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RNA-Seq analysis of pubertal mammary epithelial cells reveals novel n-3 polyunsaturated fatty acid transcriptomic changes in the fat-1 mouse model

ABSTRACT: Background/Objectives: Early exposure of nutrients during pubertal mammary gland development may reduce the risk of developing breast cancer later in life. Anticancer, n-3 polyunsaturated fatty acids (n-3 PUFA) are shown to modulate pubertal mammary gland development, however, the mechanisms of action remains unclear. Prior work focuses on effects at the whole tissue level, and little is known at the cellular level, such as mammary epithelial cells (MEC), which are implicated in cancer development. Methods: This pilot study examined the effects of lifelong n-3 PUFA exposure on the transcriptome by RNA-Seq in isolated MEC of pubertal (6–8-week-old) female fat-1 transgenic mice capable of de novo n-3 PUFA synthesis. edgeR and DESeq2 were used separately for the differential expression analysis of RNA sequencing data followed by Benjamini-Hochberg procedure for multiple testing correction. Results: Nine genes were found concordant and significantly different (p ≤ 0.05) by both DESeq2 and edgeR methods. These genes were associated with multiple pathways, suggesting that n-3 PUFA stimulates estrogen-related signalling (Mllt10, Galr3, Nrip1) and a glycolytic profile (Soga1, Pdpr, Uso1) while offering protective effects for immune and DNA damage responses (Glpd1, Garre1, Rpa1) in MEC during puberty. Conclusions: This pilot study highlights the utility of RNA-Seq to better understand mechanistic effects of specific nutrients such as n-3 PUFA in a cell-specific manner. Thus, further studies are warranted to investigate cell-specific mechanisms by which n-3 PUFA influences pubertal mammary gland development and breast cancer risk later in life.

ORGANISM(S): Mus musculus

PROVIDER: GSE281867 | GEO | 2024/11/17

REPOSITORIES: GEO

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Project description:Mastitis is an inflammation of the mammary gland (MG), usually due to bacterial infection. Although considerable attention has been paid to this economically important disease, the early stages of the host response remain poorly defined. In particular, it is unclear how mammary epithelial cells (MEC), a first barrier against pathogens, respond to infection. Indeed, it is difficult to differentiate between the contributions of MEC and infiltrating immune cells to gene expression profiles of mammary tissue during mastitis. The current investigation examines the response of MEC at the early stage of infection using a non invasive RNA sampling method taking advantage of the presence of cytoplasmic crescents contained in milk fat globules. We have recently shown that, in goats, Milk Fat Globules (MFG) provide a unique source of RNA to study the in vivo regulation of gene expression in MEC. This non invasive RNA sampling method was used during the time course of an experimental intra mammary infection (IMI) with S. aureus. Experiments were performed using ovine microarrays (Agilent) to compare gene expression patterns before infection, at 12h, 18h and 24h post-infection (PI). Furthermore, we combined this approach with laser capture microdissection of MEC isolated from frozen slides of mammary tissue to study some specific genes at the late stage of infection (30h PI). We show that at 18h PI, before the burst of somatic cells in milk, MEC play an important role in the recruitment and activation of inflammatory cells through the IL-8 signaling pathways. Then, at the late stage of infection (30h PI), the contribution of MEC in immune response changes to produce different acute phase proteins, including SAA3, serpin A1 and PTX3. These cells also express factors that contribute directly to fighting infection, such as S100A12. In summary, we demonstrate for the first time in vivo how MEC orchestrate innate immune response during an IMI with S. aureus in the goat species. We report here new opportunities to assess the dynamics of gene expression in the mammary gland, thus providing significant advances in the understanding of MEC immune capacity. Furthermore, the production of some molecules by MEC, in the early stages of IMI, could provide sensitive biomarkers for early detection and therefore, treatment of mastitis.

Project description:Mastitis is an inflammation of the mammary gland (MG), usually due to bacterial infection. Although considerable attention has been paid to this economically important disease, the early stages of the host response remain poorly defined. In particular, it is unclear how mammary epithelial cells (MEC), a first barrier against pathogens, respond to infection. Indeed, it is difficult to differentiate between the contributions of MEC and infiltrating immune cells to gene expression profiles of mammary tissue during mastitis. The current investigation examines the response of MEC at the early stage of infection using a non invasive RNA sampling method taking advantage of the presence of cytoplasmic crescents contained in milk fat globules. We have recently shown that, in goats, Milk Fat Globules (MFG) provide a unique source of RNA to study the in vivo regulation of gene expression in MEC. This non invasive RNA sampling method was used during the time course of an experimental intra mammary infection (IMI) with S. aureus. Experiments were performed using ovine microarrays (Agilent) to compare gene expression patterns before infection, at 12h, 18h and 24h post-infection (PI). Furthermore, we combined this approach with laser capture microdissection of MEC isolated from frozen slides of mammary tissue to study some specific genes at the late stage of infection (30h PI). We show that at 18h PI, before the burst of somatic cells in milk, MEC play an important role in the recruitment and activation of inflammatory cells through the IL-8 signaling pathways. Then, at the late stage of infection (30h PI), the contribution of MEC in immune response changes to produce different acute phase proteins, including SAA3, serpin A1 and PTX3. These cells also express factors that contribute directly to fighting infection, such as S100A12. In summary, we demonstrate for the first time in vivo how MEC orchestrate innate immune response during an IMI with S. aureus in the goat species. We report here new opportunities to assess the dynamics of gene expression in the mammary gland, thus providing significant advances in the understanding of MEC immune capacity.M-bM-^@M-^BFurthermore, the production of some molecules by MEC, in the early stages of IMI, could provide sensitive biomarkers for early detection and therefore, treatment of mastitis. Experiments were performed using ovine microarrays (Agilent) to compare gene expression patterns before infection, at 12h, 18h and 24h post-infection (PI). 20 sample records; mono-colour experimental design

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RNA-Seq analysis of pubertal mammary epithelial cells reveals novel n-3 polyunsaturated fatty acid transcriptomic changes in the fat-1 mouse model

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