Revealing the Action Mechanism of WISP2 in Miscarriage Based Multi-Omic Analysis and Experimental Validation

Recurrent miscarriage (RM) is a major unresolved issue in reproductive medicine. Although WISP2 is known to participate in multiple cellular processes, its function in trophoblast biology and pregnancy maintenance remains unknown. We performed single-cell RNA sequencing on chorionic villus tissues from RM patients and normal controls to identify cell-type-specific transcriptional alterations, which led to the identification of WISP2 as a candidate gene in the SCT/EVT subpopulation. Fetal membrane samples (chorion) were obtained from Qingdao Women and Children's Hospital's Surgical Outpatient Department: 20 RM patients and 20 healthy controls. Using qRT-PCR, Western blot, and immunohistochemistry to detect gene and protein expression. HTR-8 cells were used to construct WISP2 overexpressing cells, and MTT, wound healing and Transwell experiments were performed for functional experiments. Eleven cell classes in the sample were demonstrated by single-cell analysis. Among these cells were DSCs and SCT/extravillous trophoblasts (EVT). The level of WISP2 is low in SCT/EVT cells from RM patients. In chorionic villus tissues, both WISP2 and E-cadherin were significantly upregulated in the RM group relative to the control group (all p < 0.05). Immunohistochemistry confirmed elevated WISP2 and E-cadherin protein levels in RM chorionic villi (both p < 0.05), with predominant cytoplasmic and membranous localization, respectively. In HTR-8 cells, WISP2 overexpression suppressed proliferation (35%, p < 0.01), migration (45%, p < 0.01), and invasion (50%, p < 0.001), upregulated E-cadherin (p < 0.05), downregulated Vimentin (p < 0.05), and increased p-ERK (p < 0.05) without altering total ERK1/2 (p > 0.05), suggesting that WISP2 promotes an epithelial-like phenotype accompanied by ERK pathway activation. In summary, WISP2 is dysregulated in RM and suppresses trophoblast invasion and EMT in vitro, suggesting a potential role in RM pathogenesis, though in vivo validation is required.