Unknown,Transcriptomics,Genomics,Proteomics

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10X Chromium Single-cell RNA sequencing of the female reproductive tract in young, pregnant and aged C57BL/6 mice

ABSTRACT: The aim of the study was to assess the cellular composition and transcriptional changes at single-cell resolution of the female reproductive tract across the four stages of estrus cycle in young adult mice (ovary, oviduct, uterus, cervix and vagina, with spleen as a comparison) and compared this with similar analyses of decidualization (uterus) and ageing (ovary, oviduct, uterus, cervix, vagina and spleen). Vaginal smears were used to stage the estrus cycle in young mice. To collect the decidualized uteri mice were mated and uterine tissue was collected from pregnant mice at day 5.5 after mating. Tissues of aged mice were collected at the age of 18 months. All tissues were collected and digested using a collagenase and trypsin protocol to obtain single cell suspension. Libraries were prepared using the 10x scRNA-seq protocol.

INSTRUMENT(S): Illumina NovaSeq 6000

ORGANISM(S): Mus musculus

SUBMITTER: Fritjof Lammers

PROVIDER: E-MTAB-11491 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:The potential for xenoestrogens and other environmental chemicals to alter female reproductive function calls for the study of their affects on endogenous hormone-regulated gene expression pathways during uterine tissue remodeling which occurs as part of the menstrual/estrus cycle. However, our knowledge of these pathways is limited. Here, we characterize changes in the CD-1 mouse uterine transcriptome during proestrus and estrus, which are regulated by estrogen and progesterone in preparation of the uterus for pregnancy. Mice were staged beginning at 6 wk of age and uterine horns were harvested after monitoring two estrus cycles. Microarray analysis identified 2,429 genes differentially expressed in estrus compared to proestrus, indicating that the mouse uterus undergoes remarkable remodeling during the estrus cycle, affecting ~10% of all protein-coding genes. Changes in gene expression associated with structural alteration of the uterus include remodeling of the extracellular matrix, changes in cell keratins and adhesion molecules, activation of mitosis, MHC class II presentation, complement and coagulation cascades, and cytochrome P450 expression. Signaling pathways regulated during the estrus cycle, involving ligand-gated channels, Wnt and hedgehog signaling, and several transcription factors with poorly understood roles in reproductive tissues, include several genes and gene networks that have been implicated in pathological states. The information presented here builds a background for understanding of mechanisms involved in uterine tissue response to endocrine disruptors and the development of reproductive tract diseases. Beginning on postnatal day 42, female CD-1 mice were monitored for reproductive stage cycle for 2 consecutive cycles and then total RNA was isolated from uterine horns at estrus or proestrus. Samples were pooled for microarray as follows. Six litters were used to generate 2 tissue pools for estrus and 2 tissue pools for proestrus, with estrus pool 1 and proestrus pool 1 comprised of mice from the same 3 litters, and estrus pool 2 and proestrus pool 2 comprised of mice from the other 3 litters. Each pool was comprised of uterine tissue RNA from n= 4-6 individual mice. Fluorescent labeling of RNA and hybridization of the Alexa 555-labeled (green) and Alexa 647-labeled (red) RNA samples to Agilent-026655 Whole Mouse Genome Microarray 4x44K v2 (Agilent Technology, Palo Alto, CA; catalog # G4846A) were carried out, with dye swapping for each estrus cycle stage comparison.

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10X Chromium Single-cell RNA sequencing of the female reproductive tract in young, pregnant and aged C57BL/6 mice

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