Porcine Ileum Research Articles

Microplastic and the Enteric Nervous System: Effect of PET Microparticles on Selected Neurotransmitters and Cytokines in the Porcine Ileum.

Microplastic is an environmental hazard to which both animals and humans are exposed. Current reports show that it can cause inflammation, including in the gastrointestinal tract. To examine the impact on the ileum, 15 eight-week-old gilts (five individuals/group) were exposed to PET microplastics (7.6 µm-416.9 µm) at a dose of 0.1 g/day or 1 g/day for 28 days. The collected ileum fragments were investigated for the cytokine concentrations (IL-1β, IL-6, IL-8, IL-10, and TNF-α; ELISA test), neuron populations (cocaine and amphetamine-regulated transcript, galanin, neuronal nitric oxide synthase, substance P, vesicular acetylcholine transporter, and vasoactive intestinal peptide; immunofluorescence staining), and morphometric parameters (histological analysis). Under the influence of MP-PET, there was a reduction in the populations of CART- and GAL-positive neurons in the submucosal plexuses and of nNOS-, VAChT-, and VIP-positive neurons in all the plexuses. In contrast, there was an increase in GAL-positive neurons in the myenteric plexus and SP-positive neurons in all the plexuses. The concentrations of IL-1β, IL-6, IL-8, IL-10, and TNF-α did not undergo statistically significant changes under the influence of the low or high dose of MP-PET. The changes in the histological structure exclusively concerned the thinning of the mucosa and the muscularis externa. The results support the thesis that MP-PET is not neutral to the ileal cells.

Conserved B cell signaling, activation, and differentiation in porcine jejunal and ileal Peyer’s patches despite distinct immune landscapes

Peyer’s patches (PPs) are B cell-rich sites of intestinal immune induction, yet PP-associated B cell signaling, activation, and differentiation are poorly defined. Single-cell and spatial transcriptomics were completed to study B cells from porcine jejunum and ileum containing PPs. Intestinal locations had distinct immune landscapes, including more follicular B cells in ileum and increased MHC-II-encoding gene expression in jejunal B cells. Despite distinct landscapes, conserved B cell dynamics were detected across intestinal locations, including B cell signaling to CD4+ macrophages that are putative phagocytic, cytotoxic, effector cells and deduced routes of B cell activation/differentiation, including resting B cells migrating into follicles to replicate/divide or differentiate into antibody-secreting cells residing in intestinal crypts. A six-biomarker panel recapitulated transcriptomics findings of B cell phenotypes, frequencies, and spatial locations via ex vivo and in situ staining. Findings convey conserved B cell dynamics across intestinal locations containing PPs, despite location-specific immune environments. Results establish a benchmark of B cell dynamics for understanding intestinal immune induction important to promoting gut/overall health.

Swine acute diarrhea syndrome coronavirus Nsp1 suppresses IFN-λ1 production by degrading IRF1 via ubiquitin–proteasome pathway

Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a novel porcine enteric coronavirus that causes acute watery diarrhea, vomiting, and dehydration in newborn piglets. The type III interferon (IFN-λ) response serves as the primary defense against viruses that replicate in intestinal epithelial cells. However, there is currently no information available on how SADS-CoV modulates the production of IFN-λ. In this study, we utilized IPI-FX cells (a cell line of porcine ileum epithelium) as an in vitro model to investigate the potential immune evasion strategies employed by SADS-CoV against the IFN-λ response. Our results showed that SADS-CoV infection suppressed the production of IFN-λ1 induced by poly(I:C). Through screening SADS-CoV-encoded proteins, nsp1, nsp5, nsp10, nsp12, nsp16, E, S1, and S2 were identified as antagonists of IFN-λ1 production. Specifically, SADS-CoV nsp1 impeded the activation of the IFN-λ1 promoter mediated by MAVS, TBK1, IKKε, and IRF1. Both SADS-CoV and nsp1 obstructed poly(I:C)-induced nuclear translocation of IRF1. Moreover, SADS-CoV nsp1 degraded IRF1 via the ubiquitin-mediated proteasome pathway without interacting with it. Overall, our study provides the first evidence that SADS-CoV inhibits the type III IFN response, shedding light on the molecular mechanisms employed by SADS-CoV to evade the host immune response.

Modeling of culture conditions by culture system, glucose and propionic acid and their impact on metabolic profile in IPEC-J2.

The microbiological environment and their corresponding secreted metabolite spectrum are an essential modulator of the enterocyte function, effecting the whole organism. Intestinal porcine jejunal epithelial cell line (IPEC-J2) is an established in vitro model for differentiation of enterocytes in different cell culture models. An improved oxygen supply seems to be the main reason for differentiation in an air-liquid-interface culture, but this has not yet been conclusively clarified. In this context, the nutrition of the cell and its influence on the metabolism is also of crucial importance. The interest in short-chain fatty acids (SCFAs) has grown steadily in recent years due to their clinical relevance in certain diseases such as multiple sclerosis and other inflammatory diseases, but not much is known of FFAR2 and FFAR3 (free fatty acid receptor 2 and 3) in pigs. We want to address the questions: 1. about the distribution of FFAR2 and FFAR3 in vivo and in vitro in sus scrofa 2. whether there is an influence of propionic acid, glucose content and cultivation on metabolism of enterocytes? The morphological analysis of FFAR2 and FFAR3 in vivo was investigated through immunostaining of frozen sections of the porcine gut segments jejunum, ileum and colon. Both receptors are expressed along the gut and were found in the smooth muscle cells of the tunica muscularis and lamina muscularis mucosae. Furthermore, a high expression of FFAR2 and a low expression of FFAR3 in the enteric nerve system was also observed in jejunum, ileum and colon of sus scrofa. In addition, FFAR2 and FFAR3 within the vessels was investigated. FFAR3 showed a strong expression on endothelial cells of veins and lymphatic vessels but was not detectable on arteries. Furthermore, we demonstrate for the first time, FFAR2 and FFAR3 in IPEC-J2 cells on RNA- and protein level, as well as with confocal microscopy. In addition, ENO1 and NDUFA4 were investigated on RNA-level in IPEC-J2 cells as 2 important genes, which play an essential role in metabolism. Here, NDUFA4 is detected in the model animal sus scrofa as well as in the porcine cell line IPEC-J2. A potential impact of propionic acid and/or glucose and/or cultivation method on the metabolism of the cells was tested with the Seahorse analyzer. Here, a significant higher ECAR was observed in the SMC than in the OCR. In summary, we were able to show that the cultivation system appears to have a greater influence than the medium composition or nutrition of the cells. However, this can be modulated by incubation time or combination of different SCFAs.

Cholesterol 25-Hydroxylase Suppresses Swine Acute Diarrhea Syndrome Coronavirus Infection by Blocking Spike Protein-Mediated Membrane Fusion.

Swine acute diarrhea syndrome coronavirus (SADS-CoV) is an emerging porcine intestinal coronavirus that can cause acute diarrhea, vomiting, rapid weight loss, and high mortality in newborn piglets. Cholesterol 25-hydroxylase (CH25H) is a molecular mediator of innate antiviral immunity and converts cholesterol to 25-hydroxycholesterol (25HC). Previous studies have reported that CH25H and 25HC have an antiviral effect against multiple viruses. However, the interplay between SADS-CoV infection and CH25H or 25HC is still uncertain. Here, we found that CH25H and its enzymatic product 25HC restrained SADS-CoV replication by blocking membrane fusion. Our results show that CH25H was upregulated by SADS-CoV infection in vitro and in vivo, and that it was an IFN-stimulated gene in porcine ileum epithelial cells. Moreover, CH25H and CH25H mutants lacking catalytic activity can inhibit SADS-CoV replication. Furthermore, 25HC significantly suppressed SADS-CoV infection by inhibiting virus entry. Notably, we confirmed that CH25H and 25HC blocked SADS-CoV spike protein-mediated membrane fusion. Our data provide a possible antiviral therapy against SADS-CoV and other conceivable emerging coronaviruses in the future.

Comparison of the effects of normothermic machine perfusion and cold storage preservation on porcine intestinal allograft regenerative potential and viability

Intestinal transplantation (IT) is the final treatment option for intestinal failure. Static cold storage (CS) is the standard preservation method used for intestinal allografts. However, CS and subsequent transplantation induce ischemia-reperfusion injury (IRI). Severe IRI impairs epithelial barrier function, including loss of intestinal stem cells (ISC), critical to epithelial regeneration. Normothermic machine perfusion (NMP) preservation of kidney and liver allografts minimizes CS-associated IRI; however, it has not been used clinically for IT. We hypothesized that intestine NMP would induce less epithelial injury and better protect the intestine’s regenerative ability when compared with CS. Full-length porcine jejunum and ileum were procured, stored at 4 °C, or perfused at 34 °C for 6 hours (T6), and transplanted. Histology was assessed following procurement (T0), T6, and 1 hour after reperfusion. Real-time quantitative reverse transcription polymerase chain reaction, immunofluorescence, and crypt culture measured ISC viability and proliferative potential. A greater number of NMP-preserved intestine recipients survived posttransplant, which correlated with significantly decreased tissue injury following 1-hour reperfusion in NMP compared with CS samples. Additionally, ISC gene expression, spheroid area, and cellular proliferation were significantly increased in NMP-T6 compared with CS-T6 intestine. NMP appears to reduce IRI and improve graft regeneration with improved ISC viability and proliferation.

Alteration in the neurochemical profile of ENS neurons in the porcine ileum after oral exposure of glyphosate

Glyphosate-based herbicides are one of the most intensively used pollutants worldwide. The main goal of present study was to determine the changes in the neurochemical phenotype of enteric nervous system (ENS) neurons in the porcine ileum after oral glyphosate administration.Study was performed on 15 immature gilts divided into 3 groups: control (C) - animals receiving empty gelatin capsules; G1 - animals receiving a low dose of glyphosate- corresponding to the theoretical maximum daily intake (TMDI) - 0.05 mg/kg of b.w./day; G2- animals receiving a higher dose of glyphosate - corresponding to the acceptable daily intake (ADI) - 0.5 mg/kg of b.w./day) in gelatin capsules orally for 28 days. After this time, the animals were euthanized and ileum samples were collected. Frozen sections (12μm thick) were then subjected to the double immunofluorescent staining technique using primary antibody directed towards Hu C/D (used as a pan-neuronal marker) and pituitary adenylate cyclase-activating polypeptide (PACAP), neuronal nitric oxide synthase (nNOS), vesicular acetylcholine transporter (VAChT), cocaine and amphetamine-regulated transcript (CART) and appropriate secondary antibodies.Supplementation of lower dose of glyphosate increased the population of PACAP- like immunoreactive (LI) neurons in the outer submucous plexus (OSP) (from 6.34±1.24 to 14.12±1.68%), nNOS-LI neurons in the myenteric plexus (MP) (from 29.96±2.40 to 37.57±1.17%) and the inner submucous plexus (ISP) (from 12.64±1.10 to 19.76±1.34%), CART-LI neurons in the MP (from 20.44±0.81 to 29.77±1.16%) and the ISP (from 6.07±1.00 to 10.44±0.94%) and decreased the number of VAChT-LI neurons in both submucous plexuses (from 30.69±1.22 to 24.84±1.20% in the OSP and from 53.03±1.67 to 46.68±1.18% in the ISP). The higher dose of glyphosate evoke marked changes in the immunoreactivity of all neuroactive substance studied and type of enteric plexus analyzed. An increased number of PACAP-LI (from 13.75±to 40.20 to 40.20±1.62% in the MP, to 21.12 ±1.40% in the OSP and from 11.22±1.26 to 34.94±1.61% in the ISP), nNOS-LI (to 49.92±1.73% in the MP, from 19.53±1.29 to 27.69±1.29% in the OSP and to 27.92±1.26% in the ISP), CART-LI (to 35.03±1.67% in the MP, from 10.60±1.38 to 22.23±1.27% in the OSP and to14.78±1.53% in the ISP) neurons as well as a decrease in the population of VAChT-LI neurons (from 44.58±1.82 to 29.03±1.74% in the MP, to 22.24±1.96% in the OSP and to 35.87±1.42% in the ISP) were noted.Observed changes may result from neurotoxic effect of glyphosate on the ENS neurons and be part of adaptive processes of these neurons to ileum dysfunction. the National Science Centre in Poland [grant no. 2020/04/X/NZ7/00338]. Project financially supported by the Minister of Education and Science under the program entitled “Regional Initiative of Excellence” for the years 2019-2023, Project No. 010/RID/2018/19, amount of funding 12.000.000 PLN. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The effect of spring diameter on porcine ileal distraction enterogenesis.

Spring-mediated distraction enterogenesis has proven to be successful for intestinal lengthening. We aimed to evaluate the effect of spring diameter mismatch on intestinal adaptation. Juvenile mini-Yucatan pigs underwent placement of compressed nitinol springs with diameter of 10, 11, or 12mm into the ileal lumen. Pigs were euthanized on postoperative day 7. The lengths, histology, total area of blood vessels, and enteric ganglia were evaluated. All spring groups exhibited significant ileal lengthening. Across the different diameters, spring-expanded segments were similar in terms of ileal lengthening, crypt height, muscular thickness, blood vessels, and enteric ganglia area. Spring-mediated distraction enterogenesis is successful in the porcine ileum. A smaller diameter spring is as effective as a larger diameter spring in lengthening the ileum. Springs of varying diameters result in comparable structural changes in the ileum.

Generating an In Vitro Gut Model with Physiologically Relevant Biophysical Mucus Properties.

Gastrointestinal (GI) in vitro models have received lasting attention as an effective tool to model drug and nutrient absorption, study GI diseases, and design new drug delivery vehicles. A complete model of the GI epithelium should at a minimum include the two key functional components of the GI tract: mucus and the underlying epithelium. Mucus plays a key role in protecting and lubricating the GI tract, poses a barrier to orally administered therapies and pathogens, and serves as the microenvironment for the GI microbiome. These functions are reliant on the biophysical material properties of the mucus produced, including viscosity and pore size. In this study, we generated in vitro models containing Caco-2 enterocyte-like cells and HT29-MTX goblet-like cells and determined the effects of coculture and mucus layer on epithelial permeability and biophysical properties of mucus using multiple particle tracking (MPT). We found that mucus height increased as the amount of HT29-MTX goblet-like cells increased. Additionally, we found that increasing the amount of HT29-MTX goblet-like cells within culture corresponded to an increase in mucus pore size and mucus microviscosity, measured using MPT. When compared to ex vivo mucus samples from mice and pigs, we found that a 90:10 ratio of Caco-2:HT29-MTX coculture displayed similar mucus pore size to porcine jejunum and that the mucus produced from 90:10 and 80:20 ratios of cells shared mechanical properties to porcine jejunum and ileum mucus. GI coculture models are valuable tools in simulating the mucus barrier and can be utilized for a variety of applications including the study of GI diseases, food absorption, or therapeutic development.

Improving Performance and Access to Difficult-to-Reach Anatomy with a Powered Articulating Stapler.

BackgroundModern surgical staplers should provide precise placement and transection, especially in tight spaces and on thick tissue. Ideally, a stapler would move to accommodate variations in the tissue and anatomy instead of having to move the tissue around to fit the stapler. This study was undertaken to evaluate the performance characteristics of the new Echelon 3000 Stapler (ECH3). Use of the ECH3 was compared to another marketed stapler, including tests for access, seal strength, staple formation in thick tissue, and end effector stability.MethodsPelvic anatomy measurements were used to construct a virtual model of a Low Anterior Resection (LAR). Monte Carlo simulations were performed on the staplers to compare the probability of completing a transection with one or two firings. Using water infusion of stapled porcine ileum, pressure at first leak and percentage of leaks at critical pressures were measured. Rate of malformed staples was measured in thick tissue. End effector stability while firing and under moderate pressure were compared between staplers. After use, surgeons were surveyed on the functionality of the device.ResultsECH3 had a markedly higher probability of completing an LAR transection in one or two firings than the comparator stapler. Median initial leak pressure of stapled ileum was significantly higher, and rate of leaks was lower at 40 and 50 mmHg. ECH3 had fewer malformed staples for both 3.3- and 4.0-mm thick tissue. The end effector exhibited less angular movement during firing, and less deflection under a moderate load. Surgeons agreed the ECH3 provided precise placement and easy one-handed operation.ConclusionThe Echelon 3000 Stapler demonstrated improved access capability, tighter seals, fewer malformed staples, and greater end effector stability. These advantages were recognized by surgeons who evaluated the use of the device preclinically.

Generation of Porcine Ileum Through Spring-Mediated Mechanical Distraction

IntroductionShort bowel syndrome is a devastating gastrointestinal disorder in which decreased bowel length results in inadequate absorption causing nutritional deficiencies. Current treatment options are accompanied by significant morbidity. We have proposed spring-mediated distraction enterogenesis as a method to lengthen bowel with success seen in porcine jejunum. We hypothesize that spring-mediated distraction enterogenesis can be demonstrated in porcine ileum with preservation of ileal structure and function. Materials and methodsLaparotomy was performed on juvenile female mini-Yucatan pigs and a gelatin-encapsulated compressed nitinol spring was inserted into the ileal lumen and affixed proximally and distally. A control segment distal to the spring segment was marked with sutures. Postoperatively, pigs were placed on a liquid diet and euthanized on postoperative day 7. Spring and control segments were measured and processed for immunohistochemistry to evaluate for the presence of vitamin B12–intrinsic factor cotransporter, chromogranin A-producing cells, and 5-HT producing cells. ResultsAll seven pigs survived to postoperative day 7 with no adverse effects. On average, pigs gained 84.3 ± 66.4 g/d. Spring segments lengthened 1.5 ± 0.7 cm with a relative lengthening by 128% ± 56%, which was statistically significant when compared to control (P < 0.01). The average density of chromogranin-A cells in control compared to spring segments was not significantly changed (2.9 ± 1.1 cells/mm versus 3.2 ± 1.2 cells/mm, P = 0.17). Both vitamin B12–intrinsic factor cotransporter and 5-HT producing cells were present in both control and lengthened ileum. ConclusionsIntraluminal nitinol springs significantly lengthened porcine ileum. The increase in density of enteroendocrine cells may indicate enhanced endocrine function of the lengthened ileum.

Intestinal single-cell atlas reveals novel lymphocytes in pigs with similarities to human cells.

Lymphocytes can heavily influence intestinal health, but resolving intestinal lymphocyte function is challenging as the intestine contains a vastly heterogeneous mixture of cells. Pigs are an advantageous biomedical model, but deeper understanding of intestinal lymphocytes is warranted to improve model utility. Twenty-six cell types were identified in the porcine ileum by single-cell RNA sequencing and further compared with cells in human and murine ileum. Though general consensus of cell subsets across species was revealed, some porcine-specific lymphocyte subsets were identified. Differential tissue dissection and in situ analyses conferred spatial context, revealing similar locations of lymphocyte subsets in Peyer's patches and epithelium in pig-to-human comparisons. Like humans, activated and effector lymphocytes were abundant in the ileum but not periphery of pigs, suggesting tissue-specific and/or activation-associated gene expression. Gene signatures for peripheral and ileal innate lymphoid cells newly discovered in pigs were defined and highlighted similarities to human innate lymphoid cells. Overall, we reveal novel lymphocyte subsets in pigs and highlight utility of pigs for intestinal research applications.

Study of microRNA expression in Salmonella Typhimurium-infected porcine ileum reveals miR-194a-5p as an important regulator of the TLR4-mediated inflammatory response

Infection with Salmonella Typhimurium (S. Typhimurium) is a common cause of food-borne zoonosis leading to acute gastroenteritis in humans and pigs, causing economic losses to producers and farmers, and generating a food security risk. In a previous study, we demonstrated that S. Typhimurium infection produces a severe transcriptional activation of inflammatory processes in ileum. However, little is known regarding how microRNAs regulate this response during infection. Here, small RNA sequencing was used to identify 28 miRNAs differentially expressed (DE) in ileum of S. Typhimurium-infected pigs, which potentially regulate 14 target genes involved in immune system processes such as regulation of cytokine production, monocyte chemotaxis, or cellular response to interferon gamma. Using in vitro functional and gain/loss of function (mimics/CRISPR-Cas system) approaches, we show that porcine miR-194a-5p (homologous to human miR-194-5p) regulates TLR4 gene expression, an important molecule involved in pathogen virulence, recognition and activation of innate immunity in Salmonella infection.

Advanced epithelial lung and gut barrier models demonstrate passage of microplastic particles

Micro- and nanoplastics (MNP) can be found virtually everywhere around us in the biosphere and food chain, therefore humans are continuously exposed to MNP, mainly via inhalation and ingestion. Here, we have applied physiologically relevant human-based advanced in vitro models representing the lung (MucilAir™) and gut (InTESTine™ and Intestinal Explant Barrier Chip (IEBC)) to study membrane passage of various MNP and their potency to induce cytotoxic effects, barrier disturbances or pro-inflammatory cell activation. Selected MNP of various materials (polystyrene, polyethylene, nylon, car tire, and marine MNP collected from the ocean), shapes (spheres, fragments and fibers), sizes (0.05–100 μm), some of which were fluorescently labelled for tracking, were included. Without affecting cell viability, nylon fibers and (cleaned) HDPE (high density polyethylene) disrupted the MucilAir™ epithelial barrier. Luminal exposure to polystyrene particles (1 and 10 μm) and pristine HDPE fragments significantly decreased human colon tissue functionality. Furthermore, all polystyrene particles (0.05, 1 and 10 μm) affected tissue viability in porcine jejunum, ileum and colon tissue after 5 h exposure, and this was further confirmed in the IEBC after 24 h of exposure to 10 μm polystyrene particles and nylon fibers. Exposure to nylon fibers and its supernatant led to pro-inflammatory cell activation, as shown by increased IL-6 release in MucilAir™ and in human colon tissue after 96 or 24 h, respectively. Regarding transepithelial penetration of the MNP, permeability of 0.05 μm polystyrene spheres in the MucilAir™ lung cell model reached 3.6 ± 1.2% after 24 h. With 3.37 ± 0.46% after 5 h under static conditions and 5.5 ± 1.3% after 24 h under microfluidic conditions MNP permeability across intestinal tissue was highest for the largest (10 μm) polystyrene spheres. Confocal microscopy confirmed the translocation of MNP across the lung and intestinal epithelial barrier. In conclusion, we present a study revealing the passage of MNP over the epithelium of advanced in vitro models for the lung and intestine barrier. Furthermore, pro-inflammatory cell activation and disrupted barrier integrity were observed after exposure to several of the tested MNP. Future research is needed to further identify the effects of shape, size and material on these processes and subsequently the health effects of humans.

The Enigmatic N-Terminal Domain of Proglucagon; A Historical Perspective.

Enteroglucagon refers to the predominant peptide with glucagon-like immunoreactivity (GLI) that is released by the intestine into the circulation in response to nutrients. Development of a radioimmunoassay for glucagon revealed issues that were not apparent in applications of the insulin radioimmunoassay. The fact that some antisera raised against glucagon recognized glucagon-related peptides in extracts of both pancreas and gut whereas others recognized only components in the pancreas remained a mystery until it was realized that the “gut GLI cross-reactive” antisera were directed against an epitope in the N-terminal to central region of glucagon whereas the “pancreatic glucagon specific” antisera were directed against an epitope in the C-terminal region. Unlike the cross-reactive antisera, the glucagon specific antisera did not recognize components in which glucagon was extended from its C-terminus by additional amino acids. Initial attempts to purify enteroglucagon from porcine ileum led to the erroneous conclusion that enteroglucagon comprised 100 amino acids with an apparent molecular mass of 12,000 Da and was consequently given the name glicentin. Subsequent work established that the peptide constituted residues (1-69) of proglucagon (Mr 8128). In the 40 years since the structural characterization of glicentin, attempts to establish an unambiguous physiological function for enteroglucagon have not been successful. Unlike the oxyntomodulin domain at the C-terminus of enteroglucagon, the primary structure of the N-terminal domain (glicentin-related pancreatic peptide) has been poorly conserved among mammals. Consequently, most investigations of the bioactivity of porcine glicentin may have been carried out in inappropriate animal models. Enteroglucagon may simply represent an inactive peptide that ensures that the intestine does not release equimolar amounts of a hyperglycemic agent (glucagon) and a hypoglycemic agent (GLP-1) after ingestion of nutrients.

Intestinal mucosa staple line integrity and anastomotic leak pressure after healing in a porcine model.

The aim of this study was to evaluate both the intestinal mucosa staple line integrity and anastomotic leak pressure after healing in a porcine survival model. We used two suture models using two different size staples (incomplete mucosal closure model: group G [staple height 0.75mm], complete mucosal closure model: group B [staple height 1.5mm]) in the porcine ileum. Five staple lines were created in each group made in the ileum for each model, and the staple sites harvested on days 0, 2, and 7. The leak pressure at the staple site was measured at each time point. On day 0, the leak pressure for group G (79.5mmHg) was significantly lower than that for group B (182.3mmHg) (p < 0.01). On days 2 and 7, there was no significant difference between groups G and B (171mmHg and 175.5mmHg on day 2, 175.5mmHg and 175.5mmHg on day 7, p > 0.05). The histological findings in both groups showed similar healing at postoperative days 2 and 7. The integrity of the mucosal staple lines was associated with the postoperative leak pressure on day 0. However, there was no association with the leak pressure at two days or more postoperatively in a porcine model.

Cannabinoid and Cannabinoid-Related Receptors in the Myenteric Plexus of the Porcine Ileum.

Simple SummaryThe endocannabinoid system (ECS) has opened the door to novel therapeutical approaches targeting cancer, pain, anxiety, stress, and inflammatory diseases. The ECS is ubiquitously expressed in almost all members of Animalia, but its precise localization outside the central nervous system is still under investigation. In this study, the localization of the main and related cannabinoid receptors in the myenteric plexus of the porcine ileum was immunohistochemically analyzed. The myenteric plexus neurons were found to be positive for cannabinoid receptor 1 (CB1R) and the cannabinoid-related receptors transient potential vanilloid receptor 1 (TRPV1), transient potential ankyrin receptor 1 (TRPA1), and serotonin receptor 5-HT1a (5-HT1aR). In addition, the ECS receptors were also located on nerve fibers, the tunica muscularis, and the endothelium. The wide distribution of cannabinoid and cannabinoid-related receptors in the myenteric plexus provides the anatomical basis for additional investigation, suggesting the possible role of the ECS in treating pathological conditions in livestock.An important piece of evidence has shown that molecules acting on cannabinoid receptors influence gastrointestinal motility and induce beneficial effects on gastrointestinal inflammation and visceral pain. The aim of this investigation was to immunohistochemically localize the distribution of canonical cannabinoid receptor type 1 (CB1R) and type 2 (CB2R) and the cannabinoid-related receptors transient potential vanilloid receptor 1 (TRPV1), transient potential ankyrin receptor 1 (TRPA1), and serotonin receptor 5-HT1a (5-HT1aR) in the myenteric plexus (MP) of pig ileum. CB1R, TRPV1, TRPA1, and 5-HT1aR were expressed, with different intensities in the cytoplasm of MP neurons. For each receptor, the proportions of the immunoreactive neurons were evaluated using the anti-HuC/HuD antibody. These receptors were also localized on nerve fibers (CB1R, TRPA1), smooth muscle cells of tunica muscularis (CB1R, 5-HT1aR), and endothelial cells of blood vessels (TRPV1, TRPA1, 5-HT1aR). The nerve varicosities were also found to be immunoreactive for both TRPV1 and 5-HT1aR. No immunoreactivity was documented for CB2R. Cannabinoid and cannabinoid-related receptors herein investigated showed a wide distribution in the enteric neurons and nerve fibers of the pig MP. These results could provide an anatomical basis for additional research, supporting the therapeutic use of cannabinoid receptor agonists in relieving motility disorders in porcine enteropathies.

Cholesterol 25-hydroxylase suppresses porcine deltacoronavirus infection by inhibiting viral entry

Cholesterol 25-hydroxylase (CH25 H) is a key enzyme regulating cholesterol metabolism and also acts as a broad antiviral host restriction factor. Porcine deltacoronavirus (PDCoV) is an emerging swine enteropathogenic coronavirus that can cause vomiting, diarrhea, dehydration and even death in newborn piglets. In this study, we found that PDCoV infection significantly upregulated the expression of CH25H in IPI-FX cells, a cell line of porcine ileum epithelium. Overexpression of CH25H inhibited PDCoV replication, whereas CH25H silencing using RNA interference promoted PDCoV infection. Treatment with 25-hydroxycholesterol (25HC), the catalysate of cholesterol via CH25H, inhibited PDCoV proliferation by impairing viral invasion of IPI-FX cells. Furthermore, a mutant CH25H (CH25H-M) lacking hydroxylase activity also inhibited PDCoV infection to a lesser extent. Taken together, our data suggest that CH25H acts as a host restriction factor to inhibit the proliferation of PDCoV but this inhibitory effect is not completely dependent on its enzymatic activity.

Interleukin (IL)-21 Promotes the Differentiation of IgA-Producing Plasma Cells in Porcine Peyer's Patches via the JAK-STAT Signaling Pathway.

Secretory IgA is critical to prevent the invasion of pathogens via mucosa. However, the key factors and the mechanisms of IgA generation in the porcine gut are not well-understood. In this study, a panel of factors, including BAFF, APRIL, CD40L, TGF-β1, IL-6, IL-10, IL-17A, and IL-21, were employed to stimulate IgM+ B lymphocytes from porcine ileum Peyer's patches. The results showed that IL-21 significantly upregulated IgA production of B cells and facilitated cell proliferation and differentiation of antibody-secreting cells. In addition, three transcripts in porcine IgA class switch recombination (CSR), germ-line transcript α, post-switch transcript α, and circle transcript α, were first amplified by (nest-)PCR and sequenced. All these key indicators of IgA CSR were upregulated by IL-21 treatment. Furthermore, we found that IL-21 predominantly activated JAK1, STAT1, and STAT3 proteins and confirmed that the JAK-STAT signaling pathway was involved in porcine IgA CSR. Thus, IL-21 plays an important role in the proliferation and differentiation of IgA-secreting cells in porcine Peyer's patches through the JAK-STAT signaling pathway. These findings provide insights into the mucosal vaccine design by regulation of IL-21 for the prevention and control of enteric pathogens in the pig industry.

The impact of low and high doses of acrylamide on the intramural neurons of the porcine ileum

The present study was designed to assess the influence of acrylamide supplementation, in tolerable daily intake (TDI) dose and a dose ten times higher than TDI, on the neurochemical phenotype of the ENS neurons and synthesis of proinflammatory cytokines in the wall of the porcine ileum. The study was performed on 15 juvenile female Danish Landrace pigs, divided into three groups: C group- animals receiving empty gelatine capsules, LD group- animals receiving capsules with the TDI dose (0.5 μg/kg b.w./day) of acrylamide and HD group- animals receiving acrylamide in a dose ten times higher than the TDI (5 μg/kg b.w./day) in a morning meal for 28 days. It was established that supplementation of acrylamide led to an increase in substance P (SP)-, calcitonin gene-related peptide (CGRP)-, galanin (GAL)- and vesicular acetylcholine transporter (VAChT)-like immunoreactive (LI) neurons as well as a decrease in neuronal nitric oxide synthase (nNOS) -like immunoreactivity in all types of ileum intramural plexuses. Moreover, using ELISA method, an increase in the level of proinflammatory cytokines (IL-1β, IL-6 and TNF- α) was noted in the ileum wall. The results suggest that SP, CGRP, GAL, nNOS and VACHT participate in the regulation of inflammatory conditions induced by acrylamide supplementation.