Xiang pig is a native inbred line originating from a warm and humid mountainous region and is prone to infections by pathogens causing lung lesions. However, the underlying genetic factors still remain unclear. Our previous genomic resequencing comparisons found a deletion type of structural variation (SV) within the intron of gene cerebellar degeneration related protein 2 (CDR2) with a higher frequency in Xiang pigs compared with European pig breeds. This study is aimed to explore the relationship between the SV and the lung injury in Xiang pig populations. It demonstrated that the deletion genotype (MM) was dominant in Xiang pig herds and with a higher percentage of allele M than that in Xiang pig with pulmonary lesions and Large White breeds. By Sanger sequencing and RepeatMasker analysis, a SINE in 277 bp (namely SINE-277) was detected and located in the opposite orientation of CDR2 gene. The inhibit effects of SINE-277 on transcription was confirmed by EGFP reporter gene after transfected into HEK-293T cells. And the expression levels of CDR2 was increased in the lungs with MM types (P < 0.05) via both RT-qPCR and Western blotting assays. Moreover, significant differences were estimated between the SV genotypes and the lung lesion severity scores, the antibody concentrations against pathogens, and expressions of nine inflammation factor genes including NFκB. It reinforced the effects of SINE on gene CDR2 expression and might be taken as a DNA marker for the resistance/susceptibility to lung diseases in pigs.

Structural variants (SVs) are a type of mutations in animal genomes that have shown their role in the process of domestication. An insertion/ deletion of 579 bp SV in intron 21 of PIK3C3 gene in Xiang pigs was detected by genome resequencing method in our previous study. Phosphoinositide-3-kinase, class III (PIK3C3), is a critical regulator of autophagy, essential for cellular homeostasis, pathogen clearance, and immune modulation. The aim of this study was to investigate the polymorphism of intron 21 of the PIK3C3 gene of Xiang pigs and its effect on PIK3C3 expression. The polymorphism of SV PIK3C3-579 was determined by PCR. Three genotypes of this SV were detected in different breeds with uneven distribution among them. Two Short interspersed elements (SINEs) located in the SV region, opposite in the direction of PIK3C3 gene. RT-qPCR revealed that the SINE insertion significantly decreased PIK3C3 expression in the spleen of Xiang pigs and was associated with the production of the PIK3C3-circRNA. The fluorescent reporter assays confirmed that the SINE insertion significantly decreased the activity of the EGFP promoter. Additionally, The SINE deletion was associated with elevated levels of lymphocyte, monocyte, neutrophil, IFN-γ and IL-2. These data suggested that the SINE insertion could regulate the expression of PIK3C3 gene and the biogenesis of circular RNA, thereby modulating the immune response of Xiang pigs, which provides insights into the functional impact of structural variants on immune traits in this breed.

Echovirus 3 (E-3) has recently been identified as an emerging pathogen associated with both severe and mild cases of hand, foot, and mouth disease (HFMD) in Guizhou Province, China. Although these strains exhibit distinct clinical phenotypes, their underlying molecular mechanisms remain unclear. This study reports the phylogenetic and genetic features of eight E-3 strains from severe and mild HFMD cases and investigates E-3 seroprevalence in Guizhou. Eight shared nucleotide substitutions in the untranslated regions (UTRs) and six shared amino acid substitutions in the open reading frame (ORF) were identified between E-3 strains from severe and mild case groups. Nine variant-specific amino acid mutations were under positive selection and exhibited high variability. Among the 115 nucleotide substitutions in the ORF regions, 31 were specific to the severe case group and 41 to the mild case group. Recombination analysis revealed that the Guizhou E-3 strains underwent complex interserotypic recombination with echovirus 21 (E-21), enterovirus B80 (EV-B80), and echovirus 30 (E-30). E-3 strains from the severe case group demonstrated enhanced replication efficiency, higher viral titers, and greater cytopathogenicity than those from the mild case group. Analysis of 178 samples revealed a seropositivity rate of 52.25% and a geometric mean titer (GMT) of 1:28.23 for E-3, indicating the prevalence of asymptomatic infections in Guizhou Province. All Guizhou E-3 strains exhibited non-temperature-sensitive characteristics, suggesting their potential for transmissibility within the population. This study provides valuable insights for global studies on E-3-related diseases and molecular epidemiology, establishing a foundation for further investigation into the pathogenicity of E-3 variants.IMPORTANCEThis study is the first report of echovirus 3 (E-3) infections associated with severe hand, foot, and mouth disease (HFMD) cases in Guizhou Province, China. All E-3 strains showed thermotolerance and higher seroprevalence, suggesting potentially enhanced transmissibility. Several shared nucleotide substitutions and amino acid substitutions were identified in Guizhou strains from both severe and mild case groups, indicating that E-3 may have evolved through continuous genetic adaptation, potentially driving the emergence of more transmissible epidemic variants. This study revealed that the E-3 strains in the severe case group exhibited enhanced replication efficiency, higher viral titers, and increased cytopathogenicity in vitro, thereby providing evidence to support a potential association between genetic variations and clinical severity. Notably, all E-3-infected patients in the severe case group exhibited more severe clinical manifestations. These findings provide critical updates on the epidemiological trends of enteroviruses associated with HFMD and establish a foundation for a better understanding of genetic diversity, evolutionary patterns, and molecular mechanisms of enterovirus infections.

This study aimed to investigate the association of the AKR1C2-I4-sv245 polymorphism in the aldo-keto reductase family 1 member C2 (AKR1C2) gene with litter size in Xiang pigs. Polymorphism analysis demonstrated that the "ID" genotype was dominant in Xiang pigs, while "D" was the dominant allele. The DD genotype was most common in high-fertility Xiang pigs (XL), while the ID genotype was predominant in low-fertility Xiang pigs (XS). The association analysis results indicated that the litter size of the XL group in Xiang pigs was significantly higher than that of the XS group across all parity levels (P < 0.05). A significant association was observed between the AKR1C2-I4-sv245 polymorphism and the litter size in Xiang sows (P < 0.01), with the DD genotype exhibiting a higher litter size than the ID and II genotypes. Bioinformatics analysis revealed that the deleted segment contained a short interspersed element (SINE), three exon splicing enhancers, four intron splicing enhancers, and other regulatory elements associated with gene expression. Transfection of HEK-293T cells with the EGFP reporter gene confirmed the inhibitory effect of the SINE on transcription, demonstrating that its insertion significantly repressed reporter gene expression. Relative expression analysis showed that AKR1C2 mRNA and protein levels were higher in the DD genotype compared to the II genotype and in the ID genotype compared to the II genotype. Collectively, these results substantiated the regulatory role of SINE in gene expression and highlighted the potential of AKR1C2-I4-sv245 as a promising DNA marker for improving litter size in Xiang pigs.

Backfat thickness (BFT) and intramuscular fat (IMF) content reflect fat deposition and meat quality in pigs. Numerous genetic variations contribute to differences in BFT and IMF for different pig breeds. However, limited information is available on the genetic variants associated with IMF deposition in Xiang pigs. To identify the putative regulatory regions and candidate genes associated with BFT and IMF traits in Xiang pigs, we integrated the whole genomic DNA genotyping and RNA-sequencing data in Longissimus thoracis muscle from F2 individuals of Large White × Xiang pig cross and performed the expression quantitative trait loci (eQTL) analyses. We identified a total of 64,419 significant cis-variant-eGene, including 33,288 eSNPs, 7,910 eInDels, and 5,940 eSVs. Among them, 1,156 cis-eQTL loci overlapped with QTL for IMF and BFT, affecting the expressions of 541 genes, including 54 transcription factors. Enrichment analysis revealed that key candidate genes were significantly associated with lipid metabolism pathways, such as mTOR and PPAR signaling. Overlap analysis demonstrated that several eQTLs exhibited dual regulatory effects on gene expression. Notable eQTL hotspots included eSNP 12:52760182_T, eInDel 2:6807484-6,807,485, and three eSV sites, with the P2RY6 gene significantly associated with multiple variants. Our results indicated that many regions across the pig genome that affected gene expression level (eQTL) and overlapped with QTL regions associated with the fat store and lipid metabolism for IMF and BFT. These findings enhance our understanding of the genetic mechanisms underlying fat deposition in Xiang pigs and provide a theoretical framework for future functional studies and genomic selection.

BackgroundHand, foot, and mouth disease (HFMD) is caused by more than 20 different enteroviruses (EVs). The predominant EV serotypes of HFMD have been continuously changing in recent years. Guizhou Province has reported higher rates of severe and fatal cases of HFMD. However, comprehensive studies on its epidemiology, etiology, and serological characteristics have remained limited in recent years.MethodsWe collected epidemiological and laboratory data from HFMD cases between 2008 and 2023, analyzing the data by age, gender, disease severity, and EV serotypes. Clinical samples from these cases were collected to isolate EVs. The VP1 gene was amplified from isolates of enterovirus A71 (EV-A71), coxsackievirus A16 (CV-A16), coxsackievirus A6 (CV-A6), and coxsackievirus A10 (CV-A10), and the sequences were analyzed. We collected 432 serum samples from healthy individuals from 2019 to 2022 to assess antibodies against CV-A6, CV-A10, CV-A16, and EV-A71 of HFMD.ResultsA total of 513,143 HFMD cases were reported in Guizhou Province from 2008 to 2023, including 9052 (1.76%) severe cases and 193 (0.038%) deaths. In laboratory-confirmed cases, EV-A71 was the dominant serotype from 2008 to 2012; other EVs became predominant from 2013 to 2018, and CV-A6 predominated in 2019, 2022, and 2023. Interestingly, novel epidemiological patterns of CV-A6 infection were observed, with a high incidence every other year in various cities since 2019. Among 432 healthy individuals, the overall seroprevalence rates of CV-A6, CV-A10, CV-A16, and EV-A71 were 62.04%, 54.17%, 54.63%, and 64.35%, respectively. Additionally, over 70% of the participants had neutralizing antibodies (NtAbs) against at least two types of these enteroviruses. Phylogenetic analysis revealed that CV-A16 isolates clustered into the B1a or B1b evolutionary branches, while EV-A71, CV-A6, and CV-A10 isolates belonged to the C4a, D3a, and C subgenotypes, respectively.ConclusionsThis results indicate differences in the incidence of major HFMD pathogens across years, regions, and populations. Other EVs, predominantly CV-A6, have become the main pathogens causing HFMD since 2019. CV-A6, CV-A10, CV-A16, and EV-A71 exhibited relatively high seroprevalence rates. Currently, there is an urgent need to develop multivalent vaccines and implement effective measures to reduce incidence of HFMD.

BACKGROUND:A phenotype characterized by age-dependent hairless, thicker and systemic wrinkled skin has been discovered from Chinese native Xiang pig herd, which is similar to the wrinkle skin phenotype and hereditary periodic fever syndrome observed in Shar-Pei dog. Our aim was to elucidate the functional mechanisms associated with these systemic wrinkle phenotype. RESULTS:Gene expression changes in wrinkled skin were first explored via RNA-seq technology. There were 699 differentially expressed genes (DEGs) in wrinkled skin, with 272 downregulated genes and 427 upregulated genes. Most of the upregulated genes were significantly centered at pathways that generate reactive oxygen species (ROS), such as carbon and lipid metabolism, the citrate cycle and the oxidation-reduction process, leading to excessive ROS production and/or DNA damage in both the epidermal and dermal compartments of the skin. The downregulated genes were involved mainly in extracellular matrix (ECM) signaling. The most decreased genes were focused on several ECM components, including COL1A1, COL21A1, COL6A3, and ELN, likely related with ECM disorders. Moreover, 831 SVs and 26 CNVs were found in the gene regions of 335 DEGs according to the genomic resequencing data from the same wrinkled skin and control samples. These SVs and CNVs contributed to the deletion, insertion, duplication and inversion in exons, introns and regulatory regions of the DEGs. We then used F2 crossbreds by using Large White pig and the wrinkled Xiang pig as parents to verify the skin features and gene expression changes. Approximately 26% of F2 individuals exhibited a wrinkled skin phenotype, whereas the skin on F1 hybrids remained unchanged, suggesting autosomal recessive inheritance. The disorganized and thinner collagen bundles in the dermis of wrinkled skin were very similar to those of the parents, and many genes were shared between F2 pigs and the parents, including 137 DEGs corresponding to wrinkle scores, 23 oxidative stress-related genes, nine aging-related genes, and four ECM genes. CONCLUSIONS:The systemic wrinkled skin phenotype of pigs involves aberrant gene expression and genetic variations related to oxidative stress and ECM components.

Background:To address the regional heterogeneity of Alzheimer's disease, a large-scale epidemiological study of 12,421 elderly individuals was conducted in southwestern China to depict its unique risk characteristics. Methods:A total of 12,421 subjects were selected via cluster sampling from southwestern China after low quality data were filtered out. On the basis of investigations and medical imaging examinations, three groups were distinguished: the AD, mild cognitive impairment (MCI), and normal control groups. The risk factors for AD and MCI were analysed via a multivariate logistic regression model. Results:This study identifies a high burden of cognitive impairment in southwestern China, with 22.07% of adults aged ≥60 years exhibiting cognitive decline and 5.81% diagnosed with Alzheimer's disease rates surpassing national and global averages. Key risk factors included age >80 years, female sex, low education, rural residence, surgical history, and urological comorbidities. These findings underscore the need for region-specific prevention strategies, prioritizing older, less-educated rural women through combined cognitive and vascular interventions, while integrating cognitive screening into primary care in underserved areas for early detection and intervention. Conclusion:Elderly individuals in southwestern China exhibit a high prevalence of cognitive impairment, with AD associated with complex risk factors including established contributors like advanced age, dementia family history, alcohol abuse, and multisystem comorbidities-while notably identifying surgical history and urolithiasis as region specific risk signals. These findings underscore regional, environmental, and ethnic influences on AD pathogenesis, requiring tailored prevention/treatment. Future priorities include integrating brief cognitive screening into primary care, targeting high-risk groups (eg, undereducated rural elderly women), and establishing prospective cohorts to clarify causal links between urolithiasis, surgical history, and cognitive decline for refined region-adapted AD prevention.

Young rabbits are sensitive to surrounding changes and conditioned pathogens in intestine which might result in slow inflammation and diarrhea after microbial invasion. Traditional medicine herbs could provide efficacious treatment on slow infection and inflammation. The present research designed an Acanthopanax senticosus (ACS) formula consisted of five types of Chinese herbs including Acanthopanax senticosus (Rupr & Maxim) Harms (Ciwujia in Chinese), Astragalus membranceus (Fisch) Bge (Huangqi in Chinese), Indigo naturalis (Qingdai in Chinese), Houttuynia cordata Thunb (Yuxingcao in Chinese), and Glycyrrhizae radix et Rhizoma (Gancao in Chinese). The effects of ACS decoction supplement were investigated via determination of cytokines and growth performances of young rabbits, and the flora in intestinal digesta from six fragments were further explored using 16S rRNA gene sequencing technology. Compared to the control group, rabbits supplied with different doses of ACS decoction possessed lower diarrhea and death rates, together with the IL-10 concentration, while the declined IL-1β and IL-12 levels and inflammatory factor gene expressions in intestinal tissues. Additionally, ACS addition changed the diversity of flora in each segments of intestine. Functional prediction on abundances of genera enriched to seven KEGG immunity pathways. Moreover, strong correlations were determined between the abundance of bacteria with interleukins contents, and the predictive immune signaling abundances, respectively. Especially, ACS exhibited anti-inflammation effects via decreasing the abundances of Bacteroides, Clostridia_vadinBB60_group, NK4A214_group, and dgA_11_gut_group in intestine of young rabbits. In conclusion, dietary supplement with ACS exerted diarrhea-reducing effects, and improved immunity homeostasis by modulating intestinal flora diversity in young rabbits.

In this study, we compared the transcriptome of longissimus dorsi muscle between Guizhou Xiang pigs (XP) and Western commercial Large White pigs (LW), which show diffirent meat quality between them. In terms of meat quality traits, the pH 45 min, color score, backfat thickness, and intramuscular fat (IMF) content were higher in Xiang pigs than in Large White pigs (P < 0.01), while the drip loss, lean meat percentage, shear force, and longissimus dorsi muscle area of Xiang pigs were lower than that of Large White pigs (P < 0.01). Nutrients such as monounsaturated fatty acid (MUFA), total amino acids (TAA), delicious amino acids (DAA) and essential amino acids (EAA) in Xiang pigs were higher than that in Large White pigs, and the proportion of polyunsaturated fatty acid (PUFA) of Xiang pigs was significantly lower than Large White pigs (P < 0.01). Transcriptome analysis identified 163 up-regulated genes and 88 genes down-regulated in Xiang pigs longissimus dorsi muscle. Combined with the correlation analysis and quantitative trait locis (QTLs) affecting meat quality, a total of 227 DEGs were screened to be significantly associated with meat quality values. Enrichment analysis indicated that numerous members of genes were gathered in muscle development, adipogenesis, amino acid metabolism, fatty acid metabolism and synthesis. Of those, 29 genes were identified to be hub genes that might be related with the meat quality of Xiang pig, such as MYOD1, ACTB, ASNS, FOXO1, ARG2, SLC2A4, PLIN2, and SCD. Thus, we screened and identified the potential functional genes for the formation of meat quality in Xiang pigs, which provides a corresponding theoretical basis for the study of the molecular regulatory mechanism of pork quality and the improvement of pork quality.

Copy number variation (CNV) tends to occur in genetically enriched regions and is likely associated with a number of complex diseases such as skin aging. In this study, we investigated the genome-wide CNVs in 20 wrinkled skin cases (WSC) of Xiang pigs and 63 controls, and identified 7893 copy number variable regions (CNVRs). We estimated the F-statistic (Fst) at each locus and identified that 93 case-controls stratified CNVRs (Fst > = 0.15) overlapped with 87 known genes. Functional enrichment analysis showed that most of these genes were predominantly enriched in pathways and terms related to the extracellular matrix. Finally, we found that some CNVs were predicted to have high effects on genes such as VCAN, TIMP1 and FOXO1 through transcriptional amplification, transcript ablation and so on. Most of the genes overlapped with those CNVRs have been reported to be related to aging in human or animals. The copy numbers presented the positive correlations with the transcript level of the genes in skins between the cases and controls. Our results suggested that those 22 CNVRs, including 19 CNV losses and 3 CNV gains, were putatively associated with the skin wrinkle of Xiang pigs.

Mutton quality is closely related to genetic variants and gene expression alterations during growth and development, resulting in differences in nutritional values, flavor, and odor. We first evaluated and compared the composition of crude protein, crude fat, cholesterol, amino acid (AA), and fatty acid (FA) in the longissimus dorsi muscle of Guizhou black goats (GZB, n = 5) and Yunshang black goats (YBG, n = 6). The contents of cholesterol and FA related to odor in GZB were significantly lower than that in YBG, while the concentrations of umami amino acids and intramuscular fat were significantly higher in GZB. Furthermore, structural variants (SVs) in the genomes of GZB (n = 30) and YBG (n = 11) were explored. It was found that some regions in Chr 10/12/18 were densely involved with a large number of SVs in the genomes of GZB and YBG. By setting FST ≥ 0.25, we got 837 stratified SVs, of which 25 SVs (involved in 12 genes, e.g., CORO1A, CLIC6, PCSK2, and TMEM9) were limited in GZB. Functional enrichment analysis of 14 protein-coding genes (e.g., ENPEP, LIPC, ABCA5, and SLC6A15) revealed multiple terms and pathways related with metabolisms of AA, FA, and cholesterol. The SVs (n = 10) obtained by the whole genome resequencing were confirmed in percentages of 36.67 to 86.67

African swine fever (ASF) is a rapidly fatal viral haemorrhagic fever in Chinese domestic pigs. Although very high mortality is observed in pig farms after an ASF outbreak, clinically healthy and antibody-positive pigs are found in those farms, and viral detection is rare from these pigs. The ability of pigs to resist ASF viral infection may be modulated by host genetic variations. However, the genetic basis of the resistance of domestic pigs against ASF remains unclear. We generated a comprehensive set of structural variations (SVs) in a Chinese indigenous Xiang pig with ASF-resistant (Xiang-R) and ASF-susceptible (Xiang-S) phenotypes using whole-genome resequencing method. A total of 53,589 nonredundant SVs were identified, with an average of 25,656 SVs per individual in the Xiang pig genome, including insertion, deletion, inversion and duplication variations. The Xiang-R group harboured more SVs than the Xiang-S group. The F-statistics (FST) was carried out to reveal genetic differences between two populations using the resequencing data at each SV locus. We identified 2,414 population-stratified SVs and annotated 1,152 Ensembl genes (including 986 protein-coding genes), in which 1,326 SVs might disturb the structure and expression of the Ensembl genes. Those protein-coding genes were mainly enriched in the Wnt, Hippo, and calcium signalling pathways. Other important pathways associated with the ASF viral infection were also identified, such as the endocytosis, apoptosis, focal adhesion, Fc gamma R-mediated phagocytosis, junction, NOD-like receptor, PI3K-Akt, and c-type lectin receptor signalling pathways. Finally, we identified 135 candidate adaptive genes overlapping 166 SVs that were involved in the virus entry and virus-host cell interactions. The fact that some of population-stratified SVs regions detected as selective sweep signals gave another support for the genetic variations affecting pig resistance against ASF. The research indicates that SVs play an important role in the evolutionary processes of Xiang pig adaptation to ASF infection.

Background Guizhou black goat is one of the indigenous black goat breeds in the southwest region of Guizhou, China, which is an ordinary goat for mutton production. They are characterized by moderate body size, black coat, favorite meat quality with tender meat and lower odor, and tolerance for cold and crude feed. However, little is known about the genetic characteristics or variations underlying their important economic traits. Results Here, we resequenced the whole genome of Guizhou black goat from 30 unrelated individuals breeding in the five core farms. A total of 9,835,610 SNPs were detected, and 2,178,818 SNPs were identified specifically in this breed. The population structure analysis revealed that Guizhou black goat shared a common ancestry with Shaanbei white cashmere goat (0.146), Yunshang black goat (0.103), Iran indigenous goat (0.054), and Moroccan goat (0.002). However, Guizhou black goat showed relatively higher genetic diversity and a lower level of linkage disequilibrium than the other seven goat breeds by the analysis of the nucleotide diversity, linkage disequilibrium decay, and runs of homozygosity. Based on F ST and θ π values, we identified 645, 813, and 804 selected regions between Guizhou black goat and Yunshang black goat, Iran indigenous goat, and cashmere goats. Combined with the results of XP-EHH, there were 286, 322, and 359 candidate genes, respectively. Functional annotation analysis revealed that these genes are potentially responsible for the immune response (e.g., CD28 , CD274 , IL1A , TLR2 , and SLC25A31 ), humility-cold resistance (e.g., HBEGF , SOSTDC1 , ARNT , COL4A1/2 , and EP300 ), meat quality traits (e.g., CHUK , GAB2 , PLAAT3 , and EP300 ), growth (e.g., GAB2 , DPYD , and CSF1 ), fertility (e.g., METTL15 and MEI1 ), and visual function (e.g., PANK2 and NMNAT2 ) in Guizhou black goat. Conclusion Our results indicated that Guizhou black goat had a high level of genomic diversity and a low level of linkage disequilibrium in the whole genome. Selection signatures were detected in the genomic regions that were mainly related to growth and development, meat quality, reproduction, disease resistance, and humidity-cold resistance in Guizhou black goat. These results would provide a basis for further resource protection and breeding improvement of this very local breed.

The Xifeng Hot Spring is one of the eight largest hot springs in China, which is rich in radon gas and sulphur in karst scenery. Little is known about the microbiota structure in the spring. The water was collected from three sites containing the outlet of spring water discharge site (OWD), spring pool for tourist (SPT) and sewage effluent pool (SEP) in the Xifeng Hot Spring and further analyzed by culture-independent technique and culture-dependent method. A total of 57 phyla were identified from the water samples. The dominate phyla at OWD was Bacteroidetes (46.93%), while it was Proteobacteria in both sites of SEP and SPT with relative richness of 61.9% and 94.9%, respectively. Two bacteria, Deinococcus and Hymenobacter , that confirmed to be radiation-resistant, seven sulphur bacteria and three thermophilic bacteria were detected from Xifeng Hot Spring. Furthermore, it was found that genus Flavobacterium was susceptible to environmental change with abundance of 11 ~ 2825 times higher in OWD than the other two groups. Compared bacteria from the OWD group with that from 14 hot springs in six countries, total 94 unique genera bacteria were found out from the Xifeng Hot Spring including four thiometabolism-related bacteria ( Propionispira , Desulforegula , Desulfobacter and Desulfococcus ) and the thermophilic bacterium ( Symbiobacterium ). Using microbial culturing and isolation technology, sixteen strains were isolated from the water samples of three sites. The diversity of microbiota was abundant and variable along with the niche changed in conditions and surroundings. It indicated that numbers of valuable bacteria resources could be explored from the special surroundings of Xifeng Hot Spring.

Skeletal muscle is the most abundant tissue in mammals, and myogenesis and differentiation require a series of regulatory factors such as microRNAs (miRNAs). In this study, we found that miR-103-3p was highly expressed in the skeletal muscle of mice, and the effects of miR-103-3p on skeletal muscle development were explored using myoblast C2C12 cells as a model. The results showed that miR-103-3p could significantly reduce myotube formation and restrain the differentiation of C2C12 cells. Additionally, miR-103-3p obviously prevented the production of autolysosomes and inhibited the autophagy of C2C12 cells. Moreover, bioinformatics prediction and dual-luciferase reporter assays confirmed that miR-103-3p could directly target the microtubule-associated protein 4 (MAP4) gene. The effects of MAP4 on the differentiation and autophagy of myoblasts were then elucidated. MAP4 promoted both the differentiation and autophagy of C2C12 cells, which was contrary to the role of miR-103-3p. Further research revealed that MAP4 colocalized with LC3 in C2C12 cell cytoplasm, and the immunoprecipitation assay showed that MAP4 interacted with autophagy marker LC3 to regulate the autophagy of C2C12 cells. Overall, these results indicated that miR-103-3p regulated the differentiation and autophagy of myoblasts by targeting MAP4. These findings enrich the understanding of the regulatory network of miRNAs involved in the myogenesis of skeletal muscle.

The aim of the experiment was to explore the molecular mechanism of Qingdai compound preparation regulating immunity in pigs by using network pharmacology. The compounds contained in Qingdai compound preparation were screened from the Tcmsp database to obtain the active active ingredients, and the target genes for the action of the active active ingredients were predicted. The 'active ingredient-target gene' network analysis was made to screen out the main active ingredients. The immune-related genes were analyzed through Omim database and intersected with the target genes of active ingredients to obtain the porcine immune-related genes targeted by the compound preparation. The protein interaction network map (PPI) was constructed using String database and Cytoscape was used to screen the core proteins of PPI. The gene function and KEGG pathway of target gene enrichment were analyzed by David database. Molecular docking with Autodock and PyMol was performed to verify the interaction between core active ingredients and core target proteins. The results showed that Qingdai compound preparation contained 330 compounds, of which 62 were active ingredients, and 10 were main active substances, including β-sitosterol, stigmasterol, iviogenin, kaempferol, isovitexin, etc. The active ingredient acts on 64 porcine immune-related genes and 20 core target proteins such as SLC6A4, ESR1, TNF, etc. It regulates the natural immunity and specific immune system functions through IL-17 signaling pathway, TNF signaling pathway, NOD-like receptor signaling pathway, etc., and improves the resistance of porcine pigs. Qingdai compound preparation can affect the expression of IL-2, IFN-α, IFN-γ and other inflammatory factors through β-sitosterol, isovitexin and other main active ingredients. Macromolecular docking analysis showed that these active ingredients could effectively bind to target proteins, which could provide a reference for the research on the material basis and mechanism of traditional Chinese medicine to enhance immunity. At the same time, the results of network pharmacology analysis can provide a theoretical basis for a comprehensive analysis of the mechanism of Indigo Naturalis Polypill in enhancing pig immunity.

[目的]探究猪特有的细胞色素P450超家族3A亚族成员29(cytochrome P450 3A 29,CYP3A29)基因3'-UTR区核苷酸结构变异(structural variation,SV)对该基因表达的影响,解析该结构变异与香猪肉质性状形成的关系.[方法]以香猪、大白猪为研究对象,应用UCSC、miRBase、PITA、RBPsuite等软件预测CYP3A29基因3'-UTR区结构变异区间所包含的重复元件、miRNA和RBP结合位点;采用PCR方法对结构变异位点进行基因分型;计算群体基因型频率、基因频率及Hardy-Weinberg平衡状态;采用实时荧光定量PCR检测该变异对CYP3A29基因表达的影响;利用ELISA法检测香猪肌肉组织中CYP3A29蛋白含量和雄激素(T、DHT)水平,并使用SPSS 17.0软件对T、DHT含量与不同基因型进行Pearson相关性分析.[结果]生物信息学分析显示,CYP3A29基因3'-UTR的结构变异区全长303 bp,该变异区存在1个长为241 bp的短散在元件(short interspersed element,SINE),属于Pre0_SStRNA家族,位于Chr3:6 819 917-6 820 161,含有33个miRNAs和13个RBPs结合位点.基因分型显示,该结构变异在香猪、大白猪群体中呈现出丰富的多态性,表现为Ⅱ基因型(插入型)、ID基因型(杂合型)、DD基因型(双缺失).基因型群体分布频率显示,香猪中的D等位基因频率高于大白猪,该结构变异在香猪中处于Hardy-Weinberg平衡状态(P＞0.05),在大白猪中处于Hardy-Weinberg不平衡状态(P＜0.05).实时荧光定量PCR和EL1SA结果显示,CYP3A29基因ID基因型个体mRNA表达量和蛋白含量均显著高于Ⅱ和DD基因型(P＜0.05).相关性分析结果显示,香猪不同基因型个体CYP3A29基因含量与肌肉组织中T浓度呈显著相关(P＜0.05),与DHT浓度不相关(P＞0.05).香猪ID基因型个体T浓度显著高于DD和Ⅱ基因型(P＜0.05),DD基因型个体DHT浓度显著高于ID和Ⅱ基因型(P＜0.05).[结论]CYP3A29基因3'-UTR区长度为303 bp的结构变异在香猪群体中具有多态性,以缺失型D等位基因为主,即该结构变异在香猪中缺失SINE元件,导致CYP3A29基因表达上调,进而影响香猪肌肉组织中T浓度.CYP3A29基因3'-UTR结构变异与香猪肉质性状具有相关性.

皮肤是哺乳动物抵御致病因素的第 1 道防线,在皮肤衰老的过程中,整合到哺乳动物基因组中的病毒源基因序列会被重新激活,影响宿主基因的表达,激起细胞对病毒的反应,引起慢性炎症、促进细胞衰老.本研究旨在探究病毒源基因表达与皱皮香猪皮肤皱纹形成的关系,选取 12～18 月龄皱皮香猪和普通香猪各 5头进行转录组测序和生物信息学分析,采用RT-qPCR方法对皱皮香猪中差异表达的病毒源基因进行验证.2组中皮肤组织差异表达基因与病毒整合位点数据库进行比对分析,筛选出 612 个差异表达的病毒源基因.以普通香猪为对照,得到上调基因182个,下调基因430个,这些差异表达的病毒源基因主要富集在皮肤发育、炎症反应等GO条目,以及长寿调控途径、FoxO信号通路、细胞衰老等KEGG通路.经猪-人同源基因转换,与免疫、衰老、人类皮肤相关的基因取交集,筛选出高表达的候选基因 6 个:AGT、CAT、CTNNB1、JUP、KRT75和LIPE,采用RT-qPCR验证了皮肤中这 6 个基因的差异表达,与转录组测序结果趋势一致.研究结果表明,皱皮香猪皮肤皱纹的形成与病毒源基因的异常表达有紧密联系,本研究可为后期皱皮香猪皮肤褶皱形成的分子机制及衰老分子标记物选择奠定理论基础.

The biological clock plays a critical role in the reproductive system of various living organisms like swine.To examine the effects of estrus cycle on the expression of ovarian biological clock-related gene in Xiang pig,this study analyzed the expression and alternative splicing of biological clock-related genes in ovary during estrus and diestrus periods.In total,90 clock-related genes expressed in the ovaries of the Xiang pig were detected and 33 clock-related genes differentially expressed between estrous and diestrous stages.44 differential splicing events were identified from the transcripts of 34 biological clock-related genes.Furthermore,it was found 20 genes including the core clock components,arntl and cry1 were differentially regulated only at AS level and 14 genes,including per1 and clock,were differentially regulated at both expression and AS levels.We also proved that the core clock genes per1,cry1,clock and arntl and the clock-related genes,ppp1cb and ntrk1 were rhythmically expressed in Xiang pig ovaries by RT-qPCR experiments.The results demonstrated that the biological clock in the ovaries of Xiang pig might play an important role in regulating the ovarian physiological functions by the transcriptional and post-transcriptional regulation.