Gland Lactation Research Articles

Molecular complexity of mammary glands development: a review of lactogenic differentiation in epithelial cells

The mammary gland is a dynamic organ with various physiological processes like cellular proliferation, differentiation, and apoptosis during the pregnancy-lactation-involution cycle. It is essential to understand the molecular changes during the lactogenic differentiation of mammary epithelial cells (MECs, the milk-synthesizing cells). The MECs are organized as luminal milk-secreting cells and basal myoepithelial cells (responsible for milk ejection by contraction) that form the alveoli. The branching morphogenesis and lactogenic differentiation of the MECs prepare the gland for lactation. This process is governed by many molecular mediators including hormones, growth factors, cytokines, miRNAs, regulatory proteins, etc. Interestingly, various signalling pathways guide lactation and understanding these molecular transitions from pregnancy to lactation will help researchers design further research. Manipulation of genes responsible for milk synthesis and secretion will promote augmentation of milk yield in dairy animals. Identifying protein signatures of lactation will help develop strategies for persistent lactation and shortening the dry period in farm animals. The present review article discusses in details the physiological and molecular changes occurring during lactogenic differentiation of MECs and the associated hormones, regulatory proteins, miRNAs, and signalling pathways. An in-depth knowledge of the molecular events will aid in developing engineered cellular models for studies related to mammary gland diseases of humans and animals.

Comparative Histological, Histochemical and Immunohistochemical Examination of the Cranial and Caudal Gland of Each Lactating Mammary Gland Quarter of One Humped She Camel (Camelus Dromedarius)

Studying the complete histological picture of the camel mammary gland plays a major role in understanding the variations in camel milk yield and composition. Therefore, the present study aimed to give more detailed description of the structure of the lactating dromedary she-camel mammary gland. The whole udder of 9 healthy lactating she-camels (Camelus dromedaries) was dissected of the fresh carcasses just after slaughtering. Sections were dissected out and fixed in 10% neutral buffer formalin for histological and immunohistochemical examination. Other parts were fixed in glutaraldehyde for ultrastructure examination. The gross examination of the camel mammary gland revealed that it is composed of four teats. Each teat ended by two separate orifices: cranial and caudal. Each orifice leads to a separate streak canal, separate gland cistern and separate lactating glands, cranial gland, and caudal gland. The examination of the two gland cisterns pointed out that the cranial gland cistern was significantly wider than the caudal one. The histological examination of the cranial and caudal gland revealed that the caudal gland comprised an abundant amount of interstitial connective tissue than the cranial gland. In contrast, the alveoli of the cranial gland were larger in size, more active, and stretched with secretion. The ultrastructure examination showed differences in structure and activity between the secretory cells of the two glands. The immunohistochemical examination of the lactating mammary gland showed strong expression of CK5/6 in both glands. On the other hand, the two glands showed different reactions to CK8/18, Estrogen Receptors and Progesterone Receptors (PR).

Immunomodulatory Properties of Human Breast Milk: MicroRNA Contents and Potential Epigenetic Effects.

Infants who are exclusively breastfed in the first six months of age receive adequate nutrients, achieving optimal immune protection and growth. In addition to the known nutritional components of human breast milk (HBM), i.e., water, carbohydrates, fats and proteins, it is also a rich source of microRNAs, which impact epigenetic mechanisms. This comprehensive work presents an up-to-date overview of the immunomodulatory constituents of HBM, highlighting its content of circulating microRNAs. The epigenetic effects of HBM are discussed, especially those regulated by miRNAs. HBM contains more than 1400 microRNAs. The majority of these microRNAs originate from the lactating gland and are based on the remodeling of cells in the gland during breastfeeding. These miRNAs can affect epigenetic patterns by several mechanisms, including DNA methylation, histone modifications and RNA regulation, which could ultimately result in alterations in gene expressions. Therefore, the unique microRNA profile of HBM, including exosomal microRNAs, is implicated in the regulation of the genes responsible for a variety of immunological and physiological functions, such as FTO, INS, IGF1, NRF2, GLUT1 and FOXP3 genes. Hence, studying the HBM miRNA composition is important for improving the nutritional approaches for pregnancy and infant’s early life and preventing diseases that could occur in the future. Interestingly, the composition of miRNAs in HBM is affected by multiple factors, including diet, environmental and genetic factors.

Intracellular Ca2+ Signaling and Calcium Release-Activated Calcium Modulator ORAI1 Are Associated With CD4+ T Lymphocytes in Dairy Cows.

The nutritional status of dairy cows and the metabolism of specific nutrients are critical regulators of immune cell function. Around the time of parturition, mobilization of body lipid and muscle helps compensate for the decrease in nutrient intake and the increased requirements of the mammary gland for lactation. An end-result of these processes is the marked increase in circulating concentrations of fatty acids (FA), which are a major risk factor for immune dysfunction. In food animal species such as dairy cows, any disturbance in nutritional or immunological homeostasis leads to deleterious feedback loops that can further risk health, efficiency of nutrient use, and compromise availability of safe and nutritious dairy foods for humans. Despite substantial progress with respect to regulation of innate immunity, such knowledge for adaptive immunity is scarce. To help bridge this gap in knowledge, we sought to study the role of calcium release-activated calcium modulator ORAI1 activation in T cells systemic immune function in vivo. CD4+ T cells were isolated from peripheral blood of dairy cows diagnosed as healthy or with ketosis, a common metabolic disorder of FA metabolism. Results revealed that levels of intracellular Ca2+ and reactive oxygen species (ROS) along with the abundance of store-operated Ca2+ entry (SOCE) moiety increased during ketosis. Further, plasma concentrations of inflammatory cytokines were elevated, the balance of Th17/Treg cells was disrupted, mitochondrial function impaired, and the abundance of mitophagy-related proteins in CD4+ T cells altered during ketosis. Molecular characterization of the direct effects of FA was evaluated in CD4+ T cells isolated from the spleen of 1-day-old calves. Enhanced supply of FA increased intracellular Ca2+ and ROS concentrations, upregulated the abundance of proteins associated with mitochondrial dynamics and ORAI1. Intermediates of mitophagy accumulated and the balance of Th17/Treg cells also was affected by the supply of FA. These negative effects were attenuated by silencing or inhibition of ORAI1 in CD4+ T cells. Together, data indicated that physiological states that lead to increases in systemic concentrations of FA could impact adaptive immunity negatively through ORAI1 regulated intracellular Ca2+, ROS balance, and increased effector functions of Th17 cells.

Ultrastructure of the mammary gland in lactating Saanen goats

The secretory cells of the mammary gland are very diverse in their size, shape, and structure. These are highly specialized cells adapted to the synthesis, accumulation, storage, and excretion of milk. The main feature of glandular cells is the presence of secreted substances. Electron microscopy shows that substances of different secretory cells types have various electron densities. Occasionally, two or more types of secretory granules differing in size and composition can be found in one cell. The material for the morphological study was small (2‐4 mm) tissue samples of goat lactating mammary glands. Pieces were taken from deeper areas of the breast parenchyma. Ultrathin sections with a 50‐70 nm thickness were obtained for electron microscopic analysis using Leica UC7 ultramicrotome. Electron microscopy was performed using a JEOL JEM 1011 microscope. Micrographs were obtained using a Morada camera (Digital Imaging Solutions Inc.). The secretion of the mammary gland occurs according to the apocrine type. Picture of the lactocytes' cytoplasm apical part separation into the lumen of the alveoli is often observed on semi‐thin sections. Thus, numerous droplets of material secreted by lactocytes are found in the lumen of the alveoli. The shape of lactocytes in the lactating mammary gland is close to cubic. The height of the epithelial layer, generally, is 10‐12 microns (µm). The milk ducts, like the alveoli, are dilated. Most of them contain a significant amount of osmiophilic material secreted by the cells of the mammary alveoli. The cells of the epithelium of the single‐layered ducts have a flattened or cuboidal shape. The diameter of the ducts varies noticeably depending on location in the gland parenchyma. The intralobular milk ducts are 10‐20 µm in diameter. At the same time, the larger interlobular ducts reach a diameter of 40‐50 microns and are located in groups in the connective tissue stroma of the mammary gland. Notably, an increase in the mammary alveoli mass in the lactating gland is associated with a decrease in connective tissue volume and the almost complete disappearance of fat cells groups from it. The milk ducts of the teat zone in the lactating gland are strongly expanded, round, or slightly oval‐shaped in cross‐section with a diameter of 30‐60 microns. In contrast to the deeper zones of the mammary gland, we were unable to detect secreted substances in the dilated milk ducts near the teat area. The milk ducts we studied are located close to the most terminal sections of the mammary gland excretory system (gland cistern and teat canal). The milk is inevitably washed out from the ducts during chemical fixation and subsequent stages of objects preparation for histological analysis.

Transcriptome profiling of the nonlactating mammary glands of dairy goats reveals the molecular genetic mechanism of mammary cell remodeling

The mammary gland redevelops to the prepregnancy state during involution, which shows the mammary cells have the characteristics of remodeling. The rapidity and degree of mammary gland involution vary across species (e.g., between model organism mice and dairy livestock). However, the molecular genetic mechanism of involution and remodeling of goat mammary gland has not yet been clarified. This work investigated the structural changes and transcriptome characteristics of the mammary gland tissue of nonlactating dairy goats during the late lactation (LL), the dry period (DP), and late gestation (LG). Terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining revealed significant changes in the structure of the nonlactating goat mammary gland, and obvious cell apoptosis occurred at LL and DP. Sequencing identified 1,381 genes that are differentially expressed in mammary gland tissue at the 3 developmental stages. Genes related to cell growth, apoptosis, immunity, nutrient transport, synthesis, and metabolism exhibited adaptive transcriptional changes to meet the needs of a new set of mammary gland lactation functions. The significant enrichment of Gene Ontology terms such as humoral immune response, complement activation, and neutrophil-mediated immunity indicates that the innate immune system plays an important role in maintaining the health of degenerative mammary glands and eliminating apoptotic cells. The peroxisome proliferator-activated receptor signaling pathway plays an important regulatory role in lipid metabolism, especially the adaptive changes in expression of genes encoded lipid transport and enzymes, which promote the formation of milk fat during the lactation. The mammary gland development gene module revealed that pregnancy hormone receptors, cell growth factors and their receptors, and genes encoding insulin-like growth factor binding proteins regulate the physiological process of mammary gland involution through adaptive transcriptional changes. Interestingly, ERBB4 was identified as the hub gene of the network that regulates mammary gland growth and development. Overexpression of ERBB4 in mammary epithelial cells cultured in vitro can reduce cell cycle arrest in G1/S phase and apoptosis by regulating the PI3K/Akt signaling pathway and promote the proliferation of mammary epithelial cells. The gene ERBB4 also affects the expression of genes that initiate mammary gland involution and promote mammary gland remodeling. These findings contribute to an in-depth understanding of the molecular mechanisms involved in mammary gland involution and remodeling.

Aberrant RET expression affects normal mammary gland post-lactation transition, enhancing cancer potential.

ABSTRACTRET is a receptor tyrosine kinase with oncogenic potential in the mammary epithelium. Several receptors with oncogenic activity in the breast are known to participate in specific developmental stages. We found that RET is differentially expressed during mouse mammary gland development: RET is present in lactation and its expression dramatically decreases in involution, the period during which the lactating gland returns to a quiescent state after weaning. Based on epidemiological and pre-clinical findings, involution has been described as tumor promoting. Using the Ret/MTB doxycycline-inducible mouse transgenic system, we show that sustained expression of RET in the mammary epithelium during the post-lactation transition to involution is accompanied by alterations in tissue remodeling and an enhancement of cancer potential. Following constitutive Ret expression, we observed a significant increase in neoplastic lesions in the post-involuting versus the virgin mammary gland. Furthermore, we show that abnormal RET overexpression during lactation promotes factors that prime involution, including premature activation of Stat3 signaling and, using RNA sequencing, an acute-phase inflammatory signature. Our results demonstrate that RET overexpression negatively affects the normal post-lactation transition.

Determination of Risk Factors for DCIS Ductal in-situ Carcinoma of Mammary Gland

Mammary gland is altered gland present in skin whose main function in females is that it acts as a lactating gland while in males it is non-functional. When the sample of tissue of mammary gland is observed under microscope it was found that there are 2 major types of tissues which are present. These tissues are epithelial and stromal tissues. The epithelial component of the breast consists of 2 major parts: terminal duct-lobular unit (TDLU) which performs the main secretory function during lactation, and large duct system which performs the function of collection and drainage of secretions; both are interconnected to each other. During the menstrual age of a female patient there are different number of tissues (connective and adipose) is present in the mammary gland these tissues mainly perform as the support system for the mammary gland. Acini, ducts, and lobule are enclosed in intralobular stroma and is mainly formed of connective tissue(loose) myxomatous stroma and some dispersed lymphocyte. The ductal carcinoma is very serious Pathology of breast which is now a major concern in developed country and to a certain extent in developing countries also. There are many major risk factors which are responsible for the same. Some may be of lifestyle based or of genetic based. The treatment for the disease is now possible with advancement in the medical science.
 Purpose for this review article is: Determine the cause affecting factors influencing factors and pathos physiological background of development of DCIS also to describe the gene involved in alteration of cell to enhance tumorous growth and the treatment therapy involved alongside the disease.

Mammary collagen is under reproductive control with implications for breast cancer

Mammographically-detected breast density impacts breast cancer risk and progression, and fibrillar collagen is a key component of breast density. However, physiologic factors influencing collagen production in the breast are poorly understood. In female rats, we analyzed gene expression of the most abundantly expressed mammary collagens and collagen-associated proteins across a pregnancy, lactation, and weaning cycle. We identified a triphasic pattern of collagen gene regulation and evidence for reproductive state-dependent composition. An initial phase of collagen deposition occurred during pregnancy, followed by an active phase of collagen suppression during lactation. The third phase of collagen regulation occurred during weaning-induced mammary gland involution, which was characterized by increased collagen deposition. Concomitant changes in collagen protein abundance were confirmed by Masson's trichrome staining, second harmonic generation (SHG) imaging, and mass spectrometry. We observed similar reproductive-state dependent collagen patterns in human breast tissue obtained from premenopausal women. SHG analysis also revealed structural variation in collagen across a reproductive cycle, with higher packing density and more collagen fibers arranged perpendicular to the mammary epithelium in the involuting rat mammary gland compared to nulliparous and lactating glands. Involution was also characterized by high expression of the collagen cross-linking enzyme lysyl oxidase, which was associated with increased levels of cross-linked collagen. Breast cancer relevance is suggested, as we found that breast cancer diagnosed in recently postpartum women displayed gene expression signatures consistent with increased collagen deposition and crosslinking compared to breast cancers diagnosed in age-matched nulliparous women. Using publicly available data sets, we found this involution-like, collagen gene signature correlated with poor progression-free survival in breast cancer patients overall and in younger women. In sum, these findings of physiologic collagen regulation in the normal mammary gland may provide insight into normal breast function, the etiology of breast density, and inform breast cancer risk and outcomes.

Effects of a High-Grain Diet With a Buffering Agent on Milk Protein Synthesis in Lactating Goats.

Chinese dairy industries have developed rapidly, providing consumers with high-quality sources of nutrition. However, many problems have also appeared during the development process, especially the low quality of milk. To improve milk quality, a large amount of concentrated feed is usually added to the diet within a certain period of time, which increases the milk production to a certain extent. However, long-term feeding with high-concentration feed can lead to subacute rumen acidosis. Therefore, the present study aimed to determine the effect of adding a buffer on subacute rumen acidosis, and the improvement of milk production and milk quality. We also aimed to study the mechanism of promoting mammary gland lactation. A total of 12 healthy mid-lactating goats were randomly divided into two groups, they were high-grain diet group (Control) and buffering agent group. To understand the effects of high-grain diets with buffers on amino acids in jugular blood and the effects of amino acids on milk protein synthesis, Milk-Testing™ Milkoscan 4000, commercial kits, and high-performance liquid chromatography (HPLC) measurements were integrated with the milk protein rate, the amino acid concentration in jugular venous blood samples, quantitative real-time PCR, comparative proteomics, and western blotting to study differentially expressed proteins and amino acids in mammary gland tissues of goats fed high-grain diets. Feeding lactating goats with buffering agent increased the percentage of milk protein in milk, significantly increased the amino acid content of jugular blood (p < 0.05), and increase the amino acid transporter levels in the mammary gland. Compared with the high-grain group, 2-dimensional electrophoresis technology, matrix-assisted laser desorption/ionization-time of flight/time of flight proteomics analyzer, and western blot analysis further verified that the expression levels of beta casein (CSN2) and lactoferrin (LF) proteins in the mammary glands of lactating goats were higher when fed a high-grain diets and buffers. The mechanism of increased milk protein synthesis was demonstrated to be related to the activation of mammalian target of rapamycin (mTOR) pathway signals.

A Robust Mammary Organoid System to Model Lactation and Involution-like Processes.

The mammary gland is a highly dynamic tissue that changes throughout reproductive life, including growth during puberty and repetitive cycles of pregnancy and involution. Mammary gland tumors represent the most common cancer diagnosed in women worldwide. Studying the regulatory mechanisms of mammary gland development is essential for understanding how dysregulation can lead to breast cancer initiation and progression. Three-dimensional (3D) mammary organoids offer many exciting possibilities for the study of tissue development and breast cancer. In the present protocol derived from Sumbal et al., we describe a straightforward 3D organoid system for the study of lactation and involution ex vivo. We use primary and passaged mouse mammary organoids stimulated with fibroblast growth factor 2 (FGF2) and prolactin to model the three cycles of mouse mammary gland lactation and involution processes. This 3D organoid model represents a valuable tool to study late postnatal mammary gland development and breast cancer, in particular postpartum-associated breast cancer. Graphic abstract: Mammary gland organoid isolation and culture procedures.

Proteomic Analysis Reveals Proteins and Pathways Associated with Lactation in Bovine Mammary Epithelial Cell-Derived Exosomes.

Milk-derived exosomes have been reported, which are involved in many biological processes. The exosomes derived from mammary glands are not known yet, and their relationship with mammary gland lactation and the origin of milk-derived exosomes are largely unclear. The present study aimed to investigate the proteome of exosomes derived from bovine mammary epithelial cells (BMECs) and compare them with milk-derived exosomes in the database. BMEC-derived exosomes were successfully separated from the culture supernatant of BMECs by a combined ultracentrifugation approach, and the purity of exosomes was identified by western blot analysis. Liquid chromatography with tandem mass spectrometry identified 638 proteins in BMEC-derived exosomes. The MS data were deposited into the PUBLIC repository ProteomeXchange, dataset identifier(s): https://www.iprox.org/page/PSV023.html;?url=1590961453176tKpa. Gene Ontology annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that these proteins were associated with specific biological processes and molecular functions of metabolism. Cross comparison of these proteins with the protein database of milk exosomes showed that 77 common expressed proteins (CEPs) were in both BMEC- and milk-derived exosomes. The KEGG pathway analysis for these CEPs showed that they were mainly involved in signaling pathways associated with milk biosynthesis in BMECs. Among these CEPs, six proteins have been previously reported to be associated with the lactation function. The western blot analysis detected that expression of these six proteins in BMEC-derived exosomes was increased after the stimulation of methionine and β-estradiol on BMECs. In summary, the proteome of BMEC-derived exosomes reveals that they are associated with milk biosynthesis in BMECs and might be a source of milk-derived exosomes.

Surgical management of gangrenous mastitis in a 3-year-old red Sokoto doe: A case report

Gangrenous mastitis was managed in a 3-year-old lactating doe by partial mastectomy following unsuccessful medical management. The patient had progressive swelling, hardened and dark discolouration of its left half udder. There was left supra-mammary lymphadenopathy and an open wound at the proximal portion of the affected gland. The right half of the udder appeared normal and was warm to touch. Culture and sensitivity test of a swab sample from the open wound at the base of the affected mammary gland revealed Staphylococcus and Streptococcus species which were sensitive to amoxicillin, cloxicillin, gentamicin and chlortetracycline, but resistant to penicillin and streptomycin. Mastectomy was indicated because of the chronic nature of the disease and the presence of gangrene. The procedure was carried out using a standard sterile technique, and a fenestrated drainage tube was placed. The patient recovered uneventfully and was discharged 14 days post-surgery. The management protocol used in this case demonstrated that unilateral mastectomy with good post-surgical management could be used for effective management of gangrenous mastitis without complications while still preserving the unaffected gland for future lactation.

Primary Mammary Organoid Model of Lactation and Involution.

Mammary gland development occurs mainly after birth and is composed of three successive stages: puberty, pregnancy and lactation, and involution. These developmental stages are associated with major tissue remodeling, including extensive changes in mammary epithelium, as well as surrounding stroma. Three-dimensional (3D) mammary organoid culture has become an important tool in mammary gland biology and enabled invaluable discoveries on pubertal mammary branching morphogenesis and breast cancer. However, a suitable 3D organoid model recapitulating key aspects of lactation and involution has been missing. Here, we describe a robust and straightforward mouse mammary organoid system modeling lactation and involution-like process, which can be applied to study mechanisms of physiological mammary gland lactation and involution as well as pregnancy-associated breast cancer.

Analyzing the genomic and transcriptomic architecture of milk traits in Murciano-Granadina goats

BackgroundIn this study, we aimed to investigate the molecular basis of lactation as well as to identify the genetic factors that influence milk yield and composition in goats. To achieve these two goals, we have analyzed how the mRNA profile of the mammary gland changes in seven Murciano-Granadina goats at each of three different time points, i.e. 78 d (T1, early lactation), 216 d (T2, late lactation) and 285 d (T3, dry period) after parturition. Moreover, we have performed a genome-wide association study (GWAS) for seven dairy traits recorded in the 1st lactation of 822 Murciano-Granadina goats.ResultsThe expression profiles of the mammary gland in the early (T1) and late (T2) lactation were quite similar (42 differentially expressed genes), while strong transcriptomic differences (more than one thousand differentially expressed genes) were observed between the lactating (T1/T2) and non-lactating (T3) mammary glands. A large number of differentially expressed genes were involved in pathways related with the biosynthesis of amino acids, cholesterol, triglycerides and steroids as well as with glycerophospholipid metabolism, adipocytokine signaling, lipid binding, regulation of ion transmembrane transport, calcium ion binding, metalloendopeptidase activity and complement and coagulation cascades. With regard to the second goal of the study, the performance of the GWAS allowed us to detect 24 quantitative trait loci (QTLs), including three genome-wide significant associations: QTL1 (chromosome 2, 130.72-131.01 Mb) for lactose percentage, QTL6 (chromosome 6, 78.90-93.48 Mb) for protein percentage and QTL17 (chromosome 17, 11.20 Mb) for both protein and dry matter percentages. Interestingly, QTL6 shows positional coincidence with the casein genes, which encode 80% of milk proteins.ConclusionsThe abrogation of lactation involves dramatic changes in the expression of genes participating in a broad array of physiological processes such as protein, lipid and carbohydrate metabolism, calcium homeostasis, cell death and tissue remodeling, as well as immunity. We also conclude that genetic variation at the casein genes has a major impact on the milk protein content of Murciano-Granadina goats.

Histological Characteristics of the Telocytes in the Mammary Glands of She-Camel (Camelus dromedarius)

ABSTRACT Telocytes are stromal stem cells located in different organs including the lactating and non-lactating mammary glands of she-camel. Fresh speciemens of mammary gland were obtained from 10 lactating and non-lactating she-camels of 8 years old directly after slaughtering for human consumption according to the rules of the Egyptian veterinary authorities. The specimens were processed for transmission electron microscopical examination. Telocytes were found around the ducts of lactating and non-lactating mammary glands and beside the mast cells. These telocytes communicate with other cells sometimes differentiate into fibroblasts. In the lactating mammary gland telocytes were found around the acini. The telocytes were spindle and slender in shape with two long cytoplasmic extension (telopodes). These telopodes had nano-vesicles. They had rods to pear shape nuclei. In a conclusion, the telocytes are stromal stem cells in both lactating and non-lactating she-camel mammary glands

Invited review: Accelerating mammary gland involution after drying-off in dairy cattle

Bovine mammary gland involution, as a part of the reproductive cycle in dairy cows, is a very important remodeling transformation of the mammary gland for the subsequent lactation. There is considerable incentive to accelerate mammary gland involution to improve udder health, shorten the dry period, and simplify the management process by reducing dietary changes. The complex process of mammary involution is characterized by morphological changes in the epithelial cells and mammary tissue, changes in the composition of mammary secretions, and changes in the integrity of tight junctions. Involution is facilitated by elements of the immune system and several types of proteases and is coordinated by various types of hormones. This review first describes the involution process and then argues for the need to accelerate it. Last, this review focuses on various intervention methods for accelerating involution. Our aim is to provide a comprehensive overview of bovine mammary gland involution as well as potential techniques and new opinions for dry cow management.

Short communication: Differences in distribution of serotonin receptor subtypes in the mammary gland of sheep, goats, and cows during lactation and involution

Serotonin receptors (5-HTR) are present in the mammary tissue of mouse, humans, cows, and rats. In these species, serotonin is important for the mammary gland function and lactation performance. The mammary gland expression of 5-HTR in small dairy ruminants has yet to be described. In the present study, primer sequences were developed to amplify 5-HTR (1A, 1D, 1E,1B, 1F, 2A, 2B, 2C, 3a, 4, 5a, 6, and 7) using real-time quantitative PCR for the detection of mRNA expression in mammary tissue of dairy sheep, goats, and cows. The distribution of commonly expressed 5-HTR between the 3 species (1B, 1E, 2A, 2B, 4, and 7) was analyzed in the mammary tissue of late-lactation and dried-off sheep, goats, and cows using immunohistochemical staining. Real-time quantitative PCR analysis showed that the 3 studied species expressed receptors 5-HTR1B, 1E, 2A, 2B, 4, and 7. Goats and sheep expressed 5-HTR1D and 5a; 5-HTR1A and 1F were expressed only in sheep. The mammary epithelial cells were positively stained for all the studied receptors by immunohistochemistry (5-HTR1B, 1E, 2A, 2B, 4, and 7). The endothelial cells of blood vessels were positively stained for 5-HTR1B, 2A, 2B, and 7 in all the species. Additionally, 5-HTR1E was present in cow endothelium. The myoepithelial cells stained positively for 5-HTR1E in all the species, and 5-HTR4 myoepithelial staining was present only in cows and sheep. Between the lactating and dried-off mammary glands, the location of 5-HTR in the epithelial cells changed from a cytoplasmic reaction in lactating udders to a reaction in the apical region in dry udders. These results showed that the distribution of 5-HTR subtypes in the mammary gland of dairy ruminants vary among species, tissue type, and stage of gland development. These findings warrant future studies aimed at understanding whether the differences in 5-HTR subtype expression and location accounts for the differences in milk secretion and lactocyte activity among cows, goats, and sheep.

Autophagy and unfolded protein response (UPR) regulate mammary gland involution by restraining apoptosis-driven irreversible changes

The postnatal mammary gland undergoes repeated cycles of proliferation and cell death, most notably when the fully differentiated (lactating) gland dedifferentiates to a prelactation state. Accumulation of milk proteins in the secretory epithelium creates the stress signal that triggers this process (involution). How this stress is perceived, and the cellular processes that are subsequently activated, remain unclear. We now report that Unfolded Protein Response (UPR), autophagy, and apoptosis related genes cluster separately during lactation and involution in the mouse mammary gland. Time-course experiments in rodents show that autophagy and UPR signaling are tightly co-regulated at the transition from reversible to irreversible involution. Inhibition of autophagy by chloroquine or genetic deletion of one ATG7 allele enhanced progression of mammary involution into the irreversible phase, as characterized by an early/precocious induction of apoptosis. These are the first preclinical in vivo data in support of a clinical trial testing an autophagy inhibitor for prevention of intraductal breast malignancy progression to invasive breast cancer. In marked contrast, stimulation of autophagy by low dose tunicamycin treatment reduced apoptosis and extended the reversible phase of involution by sustaining the secretory epithelium. Autophagy stimulators could be used short-term to promote lactation in women experiencing difficulties or irregularities in nursing. Taken together, these data indicate that UPR and autophagy play a key role in regulating the balance between cell survival and apoptosis during normal mammary gland regression.

Transcriptome sequencing to detect the potential role of long non-coding RNAs in bovine mammary gland during the dry and lactation period

BackgroundIt is known that long non-coding RNAs (lncRNAs) play an important role in various biological processes, including cell proliferation, differentiation and apoptosis. However, their functions and profiles in lactation cycle of dairy cows are largely unknown. In this study, lncRNA-seq technique was employed to compare the expression profiles of lncRNAs and mRNAs from Chinese Holstein mammary gland in dry and lactation period.ResultTotally 3746 differentially expressed lncRNAs (DELs) and 2890 differentially expressed genes (DEGs) were identified from the dry and lactation mammary glands of Holstein cows. Functional enrichment analysis on target genes of lncRNAs indicated that these genes were involved in lactation-related signaling pathways, including cell cycle, JAK-STAT, cell adhesion, and PI3K-Akt signaling pathways. Additionally, the interaction between lncRNAs and their potential miRNAs was predicted and partly verified. The result indicated that the lactation-associated miR-221 might interact with lncRNAs TCONS_00040268, TCONS_00137654, TCONS_00071659 and TCONS_00000352, which revealed that these lncRNAs might be important regulators for lactation cycle.ConclusionThis study provides a resource for lncRNA research on lactation cycle of bovine mammary gland. Besides, the interaction between lncRNAs and the specific miRNA is revealed. It expands our knowledge about lncRNA and miRNA biology as well as contributes to clarify the regulation of lactation cycle of bovine mammary gland.