LAMTOR1/mTORC1 promotes CD276 to induce immunosuppression via PI3K/Akt/MMP signaling pathway in Clostridium perfringens-induced necrotic enteritis of laying hens.

Clostridium perfringens (C. perfringens) causes avian necrotic enteritis, leading to huge economic losses to the poultry industry. This pathogen induces host immunosuppression; however, the molecular mechanism is still unclear. Thus, we established a laying hen infection model to explore this mechanism. We randomly divided 20 one-old-day laying hens into the control and infection groups. The infection group was infected intragastrically with 1 × 109 colony-forming units of C. perfringens in 1 mL of sterile phosphate-buffered saline (PBS) once a day from d 17 to 20; the control group received the same volume of PBS without the bacterium. Twenty-four hours after the last challenge, we sacrificed the laying hens and collected the jejunum for analysis. The infection group presented alterations in blood biochemical parameters and necrotic lesion scores as well as damage to the jejunum. Proteomics revealed 427 upregulated and 291 downregulated proteins in the infection group. In the infection group, CD3, CD4, and CD8 messenger RNA expression (mRNA) expression was decreased; LAMTOR1 and mTORC1 mRNA expression was increased; CD276 protein expression was enhanced; and the PI3K/Akt/MMP pathway was activated in jejunum of laying hens. This is the first study to report CD276 expression in the jejunum related to immunosuppression in a laying hen model of necrotic enteritis. It provides some new key targets to potentially control avian necrotic enteritis.

Baicalin-aluminum complex on the regulation of IPEC-1 infected with enterotoxigenic Escherichia coli.

Enterotoxigenic Escherichia coli (ETEC) is the main bacterial cause of diarrhea in weaned piglets. Baicalin-aluminum (BA) complex is the main active ingredient of Scutellaria baicalensis Georgi extracted-aluminum complex, which has been used to treat diarrhea in weaning piglets, however the underlying mechanism remains unclear. To investigate the effects of the BA complex on the regulation of porcine intestinal epithelial (IPEC-1) cells infected with ETEC, IPEC-1 cells were incubated with an ETEC bacterial strain at a multiplicity of infection of 1 for 6 h and then treated with different concentrations of the BA complex for 6 h. ETEC infection increased the levels of cAMP and cGMP, upregulated CFTR (cystic fibrosis transmembrane conductance regulator) mRNA, and downregulated NHE4 mRNA in IPEC-1 cells. Treatment with the BA complex inhibited ETEC adhesion and the production of cAMP and cGMP, reduced CFTR mRNA expression, and increased NHE4 mRNA expression. Overall, the BA complex weakened the adhesion of ETEC to IPEC-1 cells, and inhibited cAMP/cGMP-CFTR signaling in IPEC-1 cells.

Anemoside B4 attenuates necrotic enteritis of laying hens induced by Clostridium perfringens via inhibiting NF-κB and PI3K/Akt/mTOR signalling pathways.

Poultry necrotic enteritis is an important enteric disease which might be controlled by antibiotics. However, with the excessive use of antibiotics, the phenomenon of drug resistance of Clostridium perfringens is becoming increasingly prominent. Anemoside B4 exhibits important anti-inflammatory, antioxidant and immunomodulatory effects. This study was performed to estimate the effect of Anemoside B4 on chicken necrotic enteritis induced by C. perfringens in vivo and in vitro. In the in vivo experiment we investigated the efficacy of Anemoside B4 on the growth curve, biofilm formation, haemolytic activity, virulence-related gene expression and NF-κB and PI3K/AKT/mTOR activation in Caco-2 cells induced by C. perfringens. The results showed that 12.5-50 µg/mL Anemoside B4 had no antibacterial activity but could inhibit biofilm formation, attenuate haemolytic activity and virulence-related gene expression of C. perfringens and weaken NF-κB and PI3K/Akt/mTOR activation triggered by C. perfringens in Caco-2 cells. In the in vivo experiment, 60 17-day-old healthy White Leghorns were randomly divided into six groups. The growing laying hens of the control group were fed a basic diet, and those of the five challenged groups were fed a basic diet (infection group), added 0.43 g/kg Anemoside B4 (0.43 g/kg Ane group), 0.86 g/kg Anemoside B4 (0.86 g/kg Ane group), 1.72 g/kg Anemoside B4 (1.72 g/kg Ane group) and 40 mg/kg lincomycin (lincomycin group), respectively. All challenged laying hens were infected with 1 × 109 CFU C. perfringens from day 17-20. Blood and intestinal samples were obtained, and the data demonstrated that Anemoside B4 improved the blood biochemical parameters, attenuated jejunum tissue injury, increased the spleen, thymus, bursa of fabricius index, and decreased lesion scores of the jejunum and the ileum. In the jejunum, Anemoside B4 and lincomycin downregulated the expression of IL-1ß, IL-6, IL-10, TNF-α and IFN-γ at mRNA levels. Moreover, Anemoside B4 significantly enhanced both mRNA and protein levels of tight junctions ZO-1, Claudin-1 and MUC-2 in the jejunum. Anemoside B4 weakened p-P65, p-PI3K, p-Akt and p-mTOR protein expression in the jejunum infected by C. perfringens. Diets supplemented with Anemoside B4 alleviated C. perfringens-induced necrotic enteritis in laying hens by inhibiting NF-κB and PI3K/Akt/mTOR signalling pathways and improving intestinal barrier functions.

Baicalin and probenecid protect against Glaesserella parasuis challenge in a piglet model.

Glaesserella parasuis (G. parasuis) induces vascular damage and systemic inflammation. However, the mechanism by which it causes vascular damage is currently unclear. Baicalin has important anti-inflammatory, antibacterial and immunomodulatory functions. In this study, we explored the ability of baicalin and probenecid to protect against G. parasuis challenge in a piglet model. Sixty piglets were randomly divided into a control group; an infection group; a probenecid group; and 25 mg/kg, 50 mg/kg and 100 mg/kg baicalin groups. The probenecid group and the 25 mg/kg, 50 mg/kg and 100 mg/kg baicalin groups were injected intramuscularly with 20 mg/kg body weight (BW) probenecid and 25 mg/kg BW, 50 mg/kg BW and 100 mg/kg BW baicalin, respectively. All piglets except those from the control group were injected intraperitoneally with 1 × 108 CFU of G. parasuis. The control group was injected intraperitoneally with TSB. The results showed baicalin and probenecid protected piglets against G. parasuis challenge, improved body weight and decreased temperature changes in piglets. Baicalin and probenecid attenuated IL-1ß, IL-10, IL-18, TNF-α and IFN-γ mRNA levels in the blood for 48 h, inhibited the production of the nucleosides ATP, ADP, AMP and UMP from 24 to 72 h, reduced Panx-1/P2Y6/P2X7 expression, weakened NF-kB, AP-1, NLRP3/Caspase-1 and ROCK/MLCK/MLC signalling activation, and upregulated VE-cadherin expression in the blood vessels of piglets challenged with G. parasuis. Baicalin and probenecid alleviated pathological tissue damage in piglets induced by G. parasuis. Our results might provide a promising strategy to control and treat G. parasuis infection in the clinical setting.

Baicalin attenuates PD-1/PD-L1 axis-induced immunosuppression in piglets challenged with Glaesserella parasuis by inhibiting the PI3K/Akt/mTOR and RAS/MEK/ERK signalling pathways.

Infection of piglets with Glaesserella parasuis (G. parasuis) induces host immunosuppression. However, the mechanism underlying the immunosuppression of piglets remains unclear. Activation of the PD-1/PD-L1 axis has been shown to trigger host immunosuppression. Baicalin possesses anti-inflammatory and immunomodulatory functions. However, whether baicalin inhibits PD-1/PD-L1 activation and thus alleviates host immunosuppression has not been investigated. In this study, the effect of baicalin on the attenuation of piglet immunosuppression induced by G. parasuis was evaluated. Seventy piglets were randomly divided into the control group, infection group, levamisole group, BMS-1 group, 25 mg/kg baicalin group, 50 mg/kg baicalin group and 100 mg/kg baicalin group. Following pretreatment with levamisole, BMS-1 or baicalin, the piglets were challenged with 1 × 108 CFU of G. parasuis. Our results showed that baicalin, levamisole and BMS-1 modified routine blood indicators and biochemical parameters; downregulated IL-1ß, IL-10, IL-18, TNF-α and IFN-γ mRNA expression; and upregulated IL-2 and IL-8 mRNA expression in blood. Baicalin, levamisole and BMS-1 increased the proportions of CD3+ T cells, CD3+CD4+ T cells, CD3+CD8+ T cells and CD3-CD21+ B cells in the splenocyte population, increased the proportions of CD3+ T cells, CD3+CD4+ T cells and CD3+CD8+ T cells in the blood, and inhibited PD-1/PD-L1 and TIM-3 activation. Baicalin, levamisole and BMS-1 reduced p-PI3K, p-Akt, and p-mTOR expression, the p-MEK1/2/MEK1/2 and p-ERK1/2/ERK1/2 ratios and increased RAS expression. Baicalin, levamisole and BMS-1 provided substantial protection against G. parasuis challenge and relieved tissue histopathological damage. Our findings might provide new strategies for controlling G. parasuis infection and other immunosuppressive diseases.

Quercetin Protects Blood-Brain Barrier Integrity via the PI3K/Akt/Erk Signaling Pathway in a Mouse Model of Meningitis Induced by Glaesserella parasuis.

Glaesserella parasuis (G. parasuis) causes serious inflammation and meningitis in piglets. Quercetin has anti-inflammatory and anti-bacterial activities; however, whether quercetin can alleviate brain inflammation and provide protective effects during G. parasuis infection has not been studied. Here, we established a mouse model of G. parasuis infection in vivo and in vitro to investigate transcriptome changes in the mouse cerebrum and determine the protective effects of quercetin on brain inflammation and blood-brain barrier (BBB) integrity during G. parasuis infection. The results showed that G. parasuis induced brain inflammation, destroyed BBB integrity, and suppressed PI3K/Akt/Erk signaling-pathway activation in mice. Quercetin decreased the expression of inflammatory cytokines (Il-18, Il-6, Il-8, and Tnf-α) and BBB-permeability marker genes (Mmp9, Vegf, Ang-2, and Et-1), increased the expression of angiogenetic genes (Sema4D and PlexinB1), reduced G. parasuis-induced tight junction disruption, and reactivated G. parasuis-induced suppression of the PI3K/Akt/Erk signaling pathway in vitro. Thus, we concluded that quercetin may protect BBB integrity via the PI3K/Akt/Erk signaling pathway during G. parasuis infection. This was the first attempt to explore the protective effects of quercetin on brain inflammation and BBB integrity in a G. parasuis-infected mouse model. Our findings indicated that quercetin is a promising natural agent for the prevention and treatment of G. parasuis infection.

Baicalin attenuates aflatoxin B1-induced hepatotoxicity via suppressing c-Jun-N-terminal kinase-mediated cell apoptosis.

Aflatoxin B1 (AFB1) is classified as a Class I carcinogen and common pollutant in human and animal food products. Prolonged exposure to AFB1 can induce hepatocyte apoptosis and lead to hepatotoxicity. Therefore, preventing AFB1-induced hepatotoxicity remains a critical issue and is of great significance. Baicalin, a polyphenolic compound derived from Scutellaria baicalensis Georgi, has a variety of pharmacodynamic activities, such as antiapoptotic and anticancer activities. This study systematically investigated the alleviating effect of baicalin on AFB1-induced hepatotoxicity from the perspective of apoptosis and explored the possible molecular mechanism. In the normal human liver cell line L02, baicalin treatment significantly inhibited AFB1-induced c-Jun-N-terminal Kinase (JNK) activation and cell apoptosis. In addition, the in vitro mechanism study demonstrated that baicalin alleviates AFB1-induced hepatocyte apoptosis through suppressing the translocation of phosphorylated JNK to the nucleus and decreasing the phosphorylated c-Jun/c-Jun ratio and the Bax/Bcl2 ratio. Molecular docking and drug affinity responsive target stability assays demonstrated that baicalin has the potential to target JNK. This study provides a basis for the therapeutic effect of baicalin on hepatocyte apoptosis caused by AFB1, indicating that the development of baicalin and JNK pathway inhibitors has broad application prospects in the prevention of hepatotoxicity, especially hepatocyte apoptosis.

Preparing a novel baicalin-loaded microemulsion-based gel for transdermal delivery and testing its anti-gout effect.

We previously demonstrated that baicalin had efficacy against gouty arthritis (GA) by oral administration. In this paper, a novel baicalin-loaded microemulsion-based gel (B-MEG) was prepared and assessed for the transdermal delivery of baicalin against GA. The preparation method and transdermal capability of B-MEG was screened and optimized using the central composite design, Franz diffusion cell experiments, and the split-split plot design. Skin irritation tests were performed in guinea pigs. The anti-gout effects were evaluated using mice. The optimized B-MEG comprised of 50 % pH 7.4 phosphate buffered saline, 4.48 % ethyl oleate, 31.64 % tween 80, 13.88 % glycerin, 2 % borneol, 0.5 % clove oil and 0.5 % xanthan gum, with a baicalin content of (10.42 ± 0.08) mg/g and particle size of (15.71 ± 0.41) nm. After 12 h, the cumulative amount of baicalin permeated from B-MEG was (672.14 ± 44.11) µg·cm-2. No significant skin irritation was observed following B-MEG application. Compared to the model group, B-MEG groups significantly decreased the rate of auricular swelling (P < 0.01) and number of twists observed in mice (P < 0.01); and also reduced the rate of paw swelling (P < 0.01) and inflammatory cell infiltration in a mouse model of GA. In conclusion, B-MEG represents a promising transdermal carrier for baicalin delivery and can be used as a potential therapy for GA.

PD-1/PD-L1 axis induced host immunosuppression via PI3K/Akt/mTOR signalling pathway in piglets infected by Glaesserella Parasuis.

Glaesserella parasuis, an important respiratory bacterial pathogen, causes Glässer's disease in piglets, with potential immunosuppression. We established a piglet infection model and explored the immunosuppression mechanism to improve our understanding of the host immune response to G. parasuis. Twenty piglets were randomly divided into two groups (n = 10). The infection group was intraperitoneally challenged with 2 × 108 CFU of G. parasuis in 2 mL TSB. The control group was intraperitoneally injected with equivalent TSB. After 72 h, the piglets were sacrificed, and spleen tissue was collected. PD-1/PD-L1 expression was determined. The splenocytes were isolated to detect CD3+ T, CD3+CD4+ T, CD3+CD8+ T and CD3-CD21+cell differentiation. Via data-independent acquisition (DIA), we compared the proteomics of healthy and infected spleen tissues. Glaesserella parasuis modified CD3+ T, CD3+CD4+ T, CD3+CD8+ T and CD3-CD21+ cell differentiation and PD-1/PD-L1 expression in the spleen. The infection group had 596 proteins with significant differences in expression, of which 301 were significantly upregulated and 295 downregulated. Differentially expressed proteins (DEPs) were mainly related to immune responses. This is the first study on PD-1/PD-L1 expression in the spleen associated with immunosuppression in a piglet model to explore the protein changes related to immune responses via DIA.

Baicalin inhibits monosodium urate crystal-induced pyroptosis in renal tubular epithelial cell line through Panx-1/P2X7 pathways: Molecular docking, molecular dynamics, and in vitro experiments.

Pyroptosis is a programmed cell death process that frequently occurs in many diseases, including hyperuricemic nephropathy (HN). In HN, a range of stimuli mediates inflammation, leading to the activation of inflammasomes and the production of gasdermin D (GSDMD). Baicalin (BA), a natural flavonoid renowned for its antioxidant and anti-inflammatory properties, was investigated for its role in HN in this study. Initially, HN-like inflammation and pyroptosis were induced in HK-2 cells with treatment of monosodium urate (MSU), followed by the BA treatment. The expression of pyroptosis-associated genes, Panx-1 and P2X7, at both mRNA and protein levels was assessed through real-time polymerase chain reaction (RT-qPCR) and Western blotting (WB) without or with BA treatment. The results showed that expression of Panx-1 and P2X7 at mRNA and protein levels was increased in MSU-treated HK-2 cells, which subsequently decreased upon the BA treatment. Further experiments showed that BA could combine NLRP3 inflammasome and GSDMD, destabilizing GSDMD protein. Moreover, BA protected the cell membrane from MSU-induced damage, as evidenced by Hoechst 33342 and PI double staining, lactate dehydrogenase (LDH) assays, and electron microscopy observations. These results suggest that BA is involved in the regulating Panx-1/P2X7 pathways and thus inhibits pyroptosis, highlighting its potential therapeutic effect for HN.

Baicalin Weakens the Virulence of Porcine Extraintestinal Pathogenic Escherichia coli by Inhibiting the LuxS/AI-2 Quorum-Sensing System.

Porcine extraintestinal pathogenic Escherichia coli (ExPEC) is a pathogenic bacterium that causes huge economic losses to the pig farming industry and considerably threatens human health. The quorum sensing (QS) system plays a crucial role in the survival and pathogenesis of pathogenic bacteria. Hence, it is a viable approach to prevent ExPEC infection by compromising the QS system, particularly the LuxS/AI-2 system. In this study, we investigated the effects of baicalin on the LuxS/AI-2 system of ExPEC. Baicalin at concentrations of 25, 50, and 100 µg/mL significantly diminished the survival ability of ExPEC in hostile environments and could inhibit the biofilm formation and autoagglutination ability in ExPEC. Moreover, baicalin dose-dependently decreased the production of AI-2 and down-regulated the expression level of luxS in PCN033. These results suggest that baicalin can weaken the virulence of PCN033 by inhibiting the LuxS/AI-2 system. After the gene luxS was deleted, AI-2 production in PCN033 was almost completely eliminated, similar to the effect of baicalin on the production of AI-2 in PCN033. This indicates that baicalin reduced the production of AI-2 by inhibiting the expression level of luxS in ExPEC. In addition, the animal experiment further showed the potential of baicalin as a LuxS/AI-2 system inhibitor to prevent ExPEC infection. This study highlights the potential of baicalin as a natural quorum-sensing inhibitor for therapeutic applications in preventing ExPEC infection by targeting the LuxS/AI-2 system.

Using network pharmacology and molecular docking to uncover the mechanism by which quercetin alleviates deoxynivalenol-induced porcine intestinal injury.

Deoxynivalenol is a widespread feed contaminant that leads to vomit, which results in serious symptom such as increased intestinal permeability and even intestinal mucosal necrosis. Recent studies have reported the role of quercetin in alleviating deoxynivalenol-induced intestinal injury; however, the mechanisms and targets remain unclear. Thus, we aimed to identify the mechanisms of action by using a combination of network pharmacology and molecular docking. We identified 151 quercetin targets, 235 deoxynivalenol targets and 47 porcine intestinal injury targets by searching compound database and PubMed database, among which there were two common targets. The PPI network showed that the key proteins involved are NQO1 and PPAR-γ. The PPI network showed that the key proteins involved were NQO1 and PPARG. GO analysis found that genes were enriched primarily in response to oxidative stress. The PPI network showed that the key proteins involved are NQO1 and PPAR-γ. The genes are enriched primarily in response to oxidative stress. KEGG analysis showed enrichment of the HIF, reactive oxygen species and other signaling pathways. The molecular docking results indicated key binding activity between NQO1-quercetin and PPAR-γ-quercetin. By using network pharmacology, we have revealed the potential molecular mechanisms by which quercetin alleviates deoxynivalenol-induced porcine intestinal injury, which lays the foundation for the development of drugs to treat deoxynivalenol-induced intestinal injury in pigs.

Baicalin attenuates lipopolysaccharide-induced intestinal inflammatory injury via suppressing PARP1-mediated NF-κB and NLRP3 signalling pathway.

Bacterial lipopolysaccharide (LPS) exposure is a key inducer of intestinal inflammatory injury in weaned piglets, resulting in decreased growth performance of pigs and causing severe economic losses to the swine industry; however, the mechanism of intestinal inflammatory injury is still unclear. Baicalin is one of the main active ingredients extracted from the natural plant Scutellaria baicalensis that has biological functions, including anti-inflammatory activity. The aim of this study is to investigate the effect and mechanism of baicalin intervention on intestinal inflammatory injury caused by bacterial LPS exposure. In the present study, network pharmacology, molecular docking and DARTS results identified that baicalin has the potential to target PARP1, thereby potentially regulating a series of inflammation-related pathways, including the MAPK, NF-κB and Toll-like receptor signalling pathways, which play the role of antagonizing LPS-induced intestinal inflammatory injury. Further application of the LPS-induced IPEC-J2 cell model validated the finding that baicalin could alleviate LPS-induced intestinal inflammatory injury by inhibiting the PARP1-mediated NF-κB and NLRP3 signalling pathway. These findings demonstrate that baicalin can regulate the expression of PARP1 and that PARP1 has the potential to serve as an effective therapeutic target in the LPS-induced intestinal inflammatory injury.

Baicalin inhibited PANX-1/P2Y6 signaling pathway activation in porcine aortic vascular endothelial cells infected by Glaesserella parasuis.

Glaesserella parasuis can induce endothelial barrier damage in piglets, although the mechanism by which this pathogen triggers inflammatory damage remains unclear. Baicalin possesses anti-inflammatory and anti-oxidant activities. However, whether baicalin can relieve endothelial barrier damage caused by Glaesserella parasuis infection has not yet been studied. Hence, we evaluated the ability of baicalin to counteract the changes induced by Glaesserella parasuis in porcine aortic vascular endothelial cells. The results showed that Glaesserella parasuis could upregulate the expression of pannexin 1 channel protein and promote the release of adenosine triphosphate, adenosine diphosphate, adenosine 3'-monophosphate, uridine triphosphate, uridine diphosphate, and uridine monophosphate in porcine aortic vascular endothelial cells. The expression level of purinergic receptor P2Y6 was upregulated in porcine aortic vascular endothelial cells triggered by Glaesserella parasuis. In addition, Glaesserella parasuis could activate phospholipase C-protein kinase C and myosin light chain kinase-myosin light chain signaling pathways in porcine aortic vascular endothelial cells. Baicalin could inhibit pannexin 1 channel protein expression, reduce adenosine triphosphate, adenosine diphosphate, adenosine 3'-monophosphate, uridine triphosphate, uridine diphosphate, and uridine monophosphate release, and attenuate the expression level of P2Y6 in porcine aortic vascular endothelial cells induced by Glaesserella parasuis. Baicalin could also reduce the activation of phospholipase C-protein kinase C and myosin light chain kinase-myosin light chain signaling pathways in porcine aortic vascular endothelial cells triggered by Glaesserella parasuis. Our study report that Glaesserella parasuis could promote pannexin 1 channel protein expression, induce nucleosides substance release, and P2Y6 expression in porcine aortic vascular endothelial cells and baicalin could inhibit the expression levels of pannexin 1, nucleosides substance, and P2Y6 in the porcine aortic vascular endothelial cells induced by Glaesserella parasuis, which might be served as some targets for treatment of inflammation disease caused by Glaesserella parasuis.

Integrating network pharmacology and experimental validation to explore the mechanisms of luteolin in alleviating fumonisin B1-induced intestinal inflammatory injury.

Contamination with fumonisin B1 (FB1) represents a global health problem. FB1 exposure may also trigger intestinal injury by activating inflammatory responses, leading to a reduction in production performance and economic benefits. However, the mechanism of FB1-induced intestinal inflammatory injury is still unclear. At the same time, it is urgent to develop antibiotic alternatives and therapeutic targets to alleviate antibiotic resistance and to ensure effective treatment of intestinal inflammatory injury. We combined network pharmacology and in vitro experiments to explore the core therapeutic targets and potential mechanism of luteolin in FB1-induced intestinal inflammatory injury. Network pharmacology and molecular docking revealed that nuclear factor kappa B (NF-κB) p65, extracellular signal-regulated kinase (ERK), interleukin 6 (IL-6) and IL-1ß are the important targets, and the NF-κB and ERK signalling pathways are critical in FB1-induced intestinal inflammatory injury. Besides, in vitro experiments further demonstrated that luteolin can inhibit FB1-induced intestinal inflammatory injury by inhibiting activation of the NF-κB and ERK signalling pathways and reducing the expression of IL-6 and IL-1ß in IPEC-J2 cells. We have comprehensively illustrated the potential targets and molecular mechanism by which luteolin can alleviate FB1-induced intestinal inflammatory injury. Luteolin may be an effective antibiotic alternative to prevent intestinal inflammatory injury.

Baicalin-aluminum alleviates necrotic enteritis in broiler chickens by inhibiting virulence factors expression of Clostridium perfringens.

Clostridium perfringens type A is the main cause of necrotic enteritis (NE) in chickens. Since the use of antibiotics in feed is withdrawn, it is imperative to find out suitable alternatives to control NE. Baicalin-aluminum complex is synthesized from baicalin, a flavonoid isolated from Scutellaria baicalensis Georgi. The present study investigated the effects of baicalin-aluminum on the virulence-associated traits and virulence genes expression of C. perfringens CVCC2030, it also evaluated the in vivo therapeutic effect on NE. The results showed that baicalin-aluminum inhibited bacterial hemolytic activity, diminished biofilm formation, attenuated cytotoxicity to Caco-2 cells, downregulated the expression of genes encoding for clostridial toxins and extracellular enzymes such as alpha toxin (CPA), perfringolysin O (PFO), collagenase (ColA), and sialidases (NanI, NanJ). Additionally, baicalin-aluminum was found to negatively regulate the expression of genes involved in quorum sensing (QS) communication, including genes of Agr QS system (agrB, agrD) and genes of VirS/R two-component regulatory system (virS, virR). In vivo experiments, baicalin-aluminum lightened the intestinal lesions and histological damage, it inhibited pro-inflammatory cytokines (TNF-α, IL-1ß, IL-6) expression in the jejunal and ileal tissues. Besides, baicalin-aluminum alleviated the upregulation of C. perfringens and Escherichia coli and raised the relative abundance of Lactobacillus in the ileal digesta. This study suggests that baicalin-aluminum may be a potential candidate against C. perfringens infection by inhibiting the virulence-associated traits and virulence genes expression.

Baicalin Weakens the Porcine ExPEC-Induced Inflammatory Response in 3D4/21 Cells by Inhibiting the Expression of NF-κB/MAPK Signaling Pathways and Reducing NLRP3 Inflammasome Activation.

Porcine extraintestinal pathogenic Escherichia coli (ExPEC) is a leading cause of death in pigs and has led to considerable economic losses for the pig industry. Porcine ExPEC infections often cause systemic inflammatory responses in pigs, characterized by meningitis, arthritis, pneumonia, and septicemia. Baicalin has been reported to possess potent anti-inflammatory activity, but its function in porcine ExPEC remains unknown. The aim of this study was to explore the protective effect and mechanism of baicalin against the porcine ExPEC-induced inflammatory responses in 3D4/21 cells. After treatment with baicalin, the effects on cell damage, the level of pro-inflammatory cytokines, the expression of nuclear factor-κB (NF-κB)/mitogen-activated protein kinase (MAPK) signaling pathways, and the activation of NOD-like receptor protein 3 (NLRP3) inflammasomes were examined. Our results show that baicalin significantly reduced the damage to 3D4/21 cells infected with porcine ExPEC PCN033. Further study showed that baicalin significantly reduced the transcription and expression of pro-inflammatory cytokines such as interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and interleukin-8 (IL-8). Furthermore, baicalin inhibited the phosphorylation of proteins such as P65, nuclear factor κB inhibitor α (IκBα), extracellular regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and P38 and reduced the expression levels of proteins such as NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), and caspase-1. These results reveal that baicalin reduced the damage to 3D4/21 cells by inhibiting the expression of NF-κB/MAPK signaling pathways and blocking NLRP3 inflammasome activation in 3D4/21 cells infected with porcine ExPEC. Taken together, these results suggest that baicalin may have potential as a medicine for the treatment of porcine ExPEC-infected pigs by regulating inflammatory responses. This study provides a novel potential pharmaco-therapeutic approach to preventing porcine ExPEC infection.

18ß-Glycyrrhetinic Acid Alleviates P. multocida-Induced Vascular Endothelial Inflammation by PARP1-Mediated NF-κB and HMGB1 Signalling Suppression in PIEC Cells.

Background: At present, the treatment and prevention of Pasteurella multocida infections in pigs mainly rely on antibiotics and vaccines, but inflammatory injury cannot be eliminated. The compound 18ß-glycyrrhetinic acid (GA), a pentacyclic triterpenoid extracted from Glycyrrhiza glabra L. root (liquorice) and with a chemical structure similar to that of steroidal hormones, has become a research focus because of its anti-inflammatory, antiulcer, antimicrobial, antioxidant, immunomodulatory, hepatoprotective and neuroprotective effects, but its potential for the treatment of vascular endothelial inflammatory injury by P. multocida infections has not been evaluated. This study aimed to investigate the effects and mechanisms of GA intervention in the treatment of vascular endothelial inflammatory injury by P. multocida infections. Materials and Methods: Putative targets of GA intervention in the treatment of vascular endothelial inflammatory injury by P. multocida infections were identified using network pharmacological screening and molecular docking simulation. The cell viability of PIEC cells was investigated via the CCK-8 assay. The mechanism of GA intervention in the treatment of vascular endothelial inflammatory injury by P. multocida infections were investigated using cell transfection and western blot. Results: Through network pharmacological screening and molecular docking simulation, this study found that PARP1 may be a core target for GA to exert anti-inflammatory effects. Mechanistically, GA alleviates P. multocida-induced vascular endothelial inflammation by PARP1-mediated NF-κB and HMGB1 signalling suppression. Conclusion: These findings, for the first time, demonstrate the potential therapeutic relationship among GA, PARP1 and inflammatory injury, providing a candidate drug, therapeutic targets and explanation for treating vascular endothelial inflammatory injury caused by P. multocida infection.

Baicalin and baicalein attenuate hyperuricemic nephropathy via inhibiting PI3K/AKT/NF-κB signalling pathway.

AIM: Inflammation and apoptosis are main pathological processes that lead to the development of hyperuricemic nephropathy (HN). This study aims to explore whether baicalin (BA) and baicalein (BAI) can relieve the damage through PI3K/AKT/NF-κB signal pathway and provide more reliable and precise evidence for the treatment of HN. METHODS: HN mice were induced by yeast extract with potassium oxonate (PO), and HK-2 cells were induced by monosodium urate (MSU). Molecular docking, western blot, q-PCR, and other methods were used to explore the changes of various indicators in HN mice and HK-2 cells. RESULTS: Molecular docking results showed that BA and BAI had good binding ability with PI3K, AKT, p65 and IκBα. BA and BAI significantly ameliorated the levels of renal function, decreased the p-PI3K, p-AKT and p-p65 expression, down-regulated the BAX/BCL2 and CASP3, and blunted the mRNA levels of tumour necrosis factor (TNF)-α, interleukin (IL)-1ß, and IL-18 in both renal tissue of HN mice and HK-2 cells induced by MSU. BA and BAI also decreased the oxidative stress level of MSU-induced HK-2 cells. CONCLUSION: BA and BAI were confirmed to attenuate HN through alleviating renal inflammatory and apoptosis in cells and tissues by inhibiting PI3K/AKT/NF-κB pathway. BA and BAI were expected to be developed as new anti-HN drugs.

Detection of Colistin Sulfate on Piglet Gastrointestinal Tract Microbiome Alterations.

The gut microbiome exerts important functions on host health maintenance, whereas excessive antibiotic use may cause gut flora dysfunction resulting in serious disease and dysbiosis. Colistin is a broad-spectrum antibiotic with serious resistance phenomena. However, it is unclear whether colistin alters the gastrointestinal tract microbiome in piglets. In this study, 16s rDNA-based metagenome analyses were used to assess the effects of colistin on the modification of the piglet microbiome in the stomach, duodenum, jejunum, cecum, and feces. Both α- and ß-diversity indices showed that colistin modified microbiome composition in these gastrointestinal areas. In addition, colistin influenced microbiome composition at the phylum and genus levels. At the species level, colistin upregulated Mycoplasma hyorhinis, Chlamydia trachomatis, Lactobacillus agilis, Weissella paramesenteroides, and Lactobacillus salivarius abundance, but downregulated Actinobacillus indolicus, Campylobacter fetus, Glaesserella parasuis, Moraxella pluranimalium, Veillonella caviae, Neisseria dentiae, and Prevotella disiens abundance in stomachs. Colistin-fed piglets showed an increased abundance of Lactobacillus mucosae, Megasphaera elsdenii DSM 20460, Fibrobacter intestinalis, and Unidentified rumen bacterium 12-7, but Megamonas funiformis, Uncultured Enterobacteriaceae bacterium, Actinobacillus porcinus, Uncultured Bacteroidales bacterium, and Uncultured Clostridiaceae bacterium abundance was decreased in the cecum. In feces, colistin promoted Mucispirillum schaedleri, Treponema berlinense, Veillonella magna, Veillonella caviae, and Actinobacillus porcinus abundance when compared with controls. Taken together, colistin modified the microbiome composition of gastrointestinal areas in piglets. This study provides new clinical rationalization strategies for colistin on the maintenance of animal gut balance and human public health.