Guanine nucleoside alleviates mycophenolic acid-induced toxicity in mouse embryonic stem cells

Baoshan Lin^1,2Yin Ta²Dandan Ou¹Xiaolong Liu³Xiaoqiang He⁴Zhun Rang²Dongqiang Zhang¹Wei Fu^1*Daoliang Lan^1*

• ¹Southwest Minzu University, Chengdu, China
• ²Animal Disease Prevention and Control Center of Aba Tibetan and Qiang Autonomous Prefecture, Markang, China
• ³Science Technology and Agriculture and Animal Husbandry Bureau of Hongyuan County, Hongyuan, China
• ⁴Longri stud farm, Hongyuan, China

Mycophenolic acid (MPA), an immunosuppressant widely used in organ transplantation and the treatment of autoimmune diseases, poses a teratogenic risk to mammalian fetuses. Guanine nucleoside (GUO), an important intermediate in purine metabolism, has the potential to modulate immune responses. In this study, we systematically evaluated the effects of MPA at various concentrations (0.05, 0.075, 0.1 µM) on the proliferation, apoptosis, and differentiation of mouse embryonic stem cells (PGK12.1), and investigated whether GUO at different concentrations (2, 4, 8 mM) could mitigate MPA induced toxicity. The results showed that MPA inhibited the viability of PGK12.1 cells in a dose-dependent manner, increased reactive oxygen species (ROS) levels, reduced mitochondrial membrane potential, significantly down regulated the expression of proliferation-related genes (PCNA, CCND1, CDK1) and the anti-apoptotic gene Bcl2, up regulated apoptotic genes (Bax, Caspase3), and disrupted the differentiation potential of PGK12.1 cells. Notably, treatment with 8 mM GUO significantly ameliorated these toxic effects. RNA-seq analysis revealed that pathways associated with cell proliferation, apoptosis, and differentiation (including TGF-β, PI3K-AKT, and p53) were significantly enriched in the MPA-treated group, suggesting that these signaling pathways may be involved in the response to the MPA-induced phenotype, whereas GUO may potentially counteract these effects through the regulation of pathways associated with chromatin architecture and cytoskeletal organization. This study elucidates the toxic effects of MPA on mouse embryonic stem cells and highlights the protective role of GUO, providing a foundation for further investigations into the mechanisms underlying MPA induced toxicity.