Fabienne Le Provost

An integrative multi-omic study of bovine milk production and composition in two feed restriction trials to understand adaptations to negative energy balance and to develop non-invasive biomarkers for monitoring animal energy status.

Lactation is an essential process for mammals. In sheep, the R96C mutation in suppressor of cytokine signaling 2 (SOCS2) protein is associated with greater milk production and increased mastitis sensitivity. To shed light on the involvement of R96C mutation in mammary gland development and lactation, we developed a mouse model carrying this mutation (SOCS2KI/KI). Mammary glands from virgin adult SOCS2KI/KI mice presented a branching defect and less epithelial tissue, which were not compensated for in later stages of mammary development. Mammary epithelial cell (MEC) subpopulations were modified, with mutated mice having three times as many basal cells, accompanied by a decrease in luminal cells. The SOCS2KI/KI mammary gland remained functional; however, MECs contained more lipid droplets versus fat globules, and milk lipid composition was modified. Moreover, the gene expression dynamic from virgin to pregnancy state resulted in the identification of about 3000 differentially expressed genes specific to SOCS2KI/KI or control mice. Our results show that SOCS2 is important for mammary gland development and milk production. In the long term, this finding raises the possibility of ensuring adequate milk production without compromising animal health and welfare.

Background: In dairy cows, diet is one factor that can affect their milk production and composition. However, the effect of feed restriction on milk miRNome has not yet been described. Indeed, milk is the body fluid with the highest RNA concentration, which includes numerous microRNA. Its presence in the four different milk fractions, whole milk, fat globules, mammary epithelial cells and extracellular vesicles, is still poorly documented. This study aimed to describe the effects of different feed restrictions on the miRNome composition of different milk fractions. Results: Two feed restrictions were applied to lactating dairy cows, one of high intensity and one of moderate intensity. 2,896 mature microRNA were identified in the different milk fractions studied, including 1,493 that were already known in the bovine species. Among the 1,096 microRNA that were sufficiently abundant to be informative, the abundance of 1,027 of them varied between fractions: 36 of those were exclusive to one milk fraction. Feed restriction affected the abundance of 155 microRNA, with whole milk and milk extracellular vesicles being the most affected, whereas milk fat globules and exfoliated mammary epithelial cells were little or not affected at all. The high intensity feed restriction led to more microRNA variations in milk than moderate restriction. The target prediction of known microRNA that varied under feed restriction suggested the modification of some key pathways for lactation related to milk fat and protein metabolisms, cell cycle, and stress responses. Conclusions: This study highlighted that the miRNome of each milk fraction is specific, with mostly the same microRNA composition but with variations in abundance between fractions. These specific miRNomes were affected differently by feed restrictions, the intensity of which appeared to be a major factor modulating milk miRNomes. These findings offer opportunities for future research on the use of milk miRNA as biomarkers of energy status in dairy cows, which is affected by feed restrictions.

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.

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.

Optimizing rabbit does preparation during early life to improve reproductive performance is a major challenge for breeders. Does selected for reproduction have nutritional needs, which may not be supplied with the common practice of feed restriction during rearing in commercial rabbit production. Nutrition during early life was already known to influence metabolism, reproduction and mammary gland development later in life, in particular during pregnancy. The aim of this study was to analyze four different restriction feeding strategies during post-weaning and over the pubertal periods (high or moderate restriction feeding applied from 5 to 9 weeks of age and/or restricted or ad libitum over the following 3 weeks constituting the pubertal period). Unlike food intake, which remains regular, mean body weight gain was inversely proportional to the dietary restriction applied over the considered periods. The feeding strategies in place for the four groups have no effect on the reproductive parameters of the females, as opposed to certain metabolic parameters such as cholesterolemia, that vary with dietary intake. Furthermore, restriction programs have impacted mammary tissular structures at mid-pregnancy. The expression of lipid metabolism enzymes (Fatty acid synthase N and Stearoyl co-A desaturase) is also modified in mammary epithelial cells by the dietary strategies implemented. Moreover, milk gene expression, used as differentiation markers, indicates a better mammary epithelial development regarding further lactation, in the case of the less restrictive strategies during early life period, especially the higher feeding allowance. Our results highlight the importance of investigating feeding conditions of young female rabbits and nutrition in early life rearing, in order to provide specific recommendations for optimizing lactation and thus preventing neonatal mortality of the offspring.

The mammary gland undergoes important anatomical and physiological changes from embryogenesis through puberty, pregnancy, lactation and involution. These steps are under the control of a complex network of molecular factors, in which epigenetic mechanisms play a role that is increasingly well described. Recently, studies investigating epigenetic modifications and their impacts on gene expression in the mammary gland have been performed at different physiological stages and in different mammary cell types. This has led to the establishment of a role for epigenetic marks in milk component biosynthesis. This review aims to summarize the available knowledge regarding the involvement of the four main molecular mechanisms in epigenetics: DNA methylation, histone modifications, polycomb protein activity and non-coding RNA functions.

The mouse transplantation model remains the most relevant methodology to assess the functional capacities of mammary cells and is particularly appropriate for investigations regarding mammary stem ... METHOD SUMMARY A new approach to estimate the species specificity of xenografts is described. This method is based on PCR amplification of genomic DNA samples extracted from mammary tissue that has...

In recent years, the increasing use of Next Generation Sequencing technologies to explore the genome has generated large quantities of data. For microRNAs, more and more publications have described several thousand sequences, all species included. In order to obtain a detailed description of microRNAs from the literature for three ruminant species (bovine, caprine and ovine), a new database has been created: RumimiR. To date, 2,887, 2,733 and 5,095 unique microRNAs of bovine, caprine and ovine species, respectively, have been included. In addition to the most recent reference genomic position and sequence of each microRNA, this database contains details on the animals, tissue origins and experimental conditions available from the publications. Identity with human or mouse microRNA is mentioned. The RumimiR database enables data filtering, the selection of microRNAs being based on defined criteria such as animal status or tissue origin. For ruminant studies, RumimiR supplements the widely used miRBase database by browsing and filtering using complementary criteria, and the integration of all published sequences described as novel. The principal goal of this database is to provide easy access to all ruminant microRNAs described in the literature.