Comprehensive analysis of coding and non-coding RNA transcriptomes related to hypoxic adaptation in Tibetan chickens

Background:Tibetan chickens,a unique native breed in the Qinghai-Tibet Plateau of China,possess a suite of adaptive features that enable them to tolerate the high-altitude hypoxic environment.Increasing evidence suggests that long non-coding RNAs(lncRNAs)and microRNAs(miRNAs)play roles in the hypoxic adaptation of high-altitude animals,although their exact involvement remains unclear.Results:This study aimed to elucidate the global landscape of mRNAs,lncRNAs,and miRNAs using transcriptome sequencing to construct a regulatory network of competing endogenous RNAs(ceRNAs)and thus provide insights into the hypoxic adaptation of Tibetan chicken embryos.In total,354 differentially expressed genes(DE genes),389 differentially expressed lncRNAs(DE lncRNAs),and 73 differentially expressed miRNAs(DE miRNAs)were identified between Tibetan chickens(TC)and control Chahua chickens(CH).GO and KEGG enrichment analysis revealed that several important DE miRNAs and their target DE lncRNAs and DE genes are involved in angiogenesis(including blood vessel development and blood circulation)and energy metabolism(including glucose,carbohydrate,and lipid metabolism).The ceRNA network was then constructed with the predicted DE gene-DE miRNA-DE lncRNA interactions,which further revealed the regulatory roles of these differentially expressed RNAs during hypoxic adaptation of Tibetan chickens.Conclusions:Analysis of transcriptomic data revealed several key candidate ceRNAs that may play high-priority roles in the hypoxic adaptation of Tibetan chickens by regulating angiogenesis and energy metabolism.These results provide insights into the molecular mechanisms of hypoxic adaptation regulatory networks from the perspective of coding and non-coding RNAs.

Gene expression of vascular endothelial growth factor A and hypoxic adaptation in Tibetan pig

Background： Vascular endothelial growth factor A （VEGFA） can induce endothelial cell proliferation, promote cell migration, and inhibit apoptosis. These processes play key roles in physiological blood vessel formation and pathological angiogenesis. Methods： In this study, we examined VEGFA gene expression in the heart, liver, and kidney of Tibetan pigs （-I-P）, Yorkshire pigs that migrated to high altitudes （YH）, and Yorkshire pigs that lived at low altitudes （YL）. We used PCR and Sanger sequencing to screen for single nucleotide polymorphisms （SNPs） in 5＇-flanking DNA and exons of the VEGFA gene. Quantitative real-time PCR and western blots were used to measure expression levels and PCR products were sequenced. Results： Results showed that the VEGFA mRNA and protein expression in heart, liver and kidney of TP was higher than that in YH and YL. In addition, the mRNA sequence of the pig VEGFA gene was conserved among pig breeds, and only five SNPs were found in the 5＇-flanking region of the VEGFA gene, the allele frequency distributions of the 5 SNPs were not significantly different between the TP, Yorkshire （YL）, and Diannan small-ear （DN） pig populations. Conclusion： In conclusion, the Tibetan pig showed high tissues, which suggests that the VEGFA gene may play a levels of VEGFA gene expression in several hypoxic major functional role in hypoxic adaptation.

Differential gene expression of phosphoglyceric kinase (PGK) and hypoxic adaptation in chicken

Four single-nucleotide polymorphisms (SNP) of the Phosphoglyceric Kinase (PGK) gene were discov- ered based on comparison of the sequences from an altiplano chicken breed (Tibetan chicken) and two lowland breeds (White Leghorn and Shouguang chicken). Gel-shift results indicate that one of these SNPs, an A→G mutation at position 59 in exon10, is able to bind hypoxia-induced factor-l (HIF-1), functioning as a hypoxia response element (HRE). The mutant gene results in M→T mutation at position 379 amino acid. The combined activity of this HRE and HIF-1 could increase correspondingly under a hypoxic stimulus. Hypoxia leads to increased death rates of chicken embryos; while the M→T mutation described herein is prevalent in healthy embryos grown under hypoxic conditions, thus it may repre- sent an adaptation to hypoxia. Fluorescence quantitative reverse transcription PCR results revealed that HIF-1 upregulates the transcript level of the glycolytic enzyme PGK in the brain and skeletal mus- cle of animals subjected to hypoxia. Thus, a large amount of ATP is produced by increased glycolysis, allowing the organism to meet energy metabolism demands. As such, we believe this SNP to be an adaptation to the external anoxic environment.

Small RNA Sequencing Reveals Differentially Expressed piRNAs Related to Hypoxia in Tibetan Chickens

The relevance of genetic mechanism to the phenotype of hypoxic adaptation remains elusive.Tibetan chickens typically used to investigate the mechanism for the adaptation of hypoxia and the recognition of hypoxia-related piRNA remains an open issue.The purpose of this study was to illustrate whether the piRNAs were related to hypoxic adaptation.First of all,the differentially expressed piRNAs(DEpiRNAs)were identified through RNA sequencing between the Tibetan chickens and Daheng broilers.Subsequently,the target genes of DEpiRNAs were predicted and annotated by software.The network was constructed by Cytoscape.In our study,a total of 277 DEpiRNAs(33 down-regulated,244 up-regulated)were identified in the Tibetan chickens compared with the Daheng broilers.All of the 277 DEpiRNAs predicted 36658 targeted genes.Gene Ontology(GO)analysis showed that the target genes were significantly enriched in the biological process correlated with proliferation and apoptosis of cells,including cell cycle,mitochondrial outer membrane permeabilization,and positive regulation of stress-activated mitogen-activated protein kinase cascade.Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis implicated that the DEpiRNAs were mainly involved in immune and metabolism,including natural killer cell-mediated cytotoxicity,toll-like receptor signaling pathway and fatty acid metabolism.Furthermore,a predicted network with four piRNAs acted on 11 pathways via interacting with 22 target genes,in which piR-gga-1368839 regulated metabolic pathways by acting on DHCR24.In conclusion,we determined the DEpiRNAs in the Tibetan chickens and found that these piRNAs were associated with metabolism,which may be favorable for researching the biological adaptation to hypoxic stress.

Hypoxic preconditioning in an autohypoxic animal model

Hypoxic preconditioning refers to the exposure of organisms, systems, organs, tissues or cells to moderate hypoxia/ischemia that results in increased resistance to a subsequent episode of severe hypoxia/ischemia. In this article, we review recent research based on a mouse model of repeated exposure to autohypoxia. Pre-exposure markedly increases the tolerance to or protection against hypoxic insult, and preserves the cellular structure of the brain. Furthermore, the hippocampal activity amplitude and frequency of electroencephalogram, latency of cortical somatosensory-evoked potential and spinal somatosensory-evoked potential progressively decrease, while spatial learning and memory improve. In the brain, detrimental neurochemicals such as free radicals are down-regulated, while beneficial ones such as adenosine are up-regulated. Also, antihypoxia factor（s） and gene（s） are activated. We propose that the tolerance and protective effects depend on energy conservation and plasticity triggered by exposure to hypoxia via oxygen-sensing transduction pathways and hypoxia-inducible factor-initiated cascades. A potential path for further research is the development of devices and pharma-ceuticals acting on antihypoxia factor（s） and gene（s） for the prevention and treatment of hypoxia and related syndromes.