Imprinted genes play a key role in regulating mammalian placental and embryonic development. Here, we generated glutaminyl-peptide cyclotransferase-knockout (Qpct^-/-) mice utilizing the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) platform and identified Qpct as a novel anti-angiogenic factor in regulating mouse placentation. Compared with Qpct^+/+ mice, placentae and embryos (Qpct^-/+ and Qpct^-/-) showed significant overgrowth at embryonic Day 12.5 (E12.5), E15.5, and E18.5. Using single-cell transcriptome analysis of 32 309 cells from Qpct^+/+ and Qpct^-/- mouse placentae, we identified 13 cell clusters via single-nucleus RNA sequencing (snRNA-seq) (8880 Qpct^+/+ and 13 577 Qpct^-/- cells) and 20 cell clusters via single-cell RNA sequencing (scRNA-seq) (6567 Qpct^+/+ and 3285 Qpct^-/- cells). Furthermore, we observed a global up-regulation of pro-angiogenic genes in the Qpct^-/- background. Immunohistochemistry assays revealed a notable increase in the number of blood vessels in the decidual and labyrinthine layers of E15.5 Qpct^-/+ and Qpct^-/- mice. Moreover, the elevation of multiple pairs of ligand-receptor interactions was observed in decidual cells, endothelial cells, and macrophages, promoting angiogenesis and inflammatory response. Our findings indicate that loss of maternal Qpct leads to altered phenotypic characteristics of placentae and embryos and promotes angiogenesis in murine placentae.
Animals ; Female ; Pregnancy ; Mice ; Placentation/genetics* ; Single-Cell Analysis ; Gene Expression Profiling ; Mice, Knockout ; Transcriptome ; Placenta/blood supply* ; Neovascularization, Pathologic/genetics* ; Genomic Imprinting ; Single-Cell Gene Expression Analysis ; Angiogenesis

ARNTL Transcription Factors ; deficiency ; metabolism ; Animals ; Cyclic AMP ; metabolism ; Gluconeogenesis ; Glucose ; biosynthesis ; HEK293 Cells ; Histone Deacetylases ; metabolism ; Humans ; Liver ; metabolism ; Mice ; Mice, Knockout

Objective: To explore the effect and possible mechanisms of has-miR-150-5p targeting HIF1α to regulate malignant biological behaviors of glioblastoma (GBM) U-251MG cells. Methods: Real-time quantitative PCR (RT-PCR) was used to detect the expression of miR-150-5p and hypoxia inducible factor 1 (HIF1α) in U-251MG cells. Luciferase report assay was carried out to verify the biological relationship between miR-150-5p and HIF1α and their biological functions in U-251MG cells. The protein expressions of miR150-5pand HIF1α in U-251MG cells were detected by western blotting. The ability of cell migration was detected by wound healing test and cell invasion ability was detected by transwell test. Results: After miR-150-5p mimic transfection, the mRNA expression of HIF1α was significantly reduced in U-251MG cells (P<0.01). Bioinformatics prediction and luciferase reporter assay demonstrated that miR-150-5p down-regulated HIF1α through directly binding to HIF1α 3’-untranslated region (3’-UTR) (all P<0.05). In U-251MG cells, miR-150-5p over-expression significantly inhibited HIF1α expression, cell invasion and migration (all P<0.05). Conclusion: miR150-5p inhibits cell invasion and metastasis through negative regulation of HIF1α, indicating that miR-150-5p and HIF1α were both potential therapeutic targets for glioblastoma.