Pig Tissues Research Articles

A comprehensive atlas of pig RNA editome across 23 tissues reveals RNA editing affecting interaction mRNA-miRNAs.

RNA editing is a co-transcriptional/post-transcriptional modification that is mediated by the ADAR enzyme family. Profiling of RNA editing is very limited in pigs. In this study, we collated 3813 RNA-seq data from the public repositories across 23 tissues and carried out comprehensive profiling of RNA editing in pigs. In total, 127,927 A-to-I RNA-editing sites were detected. Our analysis showed that 98.2% of RNA-editing sites were located within repeat regions, primarily within the pig-specific SINE retrotransposon PRE-1/Pre0_SS elements. Subsequently, we focused on analyzing specific RNA-editing sites (SESs) in skeletal muscle tissues. Functional enrichment analyses suggested that they were enriched in signaling pathways associated with muscle cell differentiation, including DMD, MYOD1, and CAV1 genes. Furthermore, we discovered that RNA editing event in the 3'UTR of CFLAR mRNA influenced miR-708-5p binding in this region. In this study, the panoramic RNA-editing landscape of different tissues of pigs was systematically mapped, and RNA-editing sites and genes involved in muscle cell differentiation were identified. In summary, we identified modifications to pig RNA-editing sites and provided candidate targets for further validation.

Monoclonal Antibody against Porcine LAG3 Inhibits Porcine Reproductive and Respiratory Syndrome Virus Infection

Lymphocyte activation gene 3 (LAG3) is an inhibitory receptor and the interaction between fibrinogen-like protein 1 and LAG3 can inhibit the anti-tumor effect of T cells both in vivo and in vitro, which was regarded as a new immune evasion mechanism. Porcine reproductive and respiratory syndrome (PRRS), caused by PRRSV, is an infectious disease characterized by reproductive disorders in pregnant sows and gilts and respiratory problems in pigs of all ages, seriously impacting the pig industry worldwide. In this study, monoclonal antibodies (mAbs) against porcine LAG3 (pLAG3) were developed, and one mAb (1C2) showed good reactivity with pLAG3 on PHA-activated porcine peripheral blood lymphocytes. Epitope mapping showed the epitope recognized by mAb 1C2 was located at amino acid residues 214–435 of pLAG3. LAG3 expression in the tissues of PRRSV-infected pigs was detected, using mAb 1C2 as the primary antibody, and the results revealed that PRRSV infection caused a marked increase in LAG3 expression compared to the control group. Interference of LAG3 expression on PHA-activated lymphocytes promoted PRRSV replication in the co-culture system of monocyte-derived dendritic cells and lymphocytes, whereas overexpression of LAG3 or blocking of the LAG3 signal with mAb 1C2 inhibited PRRSV replication, indicating that PRRSV infection activates the LAG3-signaling pathway, suggesting that this pathway plays an important role in PRRSV pathogenesis. The results obtained lay the foundation for subsequent research on the role of LAG3 in PRRS and other diseases with persistent infection characteristics.

Identification of genomic regions associated with fatty acid metabolism across blood, liver, backfat and muscle in pigs

BackgroundThe composition and distribution of fatty acids (FA) are important factors determining the quality, flavor, and nutrient value of meat. In addition, FAs synthesized in the body participate in energy metabolism and are involved in different regulatory pathways in the form of signaling molecules or by acting as agonist or antagonist ligands of different nuclear receptors. Finally, synthesis and catabolism of FAs affect adaptive immunity by regulating lymphocyte metabolism. The present study performed genome-wide association studies using FA profiles of blood, liver, backfat and muscle from 432 commercial Duroc pigs.ResultsTwenty-five genomic regions located on 15 Sus scrofa chromosomes (SSC) were detected. Annotation of the quantitative trait locus (QTL) regions identified 49 lipid metabolism-related candidate genes. Among these QTLs, four were identified in more than one tissue. The ratio of C20:4n-6/C20:3n-6 was associated with the region on SSC2 at 7.56–14.26 Mb for backfat, liver, and muscle. Members of the fatty acid desaturase gene cluster (FADS1, FADS2, and FADS3) are the most promising candidate genes in this region. Two QTL regions on SSC14 (103.81–115.64 Mb and 100.91–128.14 Mb) were identified for FA desaturation in backfat and muscle. In addition, two separate regions on SSC9 at 0 – 14.55 Mb and on SSC12 at 0–1.91 Mb were both associated with the same multiple FA traits for backfat, with candidate genes involved in de novo FA synthesis and triacylglycerol (TAG) metabolism, such as DGAT2 and FASN. The ratio C20:0/C18:0 was associated with the region on SSC5 at 64.84–78.32 Mb for backfat. Furthermore, the association of the C16:0 content with the region at 118.92–123.95 Mb on SSC4 was blood specific. Finally, candidate genes involved in de novo lipogenesis regulate T cell differentiation and promote the generation of palmitoleate, an adipokine that alleviates inflammation.ConclusionsSeveral SNPs and candidate genes were associated with lipid metabolism in blood, liver, backfat, and muscle. These results contribute to elucidating the molecular mechanisms implicated in the determination of the FA profile in different pig tissues and can be useful in selection programs that aim to improve health and energy metabolism in pigs.

49 Crossing-talk between micronutrient and macronutrient metabolism: Implication in health and disease

Abstract Recent goals of swine ration formulation have evolved from preventing nutritional deficiency diseases and maximizing growth performance to optimizing immune function and disease resistance with least climate impact. Thus, certain micronutrients such as selenium (Se) are widely considered to be health beneficial at dietary concentrations above nutrient requirements. During the past decade, we have conducted a number of experiments to determine impacts and mechanisms of dietary Se concentrations (0.03 to 1 or 3 mg/kg) on metabolism of carbohydrate, lipid, and protein in pigs from growing to finishing. Strikingly, pigs fed high-Se diets developed hyperinsulinemia and insulin resistance. Those pigs also had increased lipid, protein, saturated fatty acids (myristic, palmitic, stearic, and behenic acids), and aspartic acid, glutamine, and alanine and decreased polyunsaturated fatty acids (dihomo-γ-linolenic, oleic, α-linolenic, and eicosatrienoic acids) in the liver and(or) adipose tissues. The high-Se diets increased and decreased amounts of 5 and 2 selenoproteins, respectively, in tissues of pigs. These effects were associated with down-regulated insulin signaling (INSR and AKT), glycolysis (glucokinase) and gluconeogenesis (PEPCK) and up-regulated lipogenesis (FOXO1, SREBP1, ACC and FAS), and protein synthesis (mTOR and RPS6/S6). In contrast, high dietary fat intakes up- or down-regulated expressions of all selenoproteins except for TXNRD2 and TXNRD3 and showed a strong effect on expressions of DIO2, GPX6, and SELENOK. These induced selenogenome expression changes were associated with increased serum malondialdehyde and plasma concentrations of tumor necrosis factor alpha and interleukin-6, along with risk of obesity and chronic inflammation. Altogether, metabolisms of micronutrients and macronutrients in pigs are coordinated through interdependent and mutually regulated crossing talks at tissue, cellular, biochemical, and signal levels.

SP8.12 - Evaluating the pig tissue model for acute proctology simulation at General Surgery ST3 Bootcamp

Abstract Evaluating the pig tissue model for acute proctology simulation at General Surgery ST3 Bootcamp Background Acute proctology is an important skill for any general surgical trainee; however it is a skill which often is overlooked when it comes to training. Most, if not all teaching happens during patient procedures. Simulation in this area is underdeveloped and there is a lack of validated models for simulation. These models are also usually expensive and difficult to obtain. The aim of this study was to assess face validity of a low-cost model for teaching acute proctology during the ST3 general surgical bootcamp. Methods We created a model using a pig rectum and perineum. This provided two uses. Firstly, we were able to teach relevant clinical anatomy. Secondly, we were able to simulate EUA, rigid sigmoidoscopy, decompression of volvulus and haemorrhoidal banding. The models allowed for repeated use. Results Twenty-five trainees took part in this simulation. Overall, the trainees felt the rectal examination was realistic (76%), the sigmoidoscope moved realistically in the model (72%) as did the proctoscope (88%). In terms of ability, trainees felt the simulation improved their confidence in EUA (68%), rectal examination (72%) and banding (80%). A total of 80% of trainees thought the model was overall “Good or Excellent”. Conclusions Our study demonstrates that this low-cost, high-fidelity simulation model has a high face validity and may be explored for the use for training new general surgical registrars.

Myosin Vb Traffics P-glycoprotein to the Apical Membrane of Intestinal Epithelial Cells

The xenobiotic efflux pump P-glycoprotein is highly expressed on the apical membrane of the gastrointestinal tract, where it regulates the levels of intracellular substrates. P-glycoprotein is altered in disease, but the mechanisms that regulate the levels of P-glycoprotein are still being explored. The molecular motor myosin Vb (Myo5b) traffics diverse cargo to the apical membrane of intestinal epithelial cells. We hypothesized that Myo5b was responsible for the delivery of P-glycoprotein to the apical membrane of enterocytes. We used multiple murine models that lack functional Myo5b or the myosin binding partner Rab11a to analyze P-glycoprotein localization. Pig and human tissue were analyzed to determine P-glycoprotein localization in the setting of MYO5B mutations. Intestinal organoids were used to examine P-glycoprotein trafficking and to assay P-glycoprotein function when MYO5 is inhibited. In mice lacking Myo5b or the binding partner Rab11a, P-glycoprotein was improperly trafficked and had decreased presence in the brush border of enterocytes. Immunostaining of a pig model lacking functional Myo5b and human biopsies from a patient with an inactivating mutation in Myo5b also showed altered localization of intestinal P-glycoprotein. Human intestinal organoids expressing the motorless MYO5B tail domain had colocalization with P-glycoprotein, confirming that P-glycoprotein was trafficked by MYO5B in human enterocytes. Inhibition of MYO5 in human intestinal cell lines and organoids resulted in decreased P-glycoprotein capacity. Additionally, inhibition of MYO5 in human colon cancer cells diminished P-glycoprotein activity and increased cell death in response to a chemotherapeutic drug. Collectively, these data demonstrate that Myo5b is necessary for the apical delivery of P-glycoprotein.

Visceral pleura mechanics: Characterization of human, pig, and rat lung material properties

Pulmonary air leaks are amongst the most common complications in lung surgery. Lung sealants are applied to the organ surface and need to synchronously stretch with the visceral pleura, the layer of tissue which encompasses the lung parenchymal tissue. These adhesives are commonly tested on pig and rat lungs, but applied to human lungs. However, the unknown mechanics of human lung visceral pleura undermines the clinical translatability of such animal-tested sealants and the absence of how pig and rat lung visceral pleura compare to human tissues is necessary to address. Here we quantify the biaxial planar tensile mechanics of visceral pleura from healthy transplant-eligible and smoker human lungs for the first time, and further compare the material behaviors to pig and rat lung visceral pleura. Initial and final stiffness moduli, maximum stress, low-to-high strain transition, and stress relaxation are analyzed and compared between and within groups, further considering regional and directional dependencies. Visceral pleura tissue from all species behave isotropically, and pig and human visceral pleura exhibits regional heterogeneity (i.e. upper versus lower lobe differences). We find that pig visceral pleura exhibits similar initial stiffness moduli and regional trends compared to human visceral pleura, suggesting pig tissue may serve as a viable animal model candidate for lung sealant testing. The outcomes and mechanical characterization of these scarce tissues enables future development of biomimetic lung sealants for improved surgical applications. Statement of significanceSurgical lung sealants must synchronously deform with the underlying tissue and with each breath to minimize post-operative air leaks, which remain the most frequent complications of pulmonary intervention. These adhesives are often tested on pig and rat lungs, but applied to humans; however, the material properties of human lung visceral pleura were previously unexplored. Here, for the first time, the mechanics of human visceral pleura tissue are investigated, further contrasting rarely acquired donated lungs from healthy and smoking individuals, and additionally, comparing biaxial planar material characterizations to animal models often employed for pulmonary sealant development. This fundamental material characterization addresses key hindrances in the advancement of biomimetic sealants and evaluates the translatability of animal model experiments for clinical applications.

Ssc-miR-101-3p inhibits hypoxia-induced apoptosis and inflammatory response in alveolar type-II epithelial cells of Tibetan pigs via targeting FOXO3

Tibetan pigs are a unique swine strain adapted to the hypoxic environment of the plateau regions in China. The unique mechanisms underlying the adaption by Tibetan pigs, however, are still elusive. Only few studies have investigated hypoxia-associated molecular regulation in the lung tissues of animals living in the plateau region of China. Our previous study reported that ssc-miR-101-3p expression significantly differed in the lung tissues of Tibetan pigs at different altitudes, suggesting that ssc-miR-101-3p plays an important role in the adaptation of Tibetan pigs to high altitude. To understand the underlying molecular mechanism, in this study, the target genes of ssc-miR-101-3p and their functions were analyzed via various methods including qRT-PCR and GO and KEGG pathway enrichment analyses. The action of ssc-miR-101-3p was investigated by culturing alveolar type-II epithelial cells (ATII) of Tibetan pigs under hypoxic conditions and transfecting ATII cells with vectors overexpressing or inhibiting ssc-miR-101-3p. Overexpression of ssc-miR-101-3p significantly increased the proliferation of ATII cells and decreased the expression of inflammatory and apoptotic factors. The target genes of ssc-miR-101-3p were significantly enriched in FOXO and PI3K-AKT signaling pathways required to mitigate lung injury. Further, FOXO3 was identified as a direct target of ssc-miR-101-3p. Interestingly, ssc-miR-101-3p overexpression reversed the damaging effect of FOXO3 in the ATII cells. In conclusion, ssc-miR-101-3p targeting FOXO3 could inhibit hypoxia-induced apoptosis and inflammatory response in ATII cells of Tibetan pigs. These results provided new insights into the molecular mechanisms elucidating the response of lung tissues of Tibetan pigs to hypoxic stress.

Revealing the lethal effects of Pasteurella multocida toxin on multiple organ systems.

Pasteurella multocida toxin (PMT) is one of the most important virulence factors of Pasteurella multocida type D. Pasteurella multocida infection has caused enormous economic losses in the pig farming industry. Although it is well known that this bacterial infection causes progressive atrophic rhinitis, its effects on other organ tissues in pigs are unclear. In this study, PMT was expressed and purified, and the cytotoxic effects of PMT on four types of swine cells, LLC-PK1, PAM, IPEC, and ST, were investigated. LLC-PK1 exhibited the highest sensitivity to the cytotoxic effects of PMT. Our studies revealed that a PMT concentration of 0.1 μg/kg can lead to weight loss, whereas a PMT concentration of 0.5 μg/kg can lead to death in mice. PMT causes damage to the intestines, kidneys, lungs, livers, and spleens of mice. Furthermore, PMT caused acute death in pigs at treatment concentrations greater than 5 μg/kg; at PMT concentration of 2.5 μg/kg, weight loss occurred until death. PMT mainly caused damage to the hearts, lungs, livers, spleens and kidneys of pigs. The organ coefficient showed that damage to the heart and kidneys was the most severe and caused the renal pelvis and renal pyramid to dissolve and become cavitated. Pathology revealed hemorrhage in the lungs, liver, and spleen, and the kidneys were swollen and vacuolated, which was consistent with the damaged target organs in the mice. In conclusion, these findings demonstrate that PMT is extremely toxic in vitro and in vivo, causing damage to various organs of the body, especially the kidneys and lungs. This study provides a theoretical basis for the in-depth exploration of the cytotoxic effects of PMT on target organs.

3D Humanized Bioprinted Tubulointerstitium Model to Emulate Renal Fibrosis In Vitro.

Chronic kidney disease (CKD) leads to a gradual loss of kidney function, with fibrosis as pathological endpoint, which is characterized by extracellular matrix (ECM) deposition and remodeling. Traditionally, in vivo models are used to study interstitial fibrosis, through histological characterization of biopsy tissue. However, ethical considerations and the 3Rs (replacement, reduction, and refinement) regulations emphasizes the need for humanized 3D in vitro models. This study introduces a bioprinted in vitro model which combines primary human cells and decellularized and partially digested extracellular matrix (ddECM). A protocol was established to decellularize kidney pig tissue and the ddECM was used to encapsulate human renal cells. To investigate fibrosis progression, cells were treated with transforming growth factor beta 1 (TGF-β1), and the mechanical properties of the ddECM hydrogel were modulated using vitamin B2 crosslinking. The bioprinting perfusable model replicates the renal tubulointerstitium. Results show an increased Young's modulus over time, together with the increase of ECM components and cell dedifferentiation toward myofibroblasts. Multiple fibrotic genes resulted upregulated, and the model closely resembled fibrotic human tissue in terms of collagen deposition. This 3D bioprinted model offers a more physiologically relevant platform for studying kidney fibrosis, potentially improving disease progression research and high-throughput drug screening.

Abstract We008: Hallmark Maturation of the Human Pluripotent Stem Cells-derived Cardiovascular Progenitors in Myocardial Infarcted Large Animal Model

For the goal of regenerative cardiology, cellular therapy may be used to restore injured heart muscle. The molecular mechanisms behind the engraftment and maturation of these transplanted cells, however, remain poorly understood. Thus, to obtain a comprehensive understanding of the complex transcriptional landscapes within their transplanted spatial context, we utilized spatial transcriptomics in this study. We have previously demonstrated the safety and effectiveness of transplanting our cardiovascular progenitors (CVPs) produced from laminin-221 into pig hearts that had suffered myocardial infarction (MI). Here, we undertake 10x spatial transcriptomics in chronic MI pig models at 1-, 4-, and 12 weeks post-transplantation to further our understanding of these cells in vivo. With confidence, we can distinguish between the transcripts from the pigs and the humans in the infarcted region. By using human-specific anti-Ku80 antibodies for immunohistochemical labeling of the pig tissue samples, this in silico prediction was verified. The CVPs' engraftment and longevity in the infracted region were shown by the long-term identification and staining of human cells at various time points. It is interesting to note that after 12 weeks, ribosomal and mitochondrial activity were higher than at 1 and 2 weeks. These results are consistent with the hallmarks of cardiac maturation, in which cells transit from immature fetal-like cells to mature cardiomyocytes (CMs) by switching their metabolic energy source from glucose to fatty acids. Also, by comparing the heart-specific genes, including MYH6 and MYH7, we were able to quantify the myosin isoform switches in the xenograft over time. Furthermore, to delineate the fate of the transplanted cells, we performed a ligand-receptor interaction analysis. Intriguing, we identified the midkine protein as a possible secreted human paracrine factor that enhances angiogenesis in the graft. In conclusion, we have shown the successful engraftment and maturation of human CVPs toward CMs using spatial transcriptomic technology. These data will be used to understand other aspects of regenerative cardiology such as the mechanism of VTs and in vivo cellular repair mechanisms. A shiny app atlas has been generated to share our transcriptomics data for all to explore gene expression of human transplanted cells in myocardial infarcted pig hearts.

Safety assessment of electrosurgical electrodes by using mini pig tissue

Electrosurgical electrodes are the main dissecting devices widely used for surgeries throughout the world. The present study aimed to evaluate the thermal injury and safety within animals' organs following a minimally invasive electrosurgery technique with electrosurgical electrode AE40-300 (LIPO) and AE20-80 (LIFT). To ensure the effective application of electrosurgery in a clinical environment, it is crucial to minimize heat-induced injury to nearby tissues. In this study, the skin, liver, kidney, and femoral muscle dissected from 9 minipigs were used in tissue thermal spread experiments. Thermal imaging area analysis, maximum temperature, and time to reach basal temperature were evaluated. Thermography results revealed that the surgical temperature was significantly lower in the minimally invasive electrosurgery with AE40-300 (LIPO) and AE20-80 (LIFT) compared to the predicate device. In addition, AE40-300 (LIPO) and AE20-80 (LIFT) created a relatively small thermal injury area and thermal diffusion. Our results indicated that the tested devices named AE40-300 (LIPO) and AE20-80 (LIFT) reduced excessive thermal injury and could be applied to clinical use safely.

Integrating Multi-Omics Data to Identify Key Functional Variants Affecting Feed Efficiency in Large White Boars.

Optimizing feed efficiency through the feed conversion ratio (FCR) is paramount for economic viability and sustainability. In this study, we integrated RNA-seq, ATAC-seq, and genome-wide association study (GWAS) data to investigate key functional variants associated with feed efficiency in pigs. Identification of differentially expressed genes in the duodenal and muscle tissues of low- and high-FCR pigs revealed that pathways related to digestion of dietary carbohydrate are responsible for differences in feed efficiency between individuals. Differential open chromatin regions identified by ATAC-seq were linked to genes involved in glycolytic and fatty acid processes. GWAS identified 211 significant single-nucleotide polymorphisms associated with feed efficiency traits, with candidate genes PPP1R14C, TH, and CTSD. Integration of duodenal ATAC-seq data and GWAS data identified six key functional variants, particularly in the 1500985-1509676 region on chromosome 2. In those regions, CTSD was found to be highly expressed in the duodenal tissues of pigs with a high feed conversion ratio, suggesting its role as a potential target gene. Overall, the integration of multi-omics data provided insights into the genetic basis of feed efficiency, offering valuable information for breeding more efficient pig breeds.

Development of a highly reproducible GC-HRMS method for determination of Skatole in pig tissues

This study presents a sensitive and reproducible mass spectrometry method for quantifying skatole in porcine adipose tissue, muscle, and serum samples applicable for abattoirs and laboratories. Leveraging gas chromatography-high-resolution Orbitrap microscopy and microwave-assisted liquefication of the adipose tissue, the method demonstrates robust performance across key parameters. Impressive linearity (R2) values of 0.9999 and 0.9996 for adipose tissue and serum, respectively. Notably, the method exhibits a low Limit of Detection (LoD) of 0.5 ppb for adipose tissue and 0.9 ppb for serum, with corresponding Limits of Quantification (LoQ) at 1.65 ppb and 3.04 ppb, respectively. The method showed significant reproducibility of 5.9% and repeatability (RSD%) of 8.78% for adipose tissue and 4.08% for serum, with recovery rates of 90% and 87%, respectively. This streamlined method offers promising, effective quantification of boar taint compounds, emphasizing its sensitivity and reproducibility.

Lysine 2-hydroxyisobutyrylation levels determined adipogenesis and fat accumulation in adipose tissue in pigs

BackgroundExcessive backfat deposition lowering carcass grade is a major concern in the pig industry, especially in most breeds of obese type pigs. The mechanisms involved in adipogenesis and fat accumulation in pigs remain unclear. Lysine 2-hydroxyisobutyrylation (Khib), is a novel protein post-translational modification (PTM), which play an important role in transcription, energy metabolism and metastasis of cancer cells, but its role in adipogenesis and fat accumulation has not been shown.ResultsIn this study, we first analyzed the modification levels of acetylation (Kac), Khib, crotonylation (Kcr) and succinylation (Ksu) of fibro-adipogenic progenitors (FAPs), myogenic precursors (Myo) and mesenchymal stem cells (MSCs) with varied differentiation potential, and found that only Khib modification in FAPs was significantly higher than that in MSCs. Consistently, in parallel with its regulatory enzymes lysine acetyltransferase 5 (KAT5) and histone deacetylase 2 (HDAC2) protein levels, the Khib levels increased quadratically (P < 0.01) during adipogenic differentiation of FAPs. KAT5 knockdown in FAPs inhibited adipogenic differentiation, while HDAC2 knockdown enhanced adipogenic differentiation. We also demonstrated that Khib modification favored to adipogenic differentiation and fat accumulation by comparing Khib levels in FAPs and backfat tissues both derived from obese-type pigs (Laiwu pigs) and lean-type pigs (Duroc pigs), respectively. Accordingly, the expression patterns of KAT5 and HDAC2 matched well to the degree of backfat accumulation in obese- and lean-type pigs.ConclusionsFrom the perspective of protein translational modification, we are the first to reveal the role of Khib in adipogenesis and fat deposition in pigs, and provided new clues for the improvement of fat accumulation and distribution as expected via genetic selection and nutritional strategy in obese-type pigs.

Differential expression of ADRB1 causes different responses to norepinephrine in adipocytes of Duroc-Landrace-Yorkshire pigs and min pigs

Research has shown that pigs from different regions exhibit varying responses to cold stimuli. Typically, cold stimuli induce browning of white adipose tissue mediated by adrenaline, promoting non-shivering thermogenesis. However, the molecular mechanisms underlying differential response of pig breeds to norepinephrine are unclear.The aim of this study was to investigate the differences and molecular mechanisms of the effects of norepinephrine (NE) treatment on adipocytes of Min pigs (a cold-resistant pig breed) and Duroc-Landrace-Yorkshire (DLY) pigs. Real time-qPCR, western blot, and immunofluorescence were performed following NE treatment on cell cultures of adipocytes originating from Min pigs (n = 3) and DLY pigs (n = 3) to assess the expressions of adipogenesis markers, beige fat markers, and mitochondrial biogenesis markers. The results showed that NE did not affect browning of adipocytes in DLY pigs, whereas promoted browning of adipocytes in Min pigs. Further, the expression of ADRB1 (Adrenoceptor Beta 1, ADRB1) was higher in subcutaneous adipose tissue and adipocytes of Min pigs than those of DLY pigs. Overexpression of ADRB1 in DLY pig adipocytes enhanced sensitivity to NE, exhibiting decreased adipogenesis markers, upregulated beige fat markers, and increased mitochondrial biogenesis. Conversely, adipocytes treated with ADRB1 antagonist in Min pigs resulted in decreased cellular sensitivity to NE. Further studies revealed differential CpG island methylation in ADRB1 promoter region, with lower methylation levels in Min pigs compared to DLY pigs. In conclusion, differential methylation of the ADRB1 promoter region leads to different ADRB1 expression, resulting in varying responsiveness to NE in adipocytes of two pig breeds. Our results provide new insights for further analysis of the differential cold responsiveness in pig breeds from different regions.

Epidemiological investigation of Senecavirus A infection in pig herds in China from 2018 to 2021.

Senecaviurs A (SVA) infection, an emerging infectious disease in pig populations, is characterized by vesicular lesions predominantly affecting the mouth, snout, and hooves of infected pigs, similar to the symptoms of Foot and Mouth Disease Virus (FMDV). This disease first spread into China in 2015, causing great panic in the pig breeding industry. To determine the prevalence of SVA in pig herds in China from 2018 to 2021, a total of 4,901 pig tissue samples were collected from 18 provinces, autonomous regions and municipalities (P.A.M.s) for epidemiological investigation, virus isolation and genetic analysis. In 2021, the individual positive rates (IPRs) from the perspective of spatial distribution in East China, South China, Central China, North China, Southwest China, Northwest China, and Northeast China were 0, 0, 1.69, 0.94, 11.70, 3.31 and 2.21%, respectively. The herd positive rates (HPRs) were 0, 0, 9.52, 9.09, 50.00, 7.69 and 23.08%. From the perspective of temporal distribution, the IPR showed an overall downwards trend from 2018 to 2021, with only a slight increase in 2020. Moreover, the HPR decreased from 36.63 to 10.07%. From the perspective of population distribution in 2021, the IPR (2.62%) and HPR (12.00%) in apparently healthy pig herds (slaughterhouses) were greater than those in non-healthy pig herds (2.10 and 5.13%, respectively), consistent with the results in 2019. To characterize the prevalent strains, 10 SVA strains isolated from positive samples in 2019 were clustered in Clades I and VII; SVA-FJ039-2019, SVA-HuN032-2019, SVA-GX011-2019, SVA-FJ036-2019, SVA-GXF011-2019 and SVA-GXF053-2019 were clustered in Clade I; and SVA-FJ018-2019, SVA-SD069-2019, SVA-SD072-2019, and SVA-SD074-2019 were clustered in Clade VII. In conclusion, until 2021, the prevalence of SVA in pig herds in China was still relatively high, the contaminated area was still large, and there were a number of hidden infections. In the future, the epidemic status of SVA in pig herds in China must be closely monitored and the prevention and control measures must be adjusted in a timely manner.

Nested PCR optimization for detecting hepatitis E virus

The occurrence of Hepatitis E virus (HEV) infections is on the rise in developing countries, frequently linked to the consumption of undercooked meats and exposure to animal feces. Typically, HEV diagnosis relies on the immunodetection of anti-HEV antibodies and reverse transcriptase polymerase chain reaction. In this study, nested polymerase chain reaction (Nested PCR) for detecting HEV in domesticated pig and farmed wild boar samples was optimized to provide an alternative method for reliable and precise detection of HEV, particularly in animal samples. All samples were collected in Ho Chi Minh City, Vietnam. Total RNA was extracted from liver tissues of domestic pigs (n=48), rectal swabs samples (n=60), and feces samples (n=25) of farmed wild boars. Complementary DNA (cDNA) was synthesized using random hexamers. Nested PCR was performed under four different conditions: protocol 1–4, with variations in reaction components and concentrations. Two PCR programs, designated as A and B, were examined, featuring distinct cycling times and annealing temperatures for the outer and inner amplification stages. The resulting amplification products (306 bp) were visualized through gel electrophoresis. Protocol 1 and 2, when employed in conjunction with both program A and B, failed to identify any positive samples, including the positive control (0%). In contrast, protocol 3, in combination with program A, and protocol 4, paired with program B, successfully identified 28 positive results out of 133 tested samples (21%). It is noteworthy that protocol 4, when used with program B, yielded clearer and more specific bands. The study successfully optimized a nested-PCR protocol for detecting HEV in animal samples, comprising tissues, rectal swabs, and fecal samples.

Effect of the complex supplement «Gepasorbex» on the amino acid composition of muscle tissue of pigs

The modern pig breeding is focused on the constant improvement of pig housing conditions, ensuring high quality and safety of products, using effective feeding and breeding technologies, as well as the balanced consideration of ecological and economic aspects. New methods of rearing andfattening pigs affect not only the quantity, but also the quality of the slaughter raw materials obtained. Objective. To study the effect of the complex supplement "Hepasorbex" on the amino acid composition of the studied groups of pigs in the conditions of a commercial enterprise in accordance with the industrial technology. Methods. On the basis of the above, for the assessment of slaughter qualities, young animals were selected for slaughter from groups offattening young animals when piglets reached a live weight of 100 and 120 kg in the amount of 10 heads of each weight condition under the conditions of the limited liability company "Tavriiski Swyni". Next, it has been studied the effect of the complex supplement "Hepasorbex" on the amino acid composition of the muscle tissue ofpigs of experimental groups with a pre-slaughter live weight of 100 and 120 kg in the amount of 5 samples of each group. Results. It was revealed the deep changes in the animal body: in particular, in the amino acid composition of muscle tissue (replaceable, essential amino acids and their ratio) by detailed researches. Thus, 100 g of muscle tissue protein obtainedfrom the carcasses of pigs of the III research group: for pre-slaughter live weight of 100 kg, contains 9.11 g of essential amino acids and 11.43 g of replaceable amino acids, the amino acid index is 79.7 %; for pre-slaughter live weight of 120 kg - essential amino acids - 10.41 g, replaceable - 13.83 g, amino acid index - 75.27 %. The animals of the III research group are characterized by the maximum value of the protein-quality index at slaughter both at 100 kg and at 120 kg - 12.22 and 7.39, respectively, that is, "Hepasorbex" contributed to the biosynthesis of meat of better quality. Conclusions. According to the general zootechnical requirements, the protein-quality index (oxyproline, tryptophan and their ratio) of the III research group corresponds to high-quality pork (7.39 - 12.22), while the II research group (7.02 - 11.58) and the I control group (6.7 - 12.21) meet the requirements ofpork of normal quality. Keywords: technology, pigs, feed additive, amino acid rate, replaceable amino acids, essential amino acids, protein-quality index.

Tissue expression and promoter activity analysis of the porcine TNFSF11 gene

Tumour necrosis factor (TNF) superfamily member 11 (TNFSF11), also known as RANKL, plays a crucial role in regulating several physiological and pathological activities. Additionally, it is a vital factor in bone physiology, and the sex hormone progesterone regulates the expansion of stem cells and the proliferation of mammary epithelial cells. It is essential for animal growth and reproductive physiological processes. This study aimed to evaluate the tissue-specific expression characteristics and promoter activity of the TNFSF11 gene in pigs. As a result, the study examined the presence of TNFSF11 expression in the tissues of Xiangsu pigs at 0.6 and 12 months of age. Moreover, the core promoter region of TNFSF11 was also identified by utilizing a combination of bioinformatic prediction and dual-luciferase activity tests. Finally, the effect of transcription factors on the transcriptional activity of the core promoter region was determined using site-directed mutagenesis. TNFSF11 was uniformly expressed in all tissues; however, its expression in muscles was comparatively low. The core promoter region of TNFSF11 was located in the −555 to −1 region. The prediction of the transcription start site of TNFSF11 gene-2000 ∼ + 500bp showed that there was a CpG site in 17 ∼ + 487bp. Analysis of mutations in the transcription factor binding sites revealed that mutations in the Stat5b, Myog, Trl, and EN1 binding sites had significant effects on the transcriptional activity of the TNFSF11 gene, particularly following the EN1 binding site mutation (P < 0.001). This study provides insights into both the tissue-specific expression patterns of TNFSF11 in the tissues of Xiangsu pigs and the potential regulatory effects of transcription factors on its promoter activity. These results may be helpful for future research aimed at clarifying the expression and role of the porcine TNFSF11 gene.