PURPOSE: Spontaneous abortion (SA) remains a clinical challenge in early pregnancy. It has been reported that endoplasmic reticulum stress (ERS) is implicated in pregnancy-related complications. However, the precise mechanistic role of ERS in SA pathogenesis remains elusive. This study aims to explore the therapeutic potential of targeting ERS-related decidual dysfunction in SA. METHODS: An ERS model was established in both decidualized stromal cells (DSCs) and pregnant mice through tunicamycin (Tu) administration. Chromatin immunoprecipitation (ChIP) and dual-luciferase reporter assays were performed to determine the interaction between XBP1s and the transcription factor binding site (TFBS) of tumor necrosis factor receptor-associated factor 6 (TRAF6). Mitochondrial membrane potential (MMP) and mitochondrial function were assessed using JC-1 and TMRM staining following ERS induction in DSCs. The effects of XBP1s inhibitors on mitochondrial metabolism and autophagy were evaluated through RT-qPCR, Western blotting, RNA-Seq, TUNEL assays, ROS and MitoSOX detection, and histological analyses in Tu-treated DSCs and SA patients. STF-083010 (STF) or shXBP1 was utilized to assess the inhibitory effects of X-box binding protein 1 (XBP1s) on DSC function both in vitro and in vivo. RESULTS: We observed significant upregulation of XBP1s in decidual tissues from SA patients and Tu-exposed DSCs. Tu exposure significantly increased the proportion of TUNEL-positive cells and upregulated pro-inflammatory cytokines (IL-1β, TNF-α, IL-6, IL-18) in DSCs. XBP1s inhibition via shXBP1 or pharmacological inhibitor STF attenuated Tu-induced apoptosis and inflammatory cytokine expression. Notably, STF or shXBP1 treatment enhanced MMP and upregulated LC3-II expression in Tu-treated DSCs, indicating autophagy activation.Intriguingly, chloroquine (CQ)-mediated autophagy suppression exacerbated apoptosis in STF/Tu-co-treated DSCs, suggesting that XBP1s inhibition confers cytoprotection through autophagy induction. Mechanistically, XBP1s directly bound to the TFBS of TRAF6, a ubiquitin E3 ligase. TRAF6 overexpression exacerbated mitochondrial dysfunction and apoptosis while suppressing autophagy via inhibition of mTORC2/Akt pathway in Tu-treated DSCs. CONCLUSION: XBP1s inhibition restored mitochondrial homeostasis and promoted autophagy by modulating the TRAF6/mTORC2 axis under ERS conditions, providing novel mechanistic insights into SA pathogenesis and potential therapeutic targets.