Development of a droplet digital PCR assay to detect bovine alphaherpesvirus 1 in bovine semen.

BACKGROUND: Infectious bovine rhinotracheitis (IBR) caused by bovine alphaherpesvirus 1 (BoHV-1) is one of the most important contagious diseases in bovine. This is one of the most common infectious disease of cattle. This has led to high economic losses in the cattle farming industry. BoHV-1 can potentially be transmitted via semen during natural or artificial insemination (AI). Therefore, testing methods for the early diagnosis of BoHV-1 infection are urgently needed for international trade of ruminant semen. In this study, we developed a novel droplet digital PCR (ddPCR) assay for the detection of BoHV-1 DNA in semen samples. RESULTS: The ddPCR results showed that the detection limit was 4.45 copies per reaction with high reproducibility. The established method was highly specific for BoHV-1 and did not show cross-reactivity with specify the organisms (BTV, BVDV, Brucella, M . bovis). The results of clinical sample testing showed that the positivity rate of ddPCR (87.8%) was higher than that of qPCR (84.1%). CONCLUSIONS: The ddPCR assay showed good accuracy for mixed samples and could be a new added diagnostic tool for detecting BoHV-1.

Overexpression of heat shock protein 70 induces apoptosis of intestinal epithelial cells in heat-stressed pigs: A proteomics approach.

Heat stress (HS)-induced intestinal epithelial cell apoptosis may play a pivotal role in intestinal barrier dysfunction in animals. However, the underlying molecular mechanism by which HS induces apoptosis in intestinal epithelial cells is still poorly understood. Herein, a eukaryotic expression vector for an HSP70 gene was constructed and transfected into intestinal porcine epithelial cells (IPEC-J2). Afterward, functional proteomics approaches followed by liquid-chromatography-tandem mass spectrometry (LC-MS/MS) were used to identify interacting proteins. Analysis of HSP70 transfected IPEC-J2 cells revealed 246 differentially expressed proteins (DEPs), and functional annotation indicated that most DEPs were primarily related to ECM-receptor interaction, focal adhesion, and apoptosis. Furtherly, the apoptosis rate and expression levels of apoptosis-related proteins in HSP70 transfected IPEC-J2 cells were detected, we found that the expression of caspase-3, PARP, and Bax were increased, but Bcl-2 were decreased in transfected cells. Lastly, an in vitro and in vivo heat stress model were established to explore the role of HSP70 in intestinal epithelia cell apoptosis. The results of in vitrol study showed that HS-induced cellular apoptosis and increases of caspase-3, PARP, and Bax, but decreased of Bcl-2 in IPEC-J2 cells. In vivo study, the cell apoptosis were induced significantly in the duodenum, cecum, and colon of heat stressed pigs, and upregulation of HSP70 was also detected in colon tissues. Therefore, it has been shown that HSP70 plays a crucial role in heat stress-induced apoptosis and may provide new insights into the molecular mechanisms of epithelial cell apoptosis induced by heat stress in pigs.

Effect of heat stress on growth and production performance of livestock and poultry: Mechanism to prevention.

Heat stress is a widespread phenomenon in domestic animal feeding in tropical and sub-tropical areas that are subjected to a growing negative effect in livestock and poultry due to global warming. It leads to reduced food intake, retarded growth, intestinal disequilibrium, lower reproductive performance, immunity and endocrine disorders in livestock and poultry. Many studies show that the pathogenesis of heat stress is mainly related to oxidative stress, hormone secretion disorder, cytokine imbalance, cell apoptosis, cell autophagy, and abnormal cell function. Its mechanism refers to activation of mitogen-activated protein kinase (MAPK) signaling pathway and nuclear factor kappa B (NF-κB) signaling pathway, the fluctuation of tight junction protein and heat shock protein expression, and protein epigenetic modification. This manuscript reviews the mechanism of heat stress through an insight into the digestive, reproductive, immune, and endocrine system. Lastly, the progress in prevention and control techniques of heat stress has been summarized.

ERK1/2 mitogen-activated protein kinase mediates downregulation of intestinal tight junction proteins in heat stress-induced IBD model in pig.

In many mammalian species, including pigs, heat stress (HS) detrimentally leads to epithelium damage and increases intestinal permeability. However, the underlying molecular mechanisms are not thoroughly investigated yet. This study aimed to examine the RIP1/RIP3-ERK1/2 signaling pathway that regulates the expression of tight junction proteins in HS-treated pigs. In in vitro cultured intestinal porcine epithelial cells (IPEC-J2), HS induced the expression of tight junction proteins, ZO-1, claudin-1, and claudin-4, that are regulated by the ERK1/2-MAPK signaling pathway. Further, high expression of HSP70 in IPEC-J2 cells induced a significant decrease in receptor-interacting protein 1/3 (RIP1/3), phosphorylated ERK, and tight junction protein claudin-1 (P < 0.05). Necrostatin-1 (A selective inhibitor of RIPK1) suppressed the upregulation of phosphorylated ERK1/2 induced by HS, indicating that the RIP1/RIP3 regulates ERK1/2 phosphorylation in IPEC-J2 under heat stress. In addition, HS significantly damaged the intestinal morphology characterized by reduction of villus length and crypt depth in in vivo porcine model. Moreover, the expression of tight junction, ZO-1, and claudin-4 were downregulated, whereas phosphorylated p38 and ERK1/2 were upregulated in the duodenum of heat-stressed pigs. Interestingly, a decrease in ZO-1 and claudin-1 was observed in the colon, where phosphorylated ERK1/2 was similar to that in the duodenum. Our results demonstrate that RIP1/RIP3-ERK1/2 signaling pathway regulates the expression of tight junction proteins in HS-pigs. This finding further advances the intestinal barrier function's underlying mechanisms associated with signaling regulation.

Proteomic study of hypothalamus in pigs exposed to heat stress.

BACKGROUND: With evidence of warming climates, it is important to understand the effects of heat stress in farm animals in order to minimize production losses. Studying the changes in the brain proteome induced by heat stress may aid in understanding how heat stress affects brain function. The hypothalamus is a critical region in the brain that controls the pituitary gland, which is responsible for the secretion of several important hormones. In this study, we examined the hypothalamic protein profile of 10 pigs (15 ± 1 kg body weight), with five subjected to heat stress (35 ± 1 °C; relative humidity = 90%) and five acting as controls (28 ± 3 °C; RH = 90%). RESULT: The isobaric tags for relative and absolute quantification (iTRAQ) analysis of the hypothalamus identified 1710 peptides corresponding to 360 proteins, including 295 differentially expressed proteins (DEPs), 148 of which were up-regulated and 147 down-regulated, in heat-stressed animals. The Ingenuity Pathway Analysis (IPA) software predicted 30 canonical pathways, four functional groups, and four regulatory networks of interest. The DEPs were mainly concentrated in the cytoskeleton of the pig hypothalamus during heat stress. CONCLUSIONS: In this study, heat stress significantly increased the body temperature and reduced daily gain of body weight in pigs. Furthermore, we identified 295 differentially expressed proteins, 147 of which were down-regulated and 148 up-regulated in hypothalamus of heat stressed pigs. The IPA showed that the DEPs identified in the study are involved in cell death and survival, cellular assembly and organization, and cellular function and maintenance, in relation to neurological disease, metabolic disease, immunological disease, inflammatory disease, and inflammatory response. We hypothesize that a malfunction of the hypothalamus may destroy the host physical and immune function, resulting in decreased growth performance and immunosuppression in heat stressed pigs.

Magnetic molecularly imprinted specific solid-phase extraction for determination of dihydroquercetin from Larix griffithiana using HPLC.

The naturally occurring quercetin flavonoid, dihydroquercetin, is widely distributed in plant tissues and has a variety of biological activities. Herein, a magnetic molecularly imprinted solid-phase extraction was tailor made for selective determination of dihydroquercetin in Larix griffithiana using high-performance liquid chromatography. Amino-functionalized core-shell magnetic nanoparticles were prepared and characterized using scanning electron microscopy, transmission electron microscopy, vibrating sample magnetometry, and infrared spectroscopy. The polymer had an average diameter of 250 ± 2.56 nm and exhibited good stability and adsorption for template molecule, which is enriched by hydrogen bonding interaction. Multiple factors for extraction, including loading, washing, elution solvents, and extraction time, were optimized. The limit of detection was 1.23 µg/g. The precision determined at various concentration of dihydroquercetin was less than 4% and the mean recovery was between 74.64 and 101.80%. It has therefore been shown that this protocol can be used as an alternative extraction to quantify dihydroquercetin in L. griffithiana and purify quercetin flavonoid from other complex matrices.

SOCS3 control the activity of NF-κB induced by HSP70 via degradation of MyD88-adapter-like protein (Mal) in IPEC-J2 cells.

Hyperthermia in pigs induces suppressor of cytokine signaling (SOCS) 3 and SOCS4 expression in intestinal gut and causes disruption of inflammation cytokine production. These changes may affect the development of inflammatory bowel disease in heat-stressed pigs. However, the mechanisms are not well understood. Accordingly, in this study, we examined the roles of SOCS members in regulation of the nuclear factor (NF)-κB pathway and heat shock protein (HSP) 70-mediated cytokine induction in 293T human embryonic kidney cells and IPEC-J2 porcine small intestinal epithelial cells. Ectopic expression of HSP70 significantly modulated NF-κB activity (p ≤ .05). Moreover, co-expression of SOCS3 or SOCS4 with HSP70 reduced NF-κB activity, which was abolished by SOCS3 or SOCS4 knockdown with short hairpin RNA. Interestingly, MyD88-adaptor-like (Mal) protein was downregulated in cells expressing SOCS3 but not in cells expressing SOCS4. In addition, SOCS3 but not SOCS4 negatively regulated the activity of NF-κB induced by HSP70 overexpression via degradation of Mal. These findings may facilitate the development of novel SOCS3-based therapeutic strategies to control heat stress-related disorders in pigs.

Identification and functional characterization of Toll-like receptor 2-1 in geese.

BACKGROUND: Toll-like receptor 2 (TLR2), an important pattern recognition receptor, activates proinflammatory pathways in response to various pathogens. It has been reported in humans and chicken, but not in geese, an important waterfowl species in China. Since some vaccines stimulate robust immune responsesl in chicken but not in geeeses we speculated that their immune systems are different. RESULTS: In this study, we cloned the goose TLR2-1 gene using rapid amplification of cDNA ends (RACE)and showed that geese TLR2-1 encoded a 793-amino-acid protein, containing a signal secretion peptide, an extracellular leucine-rich repeat domain, a transmembrane domain and a Toll/interleukin-1 receptor signaling domain deduced from amino acid sequence. TLR2-1 shared 38.4%-93.5% homology with its homologues in other species. Tissue expression of geese TLR2-1 varied markedly, and was higher in kidney, cloacal bursa, skin and brain compared to other organs/tissues. HEK293 cells transfected with plasmids carrying goose TLR2-1 and NF-κB-luciferase responded significantly to stimulation with Mycoplasma fermentans lipopeptide. Furthermore, geese infected with Mycoplasma gallisepticum (MG) and Salmonella enteritidis (SE) showed significant upregulation of TLR2-1 in both in vivo and in vitro. CONCLUSION: Geese TLR2-1 is a functional homologue of TLR2 present in other species and plays an important role in bacterial recognition in geese.

Overexpression of the Mas1 gene mitigated LPS-induced inflammatory injury in mammary epithelial cells by inhibiting the NF-κB/MAPKs signaling pathways.

Breast infection is the primary etiology of mastitis in dairy cows, leading to a reduction in the quality of dairy products and resulting in substantial economic losses for animal husbandry. Although antibiotic treatment can eliminate the pathogenic microorganisms that induce mastitis, it cannot repair the inflammatory damage of mammary epithelial cells and blood milk barrier. Mas1 is a G protein-coupled receptor, and its role in lipopolysaccharide (LPS) -induced inflammatory injury to mammary epithelial cells has not been studied. LPS treatment of EpH4 EV cells led to a significant downregulation of Mas1 transcript levels, which attracted our great interest, suggesting that Mas1 may be an important target for the treatment of mastitis. Therefore, this study intends to verify the role of Mas1 in the inflammatory injury of EpH4 EV cells by gene overexpression technology and gene silencing technology. The findings demonstrated that the overexpression of the Mas1 gene effectively reversed the activation of the nuclear factor-κB/mitogen-activated protein kinase (NF-κB/MAPK) signaling pathways induced by LPS, while also suppressing the upregulation of pro-inflammatory mediators. Furthermore, overexpression of the Mas1 gene reversed the downregulation of zonula occludens 1 (ZO-1), Occludin, and Claudin-3 caused by LPS, suggesting that Mas1 could promote to repair the blood-milk barrier. However, the silencing of the Mas1 gene using siRNA resulted in a contrasting effect. These results indicated that Mas1 alleviated the inflammatory injury of mammary epithelial cells induced by LPS.

Lacticaseibacillus casei IB1 Alleviates DSS-Induced Inflammatory Bowel Disease by Regulating the Microbiota and Restoring the Intestinal Epithelial Barrier.

Inflammatory bowel disease (IBD) is becoming an increasingly serious health problem in humans and animals. Probiotics can inhibit the development of IBD. Due to the specificity of the strains, the function and mechanism of action of different strains are still unclear. Here, a DSS-induced colitis mouse model was utilized to investigate the ability and mechanism by which Lacticaseibacillus casei IB1 alleviates colitis. Treatment with L. casei IB1 improved DSS-induced colitis in mice, as indicated by increased body weight, colon length, and goblet cell numbers and decreased disease activity index (DAI), proinflammatory factor (TNF-α, IL-1ß, and IL-6) levels, and histopathological scores after intake of IB1. IB1 supplementation also improved the expression of tight junction proteins and inhibited the activation of the MAPK and NF-κB signaling pathways to alleviate intestinal inflammation. In addition, IB1 rebalanced the intestinal microbial composition of colitis mice by increasing the abundance of Faecalibaculum and Alistipes and decreasing the abundance of Bacteroides and Escherichia_Shigella. In summary, L. casei IB1 showed great potential for relieving colitis by regulating the microbiota and restoring the epithelial barrier. It can be used as a potential probiotic for the prevention and treatment of UC in the future.

Network pharmacology analysis of the regulatory effects and mechanisms of ALAE on sow reproduction in vivo and in vitro.

ETHNOPHARMACOLOGICAL RELEVANCE: Reproductive ability of sows is a primary element influencing the development of pig farming. Herbal extracts of Angelica sinensis (Oliv.) Diels, Astragalus mongholicus Bunge, Eucommia ulmoides Oliv., and Polypodium glycyrrhiza D.C.Eaton showed effects on improvement of reproduction in sows. AIMS OF THE STUDY: To investigate the mechanism of the treatment effects by a compound of these four Chinese herbs in a 1:1:1:1 ratio (ALAE) on endometriosis, endometritis, uterine adhesion, intrauterine growth retardation, pre-eclampsia, and its enhancement of reproductive efficiency in sows. MATERIALS AND METHODS: Active components of ALAE were identified by using ultra-performance liquid chromatography-mass spectrometry analysis and network pharmacology. Then we used the results to construct a visualization network. Key targets and pathways of ALAE involved in sow reproduction improvement were validated in sow animals and porcine endometrial epithelial cells (PEECs). RESULTS: A total of 62 active compounds were found in ALAE (41 in Polypodium glycyrrhiza D.C.Eaton, 5 in Astragalus mongholicus Bunge, 11 in Eucommia ulmoides Oliv., 5 in Angelica sinensis (Oliv.) Diels) with 563 disease-related targets (e.g. caspase-3, EGFR, IL-6) involved in EGFR tyrosine kinase inhibitor resistance, PI3K-AKT, and other signaling pathways. Molecular docking results indicated GC41 (glabridin), GC18 (medicarpin), EGFR and CCND1 are possible key components and target proteins related to reproductive improvement in sows. In PEECs, EGFR expression decreased at the mRNA and protein levels by three doses (160, 320, and 640 µg/mL) of ALAE. The phosphorylation of downstream pathway PI3K-AKT1 was enhanced. The expression of inflammatory factors (IL-6, IL-1ß), ESR1 and caspase-3 decreased through multiple pathways. Additionally, the expression levels of an anti-inflammatory factor (IL-10), angiogenesis-related factors (MMP9, PIGF, PPARγ, IgG), and placental junction-related factors (CTNNB1, occludin, and claudin1) increased. Furthermore, the total born number of piglets, the number of live and healthy litters were significantly increased. The number of stillbirths decreased by ALAE treatment in sow animals. CONCLUSIONS: Dministration of ALAE significantly increased the total number of piglets born, the numbers of live and healthy litters and decreased the number of stillbirths through improving placental structure, attenuating inflammatory response, modulating placental angiogenesis and growth factor receptors in sows. The improvement of reproductive ability may be related to activation of the EGFR-PI3K-AKT1 pathway in PEECs. Moreover, ALAE maybe involved in modulation of estrogen receptors, apoptotic factors, and cell cycle proteins.

Protective Effects of Carbonated Chitosan Montmorillonite on Vomitoxin-Induced Intestinal Inflammation.

Exposure to vomitoxin (DON) can negatively impact the intestinal health of livestock and poultry, leading to compromised nutrient absorption and utilization, resulting in slowed growth and reduced production efficiency. In this study, we synthesized carbonated chitosan montmorillonite intercalation complexes (CCM) through solution precipitation. The successful formation of intercalation complexes was confirmed by examining functional groups and surface features using infrared spectroscopy and scanning electron microscopy. To assess the impact of CCM on DON-infected mice, we established an experimental mouse model of jejunal inflammation induced by DON infection. We analyzed the effects of CCM on blood biochemical and conventional indices, jejunal inflammatory factors, pathological changes, and the expression of proteins in the MAPK pathways in DON-infected mice. Our results indicate that CCM effectively mitigates the adverse effects of DON on growth performance, jejunal injury, and the inflammatory response in mice. CCM supplementation alleviated the negative effects of DON infection on growth performance and reduced intestinal inflammation in mice. Moreover, CCM supplementation successfully inhibited the activation of the mitogen-activated protein kinase (MAPK) signaling pathway induced by DON. These findings suggest that the mitigating effect of CCM on DON-induced inflammatory injury in the murine jejunum is closely linked to the regulation of the MAPK signaling pathway.

Preliminary Proteomic Study of the Porcine Pituitary Gland under Heat Stress.

Although numerous studies have shown that the hypothalamic-pituitary-adrenal axis plays a vital role in the response to environmental stress by mediating the production of a series of hormones, the mechanism underlying these effects has not been elucidated. This study used proteomics techniques to investigate the differentially expressed proteins (DEPs) in the pituitary glands of pigs and to elucidate the potential changes in the immune-neuroendocrine system under heat stress (HS). In total, 2517 peptides corresponding to 205 proteins were detected. A comparison of the expression patterns between HSs and healthy controls revealed 56 DEPs, of which 31 were upregulated and 25 were downregulated. Ingenuity pathway analysis (IPA) was used to reveal the subcellular characteristics, functional pathways, regulatory networks, and upstream regulators of the identified proteins. The results showed that these differentially expressed proteins were involved in intercellular communication, interactions, apoptosis, nervous system development, functions, abnormalities and other functions, and in the regulatory network. Moreover, the upstream regulators of the differentially expressed proteins were mainly transcriptional regulators, hormones, and cytokines. Thus, the functional network and pathway analyses could provide insights into the complexity and dynamics of HS-host interactions and may accelerate our understanding of the mechanisms underlying HS.

Enhancing gut barrier integrity: Upregulation of tight junction proteins by chitosan oligosaccharide through the ERK1/2 signaling pathway.

OBJECTIVES: This study aimed to explore the protective mechanism of chitosan oligosaccharide (COS) against lipopolysaccharide (LPS)-induced inflammatory responses in IEC-6 cells and dextran sodium sulfate (DSS)-induced colitis in mice. METHODS: The cell inflammation model was constructed by LPS in vitro and enteritis model by DSS in vivo. RESULTS: Following LPS exposure, IEC-6 cell proliferation significantly decreased, epithelial cell integrity was compromised, and TNF-α and IL-1ß levels were increased. However, COS pretreatment reversed these changes. In vivo, DSS-treated mice exhibited evident pathological alterations, including heightened inflammatory levels and significantly decreased expression of tight junction proteins and critical proteins in the Mitogen activated proteins kinase signaling pathway. Nevertheless, COS administration notably reduced inflammatory levels and increased the expression of tight junction proteins and key proteins in the Mitogen activated proteins kinase signaling pathway. CONCLUSIONS: Our findings suggest that COS safeguards gut barrier integrity by upregulating tight junction proteins through the ERK1/2 signaling pathway. Therefore, COS has emerged as a promising candidate for novel drug interventions against inflammatory bowel disease.

Slc9a1 plays a vital role in chitosan oligosaccharide transport across the intestinal mucosa of mice.

The mechanism underlying the intestinal transport of COS is not well understood. Here, transcriptome and proteome analyses were performed to identify potential critical molecules involved in COS transport. Enrichment analyses revealed that the differentially expressed genes in the duodenum of the COS-treated mice were mainly enriched in transmembrane and immune function. In particular, B2 m, Itgb2, and Slc9a1 were upregulated. The Slc9a1 inhibitor decreased the transport efficiency of COS both in MODE-K cells (in vitro) and in mice (in vivo). The transport of FITC-COS in Slc9a1-overexpressing MODE-K cells was significantly higher than that in empty vector-transfected cells (P < 0.01). Molecular docking analysis revealed the possibility of stable binding between COS and Slc9a1 through hydrogen bonding. This finding indicates that Slc9a1 plays a crucial role in COS transport in mice. This provides valuable insights for improving the absorption efficiency of COS as a drug adjuvant.

Development of an Fe3O4 Surface-Grafted Carboxymethyl Chitosan Molecularly Imprinted Polymer for Specific Recognition and Sustained Release of Salidroside.

The choice of carrier material is critical in the study of natural drug release preparations and glycosylated magnetic molecularly imprinted materials. The stiffness and softness of the carrier material affect the efficiency of drug release and the specificity of recognition. The dual adjustable aperture-ligand in molecularly imprinted polymers (MIPs) provides the possibility of individualized design for sustained release studies. In this study, a combination of paramagnetic Fe3O4 and carboxymethyl chitosan (CC) was used to enhance the imprinting effect and improve drug delivery. A combination of tetrahydrofuran and ethylene glycol was used as a binary porogen to prepare MIP-doped Fe3O4-grafted CC (SMCMIP). Salidroside serves as the template, methacrylic acid acts as the functional monomer, and ethylene glycol dimethacrylate (EGDMA) serves as the crosslinker. Scanning and transmission electron microscopy were used to observe the micromorphology of the microspheres. The structural and morphological parameters of the SMCMIP composites were measured, including the surface area and pore diameter distribution. In an in vitro study, we found that the SMCMIP composite had a sustained release property of 50% after 6 h of release time in comparison to the control SMCNIP. The total amounts of SMCMIP released at 25 °C and 37 °C were 77% and 86%, respectively. In vitro results showed that the release of SMCMIP followed Fickian kinetics, meaning that the rate of release is dependent on the concentration gradient, with diffusion coefficients ranging from 3.07 × 10-2 cm2/s to 5.66 × 10-3 cm2/s. The results of cytotoxicity experiments showed that the SMCMIP composite did not have any harmful effects on cell growth. The survival rates of intestinal epithelial cells (IPEC-J2) were found to be above 98%. By using the SMCMIP composite, drugs may be delivered in a sustained manner, potentially leading to improved therapeutic outcomes and reduced side effects.

Corrigendum: Coral-derived endophytic fungal product, butyrolactone-I, alleviates LPS induced intestinal epithelial cell inflammatory response through TLR4/NF-κB and MAPK signaling pathways: An in vitro and in vivo studies.

[This corrects the article DOI: 10.3389/fnut.2021.748118.].

Chitosan-gentamicin conjugate attenuates heat stress-induced intestinal barrier injury via the TLR4/STAT6/MYLK signaling pathway: In vitro and in vivo studies.

Heat stress (HS) has a negative impact on animal health. A modified chitosan-gentamicin conjugate (CS-GT) was prepared to investigate its potential protective effects and mechanism of action on heat stress-induced intestinal mucosa injury in IPEC-J2 cells and mouse 3D intestinal organs in a mouse model. CS-GT significantly (P < 0.01) reversed the decline in transmembrane resistance and increased the FITC-dextran permeability of the IPEC-J2 monolayer fusion epithelium caused by heat stress. Heat stress decreased the expression of the tight binding proteins occludin, claudin1, and claudin2. However, pretreatment with CS-GT significantly increased (P < 0.01) the expression of these tight binding proteins. Mechanistically, CS-GT inhibited the activation of the TLR4/STAT6/MYLK signaling pathway induced by heat stress. Molecular docking showed that CS-GT can bind effectively with TLR4. In conclusion, CS-GT alleviates heat stress-induced intestinal mucosal damage both in vitro and in vivo. This effect is mediated, at least partly, by the inhibition of the TLR4/STAT6/MYLK signaling pathway and upregulation of tight junction proteins. These findings suggest that CS-GT may have therapeutic potential in the prevention and treatment of heat stress-related intestinal injury.

Research progress on pharmacological effects and new dosage forms of baicalin.

BACKGROUND: As a kind of flavonoid, baicalin (C21 H18 O11 ) is extracted from Scutellaria baicalensis Georgi, the extract of which can be added to animal feed in China. OBJECTIVES: The present review will describe the current understanding of the pharmacological effects of baicalin in the regulation of inflammation, oxidative stress anti-virus and anti-tumour responses. METHODS: We highlight emerging literature that the application in livestock health and performance, the biological activities, the molecular mechanisms and the dosage forms of baicalin by analysing and summarising the main points of the cited literatures. RESULTS: It is found that baicalin can improve the functions of multiple physiological systems. Baicalin has a strong anti-inflammatory effect by regulating TLR4-NFκB-MAPK signalling pathway; it also can reduce oxidative stress by regulating Nrf2-Keap1 pathway; it can inhabit many kinds of virus such as influenza virus, respiratory virus, hepacivirus and others; it can also inhibit the growth of tumour cells by blocking the cell cycle or inducing apoptosis; and new dosage forms such as cationic solid lipid nanoparticles, cyclodextrin inclusion complexes or nanocrystalline can be applied to improve the deficiency of baicalin. CONCLUSIONS: In summary, these studies have elucidated a comprehensive report on the anti-inflammatory, anti-oxidant, anti-virus and anti-tumour of baicalin, these findings thus indicated that baicalin can be used effectively to the field of animal production in future when the appropriate dosage form is determined.

Molecularly imprinted polymer-specific solid-phase extraction for the determination of 4-hydroxy-2(3H)benzoxazolone isolated from Acanthus ilicifolius Linnaeus using high-performance liquid chromatography-tandem mass spectrometry.

The minor constituent found in Acanthus ilicifolius Linnaeus, 4-hydroxy-2 (3H) benzoxazolone alkaloid (HBOA), has a range of versatile applications. Herein, a quick and straightforward method for extracting HBOA from A. ilicifolius Linnaeus was proposed. HBOA was used as a template, whereas methacrylic acid, ethylene glycol dimethacrylate, and acetonitrile were used as functional monomers, cross-linkers, and porogens, respectively. Molecularly imprinted polymers (MIPs) were synthesized by precipitation polymerization, and their adsorption isotherms, dynamics, and selective binding ability were characterized and analyzed. The results showed that the adsorption amount of the template was 90.18 mg/g. The MIPs were used as solid-phase extraction fillers and actual sample extraction columns, with a linear range of 0-100 µg/L, average recovery of 78.50-101.12%, and a relative standard deviation of 1.20-3.26%. The HBOA concentrations in the roots, stems, and leaves were 1,226, 557, and 205 µg/g, respectively. In addition, MIP-SPE was successfully used in isolating and purifying HBOA from different parts of A. ilicifolius Linnaeus, indicating its effectiveness in extracting and determining HBOA in other herbs.