The bovine lactation genome: insights into the evolution of mammalian milk

Comparison of milk protein and mammary genes in the bovine genome with those from other mammals gives insights into the evolution of lactation

Plasticity of spermatogonial stem cells

There have been significant breakthroughs over the past decade in the development and use of pluripotent stem cells as a potential source of cells for applications in regenerative medicine. It is likely that this methodology will begin to play an important role in human clinical medicine in the years to come. This review describes the plasticity of one type of pluripotent cell, spermatogonial stem cells (SSCs), and their potential therapeutic applications in regenerative medicine and male infertility. Normally, SSCs give rise to sperm when in the testis. However, both human and murine SSCs can give rise to cells with embryonic stem (ES) cell-like characteristics that can be directed to differentiate into tissues of all three embryonic germ layers when placed in an appropriate inductive microenvironment, which is in contrast to other postnatal stem cells. Previous studies have reported that SSCs expressed an intermediate pluripotent phenotype before differentiating into a specific cell type and that extended culture was necessary for this to occur. However, recent studies from our group using a tissue recombination model demonstrated that SSCs differentiated rapidly into another tissue, in this case, prostatic epithelium, without expression of pluripotent ES cell markers before differentiation. These results suggest that SSCs are capable of directly differentiating into other cell types without going through an intermediate ES cell-like stage. Because SSCs do not require reprogramming to achieve a pluripotent state, they are an attractive source of pluripotent cells for use in regenerative medicine

Uterine-specific Ezh2 deletion enhances stromal cell senescence and impairs placentation, resulting in pregnancy loss

Summary: Maternal uterine remodeling facilitates embryo implantation, stromal cell decidualization and placentation, and perturbation of these processes may cause pregnancy loss. Enhancer of zeste homolog 2 (EZH2) is a histone methyltransferase that epigenetically represses gene transcription; loss of uterine EZH2 affects endometrial physiology and induces infertility. We utilized a uterine Ezh2 conditional knockout (cKO) mouse to determine EZH2’s role in pregnancy progression. Despite normal fertilization and implantation, embryo resorption occurred mid-gestation in Ezh2cKO mice, accompanied by compromised decidualization and placentation. Western blot analysis revealed Ezh2-deficient stromal cells have reduced amounts of the histone methylation mark H3K27me3, causing upregulation of senescence markers p21 and p16 and indicating that enhanced stromal cell senescence likely impairs decidualization. Placentas from Ezh2cKO dams on gestation day (GD) 12 show architectural defects, including mislocalization of spongiotrophoblasts and reduced vascularization. In summary, uterine Ezh2 loss impairs decidualization, increases decidual senescence, and alters trophoblast differentiation, leading to pregnancy loss

The histone methyltransferase EZH2 is required for normal uterine development and function in mice

Enhancer of zeste homolog 2 (EZH2) is a rate-limiting catalytic subunit of a histone methyltransferase, polycomb repressive complex, which silences gene activity through the repressive histone mark H3K27me3. EZH2 is critical for epigenetic effects of early estrogen treatment, and may be involved in uterine development and pathologies. We investigated EZH2 expression, regulation, and its role in uterine development/function. Uterine epithelial EZH2 expression was associated with proliferation and was high neonatally then declined by weaning. Pre-weaning uterine EZH2 expression was comparable in wild-type and estrogen receptor 1 knockout mice, showing neonatal EZH2 expression is ESR1 independent. Epithelial EZH2 was upregulated by 17β-estradiol (E2) and inhibited by progesterone in adult uteri from ovariectomized mice. To investigate the uterine role of EZH2, we developed a EZH2 conditional knockout (Ezh2cKO) mouse using a cre recombinase driven by the progesterone receptor (Pgr) promoter that produced Ezh2cKO mice lacking EZH2 in Pgr-expressing tissues (e.g. uterus, mammary glands). In Ezh2cKO uteri, EZH2 was deleted neonatally. These uteri had reduced H3K27me3, were larger than WT, and showed adult cystic endometrial hyperplasia. Ovary-independent uterine epithelial proliferation and increased numbers of highly proliferative uterine glands were seen in adult Ezh2cKO mice. Female Ezh2cKO mice were initially subfertile, and then became infertile by 9 months. Mammary gland development in Ezh2cKO mice was inhibited. In summary, uterine EZH2 expression is developmentally and hormonally regulated, and its loss causes aberrant uterine epithelial proliferation, uterine hypertrophy, and cystic endometrial hyperplasia, indicating a critical role in uterine development and function

Mice lacking membrane estrogen receptor 1 are protected from reproductive pathologies resulting from developmental estrogen exposure

Both membrane and nuclear fractions of estrogen receptor 1 (ESR1) mediate 17β-estradiol (E2) actions. Mice expressing nuclear (n)ESR1 but lacking membrane (m)ESR1 (nuclear-only estrogen receptor 1 [NOER] mice) show reduced E2 responsivity and reproductive abnormalities culminating in adult male and female infertility. Using this model, we investigated whether reproductive pathologies caused by the synthetic estrogen diethylstilbestrol (DES) are mitigated by mESR1 ablation. Homozygous and heterozygous wild-type (WT and HET, respectively) and NOER male and female mice were subcutaneously injected with DES (1 mg/kg body weight [BW]) or vehicle daily from postnatal day (PND) 1-5. Uterine histology was assessed in select DES-treated females at PND 5, whereas others were ovariectomized at PND 60 and treated with E2 (10 μg/kg BW) or vehicle 2 weeks later. Neonatal DES exposure resulted in ovary-independent epithelial proliferation in the vagina and uterus of WT but not NOER females. Neonatal DES treatment also induced ovary-independent adult expression of classical E2-induced transcripts (e.g., lactoferrin [Ltf] and enhancer of zeste homolog 2 [Ezh2]) in WT but not NOER mice. At PND 90, DES-treated WT and HET males showed smaller testes and a high incidence of bacterial pyogranulomatous inflammation encompassing the testes, epididymis and occasionally the ductus deferens with spread to lumbar lymph nodes; such changes were largely absent in NOER males. Results indicate that male and female NOER mice are protected from deleterious effects of neonatal DES, and thus mESR1 signaling is required for adult manifestation of DES-induced reproductive pathologies in both sexes

Antibody-specific detection of CAIX in breast and prostate cancers.

Contains fulltext : 80121.pdf (publisher's version ) (Closed access)Carbonic anhydrase IX (CAIX) is frequently expressed in human tumors and serves as a marker for hypoxia. Further, CAIX expression is considered a predictor of poor survival in many, but not all, cancer types. Herein, we compare the specificity of two CAIX antibodies: the M75, monoclonal antibody which recognizes an epitope in the N-terminus and a commercially available polyclonal antibody generated against a C-terminal peptide (NB100-417). Western blot analysis of multiple breast cell lines revealed that the polyclonal antibody detected both membrane-bound and soluble proteins. The M75 antibody recognized only the membrane-bound species, which is presumed to be CAIX. These data were confirmed in an aggressive prostate cell line. We further compared these antibodies in prostate tumors by immunohistochemistry. Staining with NB100 was comparable to that of the M75 antibody, but only at high dilution. Otherwise, cytoplasmic staining was also noted. Two-dimensional gel electrophoresis followed by mass spectrometric analysis revealed that the cytoplasmic protein detected by NB100 is beta-tubulin. This cross-reactivity could lead to false-positives for CAIX expression in samples where cytosolic proteins are present

The bovine lactation genome: insights into the evolution of mammalian milk.

BackgroundThe newly assembled Bos taurus genome sequence enables the linkage of bovine milk and lactation data with other mammalian genomes.ResultsUsing publicly available milk proteome data and mammary expressed sequence tags, 197 milk protein genes and over 6,000 mammary genes were identified in the bovine genome. Intersection of these genes with 238 milk production quantitative trait loci curated from the literature decreased the search space for milk trait effectors by more than an order of magnitude. Genome location analysis revealed a tendency for milk protein genes to be clustered with other mammary genes. Using the genomes of a monotreme (platypus), a marsupial (opossum), and five placental mammals (bovine, human, dog, mice, rat), gene loss and duplication, phylogeny, sequence conservation, and evolution were examined. Compared with other genes in the bovine genome, milk and mammary genes are: more likely to be present in all mammals; more likely to be duplicated in therians; more highly conserved across Mammalia; and evolving more slowly along the bovine lineage. The most divergent proteins in milk were associated with nutritional and immunological components of milk, whereas highly conserved proteins were associated with secretory processes.ConclusionsAlthough both copy number and sequence variation contribute to the diversity of milk protein composition across species, our results suggest that this diversity is primarily due to other mechanisms. Our findings support the essentiality of milk to the survival of mammalian neonates and the establishment of milk secretory mechanisms more than 160 million years ago