Abstract
The mammary gland is unique in female mammals. Mammary tissue undergoes development and remodeling during lactation, a stage associated with high susceptibility to bacterial infections, inducing an inflammatory condition called mastitis. Although the immune response of the mammary gland has been the subject of intense research to improve prevention and treatment efficacy, the precise definition of its immune composition at this particular physiological stage is still missing. We combined single-cell RNA-Seq, flow cytometry, and three-dimensional confocal microscopy techniques to characterize the immune landscape of lactating murine mammary tissue. Macrophages dominated the immune cell repertoire and could be subdivided into at least two subsets: ductal and stromal macrophages. Ductal macrophages represented approximately 80% of the total CD45pos immune cells and co-expressed F4/80 and CD11c, with high levels of MHC class II molecules. They were strategically poised below the alveolar basal cells in contact with the myoepithelial cell network. Adaptive T and B lymphocytes were remarkably less numerous at this stage, which could explain the limited efficacy of vaccination against mastitis. These results support the view that new strategies to increase mammary immunity and prevent mastitis should be devised.
Highlights
Female mammals are characterized by the presence of one or several pairs of mammary glands
We used a combination of single-cell RNA sequencing, flow cytometry, and three-dimensional confocal microscopy after careful separation of the mammary glands from the nearest lymph nodes to describe the immune landscape of murine mammary tissue at the lactation stage
We set up a single-cell isolation protocol from whole mammary tissue of the fourth pair of glands of lactating C57BL/6 mice using gentle physical trituration and enzymatic digestion to determine the exact resident immune cell composition without any a priori assumption
Summary
Female mammals are characterized by the presence of one or several pairs of mammary glands. These highly specialized glands provide high-quality nutrients and passive immunity to their offspring after delivery. Lactation is the stage of highest predisposition to infection by various types of bacteria, leading to an inflammatory condition called mastitis. These infections are highly frequent and detrimental for gland function. This is true in domestic species raised for milk production, with few available treatments, except currently unwanted antibiotic administration, and current vaccine with poor efficacy. Several subsets can be distinguished, and ductal macrophages, located at the lumen interface, represent a majority of the immune cells, whereas adaptive lymphocytes are much less numerous in this tissue
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