Gut Microbiota-Derived 5-Hydroxyindoleacetic Acid Alleviates Diarrhea in Piglets via the Aryl Hydrocarbon Receptor Pathway.

With the improvement in sow prolificacy, formula feeding has been increasingly used in the pig industry. Diarrhea remains a serious health concern in formula-fed (FF) piglets. Fecal microbiota transplantation (FMT) is an efficacious strategy to reshape gut microbiota and the metabolic profile for treating diarrhea. This study aims to investigate whether FMT from breast-fed piglets could alleviate diarrhea in FF piglets. The piglets were randomly assigned to the control (CON) group, FF group, and FMT group. Our results showed that FF piglets exhibited a higher diarrhea incidence, damaged colonic morphology, and disrupted barrier function. In contrast, FMT treatment normalized the morphology and barrier function. FMT suppressed the JNK/MAPK pathway and production of proinflammatory cytokines. Additionally, FF piglets had a lower abundance of the beneficial bacterial genus Bifidobacterium compared to CON piglets. Following FMT administration, Bifidobacterium was restored. Meanwhile, 5-HIAA, a metabolite of tryptophan, and AHR-responsive CYP1A1 and CYP1B1 were upregulated. Importantly, integrated multiomics analysis revealed a strong positive correlation between Bifidobacterium and 5-HIAA. In vitro, 5-HIAA supplementation reversed the LPS-induced disruption of tight junctions and production of proinflammatory cytokines in IPEC-J2 cells. In conclusion, FMT reduced diarrhea incidence and improved growth performance. The alleviative effect of FMT on diarrhea was associated with Bifidobacterium and 5-HIAA.

Effect of Noise and Music on Neurotransmitters in the Amygdala: The Role Auditory Stimuli Play in Emotion Regulation.

Stress caused by noise is becoming widespread globally. Noise may lead to deafness, endocrine disorders, neurological diseases, and a decline in mental health. The mechanism behind noise-induced neurodevelopmental abnormalities is unclear, but apoptosis and pro-inflammatory signals may play an important role. In this study, weaned piglets were used as a model to explore noise-induced neurodevelopmental abnormalities. We hypothesized that long-term noise exposure would induce anxiety and cause acute stress, exhibited by alterations in neurotransmission in the amygdala. A total of 72 hybrid piglets (Large White × Duroc × Min Pig) were randomly divided into three groups, including noise (exposed to mechanical noise, 80-85 dB), control (blank, exposed to natural background sound, <40 dB), and music (positive control, exposed to Mozart K.448, 60-70 dB) groups. The piglets were exposed to 6 h of auditory noise daily (10:00-16:00) for 28 days. Compared with the control group, piglets exposed to noise showed more aggressive behavior. The expression of Caspase3, Caspase9, Bax, NF-κB (p56), TLR4, MYD88, I κ B α, IL-1 ß, TNF-α, and IL-12RB2 was significantly upregulated in the amygdala, while the expression of Nrf2, HO-1, CAT, and SOD was downregulated in piglets in the noise group. Cell death occurred, and numerous inflammatory cells accumulated in the amygdala of piglets in the noise group. Targeted metabolomics showed that the content of inhibitory neurotransmitter GABA was higher in the amygdala of piglets in the noise group. Compared with the noise group, piglets in the music group displayed more positive emotion-related behaviors. Compared with the noise group, the expression of genes related to apoptosis, inflammation, and oxidative damage was lower in the music group. Cells of the amygdala in the music group were also of normal morphology. Our results show that noise-induced stress causes apoptosis and neuroinflammation in the amygdala and induces anxiety during the early neonatal neural development of piglets. In contrast, to some extent, music alleviates noise-induced anxiety.

Effects of early maternal separation on the expression levels of hippocampal and prefrontal cortex genes and pathways in lactating piglets.

Introduction: In actual production, due to increased litter size when raising pigs, the management of piglets by split-suckling leads to intermittent neonatal maternal separation (MS). Early lactation is a critical period for the cognitive development of the brain of newborn piglets, and we hypothesized that intermittent MS may affect piglets' neurodevelopment and cognitive ability. Methods: To determine the effects of the MS, we selected hippocampal and prefrontal cortex (PFC) tissues from piglets for the detection of neurodevelopmental or cognitive related indicators, the control group (Con group, n = 6) was established with no MS and an experimental group (MS group, n = 6) was established with MS for 6 h/day. Piglets in the MS group were milk-supplemented during the separation period and all piglets in both treatment groups were weaned at postnatal day (PND) 35. On PND 35, three male piglets from each group were sacrificed for hippocampus and PFC samples used for reference transcriptome sequencing. Following bioinformatics analysis, Gene ontology (GO) enrichment, Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis, and candidate gene screening and pathway were performed for differentially expressed genes. Results: The results showed that a total of 1,632 differential genes were identified in the hippocampus of the MS group, including 1,077 up-regulated differential genes, 555 down-regulated differential genes, and 655 significant GO entries. Analysis of the PFC of the MS group revealed 349 up-regulated genes, 151 down-regulated differential genes, and 584 significant GO entries. Genes associated with neurodevelopment were screened for large fold differences in the hippocampus, and genes associated with cognition were screened for large fold differences in the PFC. Quantitative real-time PCR (qRT-PCR) was used to verify the sequencing data. Western blot (WB) experiments revealed that MS inhibited the neurodevelopment-related WNT signaling pathway in the hippocampus and the cognitive-related PI3K-AKT signaling pathway in the PFC. Discussion: Taken together, these findings suggest that intermittent MS may affect some cognitive functions in piglets by damaging hippocampal and PFC genes or pathways.

Effects of short-distance transportation on physiological indexes, intestinal morphology, microbial community, and the transcriptome of the jejunum in weaned piglets.

Transportation of livestock is unavoidable in animal production. A total of 72 piglets were randomly divided into the CON group and the TSG group, and the piglets in CON group were transported for two hours. The purpose of this study was to determine the effects of short-distance road transportation lasting 2 h on the jejunum of weaned piglets. Our results showed that compared with the control group, there was no impact on the growth performance of piglets in the transport group (P > 0.05). The concentrations of cortisol, heat shock protein (HSP)70, HSP90, C-reactive protein, interleukin (IL)-6, IL-8, IL-12, and interferon-γ and the activity of reactive oxygen species were increased in the jejunum of piglets in the transport group (P < 0.05 compared with the control group). The concentrations of glutathione peroxidase, claudin-1, occludin, and zonula occludens-1 showed no between-group differences (P > 0.05). Regarding intestinal morphology, the transport group showed infiltration of a small amount of lymphocytes into the jejunum mucosa epithelium that was accompanied by edema of the lamina propria, whereas the control group showed no obvious abnormalities. At the genus level, in the transport group, the 16S rRNA sequencing revealed a downward trend in the relative abundance of Lactobacillus and an upward trend in the relative abundance of Muribaculaceae_unclassified. There was also increased mRNA expression of genes associated with inflammation in the transport group, but the genes and pathways related to apoptosis were not activated. In summary, weaned piglets undergoing 2 h of short-distance road transportation showed stress and inflammatory reactions of the jejunum but did not exhibit oxidative damage or activation of the apoptosis pathway of the jejunum. Furthermore, the growth performance of the piglets was not affected by the trip.

Music with Different Tones Affects the Development of Brain Nerves in Mice in Early Life through BDNF and Its Downstream Pathways.

As a means of environmental enrichment, music environment has positive and beneficial effects on biological neural development. Kunming white mice (61 days old) were randomly divided into the control group (group C), the group of D-tone (group D), the group of A-tone (group A) and the group of G-tone (group G). They were given different tonal music stimulation (group A) for 14 consecutive days (2 h/day) to study the effects of tonal music on the neural development of the hippocampus and prefrontal cortex of mice in early life and its molecular mechanisms. The results showed that the number of neurons in the hippocampus and prefrontal cortex of mice increased, with the cell morphology relatively intact. In addition, the number of dendritic spines and the number of dendritic spines per unit length were significantly higher than those in group C, and the expressions of synaptic plasticity proteins (SYP and PSD95) were also significantly elevated over those in group C. Compared with group C, the expression levels of BDNF, TRKB, CREB, PI3K, AKT, GS3Kß, PLCγ1, PKC, DAG, ERK and MAPK genes and proteins in the hippocampus and prefrontal cortex of mice in the music groups were up-regulated, suggesting that different tones of music could regulate neural development through BDNF and its downstream pathways. The enrichment environment of D-tone music is the most suitable tone for promoting the development of brain nerves in early-life mice. Our study provides a basis for screening the optimal tone of neuroplasticity in early-life mice and for the treatment of neurobiology and neurodegenerative diseases.

Effects of early intermittent maternal separation on behavior, physiological, and growth performance in piglets.

In pig production, the management of piglets by batch lactation due to the increase in litter sizes of sows may result in intermittent early neonatal maternal separation (NMS). We speculated that NMS may affect the piglets cognitive growth performance and health. To determine the extent of the effect, 12 litters of crossbred piglets (Large White × Duroc × Min-pig) were used in this trial. Piglets in the control (Con) group (n = 6) were given a standard feeding method during lactation. Piglets in the experimental group (n = 6) were subjected to the NMS model, in which sows were led out of the enclosure with food every day (8:00-11:00 and 13:00-16:00) starting from postnatal day (PND) 7. During the separation, the piglets were supplemented with milk. All experimental piglets were weaned on PND 35. The piglets were observed for aggression, play, mutual sniffing, and exploratory behavior on PNDs 7, 8, 21, 22, 34, 35, 38, 39, 51, 52, 64, and 65. Physiological indicators, namely serum adrenaline, cortisol, interleukin (IL)-1ß, IL-4, IL-6, and tumor necrosis factor (TNF)-α were measured on PNDs 35, 38, and 65, while piglet growth performance was evaluated during suckling and 1 month after weaning. The results showed that aggressive behavior in the MS group was significantly higher than that in the Con group (P < 0.05). Playful and mutual sniffing behaviors in the MS group were significantly lower than those in the Con group except for PNDs 38 and 39 (P < 0.05). Active exploratory behavior in the MS group was significantly higher than that in the Con group on PNDs 7 and 8, and PNDs 21 and 22 (P < 0.05). The frequency of belly-nosing behavior was significantly higher in the MS group than that in the Con group except for PNDs 64 and 65 (P < 0.05). Compared with the Con group, epinephrine, IL-1ß, IL-6, and TNF-α concentrations on PNDs 35, 38, and 65 were significantly increased in the MS group (P < 0.01), while IL-4 concentration was significantly decreased (PND 35: P < 0.05; PNDs 38 and 65: P < 0.01). Compared with the Con group, the piglet diarrhea rate in the MS group during suckling was significantly increased (P < 0.01), the weaning weight was significantly decreased (P < 0.05), and it had no significant effect on the body weight at the end of the trial (P > 0.05). In conclusion, the early intermittent NMS created stress and affected the growth performance of piglets during suckling. However, the growth rate was improved by compensatory measures during late weaning.

Although management methods, such as split-suckling and foster care, in pig production can improve piglet survival rates, these methods inevitably lead to neonatal maternal separation which is an early stress on the body, and can have serious negative effects on the body. In this experiment, we investigated the effect level of neonatal maternal separation on behavior, physiology, and growth performance of piglets. The study found that early intermittent maternal separation leads to anxiety and behavioral changes in piglets, negatively affecting diarrhea rates and weaning weights in suckling piglets, but the effects on growth performance in lactating piglets can be ameliorated during the nursing period.

Selenium Alleviates Ammonia-Induced Splenic Cell Apoptosis and Inflammation by Regulating the Interleukin Family/Death Receptor Axis and Nrf2 Signaling Pathway.

Ammonia (NH3) is a harmful gas in livestock houses. So far, many researchers have demonstrated that NH3 is detrimental to animal and human organs. Selenium (Se) is one of the essential trace elements in the body and has a good antioxidant effect. However, there was little conclusive evidence that Se alleviated NH3 poisoning. To investigate the toxic mechanism of NH3 on pig spleen and the antagonistic effect of L-selenomethionine, a porcine NH3-poisoning model and an L-selenomethionine intervention model were established in this study. Our results showed that NH3 exposure increased the apoptosis rate, while L-selenomethionine supplementation alleviated the process of excessive apoptosis. Immunofluorescence staining, real-time quantitative polymerase chain reaction (qRT-PCR), and western blot results confirmed that exposure to NH3 changed the expression levels of interleukin family factors, apoptosis, death receptor, and oxidative stress factors. Our study further confirmed that excessive NH3 induced inflammatory response and mediated necroptosis leading to cell apoptosis by activating the Nrf2 signaling pathway. Excessive NH3 could mediate spleen injury through oxidative stress-induced mitochondrial dynamics disorder. L-Selenomethionine could alleviate inflammation and abnormal apoptosis by inhibiting the IL-17/TNF-α/FADD axis. Our study would pave the way for comparative medicine and environmental toxicology.

Organic Selenium Alleviates Ammonia-Mediated Abnormal Autophagy by Regulating Inflammatory Pathways and the Keap1/Nrf2 Axis in the Hypothalamus of Finishing Pigs.

Ammonia is a significant pollutant in the livestock houses and the atmospheric environment, and excessive ammonia would harm the health of livestock and breeders. Previous studies have shown that ammonia exposure could damage the tissue structure of the nervous system, but the molecular mechanism of ammonia-induced hypothalamus damage was still unclear. The purpose of this study was to determine the role of excessive ammonia in abnormal autophagy of pig hypothalamus and whether selenomethionine would have a mitigating effect on ammonia toxicity. Twenty-four 18-week pigs were randomly divided into four groups: the control group (C group), the selenium group (Se group), the ammonia + selenium group (A + Se group), and the ammonia group (A group). In our study, the expression levels of NF-κB, IL-1ß, iNOS, TNF-α, IKK-α, p-IKK-α, Nrf2, ATG5, ATG 10, ATG 12, LC3 I/II, HSP60, HSP70, and HSP90 were increased after ammonia exposure; meanwhile, IFN-γ, IKB-α, p-IKB-α, Keap1, P62, mTOR, AKT, p-AKT, PI3K, SQSTM, and Beclin1 showed decreasing trends. The results indicated that excessive ammonia inhalation inhibited the AKT/mTOR pathway to acclerated autophagy through oxidative stress-mediated inflammation in the porcine hypothalamus. L-selenomethionine could alleviate hypothalamus injury induced by ammonia exposure.

Artificial rearing alters intestinal microbiota and induces inflammatory response in piglets.

With the ongoing genetic selection for high prolificacy in sow lines and the improvements in environment and farm management, litter size has increased in recent years. Artificial rearing is becoming widely used to raise the surplus piglets in pig industry. This study aimed to investigate the changes that happened in the morphology, microbiota, mucosal barrier function, and transcriptome caused by artificial rearing in piglet colon. Two hundred and forty newborn piglets were randomly assigned into three treatments, sow rearing until weaning (CON group), artificial rearing from day 21 (AR21 group), and artificial rearing from day 7 (AR7 group). On day 35, the piglets were euthanized to collect colon samples. The results showed that the artificially reared-piglets displayed increased pre-weaning diarrhea incidence and reduced growth performance. Artificial rearing changed the diversity and structure of colonic microbiota and increased relative abundance of harmful bacteria, such as Escherichia-Shigella. In addition, the morphological disruption was observed in AR7 group, which was coincided with decreased tight junction proteins and goblet cell numbers. Moreover, the expression of TNFSF11, TNF-α, IL-1ß, TLR2, TLR4, MyD88, NF-κB, COX-2, PTGEs, iNOS, IL-2, IL-6, IL-17A, and IFN-γ was upregulated in the colon of the artificially reared-piglets, while the expression of IL-1Ra and IκBα was downregulated, indicating that artificial rearing induced inflammatory response through the activation of NF-κB pathway. Furthermore, artificial rearing regulated SLC family members, which affected solute transport and destroyed intestinal homeostasis. In conclusion, artificial rearing caused microbiota alteration, morphology disruption, the destruction of mucosal barrier function, and inflammatory response, and thus, led to subsequent increased diarrhea incidence and reduced growth performance.

Socializing Models During Lactation Alter Colonic Mucosal Gene Expression and Fecal Microbiota of Growing Piglets.

The enrichment of the social environment during lactation alleviates the stress of weaned piglets. It is significant to understand how the enriched social environment improves the weaning stress of piglets. RNA sequencing (RNA-seq) of colonic mucosa, 16S rRNA sequencing of feces, and short-chain fatty acids (SCFAs) of colonic content were used to determine the effects of social contact during lactation. In this study, thirty litter lactating piglets were divided into intermittent social contact (ISC) group that contacted with neighbors intermittently, continuous social contact (CSC) group that contacted with neighbors starting at day (D) 14 after birth, and control (CON) group in which piglets were kept in their original litter. The piglets were weaned at D35 and regrouped at D36. The colonic mucosal RNA-seq, fecal microbes, and SCFAs of colonic contents of 63-day-old piglets were analyzed. The results of RNA-seq showed that compared with the CON group, the pathways of digestion and absorption of minerals, protein, and vitamins of piglets were changed in the ISC group, whereas the pathways of retinol metabolism and nitrogen metabolism in the colonic mucosal were affected and stimulated the immune response in the CSC group. Compared with the CON group, the abundances of pernicious microorganisms (Desulfovibrio, Pseudomonas, Brevundimonas, etc.) in the CSC group and pernicious microorganisms (Desulfovibrio, Neisseria, Sutterella, etc.) and beneficial bacteria (Bifidobacterium, Megamonas, and Prevotella_9) in the ISC group were significantly higher (p < 0.05). The abundances of proinflammatory bacteria (Coriobacteriaceae_unclassified, Coprococcus_3, and Ruminococcus_2) in the CSC group were significantly increased (p < 0.05), but the abundances of SCFAs producing bacteria (Lachnospiraceae_UCG-010, Parabacteroides, Anaerotruncus, etc.) and those of anti-inflammatory bacteria (Eubacterium, Parabacteroides, Ruminiclostridium_9, and Alloprevotella) were significantly reduced (p < 0.05) in the CSC group. Compared with the CON group, the concentrations of microbial metabolites, acetate, and propionate in the colonic contents were reduced (p < 0.05) in the ISC group, whereas the concentration of acetate was reduced (p < 0.05) in the CSC group. Therefore, both ISC and CSC during lactation affected the composition of fecal microbes and changed the expression of intestinal mucosal genes related to nutrient metabolism and absorption of piglets.

The use of selenomethionine to reduce ammonia toxicity in porcine spleen by inhibiting endoplasmic reticulum stress and autophagy mediated by oxidative stress.

Ammonia (NH3) is a typical pollutant in the atmosphere and is well known for its harmful effects on plants, animals as well as human health. Previous studies have shown that NH3 exposure can cause damage to immune organs and impaired immune function in animals. Selenomethionine is a kind of organic selenium, which can not only promote the growth and development of the body, but also inhibit the generation of intracellular reactive oxygen species (ROS), and effectively improve the immune function of the body. Therefore, this study evaluated the toxic effect of NH3 exposure on spleen from a new perspective and investigated the protective effect of selenomethionine on ammonia-induced immunotoxicity. Twenty-four Large White*Duroc*Min pigs were randomly assigned to 4 groups: control group, NH3 group, selenium group, and NH3 + selenium group. Our results showed that NH3 inhalation caused autophagy in the pig spleen, a decrease in lymphocytes, and an increase in autophagic vesicles. Also, NH3 exposure led to a decrease in the activity of some antioxidant enzymes (decreased by about 50%) and a significant increase in the expression of genes related to oxidative stress and endoplasmic reticulum stress (ERS). Our results indicated that selenomethionine mitigated ammonia toxicity in pigs (alleviated about 20-55%). In summary, our findings should be of value in providing a theoretical basis for revealing the toxicity of the high-risk gas NH3, and providing a new perspective on the mechanism of Se against toxic substances.

Transcriptome Revealed Exposure to the Environmental Ammonia Induced Oxidative Stress and Inflammatory Injury in Spleen of Fattening Pigs.

Ammonia is one of the major environmental pollutants that seriously threaten human health. Although many studies have shown that ammonia causes oxidative stress and inflammation in spleen tissue, the mechanism of action is still unclear. In this study, the ammonia poisoning model of fattening pigs was successfully established. We examined the morphological changes and antioxidant functions of fattening pig spleen after 30-day exposure to ammonia. Effects of ammonia in the fattening pig spleen were analyzed from the perspective of oxidative stress, inflammation, and histone methylation via transcriptome sequencing technology (RNA-seq) and real-time quantitative PCR validation (qRT-PCR). We obtained 340 differential expression genes (DEGs) by RNA-seq. Compared with the control group, 244 genes were significantly upregulated, and 96 genes were significantly downregulated in the ammonia gas group. Some genes in Gene Ontology (GO) terms were verified and showed significant differences by qRT-PCR. The KEGG pathway revealed significant changes in the MAPK signaling pathway, which is strongly associated with inflammatory injury. To sum up, the results indicated that ammonia induces oxidative stress in pig spleen, activates the MAPK signaling pathway, and causes spleen necrosis and injury. In addition, some differential genes encoding epigenetic factors were found, which may be involved in the response mechanism of spleen tissue oxidative damage. The present study provides a transcriptome database of ammonia-induced spleen poisoning, providing a reference for risk assessment and comparative medicine of ammonia.

Stereotypic behaviors are associated with physiology and immunity differences in long-term confined sows.

Pregnant sows in the confined environment have poor welfare and frequently perform stereotypic behaviors. In order to clarify whether highly stereotypic behavior is a sign of increased stress and successfully contributes to coping with or adaptation to adverse environment, fifty pregnant sows (Large White × Landrace) housed in stalls were selected to observe behaviors and analyze physiological parameters [cortisol, major acute phase protein (Pig-MAP) and C-reactive protein (CRP)], and immunological parameters [immunoglobin A (IgA), immunoglobin G (IgG), interleukin 6 (IL-6), interleukin 10 (IL-10), tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ)] in early, middle and late gestation (27th, 62nd and 91st day). A repeated-measures analysis and Friedman test was performed to analyze the differences of behaviors and physiological and immunological parameters. The results showed that lateral lying behavior increased significantly with the progress of sows' gestation, while standing and ventral lying behaviors significantly decreased (p < 0.05). Sham-chewing, bar-biting, trough-biting and rooting behaviors significantly reduced (p < 0.05). Furthermore, there was no significant difference in physiological and immune levels in different gestational periods (p > 0.05). The results also indicated that sham-chewing behavior was positively correlated with serum cortisol, IL-6, IL-10, and negatively correlated with serum IFN-γ in each gestational period (p < 0.05). Trough-biting behavior was positively correlated with serum TNF-α in middle and late gestation (p < 0.05). Rooting behavior was positively correlated with serum IgG in each gestational period, and positively correlated with serum Pig-MAP, IL-6, and IL-10 in middle and late gestation (p < 0.05). In conclusion, the sows with a high incidence of stereotypic behaviors tried to improve stress and humoral immunity to cope with the confined environment, and long-term confined sows might be in a chronic stress state.

Cadmium induces testosterone synthesis disorder by testicular cell damage via TLR4/MAPK/NF-κB signaling pathway leading to reduced sexual behavior in piglets.

Cadmium (Cd) is a highly toxic metal pollutant that can endanger the life and health of animals. Toll-like receptor 4 (TLR4) can result in testicular cell damage by positively regulating mitogen-activated protein kinase (MAPK)/nuclear factor-kappaB (NF-κB) signaling pathway. Meanwhile, Testosterone (T) synthesis disorder can affect sexual behavior. However, the harmful influence of Cd on animal sexual behavior during its growth and development and the role of TLR4/MAPK/NF-κB signaling pathway in testicular cell damage and testosterone production remained poorly understood. Forty-two-day-old male piglets were fed with diets that contained CdCl2 (20 mg Cd/kg) for 40 days to explore the toxic effects of Cd on sexual behavior. The results showed that Cd activated TLR4, promoted MAPK (p-ERK, p-JNK, and p-p38)/NF-κB expression, induced apoptosis (Caspase-3, Cleaved Caspase3, Bax, Cyt-c, and Caspase-9 expression increased, but Bcl-2 expression decreased) and necroptosis (MLKL, RIPK1, and RIPK3 expression increased) in piglet testis. In addition, Cd exposure decreased mRNA expression of STAR, CYP11A1, 3ß-HSD, CYP17A1, and 17ß-HSD of testis and the concentrations of T and thyroid-stimulating hormone (TSH). Both the mRNA and protein expression levels of the major genes in TLR4/MAPK/NF-κB signaling pathway, apoptosis signaling pathway, and necroptosis signaling pathway increased significantly and the expression levels of testosterone decreased gradually in pig Leydig cells cultured in vitro after being treated with different concentrations of Cd. Moreover, Cd reduced sexual behavior (the parameters of sniffing, chin resting, and mounting decreased) in piglets. In conclusion, Cd induced testicular cell damage via TLR4/MAPK/NF-κB signaling pathway leading to testosterone synthesis disorder and sexual behavior reduction in piglets.

Evaluation of potential cardiotoxicity of ammonia: l-selenomethionine inhibits ammonia-induced cardiac autophagy by activating the PI3K/AKT/mTOR signaling pathway.

Ammonia is a major harmful gas in the environment of livestock and poultry. Studies have shown that excessive ammonia inhalation has adverse effects in pig heart. However, the mechanism of ammonia-induced cardiac toxicity in pigs has not been reported. L-selenomethionine is a kind of organic selenium (Se) which is easily absorbed by the body. Therefore, in this study, twenty-four 125-day-old pigs were randomly divided into 4 groups: C (control) group, A (ammonia) group, Se group (Se content: 0.5 mg kg-1), and A (ammonia) + Se group. The mechanism of ammonia-induced cardiotoxicity and the alleviating effect of L-selenomethionine were examined. The results in the A group showed as follows: a large number of myocardial fiber edema and cytoplasmic bleakness were observed in the heart; a large number of mitochondrial autophagy were observed; ATP content, ATPase activities and hematological parameters decreased significantly; Endoplasmic reticulum stress (ERS) markers (GRP78, IRE1α, ATF4, ATF6, and CHOP) were significantly induced in the mRNA and protein levels; PI3K/AKT/mTOR signaling pathway was activated; and autophagy key genes and proteins (Beclin-1, LC3, ATG3, and ATG5) were significantly up-regulated. The results of comparison between the A + Se group and the A group were as follows: the degree of edema of cardiac muscle fiber in the A + Se group was somewhat relieved; the level of mitochondrial autophagy decreased; ATP content and ATPase activities increased significantly; the mRNA and protein levels of ERS markers were significantly down-regulated; the expression level of PI3K/AKT/mTOR signaling pathway was decreased; and the mRNA and protein levels of key autophagy genes were decreased. However, the changes of these indexes in the A + Se group were still significantly different from those in the C group. Our results indicated that L-selenomethionine supplementation inhibited ammonia-induced cardiac autophagy by activating the PI3K/AKT/mTOR signaling pathway, which confirmed that L-selenomethionine could alleviate the cardiac injury caused by excessive ammonia inhalation to a certain extent. This study aims to enrich the toxicological mechanism of ammonia and provide valuable reference for future intervention of ammonia toxicity.

Cadmium exposure activates the PI3K/AKT signaling pathway through miRNA-21, induces an increase in M1 polarization of macrophages, and leads to fibrosis of pig liver tissue.

Cadmium (Cd) is a toxic substance that pollutes the environment with multiple organs. Long-term exposure to Cd can cause fibrosis in the lungs and other organs of animal body. This article explored the effects of subacute Cd exposure on pig liver fibrosis, as well as the polarization of microRNA (miRNA) and M1/M2 macrophages during this process. Based on the establishment of the pig subacute CdCl2 exposure model, we used immunofluorescence staining, Masson staining, qRT-PCR and western blotting to conduct further research. The results showed that Cd exposure can increase the expression of miRNA-21, decrease the expression of TGF-ß and SMAD7, increase the expression of PI3K/AKT signaling pathway, cause the M1/M2 imbalance and the increase of M1 polarization. Meantime, it causes the secretion of inflammatory cytokines (TNF-α, IL-1ß, and IL-6), and causes an imbalance in the expression of TIMP1, MMP2, and MMP9, which are related to the degree of fibrosis. And the expression of α-SMA, COL1 and COL3 were up-regulated. In the pig, these results indicate that liver fibrosis caused by subacute CdCl2 exposure is induced by the M1 polarization of macrophages through the PI3K/AKT signaling pathway activated by miRNA-21 signaling pathway. These research results not only enrich the theoretical basis and reference value of Cd toxicology research, but also provide new references and new research ideas for comparative medicine.

Effects of fermented wheat bran and yeast culture on growth performance, immunity, and intestinal microflora in growing-finishing pigs.

This study was conducted to evaluate the effects of feeding fermented wheat bran (FWB) and yeast culture (YC) on growth performance, immune levels, and intestinal microflora in growing-finishing pigs. In total, 96 crossbred pigs were randomly distributed into four treatments with four replicates pens and six pigs per pen. This study was performed using a 2 × 2 factor design: 1) CON (basal diet), 2) FWB (basal diet + 5% FWB), 3) YC (basal diet + 2% YC), and 4) FWB + YC (basal diet + 5% FWB + 2% YC). Dietary FWB supplementation significantly increased the average daily gain and significantly decreased the feed gain ratio of growing-finishing pigs (P < 0.05). Supplementation of FWB and YC improved the immune capacity and reduced the inflammation level of growing-finishing pigs (P < 0.05). In addition, pigs fed FWB, YC, and FWB + YC diets showed better intestinal development and morphology compared with those CON pigs. The relative abundance of Streptococcus in the FWB group was significantly lower than that in the CON group (P < 0.05), and the relative abundance of probiotics (unclassified_f_Lachnospiraceae, Turicibacter) increased significantly (P < 0.05). Furthermore, the relative abundance of probiotics (Lactobacillus, norank_f_Muribaculaceae) in the YC group was significantly increased compared with the CON group (P < 0.05). The results of this study observed positive effects of FWB and YC on growing-finishing pigs, which provides insights into the application of biological feed in swine industry.

Selenomethionine protects against ammonia-induced apoptosis through inhibition of endoplasmic reticulum stress in pig kidneys.

Ammonia (NH3) emission is a common threat to farm animals. Selenium (Se) is known for its antioxidant property and can resist several stressors affecting farm animals. The aims of this study were (Ⅰ) to determine how excess NH3 exert nephrotoxic effects in pigs and (Ⅱ) to investigate whether selenomethionine has an alleviative effect on NH3 toxicity. Two diets supplemented with different doses of Se (0.22 mg/kg or 0.50 mg/kg) and two concentrations of NH3 (< 5 mg/m3 or 89.8 mg/m3) were used in a 2 × 2 factorial design trial for a period of 30 days. The results showed that NH3 exposure caused apoptosis and increased the number of apoptotic cells in pig kidneys. Further, the activities of antioxidant enzymes were decreased, and the transcriptional and translational levels of endoplasmic reticulum stress-related genes, Bcl-2 and Caspase family members were increased under NH3 exposure. In addition, Wnt/ß-catenin signaling pathway was suppressed after NH3 treatment. Dietary supplement with selenomethionine appears to offer protection against NH3-induced kidney injury in pigs and the pathologic changes above were alleviated. Our findings provide additional insight into the mechanism of NH3 toxicity in pigs while elucidating the role of Se as a potential antidote against NH3 poisoning.

Subacute cadmium exposure promotes M1 macrophage polarization through oxidative stress-evoked inflammatory response and induces porcine adrenal fibrosis.

Cadmium (Cd) is a widely distributed environmental pollutant with immunotoxicity and endocrine toxicity. M1/M2 macrophages participate in the immune response and exert an essential influence on fibrosis. Nevertheless, whether Cd can induce porcineadrenal fibrosis by affecting the polarization of M1/M2 macrophages and its potential regulatory mechanism have not been explored. We added 20 mg/kg CdCl2 to the pig diet for 40 days to investigate the fibrogenic effect of subacute Cd exposure on the adrenal gland. The results indicated that the ACTH and CORT in serum were decreased by 15.26 % and 21.99 %, respectively. The contents of adrenal mineral elements Cd, Cr, Mn were increased up to 34, 1.93, 1.42 folds and Co, Zn, Sn were reduced by 21.57 %, 20.52 %, 15.75 %. Concurrently, the pro-oxidative indicators (LPO, MDA and H2O2) were increased by 1.85, 2.20, 2.77 folds and 3.60, 11.15, 4.11 folds upregulated mRNA levels of TLR4, NF-κB, NLRP3 were observed. Subsequently, the expression of M1 macrophages polarization markers (IL-6, iNOS, TNF-α, CCL2 and CXCL9) were raised by 2.03, 2.30, 2.35, 1.58, 1.56 folds, while M2 macrophages (IL-4, CCL24, Arg1, IL-10, MRC1) showed a 62.34 %, 31.88 %, 50.26 %, 74.00 %, 69.34 % downregulation. The expression levels of AMPK subunits and genes related to glycolysis, oxidative phosphorylation (OXPHOS) and fatty acid oxidation (FAO) were also markedly increased. Additionally, the expression level of TGF-ß1, Smad2/3 and downstream pro-fibrotic markers was obviously upregulated. Taken together, we conclude that Cd activates the oxidative stress-mediated TLR4/NF-κB/NLRP3 inflammatory signal transduction, leading to porcine adrenal fibrosis by promoting macrophage polarization toward M1.

Immunosuppression, oxidative stress, and apoptosis in pig kidney caused by ammonia: Application of transcriptome analysis in risk assessment of ammonia exposure.

Ammonia (NH3) is a recognized environmental contaminant around the world and has adverse effects on animal and human health. However, the mechanism of the renal toxicity of NH3 is not well understood. Pigs are considered an ideal model for biomedical and toxicological research because of the similarity to humans in physiological and biochemical basis. Therefore, in this study, twelve pigs were selected as research objects and randomly divided into two groups, namely the control group and the NH3 group. The formal experiment lasted 30 days. The effects of excessive NH3 inhalation on the kidney of fattening pig were evaluated by chemical analysis, ELISA, transcriptome analysis and real-time quantitative PCR (qRT-PCR) from the renal antioxidant level, renal function, blood ammonia content and gene level. Our results showed that excessive NH3 exposure could cause an increase in blood NH3 content, a reduction in renal GSH-Px, SOD and GSH, as well as an increase in MDA levels and an increase in serum creatinine, urea and uric acid levels. In addition, transcriptome analysis showed that NH3 exposure caused changes in 335 differentially expressed genes (DEGs) (including 126 up-regulated DEGs and 109 down-regulated DEGs). Some highly expressed DEGs were enriched into GO terms associated with immune function, oxidative stress, and apoptosis and were verified by qRT-PCR. The qRT-PCR results were comsistent with the transcriptome results. Our results indicated that NH3 exposure could cause changes in renal transcriptional profiles and kidney function, and induce kidney damage in the fattening pigs through oxidative stress, immune dysfunction and apoptosis. Our present study provides novel insights into the immunotoxicity mechanism of NH3 on kidney.