title: Influence of Low Dose-Radiotherapy on the Phenotype of Fibroblast-like Synoviocytes in Dependence of Initial Inflammatory Status, Gender, and Irradiation Frequency
Creators: name: Piltz, Margaux
Publisher: name: Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU)
Year: 2025

description: Background Many inflammatory musculoskeletal diseases, such as rheumatoid arthritis (RA), are char acterized by a painful swelling of joints, bone erosion, and cartilage reduction. Fibroblast-like synoviocytes (FLS) are the key effector cells that perpetuate inflammation within the synovium. In the inflammatory regions, hyperplasia takes place and FLS have a significantly lower rate of apoptosis. FLS produce components of the synovial fluid that maintain the joint homeostasis in healthy tissue. Low-dose radiotherapy (LD-RT) is known to ameliorate inflammation. It re duces bone and cartilage loss by increasing apoptosis of the FLS and reducing their growth rate in the human tumor necrosis factor-α transgenic (hTNFα tg) mice, which is an animal model of RA. The hTNF-α tg mouse model is characterized by an overexpression of TNF-α that results in the classical symptoms seen in RA patients. It has been shown that both the dose and frequency of irradiation have an impact on cells; however, the exact underlying mech anisms in LD-RT are mostly unknown. There are currently effective medications for RA, but unfortunately there are still patients who do not react to it accordingly. Therefore, LD-RT could potentially be a good additive therapy because it has been well-tested in clinical settings. Fur thermore, as females are more susceptible to RA and the gender-dependent sensitivity of ra diation is currently highly discussed, we also investigated whether there are gender effects for LD-RT. The results were compared to healthy, wild type (C57Bl/6) littermates to understand the impact of the initial inflammatory status on the radiation response of FLS. Methods The FLS of the female and male hTNFα tg and C57Bl/6 mice were isolated from the hind paws and were cultivated. 24 hours after seeding, the cells were irradiated with varying X-ray doses and frequencies (1x/2x), whereby the total dose was always 0; 0.1; 0.5; 1.0 or 2.0 Gray (Gy). The gene expression (Cxcl-1, hTNF-α, Il-6, Mmp-13, Mmp-9, Opg, Tgf-β, Cdh11, Mmp 3, Rank-l and Vcam-1) and cytokine release (hTNF-α, CXCL-1, TGF-β and IL-6) of the FLS from the mice were analysed using quantitative polymerase chain reaction (qPCR) analyses and by investigating the FLS supernatants (SN) via enzyme linked immunosorbent assay (ELISA), respectively. Results Our data showed that the untreated FLS pools display differences in terms of their cytokine release, but that these differences are dependent on the sex and the inflammatory status. In an inflammatory set-up, male-derived FLS pools secreted significantly higher properties of hTNF-α than females. Furthermore, we were able to provide a broad overview of the gene expressions of FLS after exposure to single or fractionated X-ray doses. Genes associated with osteoclastogenesis, cartilage degrading enzymes and Tgf-β appeared to be particularly impacted. Conclusion Given that we were able to demonstrate that female and male FLS pools significantly dif fered in their cytokine milieu prior to any irradiation, it has become clear how important it is to base future research upon this knowledge. It might offer a new perspective on gender-person alised treatment options, but future research would be required to further verify and validate these findings. There were no changes in gene expression or cytokine release in all healthy FLS pools when applying X-ray doses lower than 2x1.0Gy, such as 2x0.05 Gy, 2x0.25 Gy or 2x0.5 Gy, irrespective of gender. We conclude that irradiation with those doses may not cause harm to healthy surrounding FLS.