Search Results (96)

Avian pathogenic Escherichia coli (E. coli) impairs poultry production and causes substantial economic losses. This study investigated the effects of Lactobacillus reuteri postbiotics (LR) on growth performance and intestinal health of broiler chickens challenged with E. coli. A total of 180 one-day-old Arbor Acres+ broilers were allocated into three groups (six replicates per group and 10 chicks each replicate): CTR, control group; E. coli-infected group, orally challenged with a mixture of E. coli O₁, O₂, and O₇₈ at a dose of 10⁹ CFU/mL; LR + E. coli-infected group, challenged with E. coli and fed a basal diet supplemented with 100 mg/kg LR. The results showed that dietary LR significantly improved the average daily gain (ADG) in the LR + E. coli group compared to the E. coli-infected group from days 1 to 18 (p < 0.05). However, no significant differences in average daily feed intake (ADFI) or feed conversion ratio (FCR) were observed among the CTR, E. coli, and LR + E. coli groups. Infection with E. coli led to lower total antioxidant capacity in jejunum and activity of total superoxide dismutase in ileum. Moreover, dietary LR significantly alleviated the down-regulation of Mucin2 and Aquaporin-3 gene expression in jejunum and ileum caused by E. coli infection and up-regulated the gene expression of Claudin-1 and Zonula occludens 1 in the ileum. In addition, dietary LR treatment led to the up-regulation of interleukin-10 mRNA transcripts in the jejunum. Further analysis demonstrated that dietary supplementation with LR reshaped the ileal flora of birds challenged with E. coli via elevating the relative abundance of Romboutsia and Bacteroidota, while reducing the abundance of Candidatus_Arthromitus and Escherichia-Shigella. In conclusion, dietary LR supplementation improved the expression of intestinal barrier and anti-inflammatory genes and reshaped the intestinal flora in E. coli-infected broilers. Full article

This study examined the protective impact of tributyrin on heat-stressed Taihe silky fowls, providing insight into oxidative status, inflammatory response, and mucosal barrier function. Three hundred chicks were randomly assigned to 6 treatments: control (CON, 24 ± 1 °C) fed with basal diet and 5 heat stress (HS) treatments (34 ± 1 °C for 8 h/d) fed with basal diet containing 0, 0.04, 0.08, 0.16, and 0.32% tributyrin. Heat stress elevated serum malondialdehyde (MDA), tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β), D-lactate, and diamine oxidase levels, and decreased total antioxidant capacity (T-AOC) and superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), interleukin 4 (IL-4), interleukin 10 (IL-10), and immunoglobulin G (IgG) levels (p < 0.05). Compared with HS treatment, tributyrin reversed these serum changes (p < 0.05). Moreover, HS elevated jejunal and ileal MDA content and IL-1β mRNA abundance, decreased GSH-Px activity, villus height (VH), VH: crypt depth ratio, and mRNA abundance of IL-10, occludin, and zonula occludens-1 (ZO-1), and decreased cecal butyrate content (p < 0.05). Compared with HS treatment, tributyrin reduced jejunal and ileal MDA content and IL-1β mRNA abundance, increased GSH-Px activity, VH, and mRNA abundance of IL-4, IL-10, occludin, and ZO-1, and increased cecal butyrate content (p < 0.05). In conclusion, tributyrin enhanced antioxidant capacity, attenuated inflammatory responses, increased cecal butyrate content, and improved intestinal morphology and mucosal barrier function in cyclic heat-stressed Taihe silky fowls. Full article

Berberine (BBR), a benzylisoquinoline alkaloid isolated from Chinese herb Coptis chinensis, has been widely used clinically to treat intestinal infectious diseases. Recently, it has been found to have multiple pharmacological effects, including anti-inflammatory activity and immune effects in inflammatory bowel disease (IBD). However, its exact targets remain to be elucidated. In this study, we used a mouse intestinal organoid–macrophage co-culture model to investigate the anti-inflammatory effects and immune effects of BBR. Our findings demonstrated that lipopolysaccharide (LPS) induced more robust inflammatory responses and epithelium damage in the co-culture system compared to the organoid alone. BBR effectively attenuated inflammation and restored epithelial barrier integrity by suppressing M1 macrophage polarisation and infiltration, alongside upregulating the expression and organisation of tight junction protein zonula occludens-1 (ZO-1). RNA sequencing and proteomic analysis revealed that BBR disrupted organoid–macrophage interaction by inhibiting chemokine (e.g., C-X-C motif chemokine ligand 1 (CXCL1) and macrophage migration inhibitory factor (MIF)) release from epithelial cells, thereby reducing macrophage recruitment. Collectively, our study establishes the organoid–macrophage co-culture system as a more physiologically relevant model for studying epithelial–immune interactions and elucidates the multi-target mechanism of BBR, which concurrently modulates epithelial cells, macrophages, and their crosstalk. These findings lay the foundation for further exploration of the therapeutic potential of BBR in inflammatory bowel disease and the development of targeted therapies that regulate cell interactions. Full article

Natural products, such as turmeric rhizome meal (TRM), may hold the key to a sustainable solution to antimicrobial resistance rise and antibiotic prohibition in food-producing animals. This study evaluated the effects of dietary TRM at 0 (CON), 0.3 (TRM3), 0.6 (TRM6), and 0.9 g/kg (TRM9) on growth, nutrient digestibility, immunity, gut function, nutrient transport biomarkers, microbiome, and meat quality in 280 one-day-old male Ross 308 chicks over a 42-day feeding trial. Birds fed TRM indicated higher body weight gain and lower feed conversion ratio (p < 0.05). The TRM groups promoted higher (p = 0.001) serum immunoglobulin Y, immunoglobulin M, and interleukin-10 compared to the CON. Birds fed CON had higher interleukin-2 (p = 0.025), interleukin-6 (p = 0.027), and TNF-α (p = 0.008) levels compared to the TRM groups. Lactobacillus counts in jejunal villi and crypts were higher in the TRM groups than in the CON (p < 0.05). Dietary TRM increased electrogenic glucose and lysine transport, accompanied by up-regulation of claudin-5, zonula occludens 1, and mucin-2 expression (p < 0.05). In breast muscle, TRM fortification reduced malondialdehyde levels (p < 0.05) while increasing long-chain polyunsaturated fatty acids (p < 0.05). Thus, TRM is a potent, residue-free phytobiotic alternative to conventional antibiotic growth promoters in poultry systems. Full article

Oxidative stress is the key contributor to the onset of numerous diseases. Herein, we develop tea polyphenol–zinc (Tp-Zn) using a metal–polyphenol coordination strategy through a simple hybrid approach. The product is characterized by methods such as dynamic light scattering (DLS), ultraviolet–visible spectroscopy (UV–vis) and transmission electron microscopy (TEM) to evaluate the particle size and potential of Tp-Zn. Oxidative stress was induced in mice by administering diquat (25 mg/kg body weight) followed by pre-treatment with 210 mg/kg body weight tea polyphenols (TPs), 280 mg/kg body weight Tp-Zn, and 70 mg/kg body weight ZnSO₄ for 7 days. Results showed that Tp-Zn treatment significantly improved intestinal barrier function by preventing the diquat-induced down-regulation of tight junction proteins Zonula Occludens protein 1 (ZO-1) and occludin. It also mitigated liver inflammation and damage, as evidenced by reduced serum levels of Aspartate aminotransferase (AST), Alanine aminotransferase (ALT), and Alkaline phosphatase (AKP). Furthermore, Tp-Zn enhanced the antioxidant response in both the intestine and liver by up-regulating the mRNA expression of antioxidant enzymes and reducing the levels of Malondialdehyde (MDA) and reactive oxygen species (ROS) compared to the diquat group (DIQ group). Also, the detection of ROS in the small intestine confirmed Tp-Zn markedly increased intestinal Nuclear factor erythroid 2-related factor 2 (Nrf2) expression compared to the control group. This study aims to clarify that metal–polyphenol coordination with multifaceted regulation of the inflammatory microenvironment could be a novel approach for preventing or treating oxidative stress-related diseases. Full article

Glutamine (Gln), a functional amino acid, is effective in reducing weaning stress in piglets. This study aims to assess the effects of dietary Gln supplementation on intestinal morphology and functionality, as well as the growth performance of Kele and Large White hybrid weaned piglets. Forty-eight piglets aged 30 days (Kele × Large White) were randomly divided into three groups: the control group, which received a basal diet supplemented with 2.45% alanine to maintain an isonitrogenous balance; the 1% Gln group, which received the basal diet with 1.0% Gln and 1.23% alanine; and the 2% Gln group, which was given the basal diet supplemented with 2.0% Gln. Intestinal samples from 16 piglets in the control and 1% Gln groups were collected randomly on day 29 of the experiment. The results show that, compared to the control group, the 1% Gln group experienced an increase in the average daily gain (ADG) and gain-to-feed ratio (G:F). In contrast, the 2% Gln group did not demonstrate significant differences in either the ADG or G:F compared to the control group. Additionally, there were no differences in feed intake among the groups. Notably, weaned piglets in both the 1% and 2% Gln supplementation groups had reduced diarrhea rates compared to those in the control group. Furthermore, 1% Gln supplementation significantly increased villus height in both the duodenum and jejunum and the ratio of villus height to crypt depth in weaned piglets. Subsequent analyses revealed that 1% Gln supplementation increased the mRNA expression of antioxidant genes, specifically catalase and superoxide dismutase. Additionally, the mRNA levels of the intestinal tight junction genes zonula occludens-1, Claudin 1, and Occludin in the jejuna of weaned piglets were found to be elevated. In summary, incorporating 1% Gln into the diet can significantly improve intestinal functionality and promote growth in Kele and Large White hybrid weaned piglets. Full article

(1) Background: Respiratory allergens, particularly ragweed (RW) pollen and house dust mites (HDMs), are major triggers of respiratory inflammation and allergic diseases. This study investigated the impact of single- versus combined-allergen exposure on the barrier function of normal human bronchial epithelial (NHBE) cells cultured at the air–liquid interface (ALI). (2) Methods: NHBE cells were exposed to RW pollen extract (200 µg/mL), HDM extract (200 µg/mL) and their combination at varying concentrations (200 µg/mL, 100 µg/mL, 50 µg/mL, 25 µg/mL). Additional groups included a mixture of Amb a 1, Amb a 11 and Amb a 12 (100 mg/mL) and combinations of Der p 1 with the ragweed allergens (50 mg/mL, 100 µg/mL). Transepithelial electrical resistance (TEER) was recorded over 72 hours to assess barrier integrity, and immunofluorescence (IF) staining for zonula occludens-1 (ZO-1) was performed to evaluate tight junction alterations. (3) Results: TEER measurements showed a significant reduction in epithelial barrier integrity following allergen exposure, with the most pronounced disruption observed with the combined exposure to RW and HDM groups. IF staining confirmed extensive tight junction damage, highlighting their synergistic impact. (4) Conclusions: These findings emphasize the importance of assessing cumulative allergen effects, as combined exposure may exacerbate epithelial dysfunction and represent a key aspect in the management of allergic rhinitis and asthma. Full article

Irritable Bowel Syndrome (IBS) is a disorder of gut- brain interaction characterized by recurrent abdominal pain associated with altered bowel habits. The therapeutic options for IBS patients include the use of probiotics. The aim of this study was to assess the effect of a multi-strain probiotic made up by Lactobacillus rhamnosus LR 32, Bifidobacterium lactis BL 04, and Bifidobacterium longum BB 536 (Serobioma, Bromatech s.r.l., Milano, Italy) on an in vitro model of the intestinal epithelial barrier in the presence of mucosal mediators that are released by IBS patients. IBS (n = 28; IBS with predominant diarrhea, IBS-D = 10; IBS with predominant constipation, IBS-C = 9; and IBS with mixed bowel habits, IBS-M = 9) patients, diagnosed according to the Rome IV criteria, and asymptomatic controls (ACs, n = 7) were enrolled. Mucosal mediators that were spontaneously released by colonic biopsies were collected (supernatants). Two doses of Serobioma were tested with/without IBS/AC mediators. RNA was extracted from Caco-2 cells to evaluate the tight junction (TJ) expression. Serobioma (10⁶ CFU/mL) significantly reinforced the Caco-2 monolayer compared to growth medium alone (p < 0.05). IBS supernatants significantly increased Caco-2 paracellular permeability compared to the AC supernatants. The co-incubation of Caco-2 cells with IBS supernatants and Serobioma (10⁶ CFU/mL) avoided the paracellular permeability alterations that were induced by IBS supernatants alone (p < 0.001), and, in particular, IBS-D and IBS-M ones. The co-incubation of Serobioma (10⁶ CFU/mL) and IBS-D supernatants significantly increased ZO-1 expression compared to Caco-2 cells incubated with supernatants alone (p < 0.05), as confirmed via qPCR analyses. Serobioma (10⁶ CFU/mL) counteracts the paracellular permeability changes that are induced by IBS supernatants, in particular IBS-D and IBS-M supernatants, likely modulating ZO-1 expression. Full article

Background: Berberine has extremely low oral bioavailability, but shows a potent lipid-lowering effect, indicating its potential role in regulating intestinal microbiota, which has not been investigated. Methods: In the present study, five experimental diets, a control diet (Con), a high-lipid diet (HL), and high-lipid·diets·supplemented with an antibiotic cocktail (HLA), berberine (HLB), or both (HLAB) were fed to zebrafish (Danio rerio) for 30 days. Results: The HLB group showed significantly greater weight gain and feed intake than the HLA and other groups, respectively (p < 0.05). Hepatic triglyceride (TG) and total cholesterol (TC) levels, lipogenesis, and proinflammatory cytokine gene expression were significantly upregulated by the high-lipid diet, but significantly downregulated by berberine supplementation. Conversely, the expression levels of intestinal and/or hepatic farnesoid X receptor (fxr), Takeda G protein-coupled receptor 5 (tgr5), lipolysis genes, and zonula occludens 1 (zo1) exhibited the opposite trend. Compared with the HLB group, the HLAB group displayed significantly greater hepatic TG content and proinflammatory cytokine expression, but significantly lower intestinal bile salt hydrolase (BSH) activity and intestinal and/or hepatic fxr and tgr5 expression levels. The HL treatment decreased the abundance of certain probiotic bacteria (e.g., Microbacterium, Cetobacterium, and Gemmobacter) and significantly increased the pathways involved in cytochrome P450, p53 signaling, and ATP-binding cassette (ABC) transporters. The HLB group increased some probiotic bacteria abundance, particularly BSH-producing bacteria (e.g., Escherichia Shigella). Compared with the HLB group, the abundance of BSH-producing bacteria (e.g., Bifidobacterium and Enterococcus) and pathways related to Notch signaling and Wnt signaling were reduced in the HLAB group. Conclusions: This study revealed that berberine’s lipid-lowering and intestine-protective effects are closely related to the intestinal microbiota, especially BSH-producing bacteria. Full article

The distinctive clinicopathologic characteristics of OLP indicated that both microbial dysbiosis and neurogenic inflammation may be jointly involved in its progression, and transient receptor potential vanilloid receptor-1 (TRPV1) may be a crucial element. The purpose of this study was to explore how TRPV1 mediated P. melaninogenica-induced inflammation. Meanwhile, we aimed to unravel how IL-36γ dysregulated the barrier function in oral keratinocytes. Here, the expression of TRPV1, calcitonin gene-related peptide (CGRP), and its receptor receptor activity-modifying protein 1 (RAMP1) in OLP patients were detected. Prevotella melaninogenica (P. melaninogenica) was used to build a mouse model of oral chronic inflammation. Normal human oral keratinocytes (NHOKs) stimulated by P. melaninogenica were used to examine TRPV1 activation and CGRP release. To investigate the effect of exogenous CGRP on Interleukin-36 gamma (IL-36γ) expression in NHOKs and bacterial viability, P. melaninogenica and NHOKs were treated with it, respectively. Recombinant IL-36γ protein was used to probe its regulation of oral epithelial barrier function. TRPV1, CGRP, and RAMP1 were substantially expressed in OLP. P. melaninogenica increased TRPV1 expression in mice and caused the release of CGRP and an increase in pro-inflammatory cytokines via activating TRPV1 in NHOKs. Blockade of TRPV1 suppressed P. melaninogenica-induced inflammation. CGRP boosted the production of IL-36γ released by NHOKs, resulting in lower expression of zonula occludens-1 (ZO-1). Also, CGRP can decrease the viability of P. melaninogenica. Together, these findings provide fresh insight into the vital role performed by P. melaninogenica-induced functional changes in oral epithelial cells and neurons in an intricate OLP inflammatory process. Full article

Limosilactobacillus reuteri, a recognized probiotic, improves intestinal health in animals, but the mechanism remains unclear. This study investigates the mechanisms by which L. reuteri ZY15, isolated from healthy pig feces, mitigates intestinal barrier damage and inflammation caused by oxidative stress in Enterotoxigenic Escherichia coli (ETEC) K88-challenged mice. The results indicated that L. reuteri ZY15 increased antioxidant capacity by reducing serum reactive oxygen species (ROS) and superoxide dismutase (SOD) levels. L. reuteri ZY15 enhanced the intestinal barrier by upregulating mucin 1, mucin 2, occludin, zonula occludens-1 (ZO-1), and claudin-1 expressions in protein and mRNA levels. It significantly alleviated intestinal inflammation by reducing the proinflammatory cytokines interleukin-1β (IL-1β), interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and interleukin-17 (IL-17) mRNA and protein levels. Notably, L. reuteri ZY15 suppressed intestinal inflammation by inhibiting AKT/mTOR/HIF-1α/RORγt/IL-17 pathway activation. Additionally, it significantly altered the structure of gut microorganisms by enriching Akkermansia and Clostridia_UCG.014, and thereby re-establishing colonization resistance and alleviating ETEC K88-induced intestinal barrier damage and inflammation in mice. Taken together, our findings reveal the protective mechanism of L. reuteri ZY15 in mice challenged with ETEC K88 by regulating AKT/mTOR/HIF-1α/RORγt/IL-17 signaling and microbial imbalance. Leveraging these properties, live L. reuteri ZY15 offers a promising alternative treatment for Escherichia coli-induced diarrhea in weaned piglets. Full article

Background: (-)-Fenchone is a bicyclic monoterpene present in the plant species Foeniculum vulgare Mill, Thuja occidentalis L. (tuja), and Lavandula stoechas (lavender). These plants have therapeutic value in the treatment of intestinal disorders. Aim: To evaluate intestinal anti-inflammatory activity in an acute and chronic trinitrobenzene sulphonic acid (TNBS)-induced colitis model in rats. Methods: Intestinal anti-inflammatory effects were assessed using the acute and chronic TNBS-induced colitis model in rats. The mechanisms were evaluated from colonic tissue fragments of the acute and chronic models. Results: Oral administration of the (-)-fenchone (37.5–300 mg/kg) acute phase or (150 mg/kg) (p < 0.001) chronic phase reduced the macroscopic lesion score, ulcerative area, intestinal weight/length ratio, and diarrheal index in TNBS-treated animals. At a dose of 150 mg/kg, the acute and chronic phase decreased malondialdehyde (MDA) and myeloperoxidase (MPO) (p < 0.001), restored glutathione (GSH) levels and superoxide dismutase (SOD) (p < 0.001), decreased immunomarking for factor nuclear kappa B (NF-κB) and levels of interleukin (IL)-1 and tumor necrosis factor α (TNF-α), and maintained IL-10 and TGF-β basal levels. Furthermore, increased immunostaining for zonula occludens 1 (ZO-1) was observed. Conclusions: (-)-fenchone has intestinal anti-inflammatory activity related to cytoprotection of the intestinal barrier, as well as antioxidant and immunomodulatory effects. Full article

This article aims to investigate the mechanism by which Bacillus amyloliquefaciens alleviates lipopolysaccharide (LPS)-induced intestinal oxidative stress. The study involved two experimental subjects: human colorectal adenocarcinoma (Caco-2) cells and Arbor Acres broiler chickens. The experiment involving two samples was designed with the same treatment groups, specifically the control (CK) group, lipopolysaccharide (LPS) group, Bacillus amyloliquefaciens (JF) group, and JF+LPS group. In the Caco-2 experiment, we administered 2 μg/mL of LPS and 1 × 10⁶ CFU/mL of JF to the LPS and JF groups, respectively. In the broiler experiment, the LPS group (19–21 d) received an abdominal injection of 0.5 mg/kg BW of LPS, whereas the JF group was fed 1 × 10⁷ CFU/g of JF throughout the entire duration of the experiment (1–21 d). The results indicated the following: (1) JF significantly decreased the DPPH free radical clearance rate and hydrogen peroxide levels (p < 0.05). (2) JF significantly enhanced the total antioxidant capacity (T-AOC), superoxide dismutase (SOD), and glutathione peroxidase (GSH Px) activity in Caco-2 cells (p < 0.05), while concurrently reducing malondialdehyde (MDA) content (p < 0.05). (3) Compared to the CK group, JF significantly increased the mRNA expression levels of nuclear factor-erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), SOD, catalase (CAT), GSH-Px, interleukin-4 (IL-4), interleukin-10 (IL-10), Claudin, Occludin1, zonula occludens-1 (ZO-1), and mucin 2 (MUC2) in Caco-2 cells (p < 0.05), while concurrently reducing the mRNA expression of Kelch-like ECH-associated protein 1 (Keap1), tumor necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β), and interleukin-8 (IL-8) (p < 0.05). In comparison to the LPS group, the JF+LPS group demonstrated a significant increase in the mRNA expression of Nrf2, SOD, GSH-Px, and IL-4, as well as Occludin1, ZO-1, and MUC2 in Caco-2 cells (p < 0.05), alongside a decrease in the mRNA expression of Keap1, TNF-α, and IL-1β (p < 0.05). (4) In broiler chickens, the JF group significantly elevated the levels of T-AOC, CAT, and GSH-Px in the jejunum while reducing MDA content (p < 0.05). Furthermore, the CAT level in the JF+LPS group was significantly higher than that observed in the LPS group, and the levels of MDA, TNF-α, and IL-1β were significantly decreased (p < 0.05). (5) In comparison to the CK group, the JF group exhibited a significant increase in Nrf2 levels in the jejunum of broiler chickens (p < 0.05). Notably, the mRNA expression levels of IL-4, IL-10, Claudin, Occludin1, ZO-1, and MUC2 were reduced (p < 0.05), while the mRNA expression levels of Keap1, TNF-α, and IL-1β also showed a decrease (p < 0.05). Furthermore, the mRNA expression levels of Nrf2, Occludin1, ZO-1, and MUC2 in the JF+LPS group were significantly elevated compared to those in the LPS group (p < 0.05), whereas the mRNA expression levels of Keap1 and TNF-α were significantly diminished (p < 0.05). In summary, JF can enhance the intestinal oxidative stress response, improve antioxidant capacity and intestinal barrier function, and decrease the expression of inflammatory factors by regulating the Keap1/Nrf2 signaling pathway. Full article

Artemisia annua residue (AR), as the byproduct of industrial extraction of artemisinin, contains rich nutrients and active ingredients. This study was conducted to determine the effects of AR as an unconventional feed material on growth performance, immunity, and intestinal health in weaned piglets. Thirty-two piglets weaned at 21 days (7.53 ± 0.31 kg average BW) were fed with a corn–soybean basal diet (BD) and a basal diet with 1% (LAR), 2% (MAR), and 4% (HAR) AR diets for 28 days. AR diets increased the serum IgA and complement component 3 levels, superoxide dismutase activity, and villus height in the duodenum (p < 0.05). The MAR group increased the ADG, serum total protein, and mRNA expression levels of Claudin-1 in the duodenum and zonula occludens-1 (ZO-1) and the mucin 2 (MUC2) in the colon, as well as colonic Romboutsia and Anaerostipes abundances, and decreased the Proteobacteria abundance (p < 0.05). To sum up, dietary AR supplementation may enhance growth performance by improving serum immunoglobulin and antioxidant enzyme activity, intestinal morphology, tight junction protein expression, and gut microbiota of weaned piglets. Regression analysis showed that the optimal AR supplemental level for growth performance, immunity, antioxidant ability, and intestinal health of weaned piglets was 2.08% to 4.24%. Full article

Background: Stroke is one of the leading causes of death and disability worldwide. The diagnosis of stroke remains largely clinical, yet widely used stroke scoring systems and brain imaging do not satisfactorily allow the distinction of ischaemic stroke (IS) patients from stroke mimics (SMs). Blood biomarkers are promising tools that could facilitate clinical triage. Methods: This study recruited 66 patients with IS and 24 SMs. The levels of Glial fibrillary acidic protein (GFAP), Neuron-specific enolase (NSE), Neurofilament light chain (NfL) and blood-brain barrier (BBB) proteins [Occludin (OCLN), Zonula occludens 1 (ZO-1), Claudin-5] in blood serum were measured by enzyme-linked immunosorbent assay technique. Biomarker levels in IS patients and SMs were compared using the Mann–Whitney U test. Multivariable logistic regression analysis was used to evaluate the diagnostic performance of biomarkers in combination with the National Institutes of Health Stroke Scale (NIHSS) score. Results: More significant differences in circulating GFAP, NfL, OCLN, ZO-1, and Claudin-5 but not NSE were found in IS patients compared to SMs. A combination of circulating ZO-1, Claudin-5, and OCLN with NIHSS score gives the highest diagnostic accuracy, sensitivity, and specificity. Conclusions: A prediction model with circulating BBB proteins in combination with NIHSS score differentiates between IS patients and SMs. Full article