Monocyte-macrophage SAMHD1 alleviates EAMG by modulating the cGAS-STING pathway

Myasthenia gravis (MG) is an autoimmune disorder whose pathogenesis has not been fully elucidated but is primarily associated with immune system dysregulation. Growing evidence indicates that innate immunity mediated by monocyte-macrophages plays a significant role in the pathogenesis of MG. SAMHD1 is a negative regulator of innate immunity, and its involvement in MG remains unclear. This study aims to clarify the immunoregulatory mechanism of SAMHD1 in MG and to explore its potential therapeutic implications. Experimental autoimmune myasthenia gravis (EAMG) rat models were established using purified torpedo acetylcholine receptor (AChR). Peripheral blood samples from MG patients and healthy controls were collected for clinical validation. Single-cell RNA sequencing was performed on peripheral whole blood from EAMG and control rats, as well as on human (GEO database). Serum levels of AChR antibodies were quantified using ELISA, while flow cytometry was employed to detect SAMHD1 expression across different cell types and to analyze the subgroup composition of CD4+ T cells. Immunofluorescence (IF) was performed for C5b-9 and α-BTX at the neuromuscular junctions (NMJ), as well as for γH2AX on macrophages. To investigate the underlying mechanism, macrophages were selectively targeted in vivo via tail vein injection of adeno-associated virus (AAV) vectors driven by the macrophage-specific F4/80 promoter, additionally, macrophages pre-treated with 1 µM STING inhibitor H-151 for 24 h were adoptively transferred into rats. RT-qPCR and western blot analysis techniques were employed to assess mRNA and protein expression levels, respectively. Our study revealed a significant reduction of SAMHD1 in monocytes (p = 0.0286) and splenic macrophages (p = 0.0002) of EAMG rats, as well as in monocytes of MG patients (p = 0.0002). Overexpression of SAMHD1 in macrophages significantly increased body weight and grip strength, reduced clinical scores, diminished complement deposition at the NMJ in EAMG rats, and shifted CD4⁺ T cells subset distribution—specifically, decreased frequencies of Th1(5.295 ± 0.2504% v 3.817 ± 1.393%) and Th17 (4.698 ± 1.078% v 2.328 ± 0.8311%) cells and increased frequency of Treg cells (4.087 ± 0.531% v 4.892 ± 0.708%). Mechanistically, SAMHD1 overexpression alleviated macrophage-intrinsic DNA damage, suppressed cGAS-STING pathway activation, and thereby restored the proportions of Th1, Th17 and Treg cells. Conversely, activation of the cGAS-STING pathway increased the proportion of Th1 (2.510 ± 0.4636% v 6.563 ± 1.546%) and Th17 (1.237 ± 0.7205% v 5.368 ± 1.526%) cells again, reduced the number of Treg cells (6.022 ± 0.3579% v 3.908 ± 0.2902%), meanwhile, EAMG rats exhibited progressive weight loss, impaired grip strength, elevated clinical scores, and enhanced complement deposition at the NMJ. This study uncovered a previously unrecognized mechanism whereby SAMHD1 downregulation impairs DNA damage repair, thereby triggering cGAS-STING pathway activation, immune dysregulation, and subsequent exacerbation of MG pathogenesis. These findings provide new insights into the pathogenesis of MG and identify SAMHD1 and the cGAS-STING axis as promising therapeutic targets for MG intervention.