Murine embryo transfer is the final and key step needed to produce offspring after in vitro fertilization or a transgenic procedure and is accomplished surgically or by transferring embryos into the oviduct or uterine horn of a pseudopregnant mouse. During the transfer of embryos into the uterine horn, air bubbles frequently are loaded with embryos into the transfer catheter. 26 However, the role of air bubbles on murine embryonic development is unclear. In addition, the issue has not been addressed in regard to embryo transfer procedures for humans and other animals. 5,31 In mice, the onset of pregnancy is heralded by the attachment of an embryo to the luminal epithelium and its penetration into the stroma of the endometrium. In response to the steroid hormones estrogen and progesterone, the stromal cells surrounding the implantation chamber undergo a remarkable transformation. This process, known as decidualization, is an essential prerequisite for successful implantation. 32 However, an implanting embryo is not absolutely required for endometrial decidualization. 12 Decidualization can occur in response to an artificial stimulus, such as small beads or droplets of oil injected into the uterine lumen. 2,24 The endometrial tissue that forms in response to an artificial deciduogenic stimulus is called a deciduoma to distinguish it from naturally induced decidua. Air bubbles reportedly can induce decidualization in mice, rats, and hamsters, 11,28 but whether air-induced decidualization disturbs decidual gene expression, implantation, or embryonic development is unclear currently. Materials and Methods Animals. Female (age, 8 to 12 wk) and male (age, 3 to 10 mo) CD1 mice (Beijing Vital River Laboratory Animal Technology, Beijing, China) were housed under constant environmental conditions (25 ± 1 °C; relative humidity, 40% to 60%; lights on, 0600 to 1800) at our institutional animal facility. Quarterly sentinel surveillance was used to screen for a wide range of pathogens, including epizootic diarrhea of infant mice, mouse hepatitis virus, mouse norovirus, mouse parvovirus 1 and 2, Mycoplasma pulmonis, pneumonia virus of mice, Sendai virus, Helicobacter spp., fur mites, and pinworms. All results from sentinel mice tested negative throughout this study. Mice received autoclaved feed (Beijing Vital River) and reverse-osmosis-deionized water without restriction. Mice were housed in polycarbonate microisolation caging with autoclaved hardwood bedding (Beijing Vital River). Autoclaved environmental enrichment was provided weekly to all mice in the form of 5-cm squares of cotton batting that could be shredded to form nests. All procedures in this study were approved by Hebei Agricultural University Laboratory Animal Care and Use Committee. Air-induced decidualization in pseudopregnant mice. Female CD1 mice were mated with vasectomized males. The next morning the female mice were examined for vaginal plugs, and the plug-positive females were considered to be at day 0.5 of pseudopregnancy. On day 3.5, between 1500 to 1700, ...
Background Genetics evidences have long linked mosaic loss of Y-chromosome (mLOY) in peripheral leukocytes with a wide range of male age-associated diseases. However, a lack of cellular and molecular mechanistic explanations for this link has limited further investigation into the relationship between mLOY and male age-related disease. Excitingly, Sano et al. have provided the first piece of evidence directly linking mLOY to cardiac fibrosis through mLOY enriched profibrotic transforming growth factor β1 (TGF-β1) regulons in hematopoietic macrophages along with suppressed interleukin-1β (IL-1β) proinflammatory regulons. The results of this novel finding can be extrapolated to other disease related to mLOY, such as cancer, cardiac disease, and age-related macular degeneration. Results Sano et al. used a CRISPR-Cas9 gRNAs gene editing induced Y-chromosome ablation mouse model to assess results of a UK biobank prospective analysis implicating the Y-chromosome in male age-related disease. Using this in vivo model, Sano et al. showed that hematopoietic mLOY accelerated cardiac fibrosis and heart failure in male mice through profibrotic pathways. This process was linked to monocyte-macrophage differentiation during hematopoietic development. Mice confirmed to have mLOY in leukocytes, by loss of Y-chromosome genes Kdm5d, Uty, Eif2s3y, and Ddx3y, at similar percentages to the human population were shown to have accelerated rates of interstitial and perivascular fibrosis and abnormal echocardiograms. These mice also recovered poorly from the transverse aortic constriction (TAC) model of heart failure and developed left ventricular dysfunction at higher rates. This was attributed to aberrant proliferation of cardiac MEF-SK4 + fibroblasts promoted by mLOY macrophages enriched in profibrotic regulons and lacking in proinflammatory regulons. These pro-fibrotic macrophages localized to heart and eventually resulted in cardiac fibrosis via enhanced TGF-β1 and suppressed IL-1β signaling. Furthermore, treatment of mLOY mice with TGFβ1 neutralizing antibody was able to improve their cardiac function. This study by Sano et al. was able to provide a causative link between the known association between mLOY and male cardiac disease morbidity and mortality for the first time, and thereby provide a new target for improving human health. Conclusions Using a CRISPR-Cas9 induced Y-chromosome ablation mouse model, Sano et al. has proven mosaic loss of Y-chromosome in peripheral myeloid cells to have a causative effect on male mobility and mortality due to male age-related cardiac disease. They traced the mechanism of this effect to hyper-expression of the profibrotic TGF-β1 and reduced pro-inflammatory IL-1β signaling, attenuation of which could provide another potential strategy in improving outcomes against age-related diseases in men.
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