Studies on the Protective Effect of Silybin Against Low-Dose Radiation-Induced Damage to the Immune System

Abstract

With growing public concern about the health effects of low-dose radiation, numerous studies have demonstrated that low-dose radiation can cause damage to the immune system, making intervention measures essential. This study investigated the protective effects of silybin against low-dose radiation-induced immune system damage and its underlying mechanisms at both the cellular and animal levels. At the cellular level, CCK-8 assays, ROS measurements, and RT-qPCR analysis revealed that silybin alleviated the reduction in RAW264.7 cell proliferation, intracellular ROS levels, and inflammatory cytokine expression following low-dose radiation exposure. At the animal level, comparative analyses of post-irradiation body weight, peripheral blood cell counts, immune organ coefficients, spleen HE/IHC staining, and spleen immune cell numbers demonstrated that silybin mitigated the radiation-induced decrease in body weight, reduction in peripheral blood leukocyte counts, inflammatory cell infiltration in the spleen, decline in spleen immune cell numbers, and increase in cGAS protein-positive cells. These findings indicate that silybin exerts protective effects against low-dose radiation-induced immune system damage, potentially by regulating the cGAS signaling pathway to reduce radiation-induced cellular injury, thereby enhancing its radioprotective properties.

Keywords: immune damage; low-dose radiation; protection; silybin.

Figure 1

Proliferative viability of RAW264.7 cells after intervention with three natural products. (A–C) no irradiation, samples collected 48 h after intervention; (D) irradiation alone; (E–G) irradiation at 2 Gy, samples obtained 48 h post-irradiation; (H–J) irradiation at 0.2 Gy, samples acquired 48 h post-irradiation. The three natural products are silybin, pachymic acid A, and perilla proanthocyanidins. Data are presented as mean ± SEM. * p < 0.05; ** p < 0.01; *** p < 0.001; ## p < 0.01; ### p < 0.001; ns p > 0.05.

Figure 2

Evaluation of the antioxidant activity of natural products. (A,B) Three natural products—silybin, pachymic acid A, and perilla proanthocyanidins—and Vc (vitamin C) as positive controls; in vitro assay of the free radical scavenging ability of the natural products against DPPH and ABTS free radicals; (C,D) silybin and Vc as positive controls; in vitro assay of the DPPH and ABTS free radical scavenging ability of silybin; (E) flow assay of the effect of silybin on the ROS expression level in RAW264.7 cells 8 h after irradiation. Data are presented as mean ± SEM. * p < 0.05; ## p < 0.01.

Figure 3

mRNA expression of inflammatory factors in RAW264.7 cells under irradiation conditions. (A–I) Irradiation alone: samples were collected 24 and 48 h after 4 Gy, 2 Gy, and 0.2 Gy irradiation, and the IL-6, IL-1β, and TNF-α mRNA levels in RAW264.7 cells were measured. (J–L) IL-6, IL-1β, and TNF-α mRNA levels in samples collected 24 h after 0.2 Gy irradiation combined with silybin intervention. Data are presented as mean ± SEM. ## p < 0.01; ### p < 0.001; * p < 0.05; ** p < 0.01; *** p < 0.001; ns p > 0.05.

Figure 4

Changes in mouse body weight under different irradiation conditions caused by silybin: (A,B) single 2 Gy irradiation; (C,D) single 0.2 Gy irradiation; (E,F) multiple low-dose irradiation of 0.01 Gy × 20 times. Panels (A,C,E) show the mouse body weights before and after 24 h of irradiation, and panels (B,D,F) show the mouse body weights after 24 h of irradiation. For n ≥ 3, Data are presented as mean ± SEM. ## p < 0.01; ### p < 0.001; * p < 0.05; ** p < 0.01; *** p < 0.001; ns p > 0.05.

Figure 5

Changes in the blood profile of mice under different irradiation conditions caused by silybin: (A–D) single 2 Gy irradiation; (E–H) single 0.2 Gy irradiation; (I–L) multiple low-dose irradiation of 0.01 Gy × 20 times. n ≥ 3. Data are presented as mean ± SEM. # p < 0.05; ## p < 0.01; ### p < 0.001; * p < 0.05; ** p < 0.01; *** p < 0.001; ns p > 0.05.

Figure 6

Changes in the organ coefficients of immune organs in mice under different irradiation conditions in response to silybin. (A) Flowchart of the experimental design for mice receiving tissues (liver, spleen, and thymus); (B–D) single 2 Gy irradiation; (E–G) single 0.2 Gy exposure; (H–J) multiple low-dose exposures of 0.01 Gy × 20 times. n ≥ 3. Data are presented as mean ± SEM. ## p < 0.01; ### p < 0.001; ns p > 0.05.

Figure 7

Silybin effects on mouse spleen HE staining under varied irradiation conditions: (A) 2 Gy; (B) 0.2 Gy, silybin intervention concentration 5/10/20 mg/kg; (C) 0.01 Gy × 20 times, silybin intervention concentration 5/10/20 mg/kg. The red arrows indicate inflammatory cell infiltration. Magnification 40×, scale 50 μm and 20 μm.

Figure 8

Alterations in the number of splenic immune cells in mice subjected to distinct irradiation conditions caused by silybin: (A,B,G,H) 2 Gy; (C,D,I,J) 0.2 Gy; (E,F,K,L) 0.01 Gy × 20 times. n ≥ 3. Data are presented as mean ± SEM. # p < 0.05; ## p < 0.01; ### p < 0.001; * p < 0.05; ** p < 0.01; *** p < 0.001; ns p > 0.05.

Figure 9

Immunohistochemistry of splenic cGAS proteins in mice subjected to different irradiation conditions via silybin: (A,D) 2 Gy; (B,E) 0.2 Gy; (C,F) 0.01 Gy × 20 times. Magnification 40×, scale 50 μm and 20 μm. n ≥ 3. Data are presented as mean ± SEM. ## p < 0.01; ### p < 0.001; * p < 0.05; *** p < 0.001.