Obesogenic diet disrupts endometrial stromal cell differentiation in the E5.5 mouse uterus

Maternal obesity is associated with increased risk of infertility, implantation failure, miscarriage, and other pregnancy complications. While prior studies have linked obesity to uterine dysfunction and impaired endometrial biology, how obesity alters the cellular and molecular landscape of the early pregnant endometrium remains poorly understood. Here, we perform single-cell RNA sequencing of E5.5 implantation sites from control and obesogenic diet-exposed mice to generate a cellular atlas of the early decidualizing uterus. We identify obesity-associated transcriptional changes across multiple endometrial stromal cell (ESC) states and innate immune populations, including uterine natural killer cells and macrophages. Computational modeling reveals that maternal obesity likely disrupts distinct routes of ESC differentiation during decidualization and leads to shifts in ESC-derived cues known to impact immune responses. These findings provide a comprehensive resource to understand how maternal obesity reprograms the maternal–fetal interface at implantation. Experimental Design: Implantation sites were collected at E5.5 from control Chow (17% fat, 54% carbohydrate, 29% protein (kCal/g), n = 5 ) and high fat/high sucrose (HFHS) diet-exposed (45% fat, 17% sucrose, 18% carbohydrate, 20% protein (kCal/g), n = 5) C57BL/6J mice. Implantation sites were enzymatically digested into single cell suspensions, live cells were sorted by FACS using 7AAD, subjected to single-cell library construction (10X, Chromium Next GEM Single Cell 3’ Gel Bead Kit v3.1), and sequenced using a paired ends approach (and multiple lanes on flow cell in some intances) using an Illumina NextSeq 2000 instrument.  Details are accessible at https://doi.org/10.1101/2025.08.13.670199.