Bulk RNA-sequencing, single-cell RNA-sequencing analysis, and experimental validation reveal iron metabolism-related genes CISD2 and CYP17A1 are potential diagnostic markers for recurrent pregnancy loss.

BACKGROUND: Recurrent pregnancy loss (RPL) is associated with variable causes. Its etiology remains unexplained in about half of the cases, with no effective treatment available. Individuals with RPL have an irregular iron metabolism. In the present study, we identified key genes impacting iron metabolism that could be used for diagnosing and treating RPL. METHODS: We obtained gene expression profiles from the Gene Expression Omnibus (GEO) database. The Molecular Signatures Database was used to identify 14 gene sets related to iron metabolism, comprising 520 iron metabolism genes. Differential analysis and a weighted gene co-expression network analysis (WGCNA) of gene expression revealed two iron metabolism-related hub genes. Reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry were used on clinical samples to confirm our results. The receiver operating characteristic (ROC) analysis and immune infiltration analysis were conducted. In addition, we analyzed the distribution of genes and performed CellChat analysis by single-cell RNA sequencing. RESULTS: The expression of two hub genes, namely, CDGSH iron sulfur domain 2 (CISD2)and Cytochrome P450 family 17 subfamily A member 1 (CYP17A1), were reduced in RPL, as verified by both qPCR and immunohistochemistry. The Gene Ontology (GO) analysis revealed the genes predominantly engaged in autophagy and iron metabolism. The area under the curve (AUC) demonstrated better diagnostic performance for RPL using CISD2 and CYP17A1. The single-cell transcriptomic analysis of RPL demonstrated that CISD2 is expressed in the majority of cell subpopulations, whereas CYP17A1 is not. The cell cycle analysis revealed highly active natural killer (NK) cells that displayed the highest communications with other cells, including the strongest interaction with macrophages through the migratory inhibitory factor (MIF) pathway. CONCLUSIONS: Our study suggested that CISD2 and CYP17A1 genes are involved in abnormal iron metabolism, thereby contributing to RPL. These genes could be used as potential diagnostic and therapeutic markers for RPL.

A Simulation Study on the Growth of Oviduct Mucosa Cells in the Uterine Cavity Microenvironment.

OBJECTIVE: The growth of oviduct mucosa in the uterine cavity was observed by co-culture of oviduct mucosa cells and endometrial cells in different proportions to study the possibility and function of the growth of oviduct mucosa in the uterine cavity. METHODS: The extracted cells were identified by immunofluorescence with cytokeratins 19 (CK19) and vimentin. A Cell Counting Kit-8 (CCK8) experiment, cell decidualization induction, and HE staining were performed after the co-culture of two kinds of cells in different proportions. RESULTS: 1) The cells could grow normally when the two cells were co-cultured indirectly. 2) A CCK8 test of oviduct mucosa cells showed that the growth rate of each group was similar after the indirect co-culture of two kinds of cells in different proportions, which was in line with the growth law of normal cells. 3) Immunofluorescence identification of the cells showed that most of the two kinds of cells in the second passage were CK19 positive and were epithelial cells, while most of the cells in the fifth passage expressed positive vimentin antibody and were stroma cells. 4) After cell decidualization induction, the cell morphology of each group showed deciduation-like changes. 5) After decidualization, the cell morphology of each group was similar after HE staining. CONCLUSION: Oviduct mucosa cells can grow normally in the uterine environment. In the uterine environment with different degrees of endometrial loss, the growth rate of oviduct mucosa cells is not inhibited. Its morphology does not change, and it can undergo decidualization in vitro.

Discussion on operative skills in the embolization of hydrosalpinx by hysteroscopic placement of a microcoil.

This study aims to discuss the operative skills of hysteroscopic tubal embolization and reduce the occurrence of complications.Ninety-four patients were divided into group A and group B. The main surgical technique in group A: when the inner sleeve is sent to the fallopian tube and no longer accessible (but no >3 cm), remove the guide wire and put into the microcoil. But in group B, there are four major surgical techniques. First, the depth at which the guide wire enters the tube was controlled at 2 cm. Second, the inner diameter of the fallopian tube must be explored to determine the type and shape of the coils. Third, saline should be used to separate the catheter. Fourth, it is to control the release speed of the coils. The superiority of the improved operation method was confirmed by comparing the surgical failure rate, incidence of complications, and cost of surgery before and after the procedure.The reoperation rate of group A was 10% (3/30), while that of group B was 2.68% (3/112). The ectopic microcoils rate of group A was 6.67% (2/30), while that of group B was 0.89% (1/112). The microcoil damages rate of group 23.33% (7/30), while that of group B was 8.04% (9/112). All P values were <.01, and the difference was statistically significant.Hysteroscopic tubal embolization is currently a new surgical procedure to block the fallopian tubes and prevent the reverse flow of fluid in the fallopian tubes into the uterine cavity. After we improved surgical techniques, the surgical failure rate, complication rate, and operation cost of fallopian tube embolization were significantly lower than before the improved method was applied. The improved techniques led to a higher success rate.