Integrated Transcriptome and Proteome Analyses of ß-Conglycinin-Induced Intestinal Damage in Piglets.

ß-conglycinin (ß-CG) induces intestinal damage in piglets; however, its regulatory mechanisms are not fully understood. This study aimed to investigate the molecular mechanisms by which ß-CG regulates intestinal injury in piglets through downstream genes and proteins. Our findings revealed that ß-CG significantly reduced villus height while increasing the crypt depth. In addition, we analyzed the transcriptome and proteome of jejunum tissues after the ß-CG treatment. In total, 382 differentially expressed genes (DEGs) and 292 differentially expressed proteins (DEPs) were identified between the treatment and the control groups. The expression levels of DEGs and DEPs were validated by using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blotting, respectively. The findings revealed a consistent correlation between their expression levels and transcriptomic and proteomic data. In addition, Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses of DEGs and DEPs revealed their enrichment in oxidation-related GOs, as well as in lysosome-related pathways. A protein-protein interaction (PPI) regulatory network was constructed based on the DEPs. The integration of transcriptomic and proteomic analyses identified six genes that were significantly different at both the transcript and the protein levels. This study provides valuable insights into the molecular mechanisms underlying ß-CG-induced intestinal injury in piglets.

Succinylation proteomic analysis identified differentially expressed succinylation sites affecting porcine muscle fiber type function.

Muscle fiber type is a major factor in pork meat quality, however, the role of post-translational protein modifications, especially succinylation, in the regulation of muscle fiber type is not fully understood. Here we performed protein succinylation profiles of fast-type biceps femoris (BF) and slow-type soleus (SOL) muscles. A total of 4,221 succinylation sites were identified from these samples, of which 294 sites were differentially expressed. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that these succinylated proteins were mainly involved in glycolysis, tricarboxylic acid cycle, and fatty acid metabolism. Succinylation modification of the CRAT and RAB10 proteins was verified by co-immunoprecipitation. Protein-protein interaction (PPI) network analysis unveiled the interactions of these succinylated proteins that regulate pig myofiber type conversion. This investigation offers fresh perspectives into the molecular roles of protein succinylation in the regulation of pig myofiber type transformation and meat quality.

Comparative Proteomic Analysis of Glycolytic and Oxidative Muscle in Pigs.

The quality of meat is highly correlated with muscle fiber type. However, the mechanisms via which proteins regulate muscle fiber types in pigs are not entirely understood. In the current study, we have performed proteomic profiling of fast/glycolytic biceps femoris (BF) and slow/oxidative soleus (SOL) muscles and identified several candidate differential proteins among these. We performed proteomic analyses based on tandem mass tags (TMTs) and identified a total of 26,228 peptides corresponding to 2667 proteins among the BF and SOL muscle samples. Among these, we found 204 differentially expressed proteins (DEPs) between BF and SOL muscle, with 56 up-regulated and 148 down-regulated DEPs in SOL muscle samples. KEGG and GO enrichment analyses of the DEPs revealed that the DEPs are involved in some GO terms (e.g., actin cytoskeleton, myosin complex, and cytoskeletal parts) and signaling pathways (PI3K-Akt and NF-kappa B signaling pathways) that influence muscle fiber type. A regulatory network of protein-protein interaction (PPI) between these DEPs that regulates muscle fiber types was constructed, which demonstrates how three down-regulated DEPs, including PFKM, GAPDH, and PKM, interact with other proteins to potentially control the glycolytic process. This study offers a new understanding of the molecular mechanisms in glycolytic and oxidative muscles as well as a novel approach for enhancing meat quality by transforming the type of muscle fibers in pigs.

An Integrated Analysis of Lactation-Related miRNA and mRNA Expression Profiles in Donkey Mammary Glands.

Donkey milk is consumed by humans for its nutritional and therapeutic properties. MicroRNAs (miRNAs) and messenger RNAs (mRNAs) have been implicated in the regulation of milk component synthesis and mammary gland development. However, the regulatory profile of the miRNAs and mRNAs involved in lactation in donkeys is unclear. We performed mRNA-seq and miRNA-seq and constructed coexpression regulatory networks for the mammary glands during the lactating and nonlactating period of jennies. We identified 3144 differentially expressed (DE) mRNAs (987 upregulated mRNAs and 2157 downregulated mRNAs) and 293 DE miRNAs (231 upregulated miRNAs and 62 downregulated miRNAs) in the lactating group compared to the nonlactating group. The DE miRNA target mRNA were significantly associated with pathways related to RNA polymerase, glycosphingolipid biosynthesis, mRNA surveillance, ribosome biogenesis in eukaryotes, glycerophospholipid metabolism, Ras signaling, and the fly hippo signaling pathway. The mRNA-miRNA coregulation analysis showed that novel-m0032-3p, miR-195, miR-26-5p, miR-23-3p, miR-674-3p, and miR-874-3p are key miRNAs that target mRNAs involved in immunity and milk lipid, protein, and vitamin metabolism in the jenny mammary gland. Our results improve the current knowledge of the molecular mechanisms regulating bioactive milk component metabolism in the mammary glands and could be used to improve milk production in donkeys.

Comparative analysis of differentially abundant proteins between high and low intramuscular fat content groups in donkeys.

Intramuscular fat (IMF) is an important regulator that determines meat quality, and its content is closely related to flavor, tenderness, and juiciness. Many studies have used quantitative proteomic analysis to identify proteins associated with meat quality traits in livestock, however, the potential candidate proteins that influence IMF in donkey muscle are not fully understood. In this study, we performed quantitative proteomic analysis, with tandem-mass-tagged (TMT) labeling, with samples from the longissimus dorsi (LD) muscle of the donkey. A total of 585,555 spectra were identified from the six muscle samples used in this study. In total, 20,583 peptides were detected, including 15,279 unique peptides, and 2,540 proteins were identified. We analyzed differentially abundant proteins (DAPs) between LD muscles of donkeys with high (H) and low (L) IMF content. We identified 30 DAPs between the H and L IMF content groups, of which 17 were upregulated and 13 downregulated in the H IMF group. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment analysis of these DAPs revealed many GO terms (e.g., bone morphogenetic protein (BMP) receptor binding) and pathways (e.g., Wnt signaling pathway and Hippo signaling pathway) involved in lipid metabolism and adipogenesis. The construction of protein-protein interaction networks identified 16 DAPs involved in these networks. Our data provide a basis for future investigations into candidate proteins involved in IMF deposition and potential new approaches to improve meat quality in the donkey.

Cardiac valve calcification prevalence and association with neutrophil-to-lymphocyte ratio in newly diagnosed patients with non-dialysis chronic kidney disease stage 3-5.

OBJECTIVE: Cardiac valvular calcification (CVC) is the main cause of cardiovascular disease and all-cause death in patients with chronic kidney disease (CKD). However, the relationship between Neutrophil lymphocyte ratio (NLR) and CVC in patients with CKD is not clear. In this study, we aimed to investigate the prevalence of CVC in newly diagnosed patients with non-dialysis CKD stage 3-5 and evaluate the correlation between NLR and CVC. METHODS: A total of 483 newly diagnosed patients with non-dialysis CKD stage 3-5 were included. According to the presence of CVC, these patients were retrospectively divided into two groups: CVC group and non-CVC group. RESULTS: CVC was found in 80 patients (16.56 %), 53 (10.97 %) of whom had only aortic valve calcification (AVC), 18 (3.73 %) had mitral valve calcification (MVC), and 9 (1.86 %) had both AVC and MVC. The level of NLR in the CVC group was significantly higher than that in the non-CVC group (p=0.002). Multivariate logistic regression analysis showed that NLR was an independent risk factor for CVC (95% CI 1.017~1.225, p=0.020). ROC curve analysis showed that the area under the curve of NLR for predicting CVC was 0.610 (95% CI 0.543-0.676, p=0.002). The best cut-off point of NLR was 3.340, with a sensitivity of 49.4 % and a specificity of 70.0 %. CONCLUSION: CVC is not uncommon in newly diagnosed patients with non-dialysis CKD stage 3-5, and NLR is an independent risk factor for CVC (Tab. 4, Fig. 1, Ref. 34).

AMPK-PPARγ-Cidec Axis Drives the Fasting-Induced Lipid Droplet Aggregation in the Liver of Obese Mice.

Intermittent fasting is one of the most common clinical treatments for the obesity, a main risk factor of the metabolic syndrome which can lead to a variety of diseases. Fasting-induced fat mobilization alters the metabolic state of lipid in the liver, predisposing to increase the hepatic lipid droplet aggregation and triglyceride levels. However, the underlying mechanisms regarding the lipid droplet aggregation in the liver after fasting remains elusive. Here, we report that a lipid droplet surface binding protein Cidec (cell death inducing DFFA like effector C) is activated by AMPK to regulate the hepatic lipid droplet fusion following fasting in obese mice. Specifically, we found that lipid droplets were significantly aggregated in the liver of high-fat-diet and ob/ob mice after 16 and 24 h of fasting, accompanied by the dramatically up-regulated expression of Cidec. Consistently, overexpression of Cidec in the AML12 cells resulted in the intracellular lipid droplet aggregation. Furthermore, we showed that fasting caused the up-regulated expression of AMPK, which in turn activated the transcription of Cidec through the transcription factor PPARγ. Altogether, our observations reveal that fasting-induced hepatic lipid droplet aggregation is mediated by the AMPK-activated expression of Cidec via PPARγ, extending our understanding about the molecular mechanism of the impact of fasting on the obesity and providing potential targets for the treatment of human obesity.

Reactive Flash Sintering of High-Entropy Oxide (Mg, Co, Ni, Cu, Zn)1-xLixO at Room Temperature.

(Mg, Co, Ni, Cu, Zn)1-xLixO is a type of high-entropy oxide that has high ionic conductivity at room temperature and is used as a solid electrolyte. (Mg, Co, Ni, Cu, Zn)1-xLixO was successfully synthesized from precursor powder by applying reactive flash sintering for less than 4 min at room temperature (25 °C). AC and DC electric fields were independently applied to sinter ceramic samples; consequently, AC and DC electric field application resulted in relative densities that exceeded 90% and 80%, respectively. X-ray diffraction spectra of samples revealed the existence of a clear halite structure with an insignificant impurity phase, proving that (Mg, Co, Ni, Cu, Zn)1-xLixO crystals were successfully produced.

Circular RNA Profiling Identifies Novel circPPARA that Promotes Intramuscular Fat Deposition in Pigs.

Intramuscular fat (IMF) content plays an important role in pork quality. Circular RNAs (circRNAs) implicate various biological processes; however, the regulatory mechanisms and functions of circRNAs in porcine IMF remains elusive. Hence, the study assessed the circRNA expression profiling in the longissimus dorsi muscle of pigs with high (H) and low (L) IMF content to unravel their regulatory functions in improving meat quality. The RNA sequencing analysis identified 29,732 circRNAs from six sampled pigs, most of which were exon-derived. In the muscle, 336 were differentially expressed (DE) between the H and L IMF groups; 196 circRNAs were upregulated, and 140 were downregulated. Subsequent qRT-PCR validation of 10 DE circRNAs revealed expression patterns consistent with the RNA-seq data. Gene ontology and KEGG enrichment analysis revealed that most significantly enriched DE circRNAs' host genes were linked to lipid metabolism and adipogenesis processes. The circRNA-miRNA regulatory network analysis found several circRNAs targeting miRNAs associated with adipogenesis. Finally, a novel circRNA, circPPARA, was identified with the expression positively correlated with the IMF content. Detailed analysis revealed that circPPARA was formed via head-to-tail splicing and was more stable than the linear PPARA, predominantly located in the cytoplasm. Functional studies using overexpression and siRNA constructs demonstrated that circPPARA promotes differentiation and hinders the proliferation of porcine intramuscular preadipocytes. Moreover, the dual-luciferase assay revealed that circPPARA adsorbed miR-429 and miR-200b, thereby promoting intramuscular adipogenesis in pigs. Our results identified a candidate circRNA, circPPARA, that affects porcine IMF content. The study provides knowledge of the regulatory functions of circRNAs in intramuscular adipogenesis and abundant resource for future research on circRNAs in pigs.

Impact of adrenocorticotropin hormone administration on the endocrinology, estrus onset, and ovarian function of weaned sows.

Chronic stress affects the reproductive health of mammals; however, the impact of adrenocorticotropin hormone (ACTH) level elevation during chronic stress on the reproduction of weaned sows remains unclear. In this study, nine weaned sows with the same parturition date were randomly divided into control group (n = 4) and ACTH group (n = 5). Each group received intravenous administration of ACTH three times daily for 7 days. Blood samples were collected every 3 h after injection. A radioimmunoassay was used to measure the concentrations of cortisol, luteinizing hormone (LH), follicle-stimulating hormone (FSH), progesterone (P4) and estradiol-17ß (E2) in the blood. Estrus was determined according to changes in the vulva and the boar contact test. The mRNA expressions of glucocorticoid receptor, FSH receptor, LH receptor (LHR) in the corpus luteum (CL) were detected by qRT-PCR. The results showed that ACTH administration substantially delayed the initiation of estrus and the pre-ovulatory LH peak. The sows of control group ovulated within 10 days and the ovulation rate was 100%, while it was 60% in the ACTH group. Two sows of ACTH group showed pseudo-estrus. The E2 concentrations significantly decreased in the ACTH group at 36 h, 42 h and 66 h of the experimental period. The P4 concentrations in the ACTH group significantly decreased at 132, 138, and 147 h of the experimental period. ACTH significantly reduced the LHR mRNA expression in CLs. In conclusion, long-term repeated ACTH administration affects the endocrinology, estrus onset, and ovarian function of weaned sows.

Phosphoproteomic analysis identifies differentially expressed phosphorylation sites that affect muscle fiber type in pigs.

Skeletal muscle of livestock is composed of both fast- and slow-twitch muscle fibers, which are key factors in their meat quality. However, the role of protein phosphorylation in muscle fiber type is not completely understood. Here, a fast-twitch (biceps femoris, BF) and slow-twitch (soleus, SOL) muscle tissue sample was collected from three male offspring of Duroc and Meishan pigs. We demonstrate that the meat quality of SOL muscle is significantly better than that of BF muscle. We further used phosphoproteomic profiling of BF and SOL muscles to identify differences between these muscle types. A total of 2,327 phosphorylation sites from 770 phosphoproteins were identified. Among these sites, 287 differentially expressed phosphorylation sites (DEPSs) were identified between BF and SOL. GO and KEGG enrichment analysis of proteins containing DEPSs showed that these phosphorylated proteins were enriched in the glycolytic process GO term and the AMPK signaling pathway. A protein-protein interaction (PPI) analysis reveals that these phosphorylated proteins interact with each other to regulate the transformation of muscle fiber type. These analyses reveal that protein phosphorylation modifications are involved in porcine skeletal muscle fiber type transformation. This study provides new insights into the molecular mechanisms by which protein phosphorylation regulates muscle fiber type transformation and meat quality in pigs.

Long non-coding RNA Mir22hg-derived miR-22-3p promotes skeletal muscle differentiation and regeneration by inhibiting HDAC4.

Emerging studies have indicated that long non-coding RNAs (lncRNAs) play important roles in skeletal muscle growth and development. Nevertheless, it remains challenging to understand the function and regulatory mechanisms of these lncRNAs in muscle biology and associated diseases. Here, we identify a novel lncRNA, Mir22hg, that is significantly upregulated during myoblast differentiation and is highly expressed in skeletal muscle. We validated that Mir22hg promotes myoblast differentiation in vitro. Mechanistically, Mir22hg gives rise to mature microRNA (miR)-22-3p, which inhibits its target gene, histone deacetylase 4 (HDAC4), thereby increasing the downstream myocyte enhancer factor 2C (MEF2C) and ultimately promoting myoblast differentiation. Furthermore, in vivo, we documented that Mir22hg knockdown delays repair and regeneration following skeletal muscle injury and further causes a significant decrease in weight following repair of an injured tibialis anterior muscle. Additionally, Mir22hg gives rise to miR-22-3p to restrict HDAC4 expression, thereby promoting the differentiation and regeneration of skeletal muscle. Given the conservation of Mir22hg between mice and humans, Mir22hg might constitute a promising new therapeutic target for skeletal muscle injury, skeletal muscle atrophy, as well as other skeletal muscle diseases.

DNMTs Play an Important Role in Maintaining the Pluripotency of Leukemia Inhibitory Factor-Dependent Embryonic Stem Cells.

Naive pluripotency can be maintained in medium with two inhibitors plus leukemia inhibitory factor (2i/LIF) supplementation, which primarily affects canonical WNT, FGF/ERK, and JAK/STAT3 signaling. However, whether one of these three supplements alone is sufficient to maintain naive self-renewal remains unclear. Here we show that LIF alone in medium is sufficient for adaptation of 2i/L-ESCs to embryonic stem cells (ESCs) in a hypermethylated state (L-ESCs). Global transcriptomic analysis shows that L-ESCs are close to 2i/L-ESCs and in a stable state between naive and primed pluripotency. Notably, our results demonstrate that DNA methyltransferases (DNMTs) play an important role in LIF-dependent mouse ESC adaptation and self-renewal. LIF-dependent ESC adaptation efficiency is significantly increased in serum treatment and reduced in Dnmt3a or Dnmt3l knockout ESCs. Importantly, unlike epiblast stem cells, L-ESCs contribute to somatic tissues and germ cells in chimeras. L-ESCs cultured under such simple conditions as in this study would provide a more conducive platform to clarify the molecular mechanism of ESCs in in vitro culture.

Transcriptome analysis reveals the genetic basis of skeletal muscle glycolytic potential based on a pig model.

Glycolytic potential (GP) calculated based on glucose, glycogen, glucose-6-phosphate, and lactate contents is a critical factor for multiple meat quality characteristics. However, the genetic basis of glycolytic metabolism is still unclear. In this study, we constructed six RNA-Seq libraries using longissimus dorsi (LD) muscles from pigs divergent for GP phenotypic values and generated the whole genome-wide gene expression profiles. Furthermore, we identified 25,880 known and 220 novel genes from these skeletal muscle libraries, and 222 differentially expressed genes (DEGs) between the higher and lower GP groups. Notably, we found that the Lactate dehydrogenase B (LDHB) and Fructose-2, 6-biphosphatase 3 (PFKFB3) expression levels were higher in the higher GP group than the lower GP group, and positively correlated with GP and lactic acid (LA), and reversely correlated with pH value at 45 min postmortem (pH45min). Besides, LDHB and PFKFB3 expression were positively correlated with drip loss measured at 48 h postmortem (DL48h) and drip loss measured at 24 h postmortem (DL24h). Collectively, we identified a serial of DEGs as the potential key candidate genes affecting GP and found that LDHB and PFKFB3 are closely related to GP and GP-related traits. Our results lay a solid basis for in-depth studies of the regulatory mechanisms on GP and GP-related traits in pigs.

Exploring the lncRNAs Related to Skeletal Muscle Fiber Types and Meat Quality Traits in Pigs.

The alteration in skeletal muscle fiber is a critical factor affecting livestock meat quality traits and human metabolic diseases. Long non-coding RNAs (lncRNAs) are a diverse class of non-coding RNAs with a length of more than 200 nucleotides. However, the mechanisms underlying the regulation of lncRNAs in skeletal muscle fibers remain elusive. To understand the genetic basis of lncRNA-regulated skeletal muscle fiber development, we performed a transcriptome analysis to identify the key lncRNAs affecting skeletal muscle fiber and meat quality traits on a pig model. We generated the lncRNA expression profiles of fast-twitch Biceps femoris (Bf) and slow-twitch Soleus (Sol) muscles and identified the differentially expressed (DE) lncRNAs using RNA-seq and performed bioinformatics analyses. This allowed us to identify 4581 lncRNA genes among six RNA libraries and 92 DE lncRNAs between Bf and Sol which are the key candidates for the conversion of skeletal muscle fiber types. Moreover, we detected the expression patterns of lncRNA MSTRG.42019 in different tissues and skeletal muscles of various development stages. In addition, we performed a correlation analyses between the expression of DE lncRNA MSTRG.42019 and meat quality traits. Notably, we found that DE lncRNA MSTRG.42019 was highly expressed in skeletal muscle and its expression was significantly higher in Sol than in Bf, with a positive correlation with the expression of Myosin heavy chain 7 (MYH7) (r = 0.6597, p = 0.0016) and a negative correlation with meat quality traits glycolytic potential (r = -0.5447, p = 0.0130), as well as drip loss (r = -0.5085, p = 0.0221). Moreover, we constructed the lncRNA MSTRG.42019-mRNAs regulatory network for a better understanding of a possible mechanism regulating skeletal muscle fiber formation. Our data provide the groundwork for studying the lncRNA regulatory mechanisms of skeletal muscle fiber conversion, and given the importance of skeletal muscle fiber types in muscle-related diseases, our data may provide insight into the treatment of muscular diseases in humans.

Profiling and Functional Analysis of Circular RNAs in Porcine Fast and Slow Muscles.

The different skeletal muscle fiber types exhibit distinctively different physiological and metabolic properties, and have been linked to both human metabolic diseases and meat quality traits in livestock. Circular RNAs (circRNAs) are a new class of endogenous RNA regulating gene expression, but regulatory mechanisms of skeletal muscle fibers involved in circRNAs remain poorly understood. Here, we constructed circRNA expression profiles of three fast-twitch biceps femoris (Bf) and three slow-twitch soleus (Sol) muscles in pigs using RNA-seq and identified 16,342 distinct circRNA candidates. Notably, 242 differentially expressed (DE) circRNAs between Bf and Sol muscles were identified, including 105 upregulated and 137 downregulated circRNAs, and are thus potential candidates for the regulation of skeletal muscle fiber conversion. Moreover, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of host genes of DE circRNAs revealed that host genes were mainly involved in skeletal muscle fiber-related GO terms (e.g., muscle contraction, contractile fiber part, and Z disk) and skeletal muscle fiber-related signaling pathways (e.g., AMPK and cGMP-PKG). We also constructed co-expression networks of DE circRNA-miRNA-mRNA using previously acquired high-throughput sequencing mRNA and miRNA data, from which 112 circRNA-miRNA and 95 miRNA-mRNA interactions were identified. Multiple circRNAs essentially serve as a sponge for miR-499-5p, which is preferentially expressed in slow-twitch muscle and reduces the severity of Duchenne muscular dystrophy (DMD). Taken together, a series of novel candidate circRNAs involved in the growth and development of porcine skeletal muscle was identified. Furthermore, they provide a comprehensive circRNA resource for further in-depth research on the regulatory mechanisms of circRNA in the formation of skeletal muscle fiber, and may provide insights into human skeletal muscle diseases.

LncRNA 2310043L19Rik inhibits differentiation and promotes proliferation of myoblast by sponging miR-125a-5p.

Although many long non-coding RNAs (lncRNAs) have been identified in muscle, some of their physiological functions and regulatory mechanisms remain elusive. Here we report the functional identification and characterization of a novel lncRNA 2310043L19Rik (lnc-231), which is highly expressed in muscle. The expression level of lnc-231 in skeletal muscle of young mice is higher than that in aged mice. Functional analysis showed that overexpression of lnc-231 restrained differentiation and promoted proliferation of myoblast, while inhibition of lnc-231 revealed completely opposite effects in vitro. RNA molecules of lnc-231 acted mechanistically as competing endogenous RNAs (ceRNA) to target miR-125a-5p, whereas miR-125a-5p binds to the 3'-UTR of E2F3 mRNA to inhibit its function. Collectively, lncRNA 2310043L19Rik promotes proliferation and inhibits differentiation of myoblast cells by attenuating the function of miR-125a-5p.

Activin A and BMP4 Signaling Expands Potency of Mouse Embryonic Stem Cells in Serum-Free Media.

Inhibitors of Mek1/2 and Gsk3ß, known as 2i, and, together with leukemia inhibitory factor, enhance the derivation of embryonic stem cells (ESCs) and promote ground-state pluripotency (2i/L-ESCs). However, recent reports show that prolonged Mek1/2 suppression impairs developmental potential of ESCs, and is rescued by serum (S/L-ESCs). Here, we show that culturing ESCs in Activin A and BMP4, and in the absence of MEK1/2 inhibitor (ABC/L medium), establishes advanced stem cells derived from ESCs (esASCs). We demonstrate that esASCs contributed to germline lineages, full-term chimeras and generated esASC-derived mice by tetraploid complementation. We show that, in contrast to 2i/L-ESCs, esASCs display distinct molecular signatures and a stable hypermethylated epigenome, which is reversible and similar to serum-cultured ESCs. Importantly, we also derived novel ASCs (blASCs) from blastocysts in ABC/L medium. Our results provide insights into the derivation of novel ESCs with DNA hypermethylation from blastocysts in chemically defined medium.

Comprehensive analysis of circRNAs from cashmere goat skin by next generation RNA sequencing (RNA-seq).

Circular RNA (circRNA) is endogenous non-coding RNA (ncRNA) with a covalently closed circular structure. It is mainly generated through RNA alternative splicing or back-splicing. CircRNA is known in the majority of eukaryotes and very stable. However, knowledge of the circRNA involved in regulating cashmere fineness is limited. Skin samples were collected from Liaoning cashmere goats (LCG) and Inner Mongolia cashmere goats (MCG) during the anagen period. For differentially expressed circRNAs, RNA sequencing was performed, and the analysis led to an identification of 17 up-regulated circRNAs and 15 down-regulated circRNAs in LCG compared with MCG skin samples. In order to find the differentially expressed circRNAs in LCG, we carried out qPCRs on 10 candidate circRNAs in coarse type skin of LCG (CT-LCG) and fine type skin of LCG (FT-LCG). Four circRNAs: ciRNA128, circRNA6854, circRNA4154 and circRNA3620 were confirmed to be significantly differential expression in LCG. Also, a regulatory network of circRNAs-miRNAs was bioinformatically deduced and may help to understand molecular mechanisms of potential circRNA involvement in regulating cashmere fineness.

Identification of Candidate Circular RNAs Underlying Intramuscular Fat Content in the Donkey.

Intramuscular fat (IMF) content is a crucial indicator of meat quality. Circular RNAs (circRNAs) are a large class of endogenous RNAs that are involved in many physiological processes. However, the expression and function of circRNA in IMF in the donkey remains unresolved. Here we performed an expression profiling of circRNAs in the donkey longissimus dorsi muscle and identified 12,727 candidate circRNAs. Among these, 70% were derived from the exons of protein genes. Furthermore, a total of 127 differentially expressed (DE) circRNAs were identified in high (H) and low (L) IMF content groups, including 63 upregulated and 64 downregulated circRNAs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of the host genes of the DE circRNAs showed that the host genes were enriched in lipid metabolism related GO terms (e.g., fatty acid beta-oxidation using acyl-CoA dehydrogenase and MLL3/4 complex), and signaling pathways (e.g., TGF-beta and lysine degradation signaling pathway). Further analyses indicated that 127 DE circRNAs were predicted to potentially interact with miRNAs, leading to the construction of circRNA-miRNA regulatory network. Multiple circRNAs can potentially function as sponges of miRNAs that regulate the differentiation of adipocytes. Our results provide valuable expression profile information for circRNA in the donkey and new insight into the regulatory mechanisms of circRNAs in the regulation of IMF content.