Aims: Recurrent miscarriage (RM) plagues 1 %-5 % women of childbearing age. Facing the limitations of clinical treatment, its pathological mechanism remains to be clarified.
Methods: Decidual tissues of three induced abortions and three RM were collected for transcriptome sequencing. The pathological features of RM were identified by differential expression genes (DEGs) analysis, GSEA, GO and KEGG analysis, and a protein-protein interaction network was constructed for DEGs, and six algorithms were used to identify hub genes. In addition, the immune characteristics of RM patients were identified by CIBERSORT, and the correlation between them and hub genes was analyzed. Furthermore, in single-cell level, different cells were grouped according to the expression level of hub genes, and the expression ratio and abundance of hub genes in different cells and their regulation on cell function were explored.
Results: Transcriptome sequencing of patients with RM showed that a large number of genes were down-regulated, which was related to fibroblast proliferation, epithelial cell migration, female pregnancy and cell chemotaxis. Fifteen hub genes were identified by constructing a protein-protein interaction network, among which DUSP1, NR4A1 and THBS1 were involved in cell migration and chemotaxis. Immune cell infiltration analysis showed that the infiltration of T cells, macrophages and NK cells was abnormal, and there was a significant correlation with hub genes. Moreover, we found that compared with the expression of DUSP1, the non-expression of DUSP1 will reduce the extracellular matrix formation of fibroblasts and the chemotaxis of macrophages. At the same time, it is worth noting that the expression ratio and abundance of hub genes are decreased in epithelial cells, fibroblasts, macrophages and NK cells. Furthermore, single-cell analysis and in vitro and in vivo experiments show that DUSP1 and NR4A1 are low-expressed in different cells of RM patients, which is accompanied by the inhibition of fibroblast proliferation and macrophage chemotaxis. Drug prediction and screening based on hub genes show that Cinobufagin and calmidazolium are expected to be candidate drugs for RM.
Conclusion: Hub genes such as DUSP1, NR4A1 and THBS1 participate in RM by regulating epithelial cell migration, fibroblast proliferation and macrophage chemotaxis, which will provide new insight for the diagnosis and targeted therapy of RM.
Keywords: Bulk-RNA; Hub genes; Immune cell infiltration; Recurrent miscarriage; Single-cell RNA.
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