Rheumatoid Arthritis (RA) has long been a major challenge to human health, characterized by its prolonged and unremitting course with no effective cure currently available . Our research team has been dedicated to investigating the treatment and drug-resistant mechanisms of RA, with the aim of improving therapeutic outcomes. Drawing inspiration from the ancient wisdom of the renowned Chinese physician Bian Que, who introduced the concept of "preventing illness before it occurs", we have contemplated the feasibility of identifying a new target for prevention and treatment in RA before its symptoms manifest.
Throughout our extensive literature review, we have noticed that the cGAS-STING pathway should be the most widespread discussion of an inflammatory signaling pathway. Researchers worldwide have conducted extensive investigations into this signaling pathway across various diseases, making significant efforts to inhibit the activation of cGAS and STING signaling cascades. Nevertheless, moderate activation of this pathway is crucial to maintain our body's natural immune response. Blockage of this signaling cascade may lead to immunodeficiency and other complications . Therefore, we have contemplated the feasibility of identifying a point within this pathway, either upstream or downstream, to maintain the delicate equilibrium of our immune system.
As sensors of the innate immune pathway, both cytoplasmic and extracellular free DNA (cfDNA) possess the capacity to activate the cGAS pathway, triggering an immune response to eliminate senescent, damaged, mutant cells, as well as the infected foreign bacteria and viruses, thereby maintaining the homeostasis of human health . However, when cfDNA accumulates excessively, it can over-activate the cGAS pathway, resulting in severe inflammation and the onset of various diseases and complications. Recent literatures have reported a significant elevation of cfDNA levels in autoimmune diseases, such as RA and Systemic Lupus Erythematosus (SLE) [4, 5]. Additionally, it has been observed that cfDNA levels increase with age, whereas advanced age serving as a critical factor in the onset of RA . Hence, we have embarked on a journey to investigate whether the accumulation of cfDNA with advancing age is a key factor in the development of RA.
Exploration and Discovery Process
Firstly, by analyzing the blood serum samples collected from healthy volunteers, RA patients, and Osteoarthritis (OA) patients, we found that RA patients exhibited significantly lower levels of a DNA repair enzyme known as TREX1 (three prime repair exonuclease 1) and remarkedly higher levels of cGAS in comparison with healthy volunteers and OA patients. Subsequently, we further compared the gene expression level of TREX1 in young individuals, elderly individuals, and RA patients. Our findings indicated that elderly individuals had lower levels of TREX1 and higher levels of cfDNA, in comparison with young individuals. In RA patients, TREX1 was significantly down-regulated, whereas the cfDNA level was markedly elevated. Interestingly, a positive correlation was found between serum cfDNA concentrations and the disease activity (Disease Activity Score using 28 joint counts, DAS28) in RA patients, providing a strong evidence to link up DNA fragments with ageing.
Next, we investigated whether an excess of cfDNA could induce or exacerbate arthritis symptoms in rats. By injecting different doses of DNA fragments into the joint cavity and tail veins of rats, we unraveled that the DNA fragments induced the release of inflammatory factors in rats, thereby exacerbating their joint arthritis symptoms. Notably, the severity of symptoms was directly proportional to the quantity of DNA fragments injected. Furthermore, we have established the first Cre conditional TREX1 knockout rat model for evaluating the arthritic condition in rats upon the challenge of DNA fragments. Apparently, our results demonstrated that joint and tail vein injection of DNA fragments significantly exacerbated the arthritis symptoms in TREX1 knockout rats.
Concurrently, we overexpressed TREX1 by injecting AAV virus to verify its impact on rats with joint arthritis. Our findings revealed that overexpressing TREX1 alleviated the arthritis symptoms in AIA rats. These results suggested a significant association between the accumulation of excessive cfDNA and the disease conditions of RA. This further proposes that regulating TREX1 expression to eliminate excessive DNA fragments can effectively alleviate the arthritis symptoms. Regarding the increased susceptibility of elderly individuals to RA, we also validated that the downregulation of E2F1 after cell senescence, followed by a decrease in c-FOS, led to reduced sensitivity in TREX1 expression following DNA stimulation. This findings may explain why the suppression of TREX1 expression in elderly is commonly observed.
Our studies have unveiled the relationship between aging, cfDNA accumulation, and TREX1 expression. In the future, enhancing the body's ability to eliminate metabolic waste, particularly by up-regulating TREX1 expression to remove excess cfDNA accumulation, holds promise as a significant approach for preventing and treating RA. Importantly, the therapeutic potential of regulating TREX1 is not limited to RA alone; it can encompass other autoimmune diseases such as SLE and various age-related conditions like degenerative neuropathies  and atherosclerosis , all of which have been highly associated with cfDNA accumulation. Theoretically, modulating TREX1 expression may have a positive impact on these diseases. Moreover, the primary significance of regulating TREX1 expression, in the context of cfDNA as a significant activator of the cGAS-STING pathway, lies in diseases prevention. By maintaining immune balance and eliminating inflammatory risk factors before the over-activation of autoimmune mechanisms, this approach may effectively prevent a multitude of diseases triggered by inflammation, delay the aging process, and simultaneously avoid excessive immunosuppression that could lead to the issues such as undetected tumor mutations by the immune system. In summary, our works represent a promising initial step, with many intriguing possibilities awaiting further exploration.