Extracellular and intracellular inflammatory response of immune cells to radiation exposure
Authors: Didier Kaylin , Sanchez Sophie , Shahi Ankita , Capitini Christian , Schrage William 
Published: May 2024

Background: Exposure to low doses of ionizing radiation is associated with increased risk of cardiovascular disease (CVD) in a dose-dependent manner. A person exposed to low doses of radiation have a 27-29% risk of developing and dying from CVD. Low doses of radiation initiate an inflammatory process that can increase the risk of developing CVD. Inflammation initiates atherosclerosis via immune cell infiltration subsequent to endothelial cell damage. We hypothesized that CD14+ monocytes will exhibit an amplified inflammatory response, as demonstrated by significant increases in the expression of inflammatory cytokines at 48 and 72 hours post radiation exposure compared to controls which would be magnified by the inflammatory stimulus LPS. Methods: Human venous peripheral blood was collected from 5 healthy young adults (24 ± 3 years, 22 ± 1 BMI, 5 Male) using an institutional review board (IRB)-approved protocol. Women were studied in days 1-5 of menstrual cycle. Peripheral blood mononuclear cells (PBMCs) were harvested by density gradient separation and re-suspended and incubated with anti-human CD14 microbeads. CD14+ immune cells were incubated in plates, exposed to 4 gray (Gy) of x-ray radiation, and then incubated for 48 and 72 hours. CD14+ immune cells were exposed to 1:500 dilution of 0.1 mg/mL of lipopolysaccharide (LPS) for the final 8 hours of incubation. The culture media was recovered, and assayed for secreted factors, specifically for IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, IFN-γ, and TNF-α. Additionally, flow cytometry analysis was performed to quantify intracellular content of IL-10, IL-8, IFN-g, IL-4, receptor for advanced glycation end products (RAGE), and damage-associated molecular pattern (DAMP) molecules (S100A8/9) involved in inflammatory activation. All data were analyzed with two-way repeated measures ANOVA. Results: Secreted Cytokines: Cytokine expression at 48 and 72 hours did not differ between the control, radiation (4Gy), and 4Gy+LPS groups. At 48 and 72 hours, there were no significant differences observed in the expression levels of IFN-γ, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, and TNF-α in control, 4Gy, and 4Gy+LPS cells. Intracellular Content: Compared to control IL-4 content was increased at 48 and 72 hours, after exposure to 4Gy (48h: p=0.013 and 72h: p=0.008) and 4Gy+LPS (48h: p=0.022 and 72h: p=0.002). Furthermore, IL-4 was again increased at 72 hours after exposure to 4Gy+LPS compared to 48 hours (p=0.014). IL-10 content was also increased in after exposure to 4Gy and 4Gy+LPS at 72 hours compared to control (p=0.046 and p=0.007, respectively). S100A8/9 also showed a trending increase at 4Gy compared to control at 72 hours (p=0.11). No other content changed significantly. Conclusion: These data suggest that acute radiation exposure (4 Gy x-ray) has no effect on the secretome of CD14+ immune cells. However, the intracellular content of IL-4 was increased at both 48 and 72 hours, while IL-10 increased at 72 hours, indicating a shift from pro-inflammatory to anti-inflammatory intracellular content the further in time monocytes are from radiation exposure. Future work needs to test increased and repeated doses of radiation, as well as examine both x-ray and proton exposure. TRISH through NASA Cooperative Agreement NNX16AO69A and NIH HL150361. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.