Zhanqin Zhou

Identification and Profiling of microRNAs in Goat Endometrium during Embryo Implantation

Background MicroRNAs (miRNAs) are short, highly conserved small noncoding RNAs that had fundamental roles in post-transcriptional gene expression, and they are crucial for proper control of biological processes and known to participate in embryo implantation. However, miRNA expression profiles in the pre-receptive and receptive phases of the goat endometrium during embryo implantation are unknown. Results A total of 1,069 and 847 miRNAs were expressed in receptive (R) and pre-receptive (P) goat endometrium, and 632 miRNAs were co-expressed in both phases. We identified 545 (50.98%) known miRNAs in the R library and 522 (61.63%) in the P library. There were 110 up-expressed miRNAs and 33 down-expressed miRNAs in receptive endometrium compared with the pre-receptive endometrium meeting the criteria of P-values< 0.05. Moreover, GO and KEGG analysis of the target genes of the differentially expressed miRNAs revealed some candidate miRNAs, genes and pathways that may involve in the formation of the receptive endometrium. Based on stem-loop RT-qPCR, 15 miRNAs were detected and the results suggested that the majority of the miRNA expression data measured by Solexa deep sequencing could represent actual miRNA expression levels. Conclusions Our data revealed the first miRNA profile related to the biology of the goat receptive endometrium during embryo implantation, and the results suggested that a subset of miRNAs might play important roles in the formation of endometrial receptivity. Thus, elucidating the physiological roles of endometrial miRNAs will help us better understand the genetic control of embryo implantation in goats.

Characterization of the Transcriptional Complexity of the Receptive and Pre-receptive Endometria of Dairy Goats.

Endometrium receptivity is essential for successful embryo implantation in mammals. However, the lack of genetic information remains an obstacle to understanding the mechanisms underlying the development of a receptive endometrium from the pre-receptive phase in dairy goats. In this study, more than 4 billion high-quality reads were generated and de novo assembled into 102,441 unigenes; these unigenes were annotated using published databases. A total of 3,255 unigenes that were differentially expressed (DEGs) between the PE and RE were discovered in this study (P-values < 0.05). In addition, 76,729–77,102 putative SNPs and 12,837 SSRs were discovered in this study. Bioinformatics analysis of the DEGs revealed a number of biological processes and pathways that are potentially involved in the establishment of the RE, notably including the GO terms proteolysis, apoptosis, and cell adhesion and the KEGG pathways Cell cycle and extracellular matrix (ECM)-receptor interaction. We speculated that ADCY8, VCAN, SPOCK1, THBS1, and THBS2 may play important roles in the development of endometrial receptivity. The de novo assembly provided a good starting point and will serve as a valuable resource for further investigations into endometrium receptivity in dairy goats and future studies on the genomes of goats and other related mammals.

MiR-26a promoted endometrial epithelium cells (EECs) proliferation and induced stromal cells (ESCs) apoptosis via the PTEN-PI3K/AKT pathway in dairy goats†

Changes in endometrial cell morphology and function are absolutely necessary for successful embryo implantation. In this study, miR‐26a was widely expressed in dairy goats, and was found to be regulated by β‐estradiol (E2) and progesterone (P4) in endometrial epithelium cells (EECs) as well as stromal cells (ESCs). Furthermore, miR‐26a played a role in the regulation of cells proliferation and apoptosis by directly regulating PTEN and indirectly regulating the PI3K/AKT pathway in EECs but not in ESCs of dairy goats in vitro. In addition, miR‐26a regulated the expression of osteopontin (OPN), vascular endothelial growth factor (VEGF), Cyclooxygenase‐2 (COX‐2), and prolactin (PRL) in endometrial cells. Therefore, we could get a conclusion that miR‐26a had very complex and diverse functions in the endometrial cells during the development of endometrial receptivity in dairy goats. This study provided an efficient platform for studying the regulatory effect of miR‐26a on endometrial cells during the development of endometrial receptivity in dairy goats.

The developmental transcriptome landscape of receptive endometrium during embryo implantation in dairy goats

Under natural conditions, some embryos cannot implant successfully because of the dysfunction of receptive endometrium (RE). Thus, it is imperative for us to study the molecular mechanisms involved in the formation of the RE from pre-receptive endometrium (PE). In this study, the endometrium from gestational day 5 (D5, PE) and gestational day 15 (D15, RE) dairy goats were selected to systematically analyze the transcriptome using strand-specific Ribo-Zero RNA-Seq, >120 million high-quality paired-end reads were generated and 47,616 transcripts were identified in the endometrium of dairy goats. A total of 810 mRNAs were differentially expressed genes (DEGs) between the RE and PE meeting the criteria of P-values<0.05. Bioinformatics analysis of the DEGs revealed that a number of biological processes and pathways were potentially involved in the establishment of the RE, notably energy metabolism and amino acid metabolism. Furthermore, we speculated that CXCL14, IGFBP3, and LGALS15 potentially participated in the development of endometrium. Whats more, putative SNPs, InDels and AS events were identified and analyzed in the endometrium. In a word, this resulting view of the transcriptome greatly enhances the comprehensive transcript catalog and uncovers the global trends in gene expression during the formation of receptive endometrium in dairy goats.

The effects of ovariectomy on meat performance and expression of GH/IGF-I in young goats

Zhang, L., Wang, Y.-y., Fu, M.-z., Li, G., An, N., Li, S.-y. and Zhou, Z.-q. 2014. The effects of ovariectomy on meat performance and expression of GH/IGF-I in young goats. Can. J. Anim. Sci. 94: 619-626. Experiments were carried out to investigate the effects of ovariectomy on meat production efficiency and to explore the expression of GH/IGF-I in young goats. Animal performance, meat quality, levels of serum growth hormone (GH) and insulin-like growth factor 1 (IGF-I), and mRNA levels of three key genes [GH Receptor (GHR), IGF-I and IGF-I Receptor (IGF-IR)] in longissimus dorsi and biceps femoris muscles were measured. The results show that carcass weight, net meat mass, fat weight and loin eye area of ovariectomized goats were higher than those of the controls, and ovariectomized goats lost 0.40 kg of bone weight (P<0.05). There was no statistically valid difference for the color, pH, water-holding capacity, or cooking rate of meat (P>0.05) between the two groups, except for the shear value, which was significantly lower in the Ovx group than in the control group (P<0.05). The results of this research show for the first time a significant trend (P<0.05) for serum GH and IGF-I in the direction of increasing in ovariectomized goats. Furthermore, the mRNA levels of GHR, IGF-I and IGF-IR in muscle were all up-regulated, except for the IGF-I gene in biceps femoris, by ovariectomy. In summary, ovariectomy showed a beneficial promotion in animal performance, but did not reduce meat quality, and increased serum GH and IGF-I and mRNA expression levels of GHR, IGF-I and IGF-IR in young female goats.

The Effect of Deposition Se on the mRNA Expression Levels of GPxs in Goats from a Se-enriched County of China

Previous studies revealed that Se was an important regulatory factor for glutathione peroxidase (GPx) genes. However, the relationship between Se concentrations and mRNA expression levels of GPxs were unclear in goats, especially the goats living in natural Se-enriched area. Thus, the aim of this study was to determine the Se concentrations and the mRNA expression levels of GPx-1, GPx-2, GPx-3, and GPx-4 in goats from Ziyang County (ZY-H and ZY-L goats) and Baoji City (BJ-P goats), which were Se-rich region and Se-poor region in China, respectively. Atomic fluorescence spectrometry was used as an essential method to determine the Se concentrations in heart, liver, spleen, lung, kidney, longissimus, biceps femoris, and serum, and the gene expressions were quantified in mRNA samples extracted from the above tissues by real-time quantitative reverse transcription–polymerase chain reaction. We found that the Se concentrations in ZY-H and ZY-L goats were higher than that in BJ-P goats significantly (P < 0.05), and the pertinence relations of Se levels between serum and heart, liver, spleen, and kidney were significant (P < 0.05). The mRNA levels of GPx-1 in ZY-H and ZY-L goats were higher than that in BJ-P goats very significantly (P < 0.01) except for longissimus (P < 0.05). Our results indicated a significant trend for GPx-2 in the direction of increasing mRNA levels with increasing Se concentrations in goats but had no statistical significance (P > 0.05) in our experimental conditions. As to GPx-3, its mRNA expression in spleen, lung, and kidney (P < 0.05) were upregulated and were consensual to high Se contents in ZY-H goats, but no significant effects were observed in heart, liver, longissimus, and biceps femoris among our three groups (P > 0.05). The mRNA levels of GPx-4 in heart, liver, lung, and kidney of ZY-H and ZY-L goats were higher than that of BJ-P goats (P < 0.05), and the difference was very significant in lung especially (P < 0.01), but no change in spleen, longissimus, and biceps femoris (P > 0.05). In summary, these data suggested that the goats living in Ziyang County were rich in Se, and the deposition Se played important roles in the mRNA expression of GPx-1, GPx-3, and GPx-4 in certain tissues of goats differentially.

miR-182 aids in receptive endometrium development in dairy goats by down-regulating PTN expression

Increasing evidence has shown that miRNAs play important roles in endometrium development during the menstrual cycle in humans and many other animals. Our previous data indicated that miR-182 levels increase 15.55-fold and pleiotrophin (PTN) levels decrease 20.97-fold in the receptive endometrium (RE, D15) compared with the pre-receptive endometrium (PE, D5) in dairy goats. The present study shows that miR-182 is widely expressed in different tissues of dairy goats and that its expression levels are regulated by E2 and P4 in endometrial epithelium cells (EECs). We confirmed that PTN is a target of miR-182 and that miR-182 regulates the protein levels of AKT, Bcl-2, FAS, MAPK, Caspase-3 and SP1 in EECs. Furthermore, miR-182 up-regulates or maintains the expression levels of osteopontin (OPN), cyclooxygenase-2 (COX-2) and prolactin receptor (PRLR) in EECs, suggesting that miR-182 is an important regulatory factor in the construction of endometrial receptivity in dairy goats. In conclusion, miR-182 participates in the development of endometrial receptivity by down-regulating PTN and affecting the expression of select apoptosis-related genes and increasing or maintaining the expression levels of OPN, COX-2 and PRLR in the EECs of dairy goats.

Fatty acid composition and mRNA expression levels of lipid-metabolic genes in the muscles of ovariectomised young goats

This study investigated the fatty acid composition and mRNA levels of acetyl-coenzyme A carboxylase (ACC), fatty acid synthase (FAS), hormone-sensitive lipase (HSL) and lipoprotein lipase (LPL) genes in the Longissimus dorsi muscle (LD) and Biceps femoris muscle (BF) of nine ovariectomised young goats, 50 days after ovariectomy at 5 months of age, relative to nine Control goats. Ovariectomy resulted in faster growth rates (130 vs 110 g/days; P = 0.027), more intramuscular fat (3.37% vs 1.62%; P = 0.009) and heavier final weights (28.3 vs 26.6 kg; P = 0.035). The total saturated fatty acid, C15:0 in the LD and C6:0, C11:0, C12:0, C15:0 and C21:0 in the BF were lower in ovariectomised goats than that in the Control goats (P < 0.05). Ovariectomy increased the total monounsaturated fatty acids in the LD (P < 0.05) and BF (P < 0.01). In the BF, the total polyunsaturated fatty acids and C18:2 of ovariectomised goats were lower than those of the Control goats (P < 0.05), and the C20:2 content significantly decreased in the LD (P < 0.05). The LD of the ovariectomised goats expressed significantly higher FAS but lower HSL mRNA levels than those of Control goats (P < 0.01). Significantly higher ACC, FAS and LPL and lower HSL mRNA levels were observed in the BF of the ovariectomised goats than in the BF of Controls (P < 0.05). These data suggest that ovariectomy affects fatty acid composition potentially by altering the expression of lipid-metabolic genes in the muscles of young goats.

Polymorphisms of LHβ and GnRHR genes and their association with the number of embryos recovered in goats

The objective of the present study was to explore a predictor of superovulation response on the basis of associations between the number of embryos recovered and gene polymorphism. Variation in the goat LHβ and GnRHR genes was investigated using polymerase chain reaction–single-strand conformational polymorphism and DNA sequencing. Two single nucleotide polymorphisms (SNPs) were identified in the 5′-UTR of LHβ gene (A59C, P1 locus) and in the Exon 2 of GnRHR gene (T177A, P6 locus). At the P1 locus in both breeds, the frequencies of one allele were 0.46 and 0.51, respectively. At the P6 locus, the minor allele frequency was 0.23. Associations of both SNPs with the number of embryos recovered and the corpus luteum number were evaluated in Boer and Shaanbei goat breeds. Association analysis showed that both SNPs had significant (P < 0.05) effects on the number of embryos recovered and corpus luteum number. These results indicate that LHβ and GnRHR genes are potential markers for the number of embryos recovered.