Downregulation of DROSHA: Could It Affect miRNA Biogenesis in Endometriotic Menstrual Blood Mesenchymal Stem Cells?

Menstrual blood mesenchymal stem cells (MenSCs) have gained prominence in the endometriosis scientific community, given their multifunctional roles in regenerative medicine as a noninvasive source for future clinical applications. In addition, changes in post-transcriptional regulation via miRNAs have been explored in endometriotic MenSCs with a role in modulating proliferation, angiogenesis, differentiation, stemness, self-renewal, and the mesenchymal-epithelial transition process. In this sense, homeostasis of the miRNA biosynthesis pathway is essential for several cellular processes and is related to the self-renewal and differentiation of progenitor cells. However, no studies have investigated the miRNA biogenesis pathway in endometriotic MenSCs. In this study, we profiled the expression of eight central genes for the miRNA biosynthesis pathway under experimental conditions involving a two-dimensional culture of MenSCs obtained from healthy women (n = 10) and women with endometriosis (n = 10) using RT-qPCR and reported a two-fold decrease in DROSHA expression in the disease. In addition, miR-128-3p, miR-27a-3p, miR-27b-3p, miR-181a-5p, miR-181b-5p, miR-452-3p, miR-216a-5p, miR-216b-5p, and miR-93-5p, which have been associated with endometriosis, were identified through in silico analyses as negative regulators of DROSHA. Because DROSHA is essential for miRNA maturation, our findings may justify the identification of different profiles of miRNAs with DROSHA-dependent biogenesis in endometriosis.

What Do the Transcriptome and Proteome of Menstrual Blood-Derived Mesenchymal Stem Cells Tell Us about Endometriosis?

Given the importance of menstrual blood in the pathogenesis of endometriosis and the multifunctional roles of menstrual mesenchymal stem cells (MenSCs) in regenerative medicine, this issue has gained prominence in the scientific community. Moreover, recent reviews highlight how robust the integrated assessment of omics data are for endometriosis. To our knowledge, no study has applied the multi-omics approaches to endometriosis MenSCs. This is a case-control study at a university-affiliated hospital. MenSCs transcriptome and proteome data were obtained by RNA-seq and UHPLC-MS/MS detection. Among the differentially expressed proteins and genes, we emphasize ATF3, ID1, ID3, FOSB, SNAI1, NR4A1, EGR1, LAMC3, and ZFP36 genes and MT2A, TYMP, COL1A1, COL6A2, and NID2 proteins that were already reported in the endometriosis. Our functional enrichment analysis reveals integrated modulating signaling pathways such as epithelial-mesenchymal transition (↑) and PI3K signaling via AKT to mTORC1 (↓ in proteome), mTORC1 signaling, TGF beta signaling, TNFA signaling via NFkB, IL6 STAT3 signaling, and response to hypoxia via HIF1A targets (↑ in transcriptome). Our findings highlight primary changes in the endometriosis MenSCs, suggesting that the chronic inflammatory endometrial microenvironment can modulate these cells, providing opportunities for endometriosis etiopathogenesis. Moreover, they identify challenges for future research leveraging knowledge for regenerative and precision medicine in endometriosis.

Identification of suitable reference genes for mesenchymal stem cells from menstrual blood of women with endometriosis.

It has been suggested that menstrual blood-derived mesenchymal stem/stromal cells (MenMSCs) are associated with the etiopathogenesis of endometriosis and considerable effort has been invested in searching for target genes and deciphering associated molecular pathways. However, reference gene stability for proper reproducible normalization in the analyses of the expression data validation is still unexplored in this experimental context. Therefore, in this exploratory study, we used stringent case and control selection criteria and collected menstrual blood from women with a laparoscopic diagnosis of advanced endometriosis and from fertile women without endometriosis. We tested for the first time the stability of 32 candidate reference genes to achieve increased accuracy and reliable results in the quantification of gene expression and direct future experiments using reverse transcription-quantitative PCR (RT-qPCR) in MenMSCs for endometriosis studies. Using the RefFinder web tool, we recommend the EIF2B1 and POP4 reference genes for the normalization of RT-qPCR data in study designs similar to ours. Furthermore, we suggest avoiding the commonly used GAPDH and ACTB reference genes as they are unstable. This high-visibility study is capable of directing different experimental designs as MenMSCs are derived from a minimally invasive tissue source with multifunctional roles in regenerative medicine.