Single cell sequencing of live mammary cells from human milk

Researchers have explored the cellular changes that occur in human mammary tissue in lactating and non-lactating women, offering insight into the relationship between pregnancy, lactation, and breast cancer.

Breast milk in bottles

The study was led by researchers from the Wellcome-MRC Cambridge Stem Cell Institute (CSCI) and the Department of Pharmacology at the University of Cambridge.

Breast tissue is dynamic, changing over time during puberty, pregnancy, breastfeeding, and aging. The paper, published today in the journal Nature Communications, focuses on the changes that take place during lactation by investigating cells found in human milk.

This research, led by Dr Alecia-Jane Twigger of CSCI, found that the cells in milk, once thought to be dead or dying, are in fact very much alive. These living cells provide researchers with the chance to study not only the changes that occur in mammary tissues during lactation, but also insight into a potential early indicator of future breast cancer development.

“I believe that by studying human milk cells, we will be able to answer some of the most fundamental questions around mammary gland function such as: how is milk produced? Why do some women struggle to make milk? and what strategies can be employed to improve breastfeeding outcomes for women?” said Dr Alecia-Jane Twigger at the Wellcome-MRC Cambridge Stem Cell Institute who led the study.

The researchers collected voluntary breast milk samples from lactating women, as well as samples of non-lactating breast tissue donated from women who elected to have aesthetic breast reduction surgery. Using single-cell RNA sequencing analysis, the team conducted a novel comparison of the composition of the mammary cells taken using these two methods, identifying the distinctions between lactating and non-lactating human mammary glands.

Exploring the diversity between non-lactating mammary cells (NMCs)
and lactation derived mammary cells (LMCs)

Fig. 1

a Cells from non-lactating tissue (above) and human milk (below) were isolated using either mechanical dissociation and enzymatic digestion or centrifugation, for downstream analysis. b Mammary cells from both non-lactating breast tissue (top) or lactating milk cells (bottom) were cultured either in i 2D (n = 10) or ii 3D, scale bar represents 250 μm. c Representative flow cytometric profiles of immune/stromal (Draq5+/CD45+), luminal (Draq5+/EpCAM+/CD49f+/−) and myoepithelial cells (Draq5+/EpCAM/CD49f+) from i NMCs and ii LMCs. d Schematic diagram for the scRNA-seq experimental set-up for cell samples from seven non-lactating participants and nine lactating females. e Uniform manifold approximation and projection (UMAP) dimensional reduction of the mammary cells reveals distinct clusters arising from NMCs and LMCs.

While accessing breast tissue for study relies on donors already undergoing surgery, breast milk samples are much simpler to acquire. Breast milk donors are engaged via midwives or women’s networks (an undertaking made more challenging by the pandemic) and agree to share their samples over time. Typical daily production for lactating women is between 750-800ml, and the sample size for Twigger’s research is on average a mere 50ml, an amount which can contain hundreds of thousands of cells for study.

By collecting these samples donated by breastfeeding women – samples now known to contain living and viable cells – researchers have the opportunity to capture dynamic cells in a non-invasive way. This greater ease of access to breast cells can open the door to more studies on women’s health in the future.

“The first time Alecia told me that she found live cells in milk I was surprised and excited about the possibilities. We hope this finding will enable future studies into the early steps of breast cancer,” said Dr Walid Khaled, at the Wellcome-MRC Cambridge Stem Cell Institute and University of Cambridge’s Department of Pharmacology, who was also involved in the study.

This paper and its findings are part of the Human Breast Cell Atlas project funded by the MRC.

SourceUniversity of Cambridge

Twigger AJ, Engelbrecht LK, Bach K, Schultz-Pernice I, Pensa S, Stenning J, Petricca S, Scheel CH, Khaled WT. (2022) Transcriptional changes in the mammary gland during lactation revealed by single cell sequencing of cells from human milk. Nat Commun 13(1):562. [article]

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