Circular RNAs (circRNAs) are thought to play essential roles in multiple biological processes, including apoptosis, an important process in antral follicle atresia. We aimed to investigate the potential involvement of circRNAs in granulosa cell apoptosis and thus antral follicle atresia. CircRNA expression profiles were generated from porcine granulosa cells isolated from healthy antral (HA) and atretic antral (AA) follicles. Over 9632 circRNAs were identified, of which 62 circRNAs were differentially expressed (DE-circRNAs). Back-splicing, RNase R resistance, and stability of DE-circRNAs were validated, and miRNA binding sites and related target genes were predicted. Two exonic circRNAs with low false discovery rate (FDR) high fold change, miRNA binding sites, and relevant biological functions—circ_CBFA2T2 and circ_KIF16B—were selected for further characterization. qRT-PCR and linear regression analysis confirmed expression and correlation of the targeted genes—the antioxidant gene GCLC (potential target of circ_CBFA2T2) and the apoptotic gene TP53 (potential target of circ_KIF16B). Increased mRNA content of TP53 in granulosa cells of AA follicles was further confirmed by strong immunostaining of both p53 and its downstream target pleckstrin homology like domain family a member 3 (PHLDA3) in AA follicles compared to negligible staining in granulosa cells of HA follicles. Therefore, we concluded that aberrantly expressed circRNAs presumably play a potential role in antral follicular atresia.
In the swine industry pathogenic infections have a significant negative impact on neonatal survival. Piglets fed with human lysozyme, a natural antibiotic, might be more resistant to gastrointestinal infections. Here we describe the generation of transgenic swine expressing recombinant human lysozyme by somatic cell nuclear transfer. Three cloned female pigs were born, one of which expressed rhLZ at 0.32 ± 0.01 μg/ml in milk, 50-fold higher than that of the pig native lysozyme. Both the transgenic gilts and their progeny appear healthy. Introducing human lysozyme into pigs' milk has a potential to benefit the piglets by enhancing immune function and defending against pathogenic bacteria, thereby increasing the new born survival rate. This advance could be of great value to commercial swine producers.
Abstract. The objective of the present study was to investigate the effect of addition of ghrelin to in vitro culture medium on preimplantation development of porcine in vitro fertilized and parthenogenetic embryos. In Experiment 1, we sought to compare the in vitro developmental competence of IVF and parthenogenetic embryos. No significant (P<0.05) differences were detected for cleavage rate or blastocyst rate between the in vitro fertilization (IVF)-and parthenogenetic activation-derived embryos. In Experiment 2, parthenogenetic embryos were cultured in Porcine Zygote Medium-3 containing various concentrations of ghrelin. The blastocyst rate was remakably (P<0.05) increased when 5 ng/ml (PA-5) and 500 ng/ml (PA-500) of ghrelin was added to in vitro culture medium compared with the other groups. Total cell number per blastocyst was slightly promoted in the ghrelin treatment groups compared with the controls. However, the ratio of inner cell mass (ICM) cell number/total cell number was significantly reduced in the PA-50 group compared with the controls (P<0.05). In Experiment 3, we cultured in vitro fertilized embryos in Porcine Zygote Medium-3 supplemented with ghrelin at different dosages. The rate of blasotcyst formation was markedly (P<0.05) elevated when 500 ng/ml ghrelin was added to culture medium (IVF-500) compared with the controls. Increased total cell numbers (P<0.05) were observed when in vitro fertilized embryos were cultured in IVF-50 and IVF-500 compared with the controls. However, the ratio of ICM cell number/total cell number was decreased in the ghrelin treatment groups compared with the controls (P<0.05). Taken together, the results suggest that ghrelin can enhance blastocyst formation of porcine in vitro fertilized and parthenogenetic embryos while exerting a negative effect on the structural integrity of the blastocysts. Key words: Embryo culture, Ghrelin, In vitro fertilization (IVF), Parthenogenetic, Pig (J. Reprod. Dev. 53: [647][648][649][650][651][652][653] 2007) he hormone ghrelin is a 28-amino acid-residue p e p t i d e w i t h a n e s s e n t i a l n -o c t a n o y l modification at the ser 3 residue [1] that is secreted predominantly by the stomach. Substantially lower amounts are distributed in the kidneys, pancreas, bowel, placenta, pituitary, testes, hypothalamus and lung [2,3]. The structure of ghrelin is highly conserved among humans, rodents and pigs, with changes in only three residues [2,4]. Apart from its p o t e n t g r o w t h h o r m o n e ( G H ) -r e l e a s i n g competence, ghrelin promotes appetite and a positive energy balance and elicits the production of prolactin (PRL) and adrenocorticotropic hormone (ACTH) by acting through the growth hormone secretagogue receptor (GHS-R), a typical
Background Perfluorooctanoic acid (PFOA) is widely used in the manufacture of household and industrial products. It has certain toxicity and leaves many residues in the environment. Numerous studies have shown that PFOA exhibits endocrine disrupting properties and immunotoxicity and induces developmental defects. However, there is very little information regarding its toxicity on oocytes. Methods We cultured denuded oocytes in maturation medium supplemented with 0, 300, or 500 PFOA during IVM and evaluated the maturation of oocytes from the aspects of ROS(DCFH‐DA), mitochondria(MitoOrange and JC‐1), DNA damage(P‐H2AX), and cytoskeleton(β‐tubulin). Results Compared with the control group, the PFOA treatment group exhibited significantly reduced proportion of oocytes matutation. Furthermore, the DCFH‐DA test showed that PFOA significantly increased reactive oxygen species (ROS) levels. PFOA disrupted mitochondrial distribution and decreased mitochondrial function as assessed using MitoOrange and JC‐1. In addition, PFOA‐treated oocytes exhibited a significantly higher percentage of P‐H2AX, defective β‐tubulin, abnormal chromosome alignment, lower expression of the anti‐apoptotic gene Bcl‐2, and higher expression of the apoptotic genes caspase3 and Bax. Conclusion In summary, PFOA could negatively and directly affect oocyte maturation in vitro and cause oxidative stress, mitochondrial function disruption, DNA damage, cytoskeleton damage, and apoptosis.
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