Integrative ATAC-seq and RNA-seq analysis of myogenic differentiation of ovine skeletal muscle satellite cell.

Skeletal muscle satellite cells (SMSCs) play an important role in regulating muscle growth and regeneration. Chromatin accessibility allows physical interactions that synergistically regulate gene expression through enhancers, promoters, insulators, and chromatin binding factors. However, the chromatin accessibility altas and its regulatory role in ovine myoblast differentiation is still unclear. Therefore, ATAC-seq and RNA-seq analysis were performed on ovine SMSCs at the proliferation stage (SCG) and differentiation stage (SCD). 17,460 DARs (differential accessibility regions) and 3732 DEGs (differentially expressed genes) were identified. Based on joint analysis of ATAC-seq and RNA-seq, we revealed that PI3K-Akt, TGF-ß and other signaling pathways regulated SMSCs differentiation. We identified two novel candidate genes, FZD5 and MAP2K6, which may affect the proliferation and differentiation of SMSCs. Our data identify potential cis regulatory elements of ovine SMSCs. This study can provide a reference for exploring the mechanisms of the differentiation and regeneration of SMSCs in the future.

Whole-genome identification of transposable elements reveals the equine repetitive element insertion polymorphism in Chinese horses.

Transposable elements (TEs) are diverse, abundant, and complicated in genomes. They not only can drive the genome evolution process but can also act as special resources for adaptation. However, little is known about the evolutionary processes that shaped horses. In this work, 126 horse assemblages involved in most horse breeds in China were used to investigate the patterns of TE variation for the first time. By using RepeatMasker and melt software, we found that the horse-specific short interspersed repetitive elements family, equine repetitive elements (ERE1), exhibited polymorphisms in horse genomes. Phylogenetic analysis based on these ERE1 loci (minor allele frequency ≥0.05) revealed three major horse groups, namely, those in northern China, southern China, and Qinghai-Tibetan, which mirrors the result determined by SNPs to some extent. The present ERE1 family emerged ~0.26 to 1.77 Mya ago, with an activity peak at ~0.49 Mya, which matches the early stage of the horse lineage and decreases after the divergence of Equus caballus and Equus ferus przewalskii. To detect the functional ERE1(s) associated with adaptation, locus-specific branch length, genome-wide association study, and absolute allele frequency difference analyses were conducted and resulted in two common protein-coding genes annotated by candidate ERE1s. They were clustered into the vascular smooth muscle contraction (p = 0.01, EDNRA) and apelin signalling pathways (p = 0.02, NRF1). Notably, ERE1 insertion into the EDNRA gene showed a higher association with adaptation among southern China horses and other horses in 15 populations and 451 individuals (p = 4.55 e-8). Our results provide a comprehensive understanding of TE variations to analyse the phylogenetic relationships and traits relevant to adaptive evolution in horses.

A study using single-locus and multi-locus genome-wide association study to identify genes associated with teat number in Hu sheep.

The multiple teats trait is common in many species of mammals and is considered related to lactation ability in swine. However, in Hu sheep, related gene research is still relatively limited. In this study, a genome-wide association study was used to identify genetic markers and genes related to the number of teats in the Hu sheep population, a native Chinese sheep breed. A single marker method and several multi-locus methods were utilized. A total of 61 SNPs were found to be related to the number of teats. Among these, 11 SNPs and one SNP were consistently detected by two and three multi-locus models respectively. Four SNPs were concordantly identified between the single marker and multi-locus methods. We also performed quantitative real-time PCR testing of these identified candidate genes, identifying three genes with significantly different expression. Our study suggested that the LHFP, DPYSL2, and TDP-43 genes may be related to the number of teats in sheep. The combination of single and multi-locus GWAS detected additional SNPs not found with only one model. Our results provide new and important insights into the genetic mechanisms of the mammalian multiparous teat phenotype. These findings may be useful for future breeding and understanding the genetics of sheep and other livestock.

Genome-Wide Analysis of Circular RNAs Reveals circCHRNG Regulates Sheep Myoblast Proliferation via miR-133/SRF and MEF2A Axis.

As relatively new members of the non-coding RNA family, circRNAs play important roles in a variety of biological processes. However, the temporal expression pattern and the function of circRNAs during sheep skeletal muscle development remains unclear. This study aimed to identify circRNAs related to sheep skeletal muscle development and explore their roles in myoblast proliferation. The circRNA expression profiles of longissimus dorsi of sheep from F90, L30, and A3Y were obtained by the RNA-seq method. The function and mechanisms of the novel circCHRNG in muscle satellite cell proliferation were explored using CCK-8 assay, Western blot, qPCR, and dual-luciferase reporter assay. We identified 12,375 circRNAs, including 476, 133, and 233 DEcircRNAs found among three comparative groups. KEGG results showed that DEcircRNAs were enriched in muscle contraction, the regulation of cell proliferation, and the AMPK, insulin, and PI3K-Akt signaling pathways. Notably, a novel circRNA, termed circRNA CHRNG, acts as a miR-133 sponge to promote skeletal muscle satellite cell proliferation. Our study provides a systematic description of circRNAs of ovine skeletal muscle across fetal, lamb, and adult stages. GO and KEGG analyses showed that DEcircRNAs were enriched in multiple pathways associated with muscle development, such as the PI3K-Akt and AMPK signaling pathways. In addition, we propose that circCHRNG acts as a miR-133 sponge to upregulate the expression levels of SRF and MEF2A, thereby promoting myoblast proliferation.

Comprehensive analysis of lncRNAs involved in skeletal muscle development in ZBED6-knockout Bama pigs.

BACKGROUND: The mutation of insulin-like growth factor 2 (IGF2 mutation) that a single-nucleotide substitution (G→A) in the third intron of IGF2 abrogates the interaction with zinc finger BED-type containing 6 (ZBED6) and leads to increased muscle mass in pigs. IGF2 mutation knock-in (IGF2 KI) and ZBED6 knockout (ZBED6 KO) lead to changes in IGF2 expression and increase muscle mass in mice and pigs. Long noncoding RNAs (lncRNAs) may participate in numerous biological processes, including skeletal muscle development. However, the role of the ZBED6-lncRNA axis in skeletal muscle development is poorly characterized. RESULTS: In this study, we assembled transcriptomes using RNA-seq data published in previous studies by our group and identified 11,408 known lncRNAs and 2269 potential lncRNAs in seven tissues, heart, longissimus dorsi, gastrocnemius muscle, liver, spleen, lung and kidney, of ZBED6 KO (lean mass model) and WT Bama pigs. ZBED6 affected the expression of 1570 lncRNAs (differentially expressed lncRNAs [DE-lncRNAs]; log2-fold change ≥ 1, nominal p-value ≤ 0.05) in the seven examined tissues. The expressed lncRNAs (FPKM > 0.1) exhibited tissue-specific patterns in WT pigs. Specifically, 3410 lncRNAs were expressed exclusively in only one tissue. Potential functions of lncRNAs were indirectly predicted by searching their target cis- and trans-regulated protein-coding genes. LncRNAs with tissue-specific expression influence numerous genes related to tissue functions. Weighted gene coexpression network analysis (WGCNA) of 1570 DE-lncRNAs between WT and ZBED6 KO pigs was used to define the following six lncRNA modules specific to different tissues: skeletal muscle, heart, lung, spleen, kidney and liver modules. Furthermore, by conjoint analysis of longissimus dorsi data (tissue-specific expression, muscle module and DE-lncRNAs) and ChIP-PCR revealed NONSUSG002145.1 (adjusted p-values = 0.044), which is coexpressed with the IGF2 gene and binding with ZBED6, may play important roles in ZBED6 KO pig skeletal muscle development. CONCLUSIONS: These findings indicate that the identified lncRNAs may play essential roles in tissue function and regulate the mechanism of ZBED6 action in skeletal muscle development in pigs. To our knowledge, this is the first study describing lncRNAs in ZBED6 KO pigs. These results may open new research directions leading to a better understanding of the global functions of ZBED6 and of lncRNA functions in skeletal muscle development in pigs.

EPAS1 Gain-of-Function Mutation Contributes to High-Altitude Adaptation in Tibetan Horses.

High altitude represents some of the most extreme environments worldwide. The genetic changes underlying adaptation to such environments have been recently identified in multiple animals but remain unknown in horses. Here, we sequence the complete genome of 138 domestic horses encompassing a whole altitudinal range across China to uncover the genetic basis for adaptation to high-altitude hypoxia. Our genome data set includes 65 lowland animals across ten Chinese native breeds, 61 horses living at least 3,300 m above sea level across seven locations along Qinghai-Tibetan Plateau, as well as 7 Thoroughbred and 5 Przewalski's horses added for comparison. We find that Tibetan horses do not descend from Przewalski's horses but were most likely introduced from a distinct horse lineage, following the emergence of pastoral nomadism in Northwestern China ∼3,700 years ago. We identify that the endothelial PAS domain protein 1 gene (EPAS1, also HIF2A) shows the strongest signature for positive selection in the Tibetan horse genome. Two missense mutations at this locus appear strongly associated with blood physiological parameters facilitating blood circulation as well as oxygen transportation and consumption in hypoxic conditions. Functional validation through protein mutagenesis shows that these mutations increase EPAS1 stability and its hetero dimerization affinity to ARNT (HIF1B). Our study demonstrates that missense mutations in the EPAS1 gene provided key evolutionary molecular adaptation to Tibetan horses living in high-altitude hypoxic environments. It reveals possible targets for genomic selection programs aimed at increasing hypoxia tolerance in livestock and provides a textbook example of evolutionary convergence across independent mammal lineages.

Expression Profiling and Functional Characterization of miR-26a and miR-130a in Regulating Zhongwei Goat Hair Development via the TGF-ß/SMAD Pathway.

The Zhongwei goat is an important and unique goat breed indigenous to China. It has a natural hair curling phenotype at birth, but the degree of curling gradually decreases with growth. The molecular mechanism underlying the dynamic changes in the wool curvature in Zhongwei goats is poorly understood. MicroRNAs (miRNAs) play important roles in many biological processes, including hair growth and development. In this study, we selected skins from Zhongwei goats at different ages (45 and 108 days) that exhibited different levels of hair curvature and performed miRNA sequencing to explore the molecular mechanism of hair bending. In total, 28 significantly differentially expressed miRNAs (DE miRNAs) were identified in the three groups of samples between the two developmental stages. An analysis of the target genes of the above-mentioned DE miRNAs by the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses indicated that the DE miRNAs were involved in signal pathways which were previously associated with hair bending and hair follicle development, such as the TGF-ß/SMAD, PI3K-Akt, JAK-STAT, and MAPK pathways. A comprehensive analysis of the correlations between the miRNA-seq results and issued transcriptional findings indicated that SMAD1 was a target gene of miR-26a and SMAD5 was a target gene of miR-130a. Furthermore, goat dermal papilla cells were successfully isolated and purified to determine the role of miRNAs in follicle development in vitro. The study results demonstrated that miR-130a and miR-26a had significant effects on the proliferation of dermal papilla cells. In addition, the detection results of mRNA and protein levels indicate that the overexpression of miR-26a can promote the expression of related genes in the TGF-ß/SMAD pathway, while miR-130a has the opposite substitution effect. The dual luciferase report test showed that miR-26a targeted the SMAD1 gene and reduced the expression of the SMAD1 protein in hair papillary cells. Our results identified DE microRNAs which perhaps change at the time of hair straightening in Zhongwei goats and explore the role of miR-26a and miR-130a in dermal papilla cells proliferation. The present study provided a theoretical basis to explore the mechanisms underlying the Zhongwei hair growth and curly phenotype.

Genome-wide discovery of lincRNAs with spatiotemporal expression patterns in the skin of goat during the cashmere growth cycle.

BACKGROUND: Long intergenic noncoding RNAs (lincRNAs) have been recognized in recent years as key regulators of biological processes. However, lincRNAs in goat remain poorly characterized both across various tissues and during different developmental stages in goat (Capra hircus). RESULTS: We performed the genome-wide discovery of the lincRNAs in goat by combining the RNA-seq dataset that were generated from 28 cashmere goat skin samples and the 12 datasets of goat tissues downloaded from the NCBI database. We identified a total of 5546 potential lincRNA transcripts that overlapped 3641 lincRNA genes. These lincRNAs exhibited a tissue-specific pattern. Specifically, there are 584 lincRNAs expressed exclusively in only one tissue, and 91 were highly expressed in hair follicle (HF). In addition, 2350 protein-coding genes and 492 lincRNAs were differentially expressed in the skin of goat. The majority exhibited the remarkable differential expression during the transition of the goat skin from the May-June to August-October time point, which covered the different seasons. Fundamental biological processes, such as skin development, were significantly enriched in these genes. Furthermore, we identified several lincRNAs highly expressed in the HF, which exhibited not only the co-expression pattern with the key factors to the HF development but also the activated expression in the August to October time point. Intriguingly, one of spatiotemporal lincRNAs, linc-chig1598 could be a potential regulator of distal-less homeobox 3 expression during the secondary hair follicle growth. CONCLUSIONS: This study will facilitate future studies aimed at unravelling the function of lincRNAs in hair follicle development.

Exome sequencing reveals genetic differentiation due to high-altitude adaptation in the Tibetan cashmere goat (Capra hircus).

BACKGROUND: The Tibetan cashmere goat (Capra hircus), one of the most ancient breeds in China, has historically been a critical source of meat and cashmere production for local farmers. To adapt to the high-altitude area, extremely harsh climate, and hypoxic environment that the Tibetan cashmere goat lives in, this goat has developed distinct phenotypic traits compared to lowland breeds. However, the genetic components underlying this phenotypic adaptation remain largely unknown. RESULTS: We obtained 118,700 autosomal SNPs through exome sequencing of 330 cashmere goats located at a wide geographic range, including the Tibetan Plateau and low-altitude regions in China. The great majority of SNPs showed low genetic differentiation among populations; however, approximately 2-3% of the loci showed more genetic differentiation than expected under a selectively neutral model. Together with a combined analysis of high- and low-altitude breeds, we revealed 339 genes potentially under high-altitude selection. Genes associated with cardiovascular system development were significantly enriched in our study. Among these genes, the most evident one was endothelial PAS domain protein 1 (EPAS1), which has been previously reported to be involved in complex oxygen sensing and significantly associated with high-altitude adaptation of human, dog, and grey wolf. The missense mutation Q579L that we identified in EPAS1, which occurs next to the Hypoxia-Inducible Factor-1 (HIF-1) domain, was exclusively enriched in the high-altitude populations. CONCLUSIONS: Our study provides insights concerning the population variation in six different cashmere goat populations in China. The variants in cardiovascular system-related genes may explain the observed phenotypic adaptation of the Tibetan cashmere goat.

Mapping the four-horned locus and testing the polled locus in three Chinese sheep breeds.

Four-horned sheep are an ideal animal model for illuminating the genetic basis of horn development. The objective of this study was to locate the genetic region responsible for the four-horned phenotype and to verify a previously reported polled locus in three Chinese breeds. A genome-wide association study (GWAS) was performed using 34 two-horned and 32 four-horned sheep from three Chinese indigenous breeds: Altay, Mongolian and Sishui Fur sheep. The top two significant single nucleotide polymorphisms (SNPs) associated with the four-horned phenotype were both located in a region spanning positions 132.6 to 132.7 Mb on sheep chromosome 2. Similar locations for the four-horned trait were previously identified in Jacob, Navajo-Churro, Damara and Sishui Fur sheep, suggesting a common genetic component underlying the four-horned phenotype. The two identified SNPs were both downstream of the metaxin 2 (MTX2) gene and the HOXD gene cluster. For the top SNP-OAR2:g.132619300G>A-the strong associations of the AA and AG genotypes with the four-horned phenotype and the GG genotype with the two-horned phenotype indicated the dominant inheritance of the four-horned trait. No significant SNPs for the polled phenotype were identified in the GWAS analysis, and a PCR analysis for the detection of the 1.8-kb insertion associated with polled sheep in other breeds failed to verify the association with polledness in the three Chinese breeds. This study supports the hypothesis that two different loci are responsible for horn existence and number. This study contributes to the understanding of the molecular regulation of horn development and enriches the knowledge of qualitative traits in domestic animals.

MicroRNA signatures in total peripheral blood of gallbladder cancer patients.

MicroRNAs (miRNAs) are small non-coding RNA molecules that function as negative regulators of gene expression. Expression analysis of miRNAs can reflect the developmental lineage and differentiation state of the tumor. Thus, the miRNA profiles could play essential roles in gallbladder cancer (GBC). However, their role as novel biomarkers for the diagnosis of GBC is unclear yet. We assessed here for the first time whole-genome miRNA expression in peripheral blood samples of 40 GBC patients using microRNA microarray. We identified 11 miRNAs, which are significantly dysregulated in GBC patients in comparison to 40 healthy controls. Among these, the expression of let-7a, miR-21, miR-187, miR-143, miR-202, and miR-335 are in agreement with those measured by real-time PCR (p < 0.05). Furthermore, dysregulated miR-187, miR-143, and miR-202 demonstrate a significant correlation with clinicopathologic features (p < 0.05). Our results suggest a possible pathological relationship between the differential expression of miRNA in peripheral blood and GBC, and these dysregulated miRNAs could be novel tumor biomarkers for early detection of GBC.

[Molecular regulation of skeletal satellite cell's self-renewal].

Skeletal muscle possesses a remarkable ability for its regeneration and injured tissue repair. This ability depends on the activity and contributions of muscle satellite cells. Proliferating satellite cells, termed myogenic precursor cells or myoblasts, are activated and driven out of their quiescent state upon muscle injury. In this summary, we present a review to summarize the molecular regulation in skeletal satellite cells to light on the satellite cells' self-renewal mechanism.

Whole-Genome Sequencing Identifies Functional Genes for Environmental Adaptation in Chinese Sheep.

Sheep (Ovis aries), among the first domesticated species, are now globally widespread and exhibit remarkable adaptability to diverse environments. In this study, we perform whole-genome sequencing of 266 animals from 18 distinct Chinese sheep populations, each displaying unique phenotypes indicative of adaptation to varying environmental conditions. Integrating 131 environmental factors with single nucleotide polymorphism variations, we conduct a comprehensive genetic-environmental association analysis. This analysis identifies 35 key genes likely integral to the environmental adaptation of sheep. The functions of these genes include fat tail formation (HOXA10, HOXA11, JAZF1), wool characteristics (FER, FGF5, MITF, PDE4B), horn phenotypes (RXFP2), reproduction (HIBADH, TRIM71, C6H4orf22) and growth traits (ADGRL3, TRHDE). Notably, we observe a significant correlation between the frequency of missense mutations in the PAPSS2 and RXFP2 genes and variations in altitude. Our study reveals candidate genes for adaptive variation in sheep and demonstrates the diversity in the ways sheep adapt to their environment.

Two mutations at KRT74 and EDAR synergistically drive the fine-wool production in Chinese sheep.

INTRODUCTION: Fine-wool sheep are the most common breed used by the wool industry worldwide. Fine-wool sheep have over a three-fold higher follicle density and a 50% smaller fiber diameter than coarse-wool sheep. OBJECTIVES: This study aims to clarify the underlying genetic basis for the denser and finer wool phenotype in fine-wool breeds. METHOD: Whole-genome sequences of 140 samples, Ovine HD630K SNP array data of 385 samples, including fine, semi-fine, and coarse wool sheep, as well as skin transcriptomes of nine samples were integrated for genomic selection signature analysis. RESULTS: Two loci at keratin 74 (KRT74) and ectodysplasin receptor (EDAR) were revealed. Fine-scale analysis in 250 fine/semi-fine and 198 coarse wool sheep narrowed this association to one C/A missense variant of KRT74 (OAR3:133,486,008, P = 1.02E-67) and one T/C SNP in the regulatory region upstream of EDAR (OAR3:61,927,840, P = 2.50E-43). Cellular over-expression and ovine skin section staining assays confirmed that C-KRT74 activated the KRT74 protein and specifically enlarged cell size at the Huxley's layer of the inner root sheath (P < 0.01). This structure enhancement shapes the growing hair shaft into the finer wool than the wild type. Luciferase assays validated that the C-to-T mutation upregulated EDAR mRNA expression via a newly created SOX2 binding site and potentially led to the formation of more hair placodes. CONCLUSIONS: Two functional mutations driving finer and denser wool production were characterized and offered new targets for genetic breeding during wool sheep selection. This study not only provides a theoretical basis for future selection of fine wool sheep breeds but also contributes to improving the value of wool commodities.

[Isolation and biological characterization of mesenchymal stem cells from Beijing fatty chicken fetal liver].

Mesenchymal stem cells (MSCS) from chicken fetal liver are multipotent stem cells that can differentiate in vitro into various terminally differentiated cells. The majority of studies have focused on rats and mice now. Reports from other animals are less and analyses on domestic animals are few. In this study, chicken liver-derived MSCs were isolated from 7-day-old embryo of Beijing fatty chickens. Primary liver-derived MSCs were subcultured to passage 15. The surface markers of liver-derived MSCs, CD29, and CD44 were detected by immunofluorescence and the surface markers CD34 and CK19 of hematopoietic progenitor cells/hepatic oval cells were not detected. RT-PCR analysis detected positive expression of CD29, CD44, CD71, and CD73. The growth curves were typically sigmoidal. Liver-derived MSCs of different passages were successfully induced and differentiated into neuronal and osteoblast cells. The results suggest that the MSCs isolated from chicken fetal liver possess similar biological characteristics with those derived from mice, and their multilineage differentiation provides many potential applications.

Genome-Wide Signal Selection Analysis Revealing Genes Potentially Related to Sheep-Milk-Production Traits.

Natural selection and domestication have shaped modern sheep populations into a vast range of phenotypically diverse breeds. Among these breeds, dairy sheep have a smaller population than meat sheep and wool sheep, and less research is performed on them, but the lactation mechanism in dairy sheep is critically important for improving animal-production methods. In this study, whole-genome sequences were generated from 10 sheep breeds, including 57 high-milk-yield sheep and 44 low-milk-yield sheep, to investigate the genetic signatures of milk production in dairy sheep, and 59,864,820 valid SNPs (Single Nucleotide Polymorphisms) were kept after quality control to perform population-genetic-structure analyses, gene-detection analyses, and gene-function-validation analyses. For the population-genetic-structure analyses, we carried out PCA (Principal Component Analysis), as well as neighbor-joining tree and structure analyses to classify different sheep populations. The sheep used in our study were well distributed in ten groups, with the high-milk-yield-group populations close to each other and the low-milk-yield-group populations showing similar classifications. To perform an exact signal-selection analysis, we used three different methods to find SNPs to perform gene-annotation analyses within the 995 common regions derived from the fixation index (FST), nucleotide diversity (Ɵπ), and heterozygosity rate (ZHp) results. In total, we found 553 genes that were located in these regions. These genes mainly participate in the protein-binding pathway and the nucleoplasm-interaction pathway, as revealed by the GO- and KEGG-function-enrichment analyses. After the gene selection and function analyses, we found that FCGR3A, CTSK, CTSS, ARNT, GHR, SLC29A4, ROR1, and TNRC18 were potentially related to sheep-milk-production traits. We chose the strongly selected genes, FCGR3A, CTSK, CTSS, and ARNT during the signal-selection analysis to perform a RT-qPCR (Reale time Quantitative Polymerase Chain Reaction) experiment to validate their expression-level relationship with milk production, and the results showed that FCGR3A has a significant negative relationship with sheep-milk production, while other three genes did not show any positive or negative relations. In this study, it was discovered and proven that the candidate gene FCGR3A potentially contributes to the milk production of dairy sheep and a basis was laid for the further study of the genetic mechanism underlying the strong milk-production traits of sheep.

Correction: Luan et al. Identification of Critical Genes for Ovine Horn Development Based on Transcriptome during the Embryonic Period. Biology 2023, 12, 591.

In the original publication [1], there were mistakes in the order of the references, which were as follows: [...].

Identification of Critical Genes for Ovine Horn Development Based on Transcriptome during the Embryonic Period.

Horns, also known as headgear, are a unique structure of ruminants. As ruminants are globally distributed, the study of horn formation is critical not only for increasing our understanding of natural and sexual selection but also for the breeding of polled sheep breeds to facilitate modern sheep farming. Despite this, a significant number of the underlying genetic pathways in sheep horn remain unclear. In this study, to clarify the gene expression profile of horn buds and investigate the key genes in horn bud formation, RNA-sequencing (RNA-seq) technology was utilized to investigate differential gene expression in the horn buds and adjacent forehead skin of Altay sheep fetuses. There were only 68 differentially expressed genes (DEGs) identified, consisting of 58 up-regulated genes and 10 down-regulated genes. RXFP2 was differentially up-regulated in the horn buds and had the highest significance (p-value = 7.42 × 10-14). In addition, 32 DEGs were horn-related genes identified in previous studies, such as RXFP2, FOXL2, SFRP4, SFRP2, KRT1, KRT10, WNT7B, and WNT3. Further, Gene Ontology (GO) analysis showed that the DEGs were mainly enriched with regard to growth, development, and cell differentiation. Pathway analysis revealed that the Wnt signaling pathway may be responsible for horn development. Further, through combining the protein-protein interaction networks of the DEGs, it was found that the top five hub genes, namely, ACAN, SFRP2, SFRP4, WNT3, and WNT7B, were also associated with horn development. Our results suggest that only a few key genes, including RXFP2, are involved in bud formation. This study not only validates the expression of candidate genes identified at the transcriptome level in previous studies but also provides new possible marker genes for horn development, which may promote our understanding of the genetic mechanisms of horn formation.

5-methoxytryptophan induced apoptosis and PI3K/Akt/FoxO3a phosphorylation in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is a highly prevalent malignancy worldwide, and new therapeutic targets urgently need to be found to prolong patient survival. 5-methoxytryptophan (5-MTP) is a tryptophan metabolite found in animals and humans. However, the effects of 5-MTP on proliferation and apoptosis of CRC cells are currently unknown. AIM: To investigate the effects of 5-MTP on the proliferation, migration, invasion, and apoptosis abilities of CRC cells. Additionally, we seek to explore whether 5-MTP has the potential to be utilized as a drug for the treatment of CRC. METHODS: In order to evaluate the effect of 5-MTP on CRC cells, a series of experiments were conducted for evaluation. Colony formation assay and Cell Counting Kit 8 assays were used to investigate the impact of 5-MTP on the proliferation of CRC cell lines. Cell cycle assays were employed to examine the effect of 5-MTP on cellular growth. In addition, we investigated the effects of 5-MTP on apoptosis and reactive oxygen species in HCT-116 cells. To obtain a deeper understanding of how 5-MTP affects CRC, we conducted a study to examine its influence on the PI3K/Akt signaling pathway in CRC cells. RESULTS: This article showed that 5-MTP promoted apoptosis and cell cycle arrest and inhibited cell proliferation in CRC cells. In many articles, it has been reported that PI3K/Akt/FoxO3a signaling pathway is one of the most important signaling pathways involved in internal regulating cell proliferation and differentiation. Nevertheless, 5-MTP combined with PI3K/Akt/FoxO3a signaling pathway inhibitors significantly promoted apoptosis and cell cycle arrest and inhibited cell proliferation in CRC cells compared with 5-MTP alone in our study. CONCLUSION: Therefore, there is strong evidence that 5-MTP can be used as an effective medicine for CRC treatment.