Originating but free from chromosomal DNA, extrachromosomal circular DNAs (eccDNAs) are organized in circular form and have long been found in unicellular and multicellular eukaryotes. Their biogenesis and function are poorly understood as they are characterized by sequence homology with linear DNA, for which few detection methods are available. Recent advances in high-throughput sequencing technologies have revealed that eccDNAs play crucial roles in tumor formation, evolution, and drug resistance as well as aging, genomic diversity, and other biological processes, bringing it back to the research hotspot. Several mechanisms of eccDNA formation have been proposed, including the breakage-fusion-bridge (BFB) and translocation-deletion-amplification models. Gynecologic tumors and disorders of embryonic and fetal development are major threats to human reproductive health. The roles of eccDNAs in these pathological processes have been partially elucidated since the first discovery of eccDNA in pig sperm and the double minutes in ovarian cancer ascites. The present review summarized the research history, biogenesis, and currently available detection and analytical methods for eccDNAs and clarified their functions in gynecologic tumors and reproduction. We also proposed the application of eccDNAs as drug targets and liquid biopsy markers for prenatal diagnosis and the early detection, prognosis, and treatment of gynecologic tumors. This review lays theoretical foundations for future investigations into the complex regulatory networks of eccDNAs in vital physiological and pathological processes.
Male ; Female ; Animals ; Humans ; Swine ; DNA, Circular/genetics* ; Genital Neoplasms, Female ; Semen ; DNA ; Reproduction

The growth and development of skeletal muscle is an important factor affecting pork production and quality, which is elaborately regulated by many genetic and nutritional factors. MicroRNA (miRNA) is a non-coding RNA with a length of about 22 nt, which binds to the 3'UTR sequence of the mRNA of the target genes, and consequently regulates its post-transcriptional expression level. In recent years, a large number of studies have shown that miRNAs are involved in various life processes such as growth and development, reproduction, and diseases. The role of miRNAs in the regulation of porcine skeletal muscle development was reviewed, with the hope to provide a reference for the genetic improvement of pigs.
Swine ; Animals ; MicroRNAs/metabolism* ; Muscle, Skeletal/metabolism* ; Muscle Development/genetics*

Porcine epidemic diarrhea (PED) is a highly contagious disease that causes high mortality in suckling piglets. Although several licensed inactivated and live attenuated vaccines were widely used, the infection rate remains high due to unsatisfactory protective efficacy. In this study, mRNA vaccine candidates against PED were prepared, and their immunogenicity was evaluated in mice and pregnant sows. The mRNA PED vaccine based on heterodimer of viral receptor binding region (RBD) showed good immunogenicity. It elicited robust humoral and cellular immune responses in mice, and the neutralizing antibody titer reached 1:300 after a single vaccination. Furthermore, it induced neutralizing antibody level similar to that of the inactivated vaccine in pregnant sows. This study developed a new design of PED vaccine based on the mRNA-RBD strategy and demonstrated the potential for clinical application.
Pregnancy ; Swine ; Animals ; Female ; Mice ; Antibodies, Viral ; Swine Diseases/epidemiology* ; Viral Vaccines/genetics* ; Antibodies, Neutralizing ; Vaccines, Attenuated ; Diarrhea/veterinary*

The antigen gene expression level of a DNA vaccine is the key factor influencing the efficacy of the DNA vaccine. Accordingly, one of the ways to improve the antigen gene expression level of a DNA vaccine is to utilize a plasmid vector that is replicable in eukaryotic cells. A replicative DNA vaccine vector pCMVori was constructed based on the non-replicative pcDNA3.1 and the replicon of porcine circovirus 2 (PCV2) in this study. An EGFP gene was cloned into pCMVori and the control plasmid pcDNA3.1. The two recombinant vectors were transfected into PK-15 cell, and the plasmid DNA and RNA were extracted from the transfected cells. Real-time PCR was used to determine the plasmid replication efficiency of the two plasmids using plasmid before and after Bcl Ⅰ digestion as templates, and the transcription level of the Rep gene in PCV2 replicon was detected by RT-PCR. The average fluorescence intensity of cells transfected with the two plasmids was analyzed with software Image J, and the transcription level of EGFP was determined by means of real-time RT-PCR. The results showed that the replication efficiency of pCMVori in PK-15 cells incubated for 48 h was 136%, and the transcriptions of Rep and Rep' were verified by RT-PCR. The average fluorescence intensity of the cells transfected with pCMVori-EGFP was 39.14% higher than that of pcDNA3.1-EGFP, and the transcription level of EGFP in the former was also 40% higher than that in the latter. In conclusion, the DNA vaccine vector pCMVori constructed in this study can independently replicate in eukaryotic cells. As a result, the expression level of cloned target gene was elevated, providing a basis for developing the pCMVori-based DNA vaccine.
Animals ; Swine ; Circovirus/genetics* ; Vaccines, DNA/genetics* ; Replicon/genetics* ; Genetic Vectors/genetics* ; Plasmids/genetics*

OBJECTIVE: To investigate the repair method of type Ⅱc injury in the lateral meniscus popliteal tendon area based on the porcine knee joint. METHODS: Eighteen commercially available fresh porcine knee joints were randomly divided into 3 groups ( n=6). After preparing a type Ⅱc injury in the lateral meniscus popliteal tendon area, and the anterior (group A), posterior (group B), or anterior and posterior (group C) of the popliteal hiatus (PH) was sutured by vertical mattress. The tension meter was used to apply gradient tensions of 2, 4, 6, 8, and 10 N along the tibial plateau horizontally, respectively, to pull the midpoint of the lateral meniscus popliteal tendon area. The displacement values before modeling, after modeling, and after suture were recorded. The reduction value of lateral meniscus displacement and reduction rate after suture were calculated and compared between groups. RESULTS: There was no significant difference between groups ( P>0.05) in the displacement values before modeling, after modeling, and after suture under different tensions. There was no significant difference between groups A and C ( P>0.05) in the reduction value of lateral meniscus displacement and reduction rate after suture under different tensions. The reduction value of lateral meniscus displacement and reduction rate after suture in group B were lower than those in groups A and C. The reduction value of lateral meniscus displacement under tension of 2 N and the reduction rates under tensions of 2, 4, and 6 N between groups A and B showed significant differences ( P<0.05). The reduction value of lateral meniscus displacement and the reduction rate under tensions of 2, 4, and 6 N between groups B and C showed significant differences ( P<0.05). CONCLUSION Suturing the anterior area of PH is the key to repairing type Ⅱc injury of lateral meniscus popliteal tendon area.
Animals ; Humans ; Knee ; Knee Joint ; Menisci, Tibial/surgery* ; Swine ; Tendons ; Tibia

PURPOSE: Child head injury under impact scenarios (e.g. falls, vehicle crashes, etc.) is an important topic in the field of injury biomechanics. The head of piglet was commonly used as the surrogate to investigate the biomechanical response and mechanisms of pediatric head injuries because of the similar cellular structures and material properties. However, up to date, piglet head models with accurate geometry and material properties, which have been validated by impact experiments, are seldom. We aim to develop such a model for future research. METHODS: In this study, first, the detailed anatomical structures of the piglet head, including the skull, suture, brain, pia mater, dura mater, cerebrospinal fluid, scalp and soft tissue, were constructed based on CT scans. Then, a structured butterfly method was adopted to mesh the complex geometries of the piglet head to generate high-quality elements and each component was assigned corresponding constitutive material models. Finally, the guided drop tower tests were conducted and the force-time histories were ectracted to validate the piglet head finite element model. RESULTS: Simulations were conducted on the developed finite element model under impact conditions and the simulation results were compared with the experimental data from the guided drop tower tests and the published literature. The average peak force and duration of the guide drop tower test were similar to that of the simulation, with an error below 10%. The inaccuracy was below 20%. The average peak force and duration reported in the literature were comparable to those of the simulation, with the exception of the duration for an impact energy of 11 J. The results showed that the model was capable to capture the response of the pig head. CONCLUSION This study can provide an effective tool for investigating child head injury mechanisms and protection strategies under impact loading conditions.
Animals ; Swine ; Finite Element Analysis ; Skull/injuries* ; Craniocerebral Trauma/diagnostic imaging* ; Brain ; Biomechanical Phenomena ; Scalp

PURPOSE: SAM junctional tourniquet (SJT) has been applied to control junctional hemorrhage. However, there is limited information about its safety and efficacy when applied in the axilla. This study aims to investigate the effect of SJT on respiration when used in the axilla in a swine model. METHODS: Eighteen male Yorkshire swines, aged 6-month-old and weighing 55 - 72 kg, were randomized into 3 groups, with 6 in each. An axillary hemorrhage model was established by cutting a 2 mm transverse incision in the axillary artery. Hemorrhagic shock was induced by exsanguinating through the left carotid artery to achieve a controlled volume reduction of 30% of total blood volume. Vascular blocking bands were used to temporarily control axillary hemorrhage before SJT was applied. In Group I, the swine spontaneously breathed, while SJT was applied for 2 h with a pressure of 210 mmHg. In Group II, the swine were mechanically ventilated, and SJT was applied for the same duration and pressure as Group I. In Group III, the swine spontaneously breathed, but the axillary hemorrhage was controlled using vascular blocking bands without SJT compression. The amount of free blood loss was calculated in the axillary wound during the 2 h of hemostasis by SJT application or vascular blocking bands. After then, a temporary vascular shunt was performed in the 3 groups to achieve resuscitation. Pathophysiologic state of each swine was monitored for 1 h with an infusion of 400 mL of autologous whole blood and 500 mL of lactated ringer solution. T_b and T₀ represent the time points before and immediate after the 30% volume-controlled hemorrhagic shock, respectively. T₃₀, T₆₀, T₉₀ and T₁₂₀, denote 30, 60, 90, and 120 min after T₀ (hemostasis period), while T₁₅₀, and T₁₈₀ denote 150 and 180 min after T₀ (resuscitation period). The mean arterial pressure and heart rate were monitored through the right carotid artery catheter. Blood samples were collected at each time point for the analysis of blood gas, complete cell count, serum chemistry, standard coagulation tests, etc., and thromboelastography was conducted subsequently. Movement of the left hemidiaphragm was measured by ultrasonography at T_b and T₀ to assess respiration. Data were presented as mean ± standard deviation and analyzed using repeated measures of two-way analysis of variance with pairwise comparisons adjusted using the Bonferroni method. All statistical analyses were processed using GraphPad Prism software. RESULTS: Compared to T_b, a statistically significant increase in the left hemidiaphragm movement at T₀ was observed in Groups I and II (both p < 0.001). In Group III, the left hemidiaphragm movement remained unchanged (p = 0.660). Compared to Group I, mechanical ventilation in Group II significantly alleviated the effect of SJT application on the left hemidiaphragm movement (p < 0.001). Blood pressure and heart rate rapidly increased at T₀ in all three groups. Respiratory arrest suddenly occurred in Group I after T₁₂₀, which required immediate manual respiratory assistance. PaO₂ in Group I decreased significantly at T₁₂₀, accompanied by an increase in PaCO₂ (both p < 0.001 vs. Groups II and III). Other biochemical metabolic changes were similar among groups. However, in all 3 groups, lactate and potassium increased immediately after 1 min of resuscitation concurrent with a drop in pH. The swine in Group I exhibited the most severe hyperkalemia and metabolic acidosis. The coagulation function test did not show statistically significant differences among three groups at any time point. However, D-dimer levels showed a more than 16-fold increase from T₁₂₀ to T₁₈₀ in all groups. CONCLUSION In the swine model, SJT is effective in controlling axillary hemorrhage during both spontaneous breathing and mechanical ventilation. Mechanical ventilation is found to alleviate the restrictive effect of SJT on thoracic movement without affecting hemostatic efficiency. Therefore, mechanical ventilation could be necessary before SJT removal.
Male ; Animals ; Swine ; Shock, Hemorrhagic/therapy* ; Tourniquets ; Axilla ; Hemorrhage/therapy* ; Vascular Diseases ; Respiration

Early weaned piglets suffer from oxidative stress and enteral infection, which usually results in gut microbial dysbiosis, serve diarrhea, and even death. Rice bran oil (RBO), a polyphenol-enriched by-product of rice processing, has been shown to have antioxidant and anti-inflammatory properties both in vivo and in vitro. Here, we ascertained the proper RBO supplementation level, and subsequently determined its effects on lipopolysaccharide (LPS)-induced intestinal dysfunction in weaned piglets. A total of 168 piglets were randomly allocated into four groups of seven replicates (42 piglets each group, (21±1) d of age, body weight (7.60±0.04) kg, and half males and half females) and were given basal diet (Ctrl) or basal diet supplemented with 0.01% (mass fraction) RBO (RBO1), 0.02% RBO (RBO2), or 0.03% RBO (RBO3) for 21 d. Then, seven piglets from the Ctrl and the RBO were treated with LPS (100 μg/kg body weight (BW)) as LPS group and RBO+LPS group, respectively. Meanwhile, seven piglets from the Ctrl were treated with the saline vehicle (Ctrl group). Four hours later, all treated piglets were sacrificed for taking samples of plasma, jejunum tissues, and feces. The results showed that 0.02% was the optimal dose of dietary RBO supplementation based on diarrhea, average daily gain, and average daily feed intake indices in early weaning piglets. Furthermore, RBO protected piglets against LPS-induced jejunal epithelium damage, which was indicated by the increases in villus height, villus height/crypt depth ratio, and Claudin-1 levels, as well as a decreased level of jejunal epithelium apoptosis. RBO also improved the antioxidant ability of LPS-challenged piglets, which was indicated by the elevated concentrations of catalase and superoxide dismutase, and increased total antioxidant capacity, as well as the decreased concentrations of diamine oxidase and malondialdehyde in plasma. Meanwhile, RBO improved the immune function of LPS-challenged weaned piglets, which was indicated by elevated immunoglobulin A (IgA), IgM, β‍‍-defensin-1, and lysozyme levels in the plasma. In addition, RBO supplementation improved the LPS challenge-induced dysbiosis of gut microbiota. Particularly, the indices of antioxidant capacity, intestinal damage, and immunity were significantly associated with the RBO-regulated gut microbiota. These findings suggested that 0.02% RBO is a suitable dose to protect against LPS-induced intestinal damage, oxidative stress, and jejunal microbiota dysbiosis in early weaned piglets.
Male ; Female ; Swine ; Animals ; Lipopolysaccharides/toxicity* ; Antioxidants/pharmacology* ; Rice Bran Oil ; Dysbiosis ; Dietary Supplements ; Diarrhea/veterinary* ; Weaning ; Body Weight

Engineered probiotics can serve as therapeutics based on their ability of produce recombinant immune-stimulating properties. In this study, we built the recombinant Bacillus subtilis WB800 expressing antimicrobial peptide KR32 (WB800-KR32) using genetic engineering methods and investigated its protective effects of nuclear factor-E2-related factor 2 (Nrf2)‍-Kelch-like ECH-associated protein 1 (Keap1) pathway activation in intestinal oxidative disturbance induced by enterotoxigenic Escherichia coli (ETEC) K88 in weaned piglets. Twenty-eight weaned piglets were randomly distributed into four treatment groups with seven replicates fed with a basal diet. The feed of the control group (CON) was infused with normal sterilized saline; meanwhile, the ETEC, ETEC+WB800, and ETEC+WB800-KR32 groups were orally administered normal sterilized saline, 5×10¹⁰ CFU (CFU: colony forming units) WB800, and 5×10¹⁰ CFU WB800-KR32, respectively, on Days 1‍‒‍14 and all infused with ETEC K88 1×10¹⁰ CFU on Days 15‍‒‍17. The results showed that pretreatment with WB800-KR32 attenuated ETEC-induced intestinal disturbance, improved the mucosal activity of antioxidant enzyme (catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx)) and decreased the content of malondialdehyde (MDA). More importantly, WB800-KR32 downregulated genes involved in antioxidant defense (GPx and SOD1). Interestingly, WB800-KR32 upregulated the protein expression of Nrf2 and downregulated the protein expression of Keap1 in the ileum. WB800-KR32 markedly changed the richness estimators (Ace and Chao) of gut microbiota and increased the abundance of Eubacterium_rectale_ATCC_33656 in the feces. The results suggested that WB800-KR32 may alleviate ETEC-induced intestinal oxidative injury through the Nrf2-Keap1 pathway, providing a new perspective for WB800-KR32 as potential therapeutics to regulate intestinal oxidative disturbance in ETEC K88 infection.
Animals ; Swine ; Enterotoxigenic Escherichia coli ; Kelch-Like ECH-Associated Protein 1 ; Bacillus subtilis ; NF-E2-Related Factor 2 ; Antioxidants ; Oxidative Stress

Foreign body reactions induced by macrophages often cause delay or failure of wound healing in the application of tissue engineering scaffolds. This study explores the application of nanosilver (NAg) to reduce foreign body reactions during scaffold transplantation. An NAg hybrid collagen-chitosan scaffold (NAg-CCS) was prepared using the freeze-drying method. The NAg-CCS was implanted on the back of rats to evaluate the effects on foreign body reactions. Skin tissue samples were collected for histological and immunological evaluation at variable intervals. Miniature pigs were used to assess the effects of NAg on skin wound healing. The wounds were photographed, and tissue samples were collected for molecular biological analysis at different time points post-transplantation. NAg-CCS has a porous structure and the results showed that it could release NAg constantly for two weeks. The NAg-CCS group rarely developed a foreign body reaction, while the blank-CCS group showed granulomas or necrosis in the subcutaneous grafting experiment. Both matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of metalloproteinase-1 (TIMP-1) were reduced significantly in the NAg-CCS group. The NAg-CCS group had higher interleukin (IL)-10 and lower IL-6 than the blank CCS group. In the wound healing study, M1 macrophage activation and inflammatory-related proteins (inducible nitric oxide synthase (iNOS), IL-6, and interferon-‍γ (IFN-‍γ)) were inhibited by NAg. In contrast, M2 macrophage activation and proinflammatory proteins (arginase-1, major histocompatibility complex-II (MHC-II), and found in inflammatory zone-1 (FIZZ-1)) were promoted, and this was responsible for suppressing the foreign body responses and accelerating wound healing. In conclusion, dermal scaffolds containing NAg suppressed the foreign body reaction by regulating macrophages and the expression of inflammatory cytokines, thereby promoting wound healing.
Animals ; Rats ; Swine ; Interleukin-6 ; Macrophage Activation ; Tissue Inhibitor of Metalloproteinase-1 ; Wound Healing ; Foreign-Body Reaction ; Foreign Bodies ; Chitosan