Most of the development and functional differentiation in the mammary gland occur after birth. Epigenetics is defined as the stable alterations in gene expression potential that arise during ...development and proliferation. Epigenetic changes are mediated at the biochemical level by the chromatin conformation initiated by DNA methylation, histone variants, post-translational modifications of histones, non-histone chromatin proteins, and non-coding RNAs. Epigenetics plays a key role in development. However, very little is known about its role in the developing mammary gland or how it might integrate the many signalling pathways involved in mammary gland development and function that have been discovered during the past few decades. An inverse relationship between marks of closed (DNA methylation) or open chromatin (DnaseI hypersensitivity, certain histone modifications) and milk protein gene expression has been documented. Recent studies have shown that during development and functional differentiation, both global and local chromatin changes occur. Locally, chromatin at distal regulatory elements and promoters of milk protein genes gains a more open conformation. Furthermore, changes occur both in looping between regulatory elements and attachment to nuclear matrix. These changes are induced by developmental signals and environmental conditions. Additionally, distinct epigenetic patterns have been identified in mammary gland stem and progenitor cell sub-populations. Together, these findings suggest that epigenetics plays a role in mammary development and function. With the new tools for epigenomics developed in recent years, we now can begin to establish a framework for the role of epigenetics in mammary gland development and disease.
Mastitis is among the main reasons women cease breastfeeding. In farm animals, mastitis results in significant economic losses and the premature culling of some animals. Nevertheless, the effect of ...inflammation on the mammary gland is not completely understood. This article discusses the changes to DNA methylation in mouse mammary tissue caused by lipopolysaccharide-induced inflammation after in vivo intramammary challenges and the differences in DNA methylation between 1
st
and 2
nd
lactations. Lactation rank induces 981 differential methylations of cytosines (DMCs) in mammary tissue. Inflammation in 1
st
lactation compared to inflammation in 2
nd
lactation results in the identification of 964 DMCs. When comparing inflammation in 1
st
vs. 2
nd
lactations with previous inflammation history, 2590 DMCs were identified. Moreover, Fluidigm PCR data show changes in the expression of several genes related to mammary function, epigenetic regulation, and the immune response. We show that the epigenetic regulation of two successive physiological lactations is not the same in terms of DNA methylation and that the effect of lactation rank on DNA methylation is stronger than that of the onset of inflammation. The conditions presented here show that few DMCs are shared between comparisons, suggesting a specific epigenetic response depending on lactation rank, the presence of inflammation, and even whether the cells had previously suffered inflammation. In the long term, this information could lead to a better understanding of the epigenetic regulation of lactation in both physiological and pathological conditions.
Abbreviations: RRBS, reduced representation bisulphite sequencing; RT-qPCR, real-time quantitative polymerase chain reaction; MEC, mammary epithelial cells; MaSC, mammary stem cell; TSS, transcription start site; TTS, transcription termination site; UTR, untranslated region; SINE, short interspersed nuclear element; LINE, long interspersed nuclear element; CGI, CpG island; DEG, differentially expressed gene; DMC, differentially methylated cytosine; DMR, differentially methylated region; GO term, gene ontology term; MF, molecular function; BP, biological process
Mastitis is among the main reasons women cease breastfeeding, which leads to them supplementing breast milk with artificial formula. In farm animals, mastitis results in significant economic losses ...and the premature culling of some animals. Nevertheless, researchers do not know enough about the effect of inflammation on the mammary gland. This article discusses the changes to DNA methylation in mouse mammary tissue caused by lipopolysaccharide-induced inflammation (4 h post-injection of lipopolysaccharide). We analysed the expression of some genes related to mammary gland function, epigenetic regulation, and the immune response. The analysis focused on three comparisons: inflammation during the first lactation, inflammation during second lactation with no history of inflammation, and inflammation during second lactation with previous inflammation. We identified differentially methylated cytosines (DMCs), differentially methylated regions (DMRs), and some differentially expressed genes (DEGs) for each comparison. The three comparisons shared some DEGs; however, few DMCs and only one DMR were shared. These observations suggest that inflammation is one of several factors affecting epigenetic regulation during successive lactations. Furthermore, the comparison between animals in second lactation with and without inflammation, with no inflammation history during first lactation showed a different pattern compared to the other conditions in this experiment. This indicates that inflammation history plays an important role in determining epigenetic changes. The data presented in this study suggest that lactation rank and previous inflammation history are equally important when explaining mammary tissue gene expression and DNA methylation changes.
Abbreviations: RRBS, reduced representation bisulfite sequencing; RT-qPCR, real-time quantitative polymerase chain reaction; MEC, mammary epithelial cells; TSS, transcription start site; TTS, transcription termination site; UTR, untranslated region; SINE, short interspersed nuclear element; LINE, long interspersed nuclear element; CGI, CpG island; DEG, differentially expressed gene; DMC, differentially methylated cytosine; DMR, differentially methylated region; GO term, gene ontology term; MF, molecular function; BP, biological process
It has been clearly demonstrated that the maternal nutritional status during pregnancy and lactation has long-term effects on offspring health. In mammals, milk represents the first maternal support ...provided to the newborns so that its composition may play a major role in long-term programming. We therefore assessed the effects of maternal high-fat/high-sugar obesogenic (OD) or control (CD) diets on offspring growth and adiposity in the rabbit. Between 7 and 20 wk of age, the BW gain of OD milk-fed rabbits was higher than that of CD milk-fed rabbits ( < 0.05). Body fat mass measurements at 21 wk of age revealed a significant increase in body adiposity as a function of milk ingested during the neonatal period, in both female and male offspring ( < 0.05). A marked weight gain difference was observed according to the milk in both female and male offspring. Moreover, we investigated the composition in major proteins and leptin levels in milk from OD or CD diet-fed dams. Liquid chromatography-mass spectrometry analysis of individual CD skimmed milk samples enabled identification and quantification of the rabbit main milk proteins and of their main phosphorylated isoforms at 2 different stages of lactation (3 and 10 d). Here we show that the OD diet induced a reduction in the whey acidic protein content concomitantly with both an increase in serum albumin and lactoferrin contents and in the phosphorylated isoforms of the main milk proteins. Furthermore, a sharp rise in leptin levels was observed in the milk of OD diet-fed dams on Day 10 of lactation when compared with CD diet animals ( < 0.05). Taken together, these findings provide evidence that lactation is a critical window of development during which exposure to a deleterious diet is highly detrimental to long-term outcomes. Moreover, these insights suggest that it may be possible to prevent at least some of the adverse effects of inadequate maternal nutrition on the long-term metabolic outcomes of the offspring through nutritional interventions applied during the lactation period.
Alterations to the metabolic endocrine environment during early life are crucial to mammary gland development. Among these environmental parameters, the initial nutritional event after birth is the ...consumption of milk, which represents the first maternal support provided to mammalian newborns. Milk is a complex fluid that exerts effects far beyond its immediate nutritional value. The present study, therefore, aimed to determine the effect of the nutritional changes during the neonatal and prepubertal periods on the adult mammary phenotype. Newborn rabbits were suckled by dams fed a high-fat/high-sugar obesogenic (OD) or a control (CON) diet and then subsequently fed either the OD or CON diets from the onset of puberty and throughout early pregnancy. Mammary glands were collected during early pregnancy (Day 8 of pregnancy). Rabbits fed with OD milk and then subjected to an OD diet displayed an abnormal development of the mammary gland: the mammary ducts were markedly enlarged (P < 0.05) and filled with abundant secretory products. Moreover, the alveolar secretory structures were disorganized, with an abnormal aspect characterized by large lumina. Mammary epithelial cells contained numerous large lipid droplets and exhibited fingering of the apical membrane and abnormally enlarged intercellular spaces filled with casein micelles. Leptin has been shown to be involved in modulating several developmental processes. We therefore analyzed its expression in the mammary gland. Mammary leptin mRNA was strongly expressed in rabbits fed with OD milk and subjected to an OD diet by comparison with the CON rabbits. Leptin transcripts and protein were localized in the epithelial cells, indicating that the increase in leptin synthesis occurs in this compartment. Taken together, these findings suggest that early-life nutritional history, in particular through the milking period, can determine subsequent mammary gland development. Moreover, they highlight the potentially important regulatory role that leptin may play during critical early-life nutritional windows with respect to long-term growth and mammary function.
Les données récentes indiquent que plus d’un adulte sur trois dans le monde, soit 1,46 milliard de personnes, est obèse ou en surcharge pondérale. Cette obésité va avoir des conséquences délétaires ...sur la santé des individus et en particulier sur l’allaitement des femmes ainsi que sur la composition de leur lait. À plus long terme, une alimentation maternelle inappropriée peut influer sur le développement du fœtus et la santé de l’enfant, puis de l’adulte. Dans des modèles animaux, il a été montré que l’engraissement excessif, en particulier pendant la puberté, peut induire des perturbations du développement de la glande mammaire résultant à une baisse des capacités laitières. L’objectif de notre étude est de déterminer des fenêtres critiques de sensibilité à l’alimentation en ciblant la période néonatale via le lait, pouvant impacter la croissance et le développement mammaire.
Dans ces études, nous avons utilisé des lapines nourries avec un régime obésogène (OB) enrichi en sucres (+300 %) et en graisses (+300 %). Dans une première cohorte, le régime a été commencé dès la puberté. Les conséquences d’une adiposité élevée sur la lactation ont été étudiées sur une nouvelle cohorte de lapines suivant le même protocole expérimental. Afin de tester l’effet du lait sur la descendance, nous avons allaité les lapereaux dès leur naissance avec un lait issu de mères ayant été alimentées soit avec un régime OB, soit avec un régime témoin. À la puberté, ces lapereaux ont eux-mêmes été soumis soit à un régime OB, soit au régime témoin. La croissance et l’adiposité de ces lapereaux ont été suivies. Les glandes mammaires de ces animaux ont été analysées sur le plan morphologique et fonctionnel.
Trois mois après le début du régime, le poids des lapines ayant reçu le régime OB était significativement plus élevé. Cet écart résultait d’une difference dans l’adiposité entre les deux groupes d’animaux. Les glandes mammaires des lapines OB présentaient un développement modifié tant sur le plan morphologique que sur le plan fonctionnel : les canaux mammaires étaient remplis avec des produits denses. La proportion d’épithélium sécrétoire était plus importante dans le tissu mammaire de ces animaux et l’analyse histologique de l’ épithélium mammaire a révélé une abondante accumulation de protéines du lait et de lipides, rappelant un phénotype de fin de gestation. De plus, la composition du lait issu de ces lapines était altérée. Lorsque ce lait a été administré à des lapereaux, ceux-ci ont présenté une prédisposition à l’obésité caractérisée par une adiposité plus élevée. Lorsqu’il a été combiné à un régime obésogène, il a induit un développement mammaire anormal dès les stades précoces de la gestation : désorganisation des structures alvéolaires, présence de canaux hyper-dilatés contenant des protéines du lait et des globules gras, alteration de la polarité des cellules épithéliales.
Nos résultats démontrent qu’une alimentation enrichie en graisses et en sucres, commencée dès la puberté tend à favoriser un développement précoce et donc anormal de la glande mammaire. Ce type d’alimentation altère également la qualité du lait lors de la lactation. Le fait de consommer ce lait à la composition altérée aurait ainsi comme conséquence d’induire une prédisposition à une hyperadiposité et de potentialiser les effets néfastes d’un régime obésogène au niveau du développement mammaire chez la descendance.
Streptozotocin is a monofunctional alkylating agent that induces diabetes in a large variety of mammals. While multiple low doses of streptozotocin induce immune-mediated diabetes, a single high dose ...of streptozotocin causes a strictly toxic diabetes. Among mouse strains, non-obese diabetic (NOD) mice are characterized by an extreme susceptibility to high dose of streptozotocin-induced diabetes whereas C3H/Or mice are particularly resistant. We hypothesized that NOD genes involved in high dose streptozotocin-induced diabetes could be also involved in the autoimmune destruction of pancreatic beta cells that characterizes this mouse strain which is a model of Type 1 diabetes.
We carried out a whole genome linkage scan on a population of (C3H/Or x NOD) x NOD backcross 1 mice in order to identify the genetic loci involved in NOD susceptibility to high dose of streptozotocin-induced diabetes.
Two loci, in chromosome 9 (D9Mit135 marker, 48 cM) and in chromosome 11 (D11Mit286 marker, 52 cM), were associated with NOD susceptibility to high dose streptozotocin-induced diabetes, the latter being co-localized with the autoimmune diabetes-predisposing idd4 locus. Moreover, we report here that C57BL/6 mice deficient in Nitric Oxide Synthase 2 were as sensitive as wild-type C57BL/6 mice to high dose streptozotocin-induced diabetes.
Although the Nitric Oxide Synthase 2 ( Nos2) gene, localized at 45.6 cM in chromosome 11, is a good candidate gene, our results suggest that Nitric Oxide Synthase 2 activation might not be a crucial event for streptozotocin-induced destruction of pancreatic beta cells.
It has been clearly demonstrated that the maternal nutritional status during pregnancy and lactation has long-term effects on offspring health. In mammals, milk represents the first maternal support ...provided to the newborns so that its composition may play a major role in long-term programming. We therefore assessed the effects of maternal high-fat/ high-sugar obesogenic (OD) or control (CD) diets on offspring growth and adiposity in the rabbit. Between 7 and 20 wk of age, the BW gain of OD milk-fed rabbits was higher than that of CD milk-fed rabbits (P < 0.05). Body fat mass measurements at 21 wk of age revealed a significant increase in body adiposity as a function of milk ingested during the neonatal period, in both female and male offspring (P < 0.05). A marked weight gain difference was observed according to the milk in both female and male offspring. Moreover, we investigated the composition in major proteins and leptin levels in milk from OD or CD diet-fed dams. Liquid chromatography-mass spectrometry analysis of individual CD skimmed milk samples enabled identification and quantification of the rabbit main milk proteins and of their main phosphorylated isoforms at 2 different stages of lactation (3 and 10 d). Here we show that the OD diet induced a reduction in the whey acidic protein content concomitantly with both an increase in serum albumin and lactoferrin contents and in the phosphorylated isoforms of the main milk proteins. Furthermore, a sharp rise in leptin levels was observed in the milk of OD diet-fed dams on Day 10 of lactation when compared with CD diet animals (P < 0.05). Taken together, these findings provide evidence that lactation is a critical window of development during which exposure to a deleterious diet is highly detrimental to long-term outcomes. Moreover, these insights suggest that it may be possible to prevent at least some of the adverse effects of inadequate maternal nutrition on the long-term metabolic outcomes of the offspring through nutritional interventions applied during the lactation period.
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