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
DNAJC2 protein, also known as ZRF1 or MPP11, acts both as chaperone and as chromatin regulator. It is involved in stem cell differentiation and its expression is associated with various cancer ...malignancies. However, the role of Dnajc2 gene during mouse embryogenesis has not been assessed so far. To this aim, we invalidated Dnajc2 gene in FVB/Nj mice using the CrispR/Cas9 approach. We showed that this invalidation leads to the early post-implantation lethality of the nullizygous embryos. Furthermore, using siRNAs against Dnajc2 in mouse 1-cell embryos, we showed that maternal Dnajc2 mRNAs may allow for the early preimplantation development of these embryos. Altogether, these data demonstrate for the first time the requirement of DNAJC2 for early mouse embryogenesis.
•Invalidation of DNAJC2 leads to early post-implantation lethality of mouse embryos.•Dnajc2 is expressed throughout early mouse embryogenesis.•Maternal Dnajc2 mRNAs allow early preimplantation mouse embryonic development.
Preeclampsia is a disease of pregnancy involving systemic endothelial dysfunction. However, cardiovascular consequences of preeclampsia are difficult to analyze in humans. The objective of the ...present study is to evaluate the cardiovascular dysfunction induced by preeclampsia by examining the endothelium of mice suffering of severe preeclampsia induced by STOX1 overexpression. Using Next Generation Sequencing on endothelial cells of mice carrying either transgenic or control embryos, we discovered significant alterations of gene networks involved in inflammation, cell cycle, and cardiac hypertrophy. In addition, the heart of the preeclamptic mice revealed cardiac hypertrophy associated with histological anomalies. Bioinformatics comparison of the networks of modified genes in the endothelial cells of the preeclamptic mice and HUVECs exposed to plasma from preeclamptic women identified striking similarities. The cardiovascular alterations in the pregnant mice are comparable to those endured by the cardiovascular system of preeclamptic women. The STOX1 mice could help to better understand the endothelial dysfunction in the context of preeclampsia, and guide the search for efficient therapies able to protect the maternal endothelium during the disease and its aftermath.
MicroRNA (miRNA) are negative regulators of gene expression, capable of exerting pronounced influences upon the translation and stability of mRNA. They are potential regulators of normal mammary ...gland development and of the maintenance of mammary epithelial progenitor cells. This study was undertaken to determine the role of miR-30b on the establishment of a functional mouse mammary gland. miR-30b is a member of the miR-30 family, composed of 6 miRNA that are highly conserved in vertebrates. It has been suggested to play a role in the differentiation of several cell types.
The expression of miR-30b was found to be regulated during mammary gland development. Transgenic mice overexpressing miR-30b in mammary epithelial cells were used to investigate its role. During lactation, mammary histological analysis of the transgenic mice showed a reduction in the size of alveolar lumen, a defect of the lipid droplets and a growth defect of pups fed by transgenic females. Moreover some mammary epithelial differentiated structures persisted during involution, suggesting a delay in the process. The genes whose expression was affected by the overexpression of miR-30b were characterized by microarray analysis.
Our data suggests that miR-30b is important for the biology of the mammary gland and demonstrates that the deregulation of only one miRNA could affect lactation and involution.
Although conversion of the cellular form of the prion protein (PrP(C)) into a misfolded isoform is the underlying cause of prion diseases, understanding PrP(C) physiological functions has remained ...challenging. PrP(C) depletion or overexpression alters the proliferation and differentiation properties of various types of stem and progenitor cells in vitro by unknown mechanisms. Such involvement remains uncertain in vivo in the absence of any drastic phenotype of mice lacking PrP(C). Here, we report PrP(C) enrichment at the base of the primary cilium in stem and progenitor cells from the central nervous system and cardiovascular system of developing mouse embryos. PrP(C) depletion in a neuroepithelial cell line dramatically altered key cilium-dependent processes, such as Sonic hedgehog signalling and α-tubulin post-translational modifications. These processes were also affected over a limited time window in PrP(C)-ablated embryos. Thus, our study reveals PrP(C) as a potential actor in the developmental regulation of microtubule dynamics and ciliary functions.
Educators have made great strides in improving the educational experiences of students in schools today. Fifty years ago, students with special needs were hidden in the back hallways of the school ...building and forgotten for the better portion of the school day. Exposure to general education students, curriculum, and activities were unheard of; hopes of this population were not realized. Self-contained classrooms were the only option available to these students. Advocates for this special population began to propose and challenge the status quo. A review of the literature revealed there are barriers that can have a negative impact on the academic success of students with disabilities in the language arts inclusive classroom. However, current literature was missing the perceptions of the middle school principals and teachers. The purpose of this multiple case study was to examine educator and other stakeholder perceptions of the barriers inclusive classrooms pose that may impede the success of disabled students in language arts. A multiple case study design shed light on inclusive practices presently used in one independent school district; data collected from interviews of principals and teachers addressed teachers’ and principals’ concerns and provided insight into the academic disparity that exists between disabled students and their non-disabled peers in inclusive environments. Implications from this study can lead to a successful collaboration between school districts and middle school campuses that can provide mentors and other resources to motivate teachers who work in the inclusive setting to allocate resources to help students with disabilities succeed academically in language arts.
► Regulatory sequences of the
Sprn gene (5.3
Kb) were used to drive expression of a LacZ reporter gene in mice. ► Expression of the transgene was detected from 10.5
dpc onwards. ► Extra-embryonic and ...nerve cell-specific transgene expression was revealed. ► The results suggest a role for Shadoo in mouse embryo development.
The prion-like protein Shadoo has been suggested to compensate for the lack of PrP in
Prnp-knockout mice, explaining their lack of extreme phenotype. In adult mice, both PrP and Shadoo have shown overlapping expression patterns and shared functions
1. Their expression in the mouse embryo has also been suggested to be complementary, as invalidation of both genes results in embryonic lethality
2. The developmental expression profile of PrP has been described from post-implantation stages up until birth
3–5. However the spatial expression pattern of Shadoo in the developing mouse embryo is not known. We previously described the expression profile of the prion-like protein Shadoo in adult mice using
Sprn reporter mice (
Sprn-GFP and
Sprn-LacZ). Here we used these mice to describe the developmental expression of Shadoo between 10.5 and 14.5
dpc. The observed pattern in specific embryonic cell lineages and in extra-embryonic tissues is consistent with the previously reported phenotype resulting from its knockdown.
► Regulatory sequences of the Sprn gene (5.3Kb) were used to drive expression of reporter genes in mice. ► Expression of the transgenes was comparable of that of the endogene in 100% of the lines. ► ...Gonad cell-specific transgene expression was revealed. ► These results suggest a role for Shadoo in fertility and reproduction. ► These mice can be used as tools to better understand the regulation of Sprn.
The protein Shadoo (Sho) is a paralogue of prion protein, and encoded by the gene Sprn. Like prion protein it is primarily expressed in central nervous system, and has been shown to have a similar expression pattern in certain regions of the brain. We have generated reporter mice carrying a transgene encompassing the Sprn promoter, exon 1, intron 1 and the 5′-end of exon 2 driving expression of either the LacZ or GFP reporter gene to study the expression profile of Shadoo in mice. Expression of the reporter genes was analysed in brains of these transgenic mice and was shown to mimic that of the endogenous gene expression, previously described by Watts et al. 1. Consequently, the Sprn-LacZ mice were used to study the spatial expression of Sho in other tissues of the adult mouse. Several tissues were collected and stained for β-gal activity, including the thymus, heart, lung, liver, kidney, spleen, intestine, muscle, and gonads. From this array of tissues, the transgene was consistently expressed only in specific cell types of the testicle and ovary, suggesting a role for Shadoo in fertility and reproduction. These mice may serve as a useful tool in deciphering the regulation of the prion-like gene Sprn and thus, indirectly, of the Shadoo protein.
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