Gene Expression In Mammary Tissue Research Articles

Effect of Acute and Cumulative Stress on Gene Expression in Mammary Tissue and Their Interactions with Physiological Responses and Milk Yield in Saanen Goats

This study addresses the hypothesis that different acute stressors can cumulatively decrease milk yield. In fact, in a time of global warming, the impact of environmental stress and farm management practices on milk production remains unclear. In this context, our objective was to investigate the effect of acute and cumulative stress on gene expression in mammary tissue and their interactions with physiological responses and milk yield in Saanen goats. Thirty lactating goats were subjected to two treatments: (1) control (CT), in which goats were maintained following a habitual routine under comfort conditions; (2) stress (ST), in which the goats were subjected to different types of environmental stress: heat stress, adrenocorticotropic hormone administration, hoof care management, and exposure to rain. These stressors were performed sequentially, with one stress per day on four consecutive lactation days, to evaluate their effect on milk quality and milk yield. Our results showed that compared to CT goats, cumulative stress increased the gene expression of glucocorticoid receptor (GR), interferon-gamma (IFN-γ), superoxide dismutase (SOD), and catalase (CAT) in mammary tissue, which are indicators of cortisol action, inflammatory response, and antioxidant enzymes. Furthermore, the acute challenges imposed on ST goats changed their rectal temperature and respiratory frequency and increased cortisol, glucose, cholesterol, triglycerides, and high-density lipoprotein release in plasma when compared to CT goats. Although these physiological and metabolic responses restore homeostasis, ST goats showed lower milk yield and higher somatic cell count in milk than CT goats. In conclusion, the results confirmed our initial hypothesis that different acute stressors cumulatively decrease the milk yield in Saanen goats.

Effects of Dietary Probiotics and Acidifiers on the Production Performance, Colostrum Components, Serum Antioxidant Activity and Hormone Levels, and Gene Expression in Mammary Tissue of Lactating Sows.

The aims of this study were to test the effects of dietary probiotics and acidifiers on the production performance, colostrum components, serum antioxidant activity and hormone levels, and gene expression in the mammary tissue of lactating sows. Four treatments were administered with six replicates to 24 lactating sows. The control group (GC) received a basal diet, while the experimental groups received a basal diet with 200 mL/d probiotics (GP), 0.5% acidifiers (GA), and 200 mL/d probiotics + 0.5% acidifiers (GM), respectively. Compared with the GC, (1) the average weight of the piglets on the 21st day of lactation in the GM was higher (p < 0.05); (2) the colostrum fat ratio increased significantly (p < 0.05); (3) the malondialdehyde levels in GP and GM were lower (p < 0.05) on the 11th day; (4) on the 1st, 11th, and 21st days, the prolactin in GP and GM increased (p < 0.05); (5) on the 21st day, the relative expression levels of the prolactin receptor and fatty acid synthase were increased (p < 0.05). In summary, the basal diet mixed with 200 mL/d probiotics + 0.5% acidifiers could improve the production performance, colostrum components, serum antioxidant activity, and hormone levels of lactating sows.

STAT5-Driven Enhancers Tightly Control Temporal Expression of Mammary-Specific Genes

The de novo formation of milk-secreting mammary epithelium during pregnancy is regulated by prolactin through activation of the transcription factor STAT5, which stimulates the expression of several hundred mammary-specific genes. In addition to its key role in activating gene expression in mammary tissue, STAT5, which is ubiquitously expressed in most cell types, implements T cell-specific programs controlled by interleukins. However, the mechanisms by which STAT5 controls cell-specific genetic programs activated by distinct cytokines remain relatively unknown. Integration of data from genome-wide surveys of chromatin markers and transcription factor binding at regulatory elements may shed light on the mechanisms that drive cell-specific programs. Here, we have illustrated how STAT5 controls cell-specific gene expression through its concentration and an auto-regulatory enhancer supporting its high levels in mammary tissue. The unique genomic features of STAT5-driven enhancers or super-enhancers that regulate mammary-specific genes and their dynamic remodeling in response to pregnancy hormone levels are described. We have further provided biological evidence supporting the in vivo function of a STAT5-driven super-enhancer with the aid of CRISPR/Cas9 genome editing. Finally, we discuss how the functions of mammary-specific super-enhancers are confined by the zinc finger protein, CTCF, to allow exclusive activation of mammary-specific genes without affecting common neighboring genes. This review comprehensively summarizes the molecular pathways underlying differential control of cell-specific gene sets by STAT5 and provides novel insights into STAT5-dependent mammary physiology.

Effect of 17β-estradiol on milk production, hormone secretion, and mammary gland gene expression in dairy cows

Estradiol inhibits milk production in dairy cows. The present study evaluated the effect of 17β-estradiol (E2) injections on prolactin (PRL) secretion and the mammary gland response to this hormone. Eight mid-lactation cows were used in a crossover design. During each experimental period, the cows were injected daily with either E2 (2.5 mg) or soy oil (2.5 mL; control) for 7 d. For each period, blood and milk samples were collected from d -4 to 14 (relative to the first injection) to measure PRL, insulin-like growth factor-1, and cortisol concentrations. In addition, blood samples were collected during morning milking on d -4, 2, and 7 to determine the milking-induced PRL release. Mammary gland biopsies were collected on the last day of injections. Milk fat samples were collected from d 1 to 7 and on d 14. The mRNA levels of genes encoding proteins related to mammary activity (α-lactalbumin, β-casein, and acetyl-coenzyme A carboxylase), apoptosis (Bax, Bcl2, and caspase-3), PRL receptors (PRLR; long and short forms), signal transducer and activator of transcription 5 (STAT5A and STAT5B), and suppressors of cytokine signaling (SOCS2 and SOCS3) were evaluated by real-time reverse transcription PCR using RNA extracted from milk fat and mammary biopsies. Milk production was decreased moderately (about 9%) by E2 injections during the treatment period. Estradiol injections increased basal PRL levels in serum and milk but did not affect milking-induced PRL release. Estradiol injections increased the plasma concentration of insulin-like growth factor-1 but did not affect cortisol concentration during the treatment period. In mammary tissue, the expression of Bcl2 was downregulated, whereas that of STAT5A and B and the Bax:Bcl2 mRNA ratio was higher during E2 injections. The total STAT5 protein content in mammary tissue was elevated by E2 injections. We found no significant difference observed for the other genes in mammary tissue or milk fat. The present data do not support the contention that E2 injections inhibit milk production by interfering with PRL signaling, but enhanced basal PRL concentration and STAT5 gene expression in mammary tissue.

Flax meal supplementation on the activity of antioxidant enzymes and the expression of oxidative stress- and lipogenic-related genes in dairy cows infused with sunflower oil in the abomasum

The effects of abomasal infusion of sunflower oil (SO) and flax meal (FM) supplementation on activity of the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) in blood and mammary tissue and mRNA abundance of oxidative stress- and lipogenic-related genes in mammary tissue of dairy cows were examined. Eight lactating Holstein cows were assigned to a double 4×4 Latin square design with a 2×2 factorial arrangement of treatments: (1) no FM in the diet (CON); (2) diet containing 13.7g/kg FM; (3) no FM in the diet and 250g SO/d infused in the abomasum; and (4) diet containing 13.7g/kg FM and 250g SO/d infused in the abomasum. mRNA abundance of Kappa-B1 in mammary tissue tended to decrease with FM supplementation, while mRNA abundance of SOD2 tended to increase when SO was infused in the absence of FM in the diet. Fatty acid synthase, lipoprotein lipase and stearoyl-coA desaturase gene expression tended to decrease with SO infusion when cows were fed CON. Infusion of SO decreased mRNA abundance of peroxisome proliferator-activated receptorγ1 in mammary tissue of cows fed CON and sterol regulatory element binding transcription factor 1 expression decreased with SO infusion. In conclusion, FM had no effect on oxidative stress-related gene expression in mammary tissue of cows that were infused with SO in the abomasum. Limited effects were observed on mRNA abundance of antioxidant-related genes in mammary tissue whereas abomasal infusion of SO reduced lipogenic-related gene expression.

Abstract 3262: Windows of susceptibility to endocrine disruptors and gene expression in mammary tissue

Abstract Breast cancer is hormonally driven. Endocrine disruptors (EDs) are exogenous substances that mimic or antagonize endogenous hormone action and disrupt endocrine system function. While EDs have been shown to modulate the risk for mammary tumors in animal models, their effects at biologically relevant doses and during critical developmental windows remain poorly understood. We sought to examine the individual and combinatorial effects of three common EDs, diethyl phthalate, methyl paraben and triclosan in rats exposed at five distinct developmental windows of susceptibility - prenatal, neonatal, prepubertal, pubertal and parous/nulliparous. Pathological features and whole genome expression of normal rat mammary tissues were profiled to provide both qualitative and quantitative biological effects of EDs. ED exposure at levels comparable to human exposure resulted in profound changes in both gross phenotypes (e.g. reproductive mortality and mammary gland morphology) as well as in molecular genome profiles. Specifically, ED exposure appeared to hamper normal breast development and resulted in increased mortality in the offspring, possibly due to reduced milk production. Whole genome expression profiling of mammary tissues revealed that in the course of development, the number of differentially expressed genes was lower in ED-exposed rats compared to controls, suggesting developmental delay or suppression by EDs. Pathway enrichment analysis indicated that several processes including inflammation, metabolism and development were disrupted by ED exposure. Taken together, our findings reveal novel mechanisms of ED action in mammary development and strongly suggest the importance of taking into account the timing of exposure when studying the relationship between environment and disease. Citation Format: Kalpana Gopalakrishnan, Qian Li, Yula Ma, Luca Lambertini, Jia Chen, Susan L. Teitelbaum, Fabiana Manservisi, Fiorella Belpoggi, Luciano Bua, Laura Falcioni. Windows of susceptibility to endocrine disruptors and gene expression in mammary tissue. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3262. doi:10.1158/1538-7445.AM2014-3262

Short communication: Effect of heat stress during the dry period on gene expression in mammary tissue and peripheral blood mononuclear cells

Heat stress (HT) during the dry period compromises mammary gland development, decreases future milk production, and impairs the immune status of dairy cows. Our objective was to evaluate the effect of cooling HT cows during the dry period on gene expression of the mammary gland and peripheral blood mononuclear cells (PBMC). Cows were dried off 46d before their expected calving and assigned to 2 treatments, HT or cooling (CL). Cows in the CL group were cooled with sprinklers and fans whereas HT cows were not. After parturition, all cows were housed in a freestall barn with cooling. The PBMC were isolated at dry-off and at −20, 2, and 20d relative to calving from a subset of cows (HT, n=9; CL, n=10), and mammary biopsies were taken at the same intervals (HT, n=7; CL, n=6) for RNA extraction. Gene expression was assessed using a custom multiplex gene expression assay based on traditional reverse transcription-PCR. Genes involved in prolactin (PRL) signaling [PRL receptor long form, PRL receptor short form, suppressor of cytokine signaling (SOCS)2, SOCS3, IGF2, IGF binding protein 5, and cyclin D1], fatty acid metabolism (acetyl-CoA carboxylase α (ACACA) and lipoprotein lipase (LPL)], and IGF1 were evaluated in mammary tissue, and genes related to fatty acid metabolism [ACACA, fatty acid synthase (FASN), and LPL], cytokine production [IL6, IL8, and tumor necrosis factor (TNF)], and IGF1 were evaluated in PBMC. No differences were observed in PRL signaling or fatty acid metabolism gene expression in the mammary gland. In PBMC, HT cows had greater mRNA expression of IGF1 and TNF during the transition period relative to CL and upregulated IL8 and downregulated FASN mRNA expression at 2d relative to calving. We conclude that cooling HT cows during the dry period alters expression of genes involved in cytokine production and lipid metabolism in PBMC.

Reproductive Experience Alters Prolactin Receptor Expression in Mammary and Hepatic Tissues in Female Rats1

Recent studies have reported that reproductive experience in female rats alters prolactin (PRL) receptor gene expression in the brain as well as neural sensitivity to PRL. Given PRL's actions in nonneural tissues, that is, mammary tissue and liver, it was asked whether reproductive experience may also alter prolactin receptor (Prlr) gene expression in these tissues. Groups of age-matched female rats were generated with varying reproductive histories. Separate groups of primiparous (first lactation) and multiparous (second lactation) had mammary tissue and liver samples collected on Day 3 or 10 of lactation. A fifth group raised one litter to weaning and then resumed estrous cyclicity. This group and a final group of age-matched, virgin controls were killed on diestrus. Tissue was processed by quantitative PCR for expression rates of the long and short forms of Prlr mRNA as well as casein beta mRNA (mammary tissue only). Western blots were performed to quantify receptor protein content. Multiple lactations as well as lactation itself resulted in alterations in Prlr expression. Prlr gene expression in mammary tissue was increased in primiparous mothers compared with that in multiparous dams, whereas in the liver, Prlr expression was reduced during an initial lactation. In contrast, PRLR protein levels declined during lactation in mammary, but not hepatic, tissues. Overall, the results demonstrate that the prolactin receptor system is altered in nonneural tissues as a result of the female's reproductive history. The findings are discussed in the context of milk and bile production and PRL's possible role in breast cancer.

Immunomediator and milk protein gene expression in mammary tissue during endotoxin-induced mastitis

Mammary gland infection induces an inflammatory reaction which causes an increase of somatic cells in milk and activation of bacteriostatic enzymes and proteins in milk. It has been demonstrated that during spontaneously occurring subclinical mastitis the somatic milk cells, mainly macrophages, secrete cytokines, eicosanoids, acute phase proteins and other immunomediators. In contrast, the bacteriostatic protein lactoferrin is mainly secreted by mammary epithelial tissue, while major milk proteins like α-lactalbumin and κ-casein are down-regulated already during subclinical infection. In LPS-challenged quarters, tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β) and cyclooxygenase-2 expression increased to highest values at 3 h after LPS-challenge. Expression of lactoferrin, lysozyme, inducible nitric oxide synthase, l-selectin, NFκB and of the apoptotic factors caspase-3, caspase-7 and Fas (FS-7 associated surface antigen) was elevated and peaked at 6 h after challenge. No significant increase in expression of platelet-activating factor, acethylhydrolase, 5-lipoxygenase, β-integrin and insulin-like growth factor 1 was found. In the control quarters, only TNF-α, IL-1β and lysozyme increase slightly and much less than in LPS-challenged quarters whereas the other parameters tested did not change significantly. Gene expression of major milk proteins did not change significantly in response to the LPS challenge (αS1-casein (CN), αS2-CN, β-CN and β-lactoglobulin) except for α-lactalbumin which decreased in LPS-treated and control quarters and for κ-CN which decreased in the LPS-treated quarters. In conclusion, gene expression of the majority albeit not all inflammatory factors changed within hours of LPS challenge. Decreased gene expression of α-lactalbumin and κ-CN may reduce milk yield and suitability for cheese production.

Dietary lipotrope-mediated mammary carcinogenesis in female rats

Dietary deficiency of lipotropes is known to be carcinogenic in itself. This study examined 1) the susceptibility of female rats previously exposed to lipotrope-modified diets to nitrosomethylurea (NMU)-induced mammary carcinogenesis; and 2) how lipotrope-modified diets modulate DNA methylation and gene expression in mammary tissue. Female rats (36 d of age) were assigned to one of the following groups: control-synthetic diet (CSD), containing all required lipotropes; methyl-deficient diet (MDD), lacking all lipotropes; and methyl-additive diet (MAD), containing 1.5-fold the amount of each lipotrope as in CSD. Rats were injected with NMU after a 2-wk dietary treatment period, and 2 d after NMU administration all treatment groups were fed CSD for the remaining experimental period. Mammary tissues were collected from rats just prior to NMU administration. The MDD group had the shortest latency period. The number of tumors and tumor volume were greatest in the MDD rats. Dietary modification of lipotropes altered the DNA methylation pattern of ornithine decarboxylase gene. DNA was significantly hypomethylated in mammary tissues of the MDD rats. These results suggest that dietary deficiency of lipotropes led to changes in DNA methylation and enhanced NMU-induced mammary carcinogenesis.

MMTV as a model for gene expression in mammary tissue.

Studies on mechanisms of gene regulation in mammalian cells have been greatly aided by the use of viral model systems. The small-to-intermediate sized DNA viruses (SV40 and Adenovirus) have been particularly useful because their genomes are relatively simple and they rely to a great extent on components of the host cell transcription apparatus for expression of their genetic information. Members of the retrovirus family, although studied primarily because of their oncogenic activity, offer an additional utility as models of gene regulation. When integrated in the host cell chromosome, these viruses adopt a low copy template structure (provirus) more similar to host cell genes than that of the high copy DNA viruses.KeywordsGlucocorticoid ReceptorLong Terminal RepeatMouse Mammary Tumor VirusNucleosome PositionHypersensitive SiteThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.