope. Scale bar, 20 μm. The results are shown as means ± SD Statistical analyses were performed using two-tailed Student’|'s t test (∗∗∗p < 0.001).

Figure 5

Irradiation-enhanced transport of RhB into the cells was not mediated by endocytosis HT22 cells were treated with 10 μM Dynasore or left untreated for 30 min, and subsequently stained with 5 μM of RhB and immediately exposed to 0.1 THz irradiation at 33 mW/cm² for another 5 min. The cells were then imaged using a confocal laser scanning microscope. Scale bar, 20 μm. The results are shown as means ± SD Statistical analyses were performed using two-tailed Student’|'s t test (∗∗∗p < 0.001).

Figure 6

THz irradiation promoted the diffusion of the kinase inhibitors GDC0941 and H89 into neuronal cells (A) Phosphorylated AKT (p-AKT) decreased upon THz exposure in the presence of GDC-0941. HT22 cells were treated with 100 nM GDC-0941 or left untreated for 10 min and subsequently exposed to 0.1 THz irradiation at 7.8 or 33 mW/cm² for another 20 min. Then, 1 μM of insulin was added to induce an increase in p-AKT level immediately after THz irradiation. P-AKT and AKT protein levels were analyzed by western blotting. (B) THz irradiation did not affect p-AKT in the absence of GDC-0941. HT22 cells exposed to 0.1 THz irradiation at 7.8 or 33 mW/cm² for 20 min were analyzed by western blotting. (C) Phosphorylated CREB (p-CREB) decreased upon THz exposure in the presence of H89. HT22 cells were treated with 10 μM H89 or left untreated for 10 min and subsequently exposed to 0.1 THz irradiation at 7.8 or 33 mW/cm² for another 20 min. Then 10 μM forskolin was added to induce an increase of the p-CREB level immediately after THz irradiation. p-CREB and CREB protein levels were analyzed by western blotting. (D) THz irradiation did not affect p-CREB in the absence of H89. HT22 cells exposed to 0.1 THz irradiation at 7.8 or 33 mW/cm² for 20 min were analyzed by western blotting. The results are shown as means ± SD Statistical analyses were performed using ANOVA followed by Tukey’|'s post hoc test (∗∗p < 0.01, ∗∗∗p < 0.001).