Inhibiting diacylglycerol acyltransferase-1 reduces lipid biosynthesis in bovine blastocysts produced in vitro.

Diacylglycerol acyltransferase-1 (DGAT1) is one of the DGAT enzymes that catalyzes the final step in the synthesis of triacylglycerol, which is a major component of the lipid droplets in embryos. Intracellular lipids accumulated in embryos produced in vitro have been associated with reduced cryotolerance and quality. The objective of the present study was to investigate the influence of DGAT1 inhibition on embryo development, quality, and post-vitrification survival, in addition to expression profiles of selected lipid metabolism-regulating and oxidative stress genes. Bovine cumulus-oocyte complexes were matured and fertilized in vitro and were cultured in synthetic oviduct fluid (SOF) supplemented with 5% fetal calf serum (FCS) alone (Control) or with 1, 5, 10 or 50 µM DGAT1 inhibitor (A922500®; D1, D5, D10, and D50, respectively) or 0.1% dimethyl sulfoxide (CDMSO: vehicle for DGAT1 inhibitor dilution) from 54 h post-insemination until Day 8 post insemination. No differences were found in blastocyst yield on days 7 and 8 in Control, CDMSO, D10, and D50 groups. Embryos cultured with 10 or 50 µM DGAT1 inhibitor had greater mitochondrial activity (P < 0.01), and increased number of cells (P < 0.05), while the cytoplasmic lipid content was reduced (P < 0.01), the latter associated with altered expression profiles of selected genes regulating lipid metabolism or genes related with oxidative stress (transcript abundance increased for SLC2A1 and SLC2A5 and decreased for DGAT1 and GPX1). Importantly, the survival rate of blastocysts produced with 10 µM DGAT1 was higher than that of Control, CDMSO and D50 groups at 72 h after vitrification and warming (73.8 vs 57.1, 55.9 and 56.1%, respectively, P < 0.001). In conclusion, inhibition of DGAT1 synthesis in bovine embryos produced in vitro abrogates the negative effect of FCS by decreasing their lipid content, increasing mitochondria activity and improving embryo cryotolerance, as well as favoring the expression of lipid metabolism regulating and oxidative stress-related transcripts.

Effect of chronic THC administration in the reproductive organs of male mice, spermatozoa and in vitro fertilization.

The increased use of cannabis as a therapeutic drug in recent years has raised some concerns due to its potential effects on reproductive health. With regards to the male, the endocannabinoid system is involved in the spermatogenesis and in the sperm function. The chronic use of tetrahidrocannabinol (THC) has been associated with sperm anomalies, decreased sperm motility and structural changes in the testis. However, whether THC affects sperms ability to fertilize and to generate embryos remains unclear. The aim of this study was to evaluate this effect using a mice model of THC chronic treatment. For this purpose, a chronic treatment with THC was carried out. Mice were randomly allocated into two groups: an experimental group treated with a daily dose of 10 mg/kg-body weight THC for a period of 30 days and a control group treated with a vehicle. The THC-mice cortex showed a significant decrease of mRNA of Cnr1 compared to control-mice while, in the testis, the expression of Cnr1 was not affected. The weight of testis and epididymis and the histological analysis did not show any change between groups. On the other hand, no changes were observed in the sperm motility or the sperm concentration. The chronic use of THC did not generate any methylation change in the three CpG regions of Cnn1 analysed, neither in the brain nor in the embryos generated by in vitro fertilization (IVF). Finally, the embryo production by IVF was no different using spermatozoa from both THC and control mice. This work contradicts the belief that THC consumption has a negative effect on male reproductive processes.

Resveratrol-cyclodextrin complex affects the expression of genes associated with lipid metabolism in bovine in vitro produced embryos.

Antioxidants have been widely used during in vitro production to decrease the negative effect of reactive oxygen species. It was reported that the complex resveratrol-methyl ß-cyclodextrin (RV-CD) improves resveratrol's stability and bioavailability and increases its antioxidant activity. This study evaluates the effect of RV-CD during in vitro oocyte maturation (IVM) or in vitro embryo culture (IVC) on developmental competence and quantitative changes in gene expression of developmental important genes. In experiment 1, RV-CD was added to IVM media and maturation level, embryo development and oocytes, cumulus cells, and blastocysts gene expression by RT-qPCR were examined. In experiment 2, presumptive zygotes were cultured in SOF supplemented with RV-CD and embryo development and blastocysts gene expression by RT-qPCR were studied. A group without RV-CD (control- ) and a group with cyclodextrin (control+ ) were included. No differences were found in cleavage rate or blastocyst yield between groups. However, the expression of LIPE was higher in blastocysts derived from oocytes treated with resveratrol compared with control groups (p < .05). Blastocysts produced by IVC with resveratrol showed that RV-CD could modify the expression of genes related to lipid metabolism (CYP51A1, PNPLA2 and MTORC1) compared with control groups (p < .05). RV-CD in the IVM and IVC media could reduce accumulated fat by increasing lipolysis and suppressing lipogenesis of blastocysts.