Single-cell RNA-Aequencing Reveals Novel Myofibroblasts with Epithelial Cell-Like Features in the Mammary Gland of Dairy Cattle

Abstract

Background: Cow’s milk is a highly-nutritious dairy product that is widely consumed worldwide. It is secreted by the developed mammary gland (MG) of dairy cattle. However, a comprehensive understanding of cell-type diversity and cell function within bovine MG is lacking. In the current study, we used single-cell RNA sequencing to investigate the transcriptome of 24,472 high-quality MG cells isolated from newborn and adult cows. Results: Unbiased clustering analysis revealed the existence of 24 cell types, which could be divided into four categories: 9 immune, 3 epithelial, 9 fibroblast, and 3 endothelial cell types. Other cell subtypes were further identified based on re-clustering and pseudotemporal reconstruction of epithelial cells that included 3 mature luminal epithelial, 1 intermediate, and 2 progenitor cell subtypes. The individual top marker genes of these 3 mature luminal epithelial cell subtypes (L0, L1, and L5) were APOA1, STC2, and PTX3, which were further validated using immunofluorescence. Based on functional analysis, the L0, L1, and L5 cell subtypes were all involved in the upregulation of lipid metabolism, protein and hormone metabolism, and the immune response, respectively. Furthermore, we discovered a novel myofibroblast that expresses COL1A1 and CSN3, has visible epithelial-like characteristics, and shows the potential to differentiate into luminal epithelial cells, especially immune-sensing luminal cells (L5). Conclusions: We constructed the first single-cell atlas of the dairy cow MG, and our new findings of epithelial-like myofibroblast cells and their differentiation trajectories into luminal cells may provide novel insights into the development and lactogenesis in dairy cattle MGs.

Mammary cell gene expression atlas links epithelial cell remodeling events to breast carcinogenesis

“…When the lineage gene sets were compared between the two species, both commonalities and differences were apparent. Recently, scRNAseq data of dairy cattle mammary gland was reported 50 and those from other organisms could also appear in the near future. The gene lists obtained in this study would be useful to explore core gene sets for the function of the mammary gland and the differentiation machinery, together with interspecies differences and their biological meanings.…”

Recent Applications of RNA Sequencing in Food and Agriculture

“…In sea urchins, using scRNA-Seq, different cell types commonly seen during the embryo development have been identified by the selective inhibition of Delta/ Notch and Wnt responsive pathways [189]. Studying the infant and adult cattle mammary glands (MG) with scRNA-Seq, dairy scientists developed a MG-specific single-cell atlas, determined the cell-type heterogeneity, and identified a novel myofibroblast that can differentiate into luminal epithelial cells, and has potential role in lactation and immunity [190].…”