Toxoplasma gondii ‐Derived Soluble Factors Induce G2 /M Phase Accumulation and p53‐Associated Mitochondrial Apoptosis in Human Umbilical Cord Mesenchymal Stem Cells

ABSTRACT

Toxoplasma gondii infection poses potential risks to stem cell‐based therapeutic applications; however, the effects of parasite‐derived soluble factors on mesenchymal stem cells remain unclear. Here, we investigated the impact of T. gondii ‐derived soluble factors (Tg‐SFs) on human umbilical cord‐derived mesenchymal stem cells (hUC‐MSCs). hUC‐MSCs were exposed to Tg‐SFs generated in a Transwell co‐culture system, and cellular phenotypes and p53 signalling were analysed using integrated functional and molecular approaches. Tg‐SFs induced upper‐chamber multiplicity of infection (MOI)‐dependent cytotoxicity in hUC‐MSCs, accompanied by cytoskeletal remodelling and nuclear condensation. Tg‐SFs exposure caused marked G2/M phase accumulation, which was accompanied by reduced expression of Cyclin A2, Cyclin B1, and Cyclin E2 and increased p21 expression. In addition, Tg‐SFs exposure reduced phosphorylation of CDK1 at Thr161. Tg‐SFs‐exposed hUC‐MSCs underwent significant apoptosis, as evidenced by PARP and Caspase‐3 cleavage. Mechanistically, Tg‐SFs promoted accumulation of total p53 protein and selective phosphorylation of p53 at Ser15 and Ser392, and modulated Bcl‐2 family proteins by upregulating Bax, BID, Bak, Bad, and Puma while downregulating Bcl‐xL and Mcl‐1. Pharmacological inhibition of p53 with PFT‐α and siRNA‐mediated p53 knockdown partially reversed these effects, indicating that p53 signalling contributes substantially, but not exclusively, to Tg‐SFs‐induced mitochondrial apoptosis. These findings indicate that T. gondii ‐derived soluble factors impair hUC‐MSC viability through G2/M phase accumulation and p53‐associated mitochondrial apoptosis, highlighting a potential safety consideration for MSC‐based applications under parasite‐associated conditions.