Specific microRNAs in stallion spermatozoa are potential biomarkers of high functionality.

Males of some species, from horses to humans, require medical help for subfertility problems. There is an urgent need for novel molecular assays that reflect spermatozoal function. In the last 25 years, studies examined RNAs in spermatozoa as a window into gene expression during their development and, more recently, for their functions in early embryo development.

The Use of Diets in the Diagnosis and Treatment of Common Gastrointestinal Diseases in Dogs and Cats.

The nutritional health of dogs and cats is important to pet owners around the world. Nutrition is inextricably linked to the health of the gastrointestinal system and vice versa. Gastrointestinal signs, such as vomiting, diarrhea, anorexia, or weight loss, are one of the most common reasons that dog and cat owners make non-routine appointments with veterinarians.

Conservation of Glutathione Transferase mRNA and Protein Sequences Similar to Human and Horse Alpha Class GST A3-3 across Dog, Goat, and Opossum Species.

The glutathione transferase A3-3 (GST A3-3) homodimeric enzyme is the most efficient enzyme that catalyzes isomerization of the precursors of testosterone, estradiol, and progesterone in the gonads of humans and horses. However, the presence of GST A3-3 orthologs with equally high ketosteroid isomerase activity has not been verified in other mammalian species, even though pig and cattle homologs have been cloned and studied. Identifying genes is a challenge because of multiple gene duplications (e.

Glutathione Transferases as Efficient Ketosteroid Isomerases.

In addition to their well-established role in detoxication, glutathione transferases (GSTs) have other biological functions. We are focusing on the ketosteroid isomerase activity, which appears to contribute to steroid hormone biosynthesis in mammalian tissues. A highly efficient GST A3-3 is present in some, but not all, mammals.

Transcriptomic analysis of the honey bee (Apis mellifera) queen spermathecae reveals genes that may be involved in sperm storage after mating.

Honey bee (Apis mellifera) queens have a remarkable organ, the spermatheca, which successfully stores sperm for years after a virgin queen mates. This study uniquely characterized and quantified the transcriptomes of the spermathecae from mated and virgin honey bee queens via RNA sequencing to identify differences in mRNA levels based on a queen's mating status. The transcriptome of drone semen was analyzed for comparison.

Identification and quantification of coding and long non-coding RNAs in stallion spermatozoa separated by density.

Background: It is not unusual for stallions to have fertility problems. For many, artificial insemination with more dense spermatozoa (isolated by density gradient centrifugation) results in greater pregnancy rates compared with the rates when using unfractionated spermatozoa. RNAs in spermatozoa delivered to the oocyte at conception are required for embryo development.

Dexamethasone downregulates expression of several genes encoding orphan nuclear receptors that are important to steroidogenesis in stallion testes.

Glucocorticoids impair testosterone synthesis by an unknown mechanism. Stallions treated with the synthetic glucocorticoid dexamethasone had testes collected at 6 or 12 hours postinjection. The testicular expression of selected genes encoding nuclear receptors and steroidogenic enzymes was measured.

Characterization of equine GST A3-3 as a steroid isomerase.

Glutathione transferases (GSTs) comprise a superfamily of enzymes prominently involved in detoxication by making toxic electrophiles more polar and therefore more easily excretable. However some GSTs have developed alternative functions. Thus, a member of the Alpha class GSTs in pig and human tissues is involved in steroid hormone biosynthesis, catalyzing the obligatory double-bond isomerization of Δ-androstene-3,17-dione to Δ-androstene-3,17-dione and of Δ-pregnene-3,20-dione to Δ-pregnene-3,20-dione on the biosynthetic pathways to testosterone and progesterone.

Dexamethasone acutely down-regulates genes involved in steroidogenesis in stallion testes.

In rodents, livestock and primate species, a single dose of the synthetic glucocorticoid dexamethasone acutely lowers testosterone biosynthesis. To determine the mechanism of decreased testosterone biosynthesis, stallions were treated with 0.1mg/kg dexamethasone 12h prior to castration.

Marinobufagenin regulates permeability and gene expression of brain endothelial cells.

Marinobufagenin (MBG) is a cardiotonic steroid that increases in the circulation in preeclampsia. Preeclampsia and eclampsia are associated with cerebral edema. Therefore, we examined the effects of MBG on human brain microvascular endothelial cells (HBMEC) in vitro.

CatSper and the relationship of hyperactivated motility to intracellular calcium and pH kinetics in equine sperm.

In vitro fertilization does not occur readily in the horse. This may be related to failure of equine sperm to initiate hyperactivated motility, as treating with procaine to induce hyperactivation increases fertilization rates. In mice, hyperactivated motility requires a sperm-specific pH-gated calcium channel (CatSper); therefore, we investigated this channel in equine sperm.

Estradiol up-regulates expression of the A + U-rich binding factor 1 (AUF1) gene in the sheep uterus.

The A+U-rich binding factor 1 (AUF1 or HNRPD) gene produces predominant RNA binding proteins. The AUF1 transcript is alternatively spliced to produce four protein isoforms that stabilize or destabilize hundreds of mRNAs. Previously, we discovered that estradiol (E2) treatment of ovariectomized sheep increased concentrations of AUF1p45 protein which stabilized estrogen receptor alpha (ER) mRNA in the uterus.

Production and characterization of agonistic monoclonal antibodies against chicken CD40.

CD40 is mainly expressed by professional antigen-presenting cells (APCs). Its ligand, CD40L, is transiently expressed on activated CD4(+) T-cells. CD40-CD40L interactions mediate T-cell help to APCs and provide crucial signals for affinity maturation and B-cell class switching.

Transforming growth factor beta (TGFB) signaling is activated during porcine implantation: proposed role for latency-associated peptide interactions with integrins at the conceptus-maternal interface.

The process of implantation is mediated by a complex network of signaling and adhesive factors. In the pig, latent and active transforming growth factor beta (TGFB), TGFB receptors (TGFBR), and integrins (ITGs) are present during the peri-implantation period. TGFB signals via TGFBR and activates downstream effector SMAD proteins 2 and 3 (p-SMAD2/3).

In vitro culture of precision-cut testicular tissue as a novel tool for the study of responses to LH.

In vitro culture systems are valuable tools for investigating reproductive mechanisms in the testis. Here, we report the use of the precision-cut in vitro system using equine testicular slices. Testes were collected from immature light breed stallions (n=3) and cut into slices (mean slice weight= 13.

Intrauterine infusion of latency-associated peptide (LAP) during early porcine pregnancy affects conceptus elongation and placental size.

In the pig, transforming growth factor beta (TGFB), TGFB receptors (TGFBRs), and integrins are present during the peri-implantation period. Latency-associated peptide (LAP), a part of latent TGFB, can bind to integrin heterodimers via its Arg-Gly-Asp (RGD) sequence; therefore, ligand-receptor interactions between TGFB and TGFBRs, along with LAP and integrin heterodimers, may be functional in mediating events supporting conceptus elongation and attachment. With the use of surgically implantable osmotic pumps, we were able to maintain pregnancy with the aim of mechanistically altering in vivo receptor-ligand interactions involving TGFB with TGFBRs and LAP with integrins during porcine pregnancy.

Estradiol up-regulates AUF1p45 binding to stabilizing regions within the 3'-untranslated region of estrogen receptor alpha mRNA.

Estradiol up-regulates expression of the estrogen receptor alpha gene in the uterus by stabilizing estrogen receptor alpha mRNA. Previously, we defined two discrete minimal estradiol-modulated stability sequences (MEMSS) within the extensive 3'-untranslated region of estrogen receptor alpha mRNA with an in vitro stability assay using cytosolic extracts from sheep uterus. We report here that excess MEMSS RNA inhibited the enhanced stability of estrogen receptor alpha mRNA in extracts from estradiol-treated ewes compared with those from control ewes.

Steroid hormones acutely regulate expression of a Nudix protein-encoding gene in the endometrial epithelium of sheep.

Steroid hormones regulate endometrial gene expression to meet the needs of developing embryos. Our hypothesis is that steroid hormones transiently induce expression of genes in the endometrial epithelium to make the uterine environment different between the earliest days of pregnancy. We identified one such gene product using differential display-polymerase chain reactions.

Functional effects of transforming growth factor beta on adhesive properties of porcine trophectoderm.

In pigs, expression and amounts of biologically active TGFbetas at the conceptus-maternal interface increase significantly as conceptuses elongate and begin the implantation process. Before their activation, secreted TGFbetas are noncovalently associated with their respective, isoform-specific latency-associated peptides (LAPs), which contain the Arg-Gly-Asp (RGD) amino acid sequence that serves as a ligand for numerous integrins. Objectives of this study were to determine whether TGFbeta1 increases production of fibronectin by porcine trophectoderm, whether porcine trophectoderm adheres specifically to fibronectin and LAP, and whether functional interactions between porcine trophectoderm and the two TGFbeta-associated proteins, fibronectin and LAP, are integrin mediated.

Steroid hormones regulate gene expression posttranscriptionally by altering the stabilities of messenger RNAs.

Hormones exert powerful effects on reproductive physiology by regulating gene expression. Recent discoveries in hormone action emphasize that regulation of gene expression is not restricted to their alterations of the rate of gene transcription. On the contrary, hormonal effects on the stability of a specific mRNA can profoundly alter its steady-state concentration.

Cell-specific expression of estrogen-responsive genes in the uteri of cyclic, early pregnant and ovariectomized ewes.

A single physiological dose of estradiol up-regulates estrogen receptor-alpha(ER), progesterone receptor (PR), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), c-fos, cyclophilin, and actin mRNAs in the endometrium of ovariectomized ewes. Therefore, we hypothesized that these genes would be up-regulated by the preovulatory surge of estrogen which occurs on the evening of Day 15 in the estrous cycle of sheep. ER and PR mRNA concentrations increased between Day 15 and Day 1 in cyclic ewes in most endometrial epithelial cells, while GAPDH mRNA increased in epithelial and stromal cells in the deep endometrium.

Gene expression in the spermatogenically inactive "dark" and maturing "light" testicular tissues of the prepubertal colt.

In the testis of the 1.5-year-old horse, spermatogenesis initiates locally in grossly light, central areas that contrast with grossly dark, peripheral areas that are as yet inactive in spermatogenesis. Gene expression was compared between "light" and "dark" tissues of 1.

Myometrial effects of selective estrogen receptor modulators on estradiol-responsive gene expression are gene and cell-specific.

We examined in vivo effects of selective estrogen receptor modulators (SERMs) 4-OH-tamoxifen (Tam), GW 5638 (GW) and EM-800 (EM) on myometrial gene expression. The uteri of ovariectomized ewes were infused with 10(-7)M of one SERM via indwelling catheters for 24h preceding hysterectomy. Half of the ewes in each SERM group received an intramuscular injection of 50 microg 17beta-estradiol (E2) 18 h prior to hysterectomy.

Endometrial effects of selective estrogen receptor modulators (SERMs) on estradiol-responsive gene expression are gene and cell-specific.

Three selective estrogen receptor modulator (SERM) drugs which included 4-OH-tamoxifen (Tam), EM-800 (EM) and GW 5638 (GW) were investigated to determine their ability to inhibit estradiol-responsive gene expression in sheep endometrium. The uteri of ovariectomized ewes (10 ewes per SERM group) were infused with 10(-7)M SERMs for 24h prior to hysterectomy. Five ewes from each group received 50 microg 17beta-estradiol (E2) and the remaining five ewes received vehicle 18 h prior to hysterectomy.

Estradiol and a selective estrogen receptor modulator affect steroid hormone receptor messenger RNA levels and turnover in explant cultures of sheep endometrium.

Estrogens upregulate estrogen receptor (ER) and progesterone receptor (PR) gene expression in endometrium immediately before ovulation to prepare it for nurturing embryos. Most in vitro model systems have lost the ability to upregulate expression of the ER gene in response to estradiol (E2) or the ability to express the ER gene at all. Here, we used explant cultures from control and E2-treated ewes and assessed expression of four genes (ER, PR, glyceraldehyde 3-phosphate dehydrogenase [GAPDH], and cyclophilin [CYC] genes) that are upregulated by E2 in vivo on Northern blots.