| Chinese pig breeds have a long history of domestication,and native pig breeds have rich phenotypes.As a high-quality germplasm resource in China,Tibetan pig has strong biological adaptability and stress resistance,and is different from other pig breeds.The liver is the metabolic core of the whole body.During pregnancy,a series of physiological changes in the body are inseparable from the participation of the liver.Studies have shown that the liver will undergo adaptive changes during pregnancy to maintain the homeostasis of the body during pregnancy.Changes in the liver during pregnancy include,but are not limited to,hepatocyte proliferation and hypertrophy,lipid and glucose metabolism,etc.In order to explore the characteristics of liver morphology and genetic changes of Tibetan pigs during pregnancy,the liver of female young Tibetan pigs(control group)and pregnant female Tibetan pig(experimental group)were used to observe the histomorphological changes of liver by HE staining.Using RNA-seq technology,eight sequencing libraries of Tibetan pig liver were constructed in two periods.The key m RNA and lnc RNA related to liver adaptability were screened by bioinformatics and verified by m RNA-lnc RNA and RT-q PCR.In addition,the public database was used to analyze the transcriptome information of the livers of other pig breeds and compared with the livers of Tibetan pigs to determine the differences between them.The main results are as follows:(1)The results of HE sections showed that there were some “fat holes” in the liver sections of both groups,which were slightly more than those in the young group,and the staining of hepatocytes in the pregnant group was slightly more than that in the young group.compared with the young group,the number of hepatic plate cells between central vein and portal vein in pregnant Tibetan pigs was significantly more than that in young Tibetan pigs,indicating that adaptive changes may have taken place in the tissue structure of the liver under pregnancy stimulation.(2)The sequencing library of Tibetan pig liver was constructed by RNA sequencing technology,and a total of 94.81 G effective data were obtained,and the data were compared and spliced by software such as STAR and String Tie,and a total of 358 differentially expressed m RNA and 169 differential lnc RNA were obtained after differential analysis.5 differentially expressed m RNA and 5 lnc RNA were randomly selected,and the sequencing results were verified by real-time fluorescence quantitative PCR.(3)GO analysis of differentially expressed m RNA showed that the differential genes were mainly enriched in the biological processes related to lipid storage and metabolism,cell cycle and immunity,while KEGG analysis showed that more genes were enriched in PI3K-Akt,PPAR,AMPK,cholesterol metabolism,Toll-like receptor signals and other pathways,which were mostly related to cell growth and metabolism.The differential genes FDFT1,SQLE,MSMO1 and LSS maybe related to lipid accumulation in the liver during pregnancy,CRP,CCR7,TLR2 C and D86 may be related to the establishment of immune tolerance in the liver during pregnancy.PPI interaction analysis showed that the core network with the highest differential protein score was related to cell mitosis,and the differential genes CHEK1,RFC4,RAD51,MCM3,CHAF1 B and PRIM1 were involved in the process of DNA replication and cell cycle.(4)All genes within the upstream and downstream 100 kb of differentially expressed lnc RNA were predicted as target genes of lnc RNA by cis.The results showed that there was a cis-regulatory relationship between 128 differential lnc RNA and 284 m RNA.GO enrichment analysis of these target genes showed that these target genes were significantly enriched in122 biological processes,including DNA metabolism,autophagy,lipid biosynthesis and organ regeneration,in addition,co-expression analysis predicted trans target genes of lnc RNA,96 lnc RNA and 179 m RNA formed 1179 pairs of lnc RNA-m RNA co-expression pairs,of which1148 pairs were positively correlated(PCC ≥ 0.95),31 pairs were negatively correlated(PCC≤-0.95).Using Cytoscape software to construct lnc RNA-m RNA visual interaction network,it is found that ENSSSCT00000093472 and ENSSSCT00000087990 can not only affect multiple target genes through trans,but also interact with target genes through cis,and target genes such as GPSM3 and PLPP2 also interact with multiple lnc RNA.(5)After comparing the transcriptomes of Tibetan pigs with "Ningxiang pig","Songliao black pig","Yanan pig" and "Wuzhishan pig",it was found that there were differences in metabolism and immunity between Tibetan pig and "Ningxiang pig".Among them,genes related to glucose metabolism in TCA cycle and lipid metabolism in PPAR pathway are upregulated,and many genes in T17 cell differentiation pathway are also activated.To sum up,physiological changes have taken place in the liver of pregnant Tibetan pigs.transcriptome analysis showed that differentially expressed m RNA in liver tissue was enriched in the biological processes related to lipid storage and metabolism,cell cycle and immunity;through the prediction of cis and trans target genes and the construction of lnc RNA-m RNA visual interaction network,it was found that some differentially expressed lnc RNA may be involved in the adaptive regulation of liver during pregnancy.In the comparison between Tibetan pig liver and other pig transcriptional groups,it was found that there were great differences in metabolism and immunity.This study provides basic data for the regulation mechanism and expression pattern of key genes in the liver of pregnant Tibetan pigs,and data mining helps to understand the changes of liver transcriptome among different pig breeds and provides a theoretical basis for further research in related fields.and has a certain social significance in the protection of native germplasm resources. |
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