P-482 Single-cell atlas of human menstrual immune cycle reveal the key role of CD11c+ NK1 in embryo implantation

Abstract Study question How does the endometrial immune microenvironment dynamically change throughout the menstrual cycle to prepare for embryo implantation? Summary answer The dramatic accumulation and the acquisition of ability to response to EVTs of CD11c+ NK1 cell after the ovulation were prepared for embryo implantation. What is known already The human endometrium is a dynamic tissue that undergoes cyclic remodeling under the control of gonadal hormones. Recent scRNA-seq studies have provided insights into the cellular and molecular heterogeneity of the endometrial microenvironment, however, a comprehensive atlas of immune cell dynamics across the entire menstrual cycle is still lacking, hindering our understanding of the physiological immune changes that occur in the preparation for embryo implantation and growth. Study design, size, duration We collected endometrial biopsies from 55 healthy donors which cover the entire menstrual cycle. The obtained endometrium was tissue dissociated and sorted CD45+ leukocytes for single-cell analysis. Compositional transcriptomic features of endometrial immune cells at each time point across menstrual cycle were comparatively analyzed. Finally, combing with technology of mIF, we revealed a comprehensive map of temporal dynamics of human endometrial immune microenvironment across menstrual cycle, and provided an in-depth insight of CD11c+ uNK1 cells. Participants/materials, setting, methods Participants: 55 healthy donors in her natural menstrual cycle. Methods: The standard histological criteria, B-scan ultrasonography, combined with the hormone level were used to accurately determine the developmental stage of endometrial tissue. Single cell transcriptome libraries were prepared using the Massively Parallel Single-Cell RNA-seq method (MARS-seq). MARS-seq data were analyzed using the MetaCell package. Compositional analysis, differential expression analysis, Hotspot analysis and GSEA were used to map the temporal dynamics of human endometrial immune microenvironment. Main results and the role of chance Our analysis identified 40 populations of immune subsets. Among them, we identified two major population of resident NK cells: CD11c+ NK1 and CD103+ NK2. We observed the CD11c+ uNK1, which expressed higher level of KLRC, KIRs and LILRB1 that can bind to HLA-E, HLA-C or HLA-G molecules, can directly interact with EVTs. Specially, during the menstrual cycle, the proportion of NK1 cells remained constant during the proliferative phase and dramatically increase at the secretory phase, while CD103+ NK2 cells showed a sharp decrease during the secretory phase. Furthermore, we found the proliferation capacity changes of two NK subsets, and the differentiation capacity of CD11c+ NK1 to CD103+ NK2 cell might be attributed to proportion change waves of NK subsets during the menstrual cycle. Taken together, the function of CD11c+ NK1 cell to response to EVTs indicated its important regulatory role in the process of embryo implantation, and the dramatic accumulation of CD11c+ NK1 cell after the ovulation was prepared for this process. Limitations, reasons for caution It is possible that the compositional and functional change of CD11c+ NK1 cell during the menstrual cycle lead the embryo implantation failure and pregnancy failure. And future research is required to explore the relationship between these two events. Wider implications of the findings Our findings shed light on the molecular and cellular mechanisms underlying endometrial receptivity, and these findings may offer potential targets for the intervention of infertility. Trial registration number No