A New Insight on the Radioprotective Potential of Epsilon-Aminocaproic Acid

Abstract

Background and objectives: The aim of the study was to scrutinize the ability of epsilon-aminocaproic acid (EACA) to prevent radiation-induced damage to human cells. Materials and Methods: Human peripheral blood mononuclear cells (PBMCs) were exposed to ionizing radiation at three low doses (22.62 mGy, 45.27 mGy, and 67.88 mGy) in the presence of EACA at the concentration of 50 ng/mL. Results: EACA was able to prevent cell death induced by low-dose X-ray radiation and suppress the formation of reactive oxygen species (ROS). EACA also demonstrated a capacity to protect DNA from radiation-induced damage. The data indicated that EACA is capable of suppression of radiation-induced apoptosis. Comparative tests of antioxidative activity of EACA and a range of free radical scavengers showed an ability of EACA to effectively inhibit the generation of ROS. Conclusions: This study showed that the pretreatment of PBMCs with EACA is able to protect the cells from radiation-elicited damage, including free radicals' formation, DNA damage, and apoptosis.

Keywords: DNA damage; apoptosis; epsilon-aminocaproic acid; protection; radiation; reactive oxygen species.

Figure 1

Cytotoxic effect of various doses of epsilon-aminocaproic acid (EACA) on cells viability presented as a normalized ratio to the control. Error bars in the graphs indicate the standard error of the mean (SEM) for n = 3 independent experiments. **—p ≤ 0.01 compared to the control.

Figure 2

Effect of epsilon-aminocaproic acid (EACA) on cell viability after X-ray exposure (presented as %). The measurements have been done 12 h after exposure to radiation. Normal peripheral blood mononuclear cells (PBMCs) were subjected to three radiation doses: 22.62 mGy, 45.27 mGy, and 67.88 mGy in the presence of EACA (50 ng/mL). Error bars in the graphs indicate the standard error of the mean (SEM) for n = 3 independent experiments.

Figure 3

Impact of epsilon-aminocaproic acid (EACA) on production of reactive oxygen species (ROS) presented as a normalized ratio to the control. The measurements were done immediately after exposure to X-ray radiation. Normal peripheral blood mononuclear cells (PBMCs) were subjected to three radiation doses: 22.62 mGy, 45.27 mGy, and 67.88 mGy in the presence or absence of EACA (50 ng/mL). Error bars in the graphs indicate the standard error of the mean (SEM) for n = 3 independent experiments. **—p ≤ 0.01 compared to the control.

Figure 4

Effect of epsilon-aminocaproic acid (EACA) on DNA damage induced by radiation (%). The measurements were done immediately after exposure to X-ray radiation. Normal peripheral blood mononuclear cells (PBMCs) were subjected to three radiation doses: 22.62 mGy, 45.27 mGy, and 67.88 mGy in the presence or absence of EACA (50 ng/mL). DNA damage is presented as a percentage (%). Error bars in the graphs indicate the standard error of the mean (SEM) for n = 3 independent experiments. **—p ≤ 0.01 compared to the control; *—p ≤ 0.05 compared to the control.

Figure 5

Images of DNA comets of PBMCs subjected to X-ray radiation in the presence or absence of EACA (50 ng/mL). The DNA-comet assay was performed immediately after exposure to radiation. (A) DNA comets of control group; (B) DNA comets of cells subjected to hydrogen peroxide (positive control); (C) DNA comets of cells subjected to hydrogen peroxide and EACA; (D) DNA comets of cells exposed to 22.62 mGy X-ray; (E) DNA comets of cells exposed to 22.62 mGy X-ray and EACA; (F) DNA comets of cells exposed to 45.27 mGy X-ray; (G) DNA comets of cells exposed to 45.27 mGy X-ray and EACA; (H) DNA comets of cells exposed to 67.88 mGy X-ray; (I) DNA comets of cells exposed to 67.88 mGy X-ray and EACA.

Figure 6

Effect of epsilon-aminocaproic acid (EACA) on the prevention of radiation-induced apoptosis (%). The measurements were done 6 h after exposure to X-ray radiation. Normal peripheral blood mononuclear cells (PBMCs) were subjected to three radiation doses: 22.62 mGy, 45.27 mGy, and 67.88 mGy in the presence or absence of EACA (50 ng/mL). The number of apoptotic cells (Annexin V labeling) is presented as a percentage (%). Error bars in the graphs indicate the standard error of the mean (SEM) for n = 3 independent experiments. **—p ≤ 0.01 compared to the control.

Figure 7

Microscopic images of apoptotic PBMCs (600×). The measurements were done 6 h after exposure to radiation. (A) control group; (B) cells subjected to hydrogen peroxide (positive control); (C) cells subjected to hydrogen peroxide and EACA; (D) cells exposed to 22.62 mGy X-ray; (E) cells exposed to 22.62 mGy X-ray and EACA; (F) cells exposed to 45.27 mGy X-ray; (G) cells exposed to 45.27 mGy X-ray and EACA; (H) cells exposed to 67.88 mGy X-ray; (I) cells exposed to 67.88 mGy X-ray and EACA. Staining by Annexin V (green color) indicates early apoptosis stage; double staining by Propidium Iodide (red) and Annexin V (green) reflects the late apoptosis phase.

Figure 8

Comparison of anti-ROS activity of EACA (50 ng/mL) and free radical scavengers in the presence of hydrogen peroxide (concentration of 1.0 mM). ROS production is presented as a ratio normalized to the control group. Abbreviations: epsilon-aminocaproic acid (EACA), N-acetyl-cysteine (NAC), caffeic acid, p-Coumaric acid, gallic acid, quercetin, ascorbic acid, lipoic acid, polydatin, and Vit E (tocopherol). Error bars in the graphs indicate the standard error of the mean (SEM) for n = 3 independent experiments. **—p ≤ 0.01 compared to the control. In addition, statistical comparison between the hydrogen peroxide group and free radical scavengers’ groups have been done.