Pig Intestine Research Articles

Structure, function, and potential application of F18 in vaccine development

Post-weaning diarrhea is one of the most common diseases in pigs, affecting pigs in the first two weeks after weaning, causing significant economic losses. The main cause of this disease is Enterotoxigenic Escherichia coli (ETEC), which is characterized by two pathogenic factors: fimbriae and enterotoxins. These fimbriae are considered to mediate bacterial adhesion through binding to the receptor present in the pig intestine. Several studies indicate that F4 and F18 fimbriae account for a high percentage of the fimbriae in ETEC and are detected in most of the isolated ETEC strains. Currently, oral vaccines are a potential preventive measure used with the ability to generate IgA, stimulate the mucosal immune system, and be widely noticed by researchers. However, the vaccines are only effective against ETEC-F4 and less effective against ETEC-F18, so more research is needed to improve the effectiveness of ETEC-F18 vaccines in the future. In this review, we presented the structure, function, and potential application of the F18 in vaccine development.

Metagenomic analysis fecal microbiota of dysentery-like diarrhoea in a pig farm using next-generation sequencing.

Porcine enteric diseases including swine dysentery involves a wide range of possible aetiologies and seriously damages the intestine of pigs of all ages. Metagenomic next-generation sequencing is commonly used in research for detecting and analyzing pathogens. In this study, the feces of pigs from a commercial swine farm with dysentery-like diarrhea was collected and used for microbiota analysis by next-generation sequencing. While Brachyspira spp. was not detected in diarrheal pig fecal samples, indicating that the disease was not swine dysentery. The quantity of microbial population was extremely lowered, and the bacterial composition was altered with a reduction in the relative abundance of the probiotics organisms, Firmicutes and Bacteroidetes, with an increase in pathogens like Fusobacterium and Proteobacteria, in which the specific bacteria were identified at species-level. Viral pathogens, porcine circovirus type 2, porcine lymphotropic herpesviruses 1, and porcine mastadenovirus A were also detected at pretty low levels. Carbohydrate-active enzymes (CAZy) analysis indicated that the constitute of Firmicutes and Bacteroidete were also changed. Further, the Kyoto Encyclopedia of Genes and Genomes (KEGG) alignment analysis indicated that the microbiota of diarrheal pigs had a lower ability in utilizing energy sources but were enriched in multi-drug resistance pathways. Comprehensive Antibiotic Resistance Database (CARD) and Virulence Factors of Pathogenic Bacteria (VFDB) analysis indicated that genes for elfamycin and sulfonamide resistance and the iron uptake system were enriched in diarrheal pigs. This revealed potential bacterial infection and can guide antibiotic selection for treating dysentery. Overall, our data suggested that alterations in both the population and functional attributes of microbiota in diarrheal pigs with decreased probiotic and increased pathogenic microorganisms. These results will help elucidate the mechanism of dysentery-like diarrhea and the development of approaches to control the disease.

Combined effects of host genetics and diet on porcine intestinal fungi and their pathogenic genes.

As research on gut microbes progresses, it becomes increasingly clear that a small family of microbiota--fungi, plays a crucial role in animal health. However, little is known about the fungal composition in the pig intestine, especially after a dietary fiber diet and hybrid genetics, and the changes in host pathogenicity-associated genes they carry. The purpose of this study is to investigate the effects of diet and genetics on the diversity and structure of porcine intestinal fungi and to describe, for the first time, the host pathogenicity-related genes carried by porcine intestinal fungi. Samples of colonic contents were collected for metagenomic analysis using a 3 × 2 parsing design, where three pig breeds (Taoyuan, Duroc, and crossbred Xiangcun) were fed high or low fiber diets (n = 10). In all samples, we identified a total of 281 identifiable fungal genera, with Ascomycota and Microsporidia being the most abundant fungi. Compared to Duroc pigs, Taoyuan and Xiangcun pigs had higher fungal richness. Interestingly, the fiber diet significantly reduced the abundance of the pathogenic fungus Mucor and significantly increased the abundance of the fiber digestion-associated fungus Neocallimastix. Pathogenic fungi exert their pathogenicity through the genes they carry that are associated with host pathogenicity. Therefore, we obtained 839 pathogenicity genes carried by the spectrum of fungi in the pig intestine by comparing the PHI-base database. Our results showed that fungi in the colon of Taoyuan pigs carried the highest abundance of different classes of host pathogenicity-related genes, and the lowest in Duroc pigs. Specifically, Taoyuan pigs carried high abundance of animal pathogenicity-related genes (CaTUP1, CPAR2_106400, CaCDC35, Tfp1, CaMNT2), and CaTUP1 was the key gene for Candida pathogenicity. The intestinal fungal composition of crossbred Xiangcun pigs and the abundance of host pathogenicity-associated genes they carried exhibited a mixture of characteristics of Taoyuan and Duroc pigs. In conclusion, our results provide the first comprehensive report on the effects of dietary fiber and genetics on the composition of intestinal fungi and the host-associated pathogenicity genes they carry in pigs. These findings provide a reference for subsequent pig breeding and development of anti-pathogenic fungal drugs.

Combined indocyanine green and medical glue enables stable and precise position in animal studies: promising for fluorescence-guided pulmonary ground glass nodule resection.

Accurate preoperative localization of pulmonary nodules is crucial for surgical treatment. The use of indocyanine green (ICG) for localization is prone to thoracic contamination and spread, resulting in the eventual failure of localization. By using medical glue combined with ICG, we can accurately and permanently locate various tissues in animal study, which can provide evidences for clinical translations. A series of medical glue and ICG volume ratios of 2:3, 3:3, 4:3, 6:3, and 9:3 were mixed and injected immediately into subcutaneous tissues of BALB/c nude mice; either medical glue or ICG was injected singly in the control group. Fluorescence intensity over time and boundary sharpness were investigated to determine the optimal ratio. Then, fluorescence guided resection of tissue was performed ex vivo on the pig intestine utilizing optimal ratio. Further, localization agents with the optimal ratio were injected into the organs of living mice, and fluorescence imaging for accurate positioning was performed 24 hours later. The localization agents with a volume ratio of 4:3 showed the best boundary sharpness and the strongest photostability. With the guidance of fluorescence navigation, the marked tissues were accurately separated and removed from the surrounding tissue both on mice and on pig intestines. In the organs of living mice, the localization agents (ratio 4:3) realized accurate positioning of marked tissues. Additionally, the medical glue limited the diffusion of ICG, promising to enable more stable and precise positioning of the nodules during surgery. The combination of ICG and medical glue presents a superior approach when compared to the individual use of either ICG or medical glue. This technique offers enhanced precision and durability and sealed the wound, thereby mitigating the risk of pneumothorax following puncture procedures. This innovative technique optimizes the properties of medical adhesive to augment tissue density while harnessing the real-time fluorescent endoscopic marking capabilities of ICG during surgical interventions. By employing this innovative technique, it holds significant promise for augmenting the accuracy of pulmonary nodule localization in thoracoscopic surgery within future clinical applications.

Mucous non-goblet cells in the small intestine of guinea pigs (Cavia porcellus): a histological and histochemical study.

The covering and glandular epithelium of the small intestine in guinea pigs (Cavia porcellus) include some mucus-secreting cells. Goblet cells are specific cells for mucus secretion with a distinctive cup-like appearance due to the accumulation of mucin in the apical pole. The deep crypt secretory (DCS) cells were identified in a limited array of species and only recently were noticed in the large intestine in mice, guinea pigs, humans, monkeys, and pigs. Our study focuses on the microscopical and histochemical features of the DCS cells in the small intestine of guinea pigs. The samples from the small intestine were collected from five fully grown guinea pigs that were presented to the Hospital of the Faculty of Veterinary Medicine Cluj-Napoca (Romania) with severe lesions resulted from domestic activities. The collected tissue samples underwent fixation in 10% buffered formalin and were later processed by standard paraffin technique. Mucous substances were detected using the Periodic Acid-Schiff and Alcian-Blue histochemical stain methods. The intestinal samples of the guinea pigs assessed had a standard microanatomical structure. As regards the mucous-secreting cells from the small intestine, two cell types were identified, i.e. the goblet cells and DCS cells. DCS cells were only detected in the deep parts of the Lieberkühn glands from the jejunum and ileum, and were different morphologically and histochemically from the regular goblet cells. Our study managed to describe for the first time in guinea pigs, the existence of DCS cells in the jejunum and ileum of the small intestine, but not in the duodenum.

A combined pig model to determine the net absorption of volatile fatty acids in the large intestine under different levels of crude fiber.

This study aimed to develop a combined model to quantify the net absorption of volatile fatty acids (VFA) in the large intestine (LI) of pigs. Fifteen female growing pigs (Duroc × Large White × Landrace) were ranked by body weight (30 ± 2.1 kg) on day 0 and assigned to one of three treatments, namely the basal diet containing different crude fiber (CF) levels (LCF: 3.0% CF, MCF: 4.5% CF, and HCF: 6.0% CF). The pigs were implanted with the terminal ileum fistula and the cannulation of the ileal mesenteric vein (IMV), portal vein (PV), and left femoral artery (LFA) from days 6 to 7. [13 C]-Labeled VFA and P-aminohippuric acid were constantly perfused into the terminal ileum fistula and the cannulation of the IMV (day 15), respectively. Blood samples were collected from the PV and the LFA during perfusion (5 h), and LI samples were collected. The net flux of [12 C]-acetic acid in the PV was greater for LCF versus MCF (p = 0.045), but no difference was observed in the net flux of [12 C]-propionic acid (p = 0.505) and [12 C]-butyric acid (p = 0.35) in the PV among treatments. The deposition of [12 C]-acetic acid in the LI was greater for LCF versus MCF (p = 0.014), whereas the deposition of [12 C]-propionic acid (p = 0.007) and [12 C]-butyric acid (p = 0.037) in the LI was greater for LCF versus HCF. In conclusion, this pig model was found conducive to study the net absorption of VFAs in the LI, and LCF had more net absorption of VFAs in the LI than MCF and HCF.

Methionine influences the profile of intestinal antibiotic resistome through inhibiting the growth of Escherichia coli

Antibiotic resistance genes (ARGs) are a new type of environmental pollutant. However, studies have mainly focused on the distribution characteristics of ARGs in the livestock environment, lacking of studies on the composition of ARGs in the intestinal tract of animals and the effect of nutrients on intestinal ARGs and microbial communities. Reducing antimicrobial resistance and maintaining optimal animal health and performance are urgently needed. Methionine is an essential amino acid which plays a critical role in the growth and reproductive performance of animals. In this study, feeding experiment, in vitro fermentation and bacterial culture experiment were performed to explore the influence of methionine on the intestinal resistome of sows. We found that dietary 0.2 % methionine supplementation decreased the total abundance of intestinal ARGs, which was further confirmed by in vitro fecal microbial fermentation of sows. Metagenome binning analysis identified that Escherichia coli was the major ARG host, which carried 60–113 ARGs and 134–286 virulence factors, indicating that Escherichia coli in the pig intestine is not only a core ARG host, but also an important pathogen. In addition, we found that methionine supplementation inhibited the growth of Escherichia coli, indicating that dietary methionine may reduce the resistome risk in sow intestine by inhibiting core ARG hosts such as Escherichia coli. These findings reveal that dietary methionine application plays a critical role in intestinal antibiotic resistance, providing a new idea for preventing and controlling environmental pollution by antibiotic-resistant microbes.

Synthesis of glycine from 4-hydroxyproline in tissues of neonatal pigs.

Glycine from sow's milk only meets 20% of the requirement of suckling piglets. However, how glycine is synthesized endogenously in neonates is not known. This study determined glycine synthesis from 4-hydroxyproline (an abundant amino acid in milk and neonatal blood) in tissues of sow-reared piglets with normal birth weights. Piglets were euthanized at 0, 7, 14 and 21 days of age, and their tissues were used to determine glycine synthesis from 0 to 5 mM 4-hydroxyproline, activities and mRNA expression of key glycine-synthetic enzymes, and their cell-specific localization. Activities of 4-hydroxyproline oxidase (OH-POX), proline oxidase (POX), serine hydroxymethyltransferase (SHMT), threonine dehydrogenase (TDH), alanine:glyoxylate transaminase (AGT), and 4-hydroxy-2-oxoglutarate aldolase (HOA) occurred in the kidneys and liver from all age groups of piglets, and in the pancreas of 7- to 21-day-old piglets. Activities of OH-POX and HOA were absent from the small intestine of newborn pigs but present in the small intestine of 7- to 21-day-old piglets and in the skeletal muscle of 14- to 21-day-old piglets. Between days 0 and 21 of age, the enzymatic activities of OH-POX, AGT, and HOA decreased in the liver and kidneys but increased in the pancreas and small intestine with age. The mRNA levels of these three enzymes changed in a manner similar to their enzymatic activities. In contrast to OH-POX, AGT, and HOA, the enzymatic activities of POX, SHMT, and TDH were present in the kidneys, liver, and intestine of all age groups of piglets. Glycine was synthesized from 0.1 to 5 mM 4-hydroxyproline in the liver and kidney from 0- to 21-day-old piglets, as well as the pancreas, small intestine, and skeletal muscle from 14- to 21-day-old piglets in a concentration-dependent manner. Collectively, our findings indicate that 4-hydroxyproline is used for the synthesis of glycine in tissues of piglets to compensate for the deficiency of glycine in milk.

Effect of Particle Size of Wheat and Barley Grain on the Digestibility and Fermentation of Carbohydrates in the Small and Large Intestines of Growing Pigs

The objective of this investigation was to study the effects of different cereal types, barley and wheat, with different particle sizes (PS) on the recovery of ileal digesta and fecal excretion, digestion of nutrients and fiber components, mean transit time (MTT), and short-chain fatty acid content and composition in growing pigs studied in two experiments. Five barrows with ileal cannulas (initial BW 35.9 ± 1.5 kg) in Experiment 1 and thirty-two castrated pigs (30.8 ± 1.3 kg) in Experiment 2 were fed four different diets: barley fine, barley coarse, wheat fine and wheat coarse diets. The cereal type and PS did not influence the relative weight of the small and large intestines and pH of digesta, whereas MTT in the large intestine of pigs fed the coarse barley diet was lower compared to pigs fed other diets (p < 0.05). Pigs fed the coarse barley diet had lower apparent ileal digestibility (AID) and apparent total tract digestibility (ATTD) of nutrients and fiber (p < 0.05), whereas pigs fed the fine barley diet had similar AID and ATTD to pigs fed wheat fine and coarse diets (p < 0.05). In conclusion, the barley diet was more influenced by PS in comparison to wheat, thereby inducing lower AID and ATTD of nutrient.

Changes caused by low doses of glyphosate in the population of substance P - like immunoreactive enteric neurons in the small intestine in pigs

Food safety has become a priority for highly developed societies. Glyphosate is an active ingredient of the most widely used herbicide formulations in protecting agricultural and horticultural crops. The aim of the current study was to determine how administration of glyphosate affects the quantitative population of substance P (SP)- synthesizing neurons in the porcine small intestine.Fifteen sexually immature gilts divided into 3 groups were used: control (C) - animals receiving empty gelatin capsules; E1 - animals receiving a low dose of glyphosate - corresponding to the theoretical maximum daily intake (TMDI) - 0.05 mg/kg of b.w./day; E2 - 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 with morning feeding. After 4 weeks period of glyphosate supplementation, all pigs were euthanized and small intestine samples were collected for further examination. 12 μm-thick frozen intestinal sections were then double-stained by incubating with primary (HuC/D- used as a pan neuronal marker and SP) as well as secondary (Alexa Fluor 488 and 546) antisera.Glyphosate supplementation increased the number of intramural neurons showing immunoreactivity to SP. In the duodenum, both doses of glyphosate evoked marked changes in population of SP-like immunoreactive (LI) neurons in the outer submucous plexus (OSP) (from 22.36±1.12% to 26.65±0.96% in the E2 group and to 38.53±1.44% in the E2 group) as well as in the inner submucous plexus (ISP) (from 12.30±0.89% to 20.21±2.34% and 35.71±1.90%). In the myenteric plexus (MP) administration of higher dose of glyphosate led to an increase in the number of SP-LI neurons (from 1.22±0.15% to 6.75±0.84%). In the jejunum, an increased number of SP-positive neurons was noted in all type of enteric plexuses and both experimental groups (in the MP: from 1.25±0.20% in the C group to 3.27±0.30% in the E1 group and to 5.41±0.26% in the E2 group; in the OSP: from 23.03±1.43% to 31.25±1.34% and 42.26±1.04%; in the ISP: from 12.35±0.70% to 16.36±0.68% and 43.06±1.21%). In the ileum, the greatest increase was recorded in the ISP (from 20.22±0.96% to 31.16±1.84% in the E1 group and to 43.11±1.34% in the E2 group) and in the OSP (from 15.63±0.79% to 22.10±1.52% and 35.93±2.27%). Whereas in the MP only in the E2 group an increase was observed (from 1.01±0.19% to 6.11±0.53%).Neurons in the ENS underwent adaptive changes in response to the glyphosate intoxication, which manifested themselves as increased population of cells displaying SP- immunoreactivity. SP engaged in defensive reactions of neurons and conduction of sensory stimuli may play an important role in protecting the ENS neurons against the harmful effect of glyphosate. the National Science Centre in Poland [grant no. 2020/04/X/NZ7/00338] 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.

A lumen-adapted navigation scheme with spatial awareness from monocular vision for autonomous robotic endoscopy

Lumen is a common anatomical condition in endoscopic therapy or diagnostics. The navigation of autonomous robotic endoscopy usually involves vision techniques such as detection of lumen center or anatomically specific contour features. However, these methods may fail to achieve smooth interventions and result in large conveying force without spatial awareness of the tissue state. In this paper, a novel navigation pipeline based on a spatial-aware monocular vision is proposed. The spatial awareness pipeline starts with a data-driven depth estimation technique for reconstructing real-time approximate tissue surface. The spatial shape of the lumen described by the skeleton is then extracted using digital topology. We modify the skeleton to get a smooth pathway, and design an adaptive autonomous control strategy using geometric information from the pathway. We conduct experiments on a colon phantom and ex vivo pig intestines. We test turning performance of several bending segments with different angles in phantom, as well as the overall performance of a long-range intervention task in the phantom and pig intestines. The results show our navigation scheme achieve smoother intervention with lower conveying force. The proposed navigation method with spatial awareness can effectively improve the fluency of autonomous robotic endoscopy.

Effect of microRNA, miR-215, on gene expression and metabolism of porcine intestinal epithelial cells

MiR-215 maintains a high level of expression in healthy pig intestine and is decreased in lesions or injuries. To clarify the influence of miR-215 on the porcine intestine, miR-215 mimic/inhibitor was transfected into the intestinal porcine epithelial cell line, J2 (IPEC-J2) to overexpress or suppress the expression of miR-215. The treated cells were subjected to various functional assays. The effect of miR-215 on global gene expression was detected using high-throughput sequencing on the BGISEQ500 platform; cell proliferation was evaluated using a Cell Counting Kit-8 (CCK-8); and the role of miR215 on cellular metabolism was tested using non-targeted metabonomics and liquid chromatography with tandem mass spectrometry (LC-MS/MS) analysis. Our results revealed that miR-215 promoted cell proliferation in IPEC-J2, up-regulated the expression of the ribosomal protein family, and downregulated the expression of oncogenesis-related genes and tumour-suppressor genes. Results of the RNA sequencing and the metabonomics detection reveal that miR-215 affects the PI3K-Akt signalling pathway. Our results contribute to further research on miR-215 in the regulation of barrier function and metabolism in the porcine intestine.

Efek Antelmintik Ekstrak Metanol Kulit Batang Nangka (Artocarpus heterophyllus) terhadap Cacing Ascaris lumbricoides

Jackfruit (Artocarpus heterophyllus) is a type of plant that grows in Indonesia which functions as a food ingredient and as traditional medicine. Generally, this genus contains prenylated flavonoid compounds. This study aimed to determine the anthelmintic effect of jackfruit bark extract against Ascaris lumbricoides worms by utilizing total maceration with methanol solvent as the extraction method, which yielded 6.61%. Roundworm Ascaris lumbricoides are found in pig intestines. The samples were divided into 5 groups, including positive control of 250 mg combantrin solution, negative control of NaCMC and a treatment group of jackfruit bark extract with a concentration of 1%, 3%, and 5%. Each group consisted of three worms, incubated at 37oC for 24 hours with a 6 hours interval. The results showed that at a concentration of 5%, a jackfruit bark extract was efficient as an anthelmintic caused two worms to lyse at 6 hours and one at 12 hours.

Effect of phage vB_EcoM_FJ1 on the reduction of ETEC O9:H9 infection in a neonatal pig cell line

Enterotoxigenic Escherichia coli (ETEC) colonizes the intestine of young pigs causing severe diarrhoea and consequently bringing high production costs. The rise of antibiotic selective pressure together with ongoing limitations on their use, demands new strategies to tackle this pathology. The pertinence of using bacteriophages as an alternative is being explored, and in this work, the efficacy of phage vB_EcoM_FJ1 (FJ1) in reducing the load of ETEC EC43-Ph (serotype O9:H9 expressing the enterotoxin STa and two adhesins F5 and F41) was assessed. Foreseeing the oral application on piglets, FJ1 was encapsulated on calcium carbonate and alginate microparticles, thus preventing phage release under adverse conditions of the simulated gastric fluid (pH 3.0) and allowing phage availability in simulated intestinal fluid (pH 6.5). A single dose of encapsulated FJ1, provided to IPEC-1 cultured cells (from intestinal epithelium of piglets) previously infected by EC43, provided bacterial reductions of about 99.9% after 6 h. Although bacteriophage-insensitive mutants (BIMs) have emerged from treatment, the consequent fitness costs associated with this new phenotype were demonstrated, comparatively to the originating strain. The higher competence of the pig complement system to decrease BIMs’ viability, the lower level of colonization of IPEC-1 cells observed with these mutants, and the increased survival rates and health index recorded in infected Galleria mellonella larvae supported this observation. Most of all, FJ1 established a proof-of-concept of the efficiency of phages to fight against ETEC in piglet intestinal cells.

Whole-Genome Sequencing of Pig Gut Lactobacillus amylovorus CICC 6090.

Lactobacillus amylovorus has been reported to reduce weight and fat content in humans. To identify and understand the mechanism underlying this process, the complete genome of CICC 6090, which was isolated from pig intestines, was sequenced using PromethION and next-generation sequencing.

Evaluation of dietary curcumin nanospheres as phytobiotics on growth performance, serum biochemistry, nutritional composition, meat quality, gastrointestinal health, and fecal condition of finishing pigs.

Curcumin is a bioactive functional feeding stimulant that is widely used as an additive in cuisine and animal feeds. Owing to its hydrophobic nature and low bioavailability, the nanoformulation of curcumin has recently received special attention from researchers. In this study, we investigated the effects of curcumin nanospheres (CN) on the growth performance, serum biochemistry, meat quality, intestinal immunohistochemistry, fecal malodors and microbes in finishing pigs. A total of 90 crossbred pigs (Duroc × [Yorkshire × Landrace]) with an average initial body weight of 73.77 ± 0.08 kg were randomized into 3 dietary groups in triplicate pens (10 pigs in each pen): control (CON) without supplementation of CN and the pigs in the remaining two groups were supplemented with CN at 1.0 (CN1) and 2.0 (CN2) mL/kg diet for a 40-day long experiment. The results showed that pigs fed the higher CN supplemented diet (CN2) had significantly higher final weight (FW) and weight gain (WG) than those fed the CON diet, and no significant differences were observed in the feed conversion ratio (FCR) and average daily feed intake (ADFI) after 28 days. At the end of the experiment, pigs fed the CN supplemented diet showed no significant difference in WG, ADFI or FCR compared to those on the CON diet. Overall, at the termination of the 40-day feeding trial, dietary CN had a significant effect on FW and WG, except for ADFI and FCR, in finishing pigs. After 40 days of the feeding trial, serum biochemical parameters such as glutamic-pyruvic transaminase, glutamic-oxaloacetic transaminase, triglycerides, and total cholesterol levels were significantly decreased in pigs fed the CN supplemented diet. However, high density lipoprotein levels were significantly increased in pigs fed the CN diets. Protein and lipid contents, as well as yellowness and lightness of the neck and longissimus dorsi muscles were not significantly affected by CN supplementation; however, there was a tendency to increase the redness of the longissimus dorsi muscle in pigs fed the CN2 supplemented diet compared to the CON diet. Meat grading and carcass weight significantly increased in pigs fed a higher CN supplemented diet. Fecal Escherichia coli and ammonia gas were significantly depleted in pigs fed CN diets. Histomorphological parameters, such as villus height, crypt depth and goblet cells in the jejunum of the intestine were significantly increased in pigs fed CN diet. Immunohistochemical staining showed that pro-inflammatory cytokine like tumor necrosis factor-α expression was reduced in pigs fed CN supplemented diets compared to the CON diet; however, antibodies such as immunoglobulin A and tight junction proteins such as claudin 3 were highly expressed in the intestine of pigs fed the CN diets. Overall, the results demonstrate the potential of dietary curcumin nanospheres as a nanobiotechnology tool as well as an effective feed additive for improving the performance and health status of finishing pigs.

Validation of Recombinant Heparan Sulphate Reagents for CNS Repair.

Therapies that target the multicellular pathology of central nervous system (CNS) disease/injury are urgently required. Modified non-anticoagulant heparins mimic the heparan sulphate (HS) glycan family and have been proposed as therapeutics for CNS repair since they are effective regulators of numerous cellular processes. Our in vitro studies have demonstrated that low-sulphated modified heparan sulphate mimetics (LS-mHeps) drive CNS repair. However, LS-mHeps are derived from pharmaceutical heparin purified from pig intestines, in a supply chain at risk of shortages and contamination. Alternatively, cellular synthesis of heparin and HS can be achieved using mammalian cell multiplex genome engineering, providing an alternative source of recombinant HS mimetics (rHS). TEGA Therapeutics (San Diego) have manufactured rHS reagents with varying degrees of sulphation and we have validated their ability to promote repair in vitro using models that mimic CNS injury, making comparisons to LS-mHep7, a previous lead compound. We have shown that like LS-mHep7, low-sulphated rHS compounds promote remyelination and reduce features of astrocytosis, and in contrast, highly sulphated rHS drive neurite outgrowth. Cellular production of heparin mimetics may, therefore, offer potential clinical benefits for CNS repair.

Culture of Piglet Intestinal 3D Organoids from Cryopreserved Epithelial Crypts and Establishment of Cell Monolayers.

Intestinal organoids are increasingly being used to study the gut epithelium for digestive disease modeling, or to investigate interactions with drugs, nutrients, metabolites, pathogens, and the microbiota. Methods to culture intestinal organoids are now available for multiple species, including pigs, which is a species of major interest both as a farm animal and as a translational model for humans, for example, to study zoonotic diseases. Here, we give an in-depth description of a procedure used to culture pig intestinal 3D organoids from frozen epithelial crypts. The protocol describes how to cryopreserve epithelial crypts from the pig intestine and the subsequent procedures to culture 3D intestinal organoids. The main advantages of this method are (i) the temporal dissociation of the isolation of crypts from the culture of 3D organoids, (ii) the preparation of large stocks of cryopreserved crypts derived from multiple intestinal segments and from several animals at once, and thus (iii) the reduction in the need to sample fresh tissues from living animals. We also detail a protocol to establish cell monolayers derived from 3D organoids to allow access to the apical side of epithelial cells, which is the site of interactions with nutrients, microbes, or drugs. Overall, the protocols described here is a useful resource for studying the pig intestinal epithelium in veterinary and biomedical research.

Sorghum tannin extract impedes in vitro digestibility and fermentability of nutrients in the simulated porcine gastrointestinal tract.

The site and extent of digestion of sorghum nutrients affected by tannins in the intestine are not clarified. Porcine small intestine digestion and large intestine fermentation were simulated in vitro to determine the effects of sorghum tannin extract on the digestion and fermentation characteristics of nutrients in the mimicked porcine gastrointestinal tract. In experiment 1, low-tannin sorghum grain without or with 30 mg/g sorghum tannin extract were digested by porcine pepsin and pancreatin to measure in vitro digestibility of nutrients. In experiment 2, the lyophilized porcine ileal digesta from 3 barrows (Duroc × Landrace × Yorkshire, 27.75 ± 1.46 kg) fed the low-tannin sorghum grain without or with 30 mg/g sorghum tannin extract and the undigested residues from experiment 1 were, individually, incubated with fresh pig cecal digesta as inoculums for 48 h to simulate the porcine hindgut fermentation. The results revealed that sorghum tannin extract decreased in vitro digestibility of nutrients both by pepsin hydrolysis or pepsin-pancreatin hydrolysis (P < 0.05). Although enzymatically unhydrolyzed residues provided more energy (P = 0.09) and nitrogen (P < 0.05) as fermentation substrates, the microbial degradation of nutrients from unhydrolyzed residues and porcine ileal digesta were both decreased by sorghum tannin extract (P < 0.05). Regardless of unhydrolyzed residues or ileal digesta as fermentation substrates, microbial metabolites including the accumulative gas production excluding the first 6 h, total short-chain fatty acid and microbial protein content in the fermented solutions were decreased (P < 0.05). The relative abundances of Lachnospiraceae AC2044 and NK4A136 and Ruminococcus_1 was decreased by sorghum tannin extract (P < 0.05). In conclusion, sorghum tannin extract not only directly decreased the chemical enzymatic digestion of nutrients in the simulated anterior intestine, but also directly inhibited the microbial fermentation including microbial diversities and metabolites in the simulated posterior intestine of pigs. The experiment implies that the decreased abundances of Lachnospiraceae and Ruminococcaceae by tannins in the hindgut may weaken the fermentation capacity of microflora and thus impair the nutrient digestion in the hindgut, and ultimately reduce the total tract digestibility of nutrients in pigs fed high tannin sorghum.

Immunodetection of P2X2 Receptor in Enteric Nervous System Neurons of the Small Intestine of Pigs.

Extracellular adenosine 5'-triphosphate (ATP) is one of the best-known and frequently studied neurotransmitters. Its broad spectrum of biological activity is conditioned by the activation of purinergic receptors, including the P2X2 receptor. The P2X2 receptor is present in the central and peripheral nervous system of many species, including laboratory animals, domestic animals, and primates. However, the distribution of the P2X2 receptor in the nervous system of the domestic pig, a species increasingly used as an experimental model, is as yet unknown. Therefore, this study aimed to determine the presence of the P2X2 receptor in the neurons of the enteric nervous system (ENS) of the pig small intestine (duodenum, jejunum, and ileum) by the immunofluorescence method. In addition, the chemical code of P2X2-immunoreactive (IR) ENS neurons of the porcine small intestine was analysed by determining the coexistence of selected neuropeptides, i.e., vasoactive intestinal polypeptide (VIP), substance P (sP), and galanin. P2X2-IR neurons were present in the myenteric plexus (MP), outer submucosal plexus (OSP), and inner submucosal plexus (ISP) of all sections of the small intestine (duodenum, jejunum, and ileum). From 44.78 ± 2.24% (duodenum) to 63.74 ± 2.67% (ileum) of MP neurons were P2X2-IR. The corresponding ranges in the OSP ranged from 44.84 ± 1.43% (in the duodenum) to 53.53 ± 1.21% (in the jejunum), and in the ISP, from 53.10 ± 0.97% (duodenum) to 60.57 ± 2.24% (ileum). Immunofluorescence staining revealed the presence of P2X2-IR/galanin-IR and P2X2-IR/VIP-IR neurons in the MP, OSP, and ISP of the sections of the small intestine. The presence of sP was not detected in the P2X2-IR neurons of any ganglia tested in the ENS. Our results indicate for the first time that the P2X2 receptor is present in the MP, ISP, and OSP neurons of all small intestinal segments in pigs, which may suggest that its activation influences the action of the small intestine. Moreover, there is a likely functional interaction between P2X2 receptors and galanin or VIP, but not sP, in the ENS of the porcine small intestine.