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Intestinal permeabilization is central to the pathophysiology of chronic gut inflammation. This study investigated the efficacy of glucoraphanin (GR), prevalent in cruciferous vegetables, particularly broccoli, and its derivative sulforaphane (SF), in inhibiting tumor necrosis factor alpha (TNFα)-induced Caco-2 cell monolayers inflammation and permeabilization through the regulation of redox-sensitive events. TNFα binding to its receptor led to a rapid increase in oxidant production and subsequent elevation in the mRNA levels of NOX1, NOX4, and Duox2. GR and SF dose-dependently mitigated both these short- and long-term alterations in redox homeostasis. Downstream, GR and SF inhibited the activation of the redox-sensitive signaling cascades NF-κB (p65 and IKK) and MAPK ERK1/2, which contribute to inflammation and barrier permeabilization. GR (1 μM) and SF (0.5-1 μM) prevented TNFα-induced monolayer permeabilization and the associated reduction in the levels of the tight junction (TJ) proteins occludin and ZO-1. Both GR and SF also mitigated TNFα-induced increased mRNA levels of the myosin light chain kinase, which promotes TJ opening. Molecular docking suggests that although GR is mostly not absorbed, it could interact with extracellular and membrane sites in NOX1. Inhibition of NOX1 activity by GR would mitigate TNFα receptor downstream signaling and associated events. These findings support the concept that not only SF, but also GR, could exert systemic health benefits by protecting the intestinal barrier against inflammation-induced permeabilization, in part by regulating redox-sensitive pathways. GR has heretofore not been viewed as a biologically active molecule, but rather, the benign precursor of highly active SF. The consumption of GR and/or SF-rich vegetables or supplements in the diet may offer a means to mitigate the detrimental consequences of intestinal permeabilization, not only in disease states but also in conditions characterized by chronic inflammation of dietary and lifestyle origin.

Endothelial cells of mammalian blood vessels have multiple levels of heterogeneity along the vascular tree and among different organs. Further heterogeneity results from blood flow turbulence and variations in shear stress. In the aorta, vascular endothelial protein tyrosine phosphatase (VE-PTP), which dephosphorylates tyrosine kinase receptor Tie2 in the plasma membrane, undergoes downstream polarization and endocytosis in endothelial cells exposed to laminar flow and high shear stress. VE-PTP sequestration promotes Tie2 phosphorylation at tyrosine992 and endothelial barrier tightening. The present study characterized the heterogeneity of VE-PTP polarization, Tie2-pY992 and total Tie2, and claudin-5 in anatomically defined regions of endothelial cells in the mouse descending thoracic aorta, where laminar flow is variable and IgG extravasation is patchy. We discovered that VE-PTP and Tie2-pY992 had mosaic patterns, unlike the uniform distribution of total Tie2. Claudin-5 at tight junctions also had a mosaic pattern, whereas VE-cadherin at adherens junctions bordered all endothelial cells. Importantly, the amounts of Tie2-pY992 and claudin-5 in aortic endothelial cells correlated with downstream polarization of VE-PTP. VE-PTP and Tie2-pY992 also had mosaic patterns in the vena cava, but claudin-5 was nearly absent and extravasated IgG was ubiquitous. Correlation of Tie2-pY992 and claudin-5 with VE-PTP polarization supports their collective interaction in the regulation of endothelial barrier function in the aorta, yet differences between the aorta and vena cava indicate additional flow-related determinants of permeability. Together, the results highlight new levels of endothelial cell functional mosaicism in the aorta and vena cava, where blood flow dynamics are well known to be heterogeneous.

Formation of fluid-filled lumina by epithelial tissues is essential for organ development. How cells control the hydraulic and cortical forces to control lumen morphology is not well understood. Here, we quantified the mechanical role of tight junctions in lumen formation using MDCK-II cysts. We found that the paracellular ion barrier formed by claudin receptors is not required for the hydraulic inflation of a lumen. However, the depletion of the zonula occludens scaffold resulted in lumen collapse and folding of apical membranes. Combining quantitative measurements of hydrostatic lumen pressure and junctional tension with modeling enabled us to explain lumen morphologies from the pressure-tension force balance. Tight junctions promote lumen inflation by decreasing cortical tension via the inhibition of myosin. In addition, our results suggest that excess apical area contributes to lumen opening. Overall, we provide a mechanical understanding of how epithelial cells use tight junctions to modulate tissue and lumen shape. [Display omitted] • Depletion of ZO scaffold causes lumen collapse and apical membrane folding • The claudin permeability barrier is not necessary for hydraulic lumen inflation • Tight junctions promote lumen inflation by decreasing cortical tension • Lumen shape controlled by pressure-tension force balance and apical area constraints Mukenhirn et al. show that epithelial cells use tight junctions to control lumen shape, and the depletion of the tight junction scaffold leads to lumen collapse and increased cortical tension. Claudin-depleted MDCK-II cysts maintain lumen pressure and shape despite high ion permeability. [ABSTRACT FROM AUTHOR]

Background: Glioblastoma multiforme (GBM) is an aggressive tumor, difficult to treat pharmacologically because of the blood-brain barrier (BBB), which is rich in ATP-binding cassette (ABC) transporters and tight junction (TJ) proteins. The BBB is disrupted within GBM bulk, but it is competent in brain-adjacent-to-tumor areas, where eventual GBM foci can trigger tumor relapse. How GBM cells influence the permeability of BBB is poorly investigated. Methods: To clarify this point, we co-cultured human BBB models with 3 patient-derived GBM cells, after separating from each tumor the stem cell/neurosphere (SC/NS) and the differentiated/adherent cell (AC) components. Also, we set up cultures of BBB cells with the conditioned medium of NS or AC, enriched or depleted of IL-6. Extracellular cytokines were measured by protein arrays and ELISA. The intracellular signaling in BBB cells was measured by immunoblotting, in the presence of STAT3 pharmacological inhibitor or specific PROTAC. The competence of BBB was evaluated by permeability assays and TEER measurement. Results: The presence of GBM cells or their conditioned medium increased the permeability to doxorubicin, mitoxantrone and dextran-70, decreased TEER, down-regulated ABC transporters and TJ proteins at the transcriptional level. These effects were higher with AC or their medium than with NS. The secretome analysis identified IL-6 as significantly more produced by AC than by NS. Notably, AC-conditioned medium treated with an IL-6 neutralizing antibody reduced the BBB permeability to NS levels, while NS-conditioned medium enriched with IL-6 increased BBB permeability to AC levels. Mechanistically, IL-6 released by AC GBM cells activated STAT3 in BBB cells. In turn, STAT3 down-regulated ABC transporter and TJ expression, increased permeability and decreased TEER. The same effects were obtained in BBB cells treated with STA-21, a pharmacological inhibitor of STAT3, or with a PROTAC targeting STAT3. Conclusions: Our work demonstrates for the first time that the degree of GBM differentiation influences BBB permeability. The crosstalk between GBM cells that release IL-6 and BBB cells that respond by activating STAT3, controls the expression of ABC transporters and TJ proteins on BBB. These results may pave the way for novel therapeutic tools to tune BBB permeability and improve drug delivery to GBM. [ABSTRACT FROM AUTHOR]

Cell junctions, including anchoring, occluding and communicating junctions, play an indispensable role in the structural and functional organization of multicellular tissues, including in liver. Specifically, hepatic cell junctions mediate intercellular adhesion and communication between liver cells. The establishment of the hepatic cell junction network is a prerequisite for normal liver functioning. Hepatic cell junctions indeed support liver‐specific features and control essential aspects of the hepatic life cycle. This review paper summarizes the role of cell junctions and their components in relation to liver physiology, thereby also discussing their involvement in hepatic dysfunctionality, including liver disease and toxicity. [ABSTRACT FROM AUTHOR]

Humans can be exposed to per- and polyfluoroalkyl substances (PFASs) via many exposure routes, including diet, which may lead to several adverse health effects. So far, little is known about PFAS transport across the human intestinal barrier. In the current study, we aimed to assess the transport of 5 PFASs (PFOS, PFOA, PFNA, PFHxS and HFPO-DA) in a human induced pluripotent stem cell (hiPSC)-derived intestinal epithelial cell (IEC) model. This model was extensively characterized and compared with the widely applied human colonic adenocarcinoma cell line Caco-2 and a human primary IEC-based model, described to most closely resemble in vivo tissue. The hiPSC-derived IEC layers demonstrated polarized monolayers with tight junctions and a mucus layer. The monolayers consisted of enterocytes, stem cells, goblet cells, enteroendocrine cells, and Paneth cells that are also present in native tissue. Transcriptomics analysis revealed distinct differences in gene expression profiles, where the hiPSC-derived IECs showed the highest expression of intestinal tissue-specific genes relative to the primary IEC-based model and the Caco-2 cells clustered closer to the primary IEC-based model than the hiPSC-derived IECs. The order of PFAS transport was largely similar between the models and the apparent permeability (Papp) values of PFAS in apical to basolateral direction in the hiPSC-derived IEC model were in the following order: PFHxS > PFOA > HFPO-DA > PFNA > PFOS. In conclusion, the hiPSC-derived IEC model highly resembles human intestinal physiology and is therefore a promising novel in vitro model to study transport of chemicals across the intestinal barrier for risk assessment of chemicals. [ABSTRACT FROM AUTHOR]

Proteins and peptides have been increasingly developed as pharmaceuticals owing to their high potency and low side effects. However, their administration routes are confined to injections, such as intra-muscular and intra-venous injections, making patient compliance a challenge. Hence, non-injectable delivery systems are crucial to expanding the clinical use of proteins and peptides. In this context, two choline-based ionic liquids (ILs), namely, choline geranic acid ([Ch][Ger]) and choline citric acid ([Ch][Cit]), have been identified as promising agents for enhancing the permeation and prolonging the retention time of glucagon (GC) after intra-nasal administration. Notably, intra-nasal delivery of GC via ILs (GC/ILs) elicited rapid and smooth reversal of acute hypoglycaemia without leading to rebound hyperglycaemia in type 1 diabetic rats subjected to insulin induction. In addition, ILs could improve the transcellular transport of GC through electrostatic interaction. ILs could also transiently open inter-cellular tight junctions transiently to facilitate the paracellular transport of GC. Safety tests indicated that continuous intra-nasal delivery of ILs led to reversible changes, such as epithelial cell inflammation, goblet cell overgrowth, and impacts on the distribution of nasal cilia. However, these changes could be alleviated by the innate self-repair ability of mucosal epithelial cells. This study highlights the considerable potential of ILs for long-term nasal delivery of biomacromolecules. [Display omitted] • Intra-nasal delivery of glucagon in ionic liquids (GC/ILs) for acute hypoglycaemia rescue • ILs improved the stability and solubility of GC through hydrogen bonding interactions. • ILs enhanced the paracellular transport of GC as a novel transient permeation enhancer. • The water content of ILs significantly affected the nasal retention and efficacy of GC/ILs. • The long-term regimen of nasal delivery of GC/ILs showed a favourable safety profile. [ABSTRACT FROM AUTHOR]

Background: Altered gut metabolites, especially short-chain fatty acids (SCFAs), in feces and plasma are observed in patients with Parkinson's disease (PD). Objective: We aimed to investigate the colonic expression of two SCFA receptors, free fatty acid receptor (FFAR)2 and FFAR3, and gut barrier integrity in patients with PD and correlations with clinical severity. Methods: In this retrospective study, colonic biopsy specimens were collected from 37 PD patients and 34 unaffected controls. Of this cohort, 31 participants (14 PD, 17 controls) underwent a series of colon biopsies. Colonic expression of FFAR2, FFAR3, and the tight junction marker ZO-1 were assayed by immunofluorescence staining. The You Only Look Once (version 8, YOLOv8) algorithm was used for automated detection and segmentation of immunostaining signal. PD motor function was assessed with the Movement Disorder Society (MDS)-Unified Parkinson's Disease Rating Scale (UPDRS), and constipation was assessed using Rome-IV criteria. Results: Compared with controls, PD patients had significantly lower colonic expression of ZO-1 (p < 0.01) and FFAR2 (p = 0.01). On serial biopsy, colonic expression of FFAR2 and FFAR3 was reduced in the pre-motor stage before PD diagnosis (both p < 0.01). MDS-UPDRS motor scores did not correlate with colonic marker levels. Constipation severity negatively correlated with colonic ZO-1 levels (r = -0.49, p = 0.02). Conclusions: Colonic expression of ZO-1 and FFAR2 is lower in PD patients compared with unaffected controls, and FFAR2 and FFAR3 levels decline in the pre-motor stage of PD. Our findings implicate a leaky gut phenomenon in PD and reinforce that gut metabolites may contribute to the process of PD. [ABSTRACT FROM AUTHOR]

The consumption of fructose is increasing day by day. Understanding the impact of increasing fructose consumption on the small intestine is crucial since the small intestine processes fructose into glucose. ∆9-Tetrahydrocannabinol (THC), a key cannabinoid, interacts with CB1 and CB2 receptors in the gastrointestinal tract, potentially mitigating inflammation. Therefore, this study aimed to investigate the effects of the high-fructose diet (HFD) on the jejunum of rats and the role of THC consumption in reversing these effects. Experiments were conducted on Sprague–Dawley rats, with the experimental groups as follows: control (C), HFD, THC, and HFD + THC. The HFD group received a 10% fructose solution in drinking water for 12 weeks. THC groups were administered 1.5 mg/kg/day of THC intraperitoneally for the last four weeks. Following sacrification, the jejunum was evaluated for mucus secretion capacity. IL-6, JNK, CB2 and PCNA expressions were assessed through immunohistochemical analysis and the ultrastructural alterations via transmission electron microscopy. The results showed that fructose consumption did not cause weight gain but triggered inflammation in the jejunum, disrupted the cell proliferation balance, and increased mucus secretion in rats. Conversely, THC treatment displayed suppressed inflammation and improved cell proliferation balance caused by HFD. Ultrastructural examinations showed that the zonula occludens structures deteriorated in the HFD group, along with desmosome shrinkage. Mitochondria were found to be increased due to THC application following HFD. In conclusion, the findings of this research reveal the therapeutic potential of THC in reversing HFD-related alterations and provide valuable insights for clinical application. [ABSTRACT FROM AUTHOR]

Introduction: Tight junctions (TJs) and their constituent proteins play pivotal roles in cellular physiology and anatomy by establishing functional boundaries within and between neighboring cells. While the involvement of TJ proteins, such as claudins, in cancer is extensively studied, studies highlighting their interaction with immune system are still meager. Studies indicate that alterations in cytokines and immune cell populations can affect TJ proteins, compromising TJ barrier function and exacerbating pro-inflammatory conditions, potentially leading to epithelial cell malignancy. Disrupted TJs in established tumors may foster a pro-tumor immune microenvironment, facilitating tumor progression, invasion, epithelial-to-mesenchymal transition and metastasis. Although previous literature contains many studies describing the involvement of TJs in pathogenesis of malignancies their role in modulating the immune microenvironment of tumors is just beginning to be unleashed. Areas covered: This article for the first time attempts to discern the importance of interaction between TJs and immune microenvironment in malignancies. To achieve the above aim a thorough search of databases like PubMed and Google Scholar was conducted to identify the recent and relevant articles on the topic. Expert opinion: Breaking the vicious cycle of dysbiosis/infections/chemical/carcinogen-induced inflammation-TJ remodeling-malignancy-TJ dysregulation-more inflammation can be used as a strategy to complement the effect of immunotherapies in various malignancies. [ABSTRACT FROM AUTHOR]

The prolonged exposure to arsenic results in intestinal barrier dysfunction, which is strongly concerned with detrimental processes such as oxidative stress and the inflammatory response. Ferulic acid (FA), as a phenolic acid, possesses the capability to mitigate arsenic‐induced liver damage and cardiotoxic effects dependent on inhibition of oxidative stress and inflammatory responses. FA can mitigate testicular tissue damage and alveolar epithelial dysfunction, the mechanism of which may rely on nuclear factor erythroid 2‐related factor 2/heme oxygenase 1 (Nrf2/HO‐1) activation and nuclear factor‐kappa B (NF‐κB) pathway blocking. Based on the antioxidant and anti‐inflammatory properties of FA, we speculated that FA might have the potential to inhibit arsenic‐induced intestinal damage. To confirm this scientific hypothesis, mice exposed to sodium arsenite were treated with FA to observe colonic histopathology and TJ protein levels, and oxidative stress and TJ protein levels in Caco‐2 cells exposed to sodium arsenite were assessed after FA intervention. In addition, molecular levels of NF‐κB and Nrf2/HO‐1 pathway in colon and Caco‐2 cells were also detected. As shown in our data, FA inhibited arsenic‐induced colon injury, which was reflected in the improvement of mucosal integrity, the decrease of down‐regulated expression of tight junction (TJ) proteins (Claudin‐1, Occludin, and ZO‐1) and the inhibition of oxidative stress. Similarly, treatment with FA attenuated the inhibitory effect of arsenic on TJ protein expression in Caco‐2 cells. In addition to suppressing the activation of NF‐κB pathway, FA retrieved the activation of Nrf2/HO‐1 pathway in colon and intestinal epithelial cells induced by arsenic. In summary, our findings propose that FA has the potential to mitigate arsenic‐induced intestinal damage by preserving the integrity of intestinal epithelial TJs and suppressing oxidative stress. These results lay the groundwork for the potential use of FA in treating colon injuries caused by arsenic. [ABSTRACT FROM AUTHOR]

Reperfusion after cerebral ischemia leads to secondary damage to the nervous system, called cerebral ischemia/reperfusion injury (CIRI). The blood–brain barrier (BBB) consists of endothelial cells and tight junction (TJ) proteins, and its disruption aggravates CIRI. Two GSE datasets identified Tissue Factor Pathway Inhibitor 2 (TFPI2) as a differentially upregulated gene (Log2FC > 1, p < 0.01) in the cerebral cortex of ischemic rats, and TFPI2 affects angiogenesis of endothelial cells. Moreover, genes (c-Jun, c-Fos, FosL1) encoding subunits of Activator Protein-1 (AP-1), a transcription factor involved in IRI, were highly expressed in ischemic samples. Thus, the effects of the AP-1/TFPI2 axis on CIRI were explored. We determined increased TFPI2 expression in the cerebral cortex of rats receiving middle cerebral artery occlusion (MCAO) for 90 min and reperfusion (R) for 48 h. Then AAV2-shTFPI2 particles (5 × 1010 vg) were injected into the right lateral ventricle of rats 3 weeks before MCAO/R. TFPI2 knockdown decreased infarct size and neuronal injury in ischemic rats. It improved BBB integrity, demonstrated by reduced FITC-dextran leakage in brain tissues of MCAO/R-operated rats. Furthermore, it increased the expression of TJ proteins (Occludin, Claudin-5, TJP-1) in the cerebral cortex of rats with CIRI. Consistently, we found that TFPI2 knockdown mitigated cell damage in mouse endothelial bEND.3 cells with oxygen and glucose deprivation (ODG) for 6 h and reoxygenation (R) for 18 h (OGD/R) treatment. High co-expression of c-Jun and c-Fos significantly elevated TFPI2 promoter activity. c-Jun knockdown inhibited TFPI2 expression in OGD/R-treated bEND.3 cell. Collectively, our findings demonstrate that inhibition of the AP-1/TFPI2 axis alleviates CIRI. [ABSTRACT FROM AUTHOR]

Oleanolic acid (OA) is a naturally occurring pentacyclic triterpene compound that has been reported to cause cholestatic liver injury. However, the regulation and pathogenic role of bile acids in OA‐induced development of cholestatic liver injury remains largely unclear. Farnesoid X receptor (FXR) is a metabolic nuclear receptor that plays an important role in bile acid homeostasis in the liver by regulating efflux transporters bile salt export pump (BSEP) and multidrug resistance‐associated protein 2 (MRP2). The aim of this study was to investigate the effect of OA on hepatocyte tight junction function and determine the role of FXR, BSEP, and MRP2 in the mechanism of impairment of transport of bile acids induced by OA. Both in vivo and in vitro models were used to characterize the OA‐induced liver injury. The liquid chromatography–tandem mass spectrometry (LC‐MS) was employed to characterize the efflux function of the transporters, and the results showed that OA caused a blockage of bile acids efflux. OA treatment resulted in decreased expression levels of the tight junction proteins zonula occludens‐1 and occludin. Immunofluorescence results showed that OA treatment significantly reduced the number of bile ducts and the immunofluorescence intensity. Pretreatment with agonists of FXR and MRP2, respectively, in animal experiments attenuated OA‐induced liver injury, while pretreatment with inhibitors of BSEP and MRP2 further aggravated OA‐induced liver injury. These results suggest that OA inhibits FXR‐mediated BSEP and MRP2, leading to impaired bile acid efflux and disruption of tight junctions between liver cells, resulting in liver damage. Oleanolic acid (OA) is a naturally occurring pentacyclic triterpene compound that has been reported to cause cholestatic liver injury. In this study, OA inhibits FXR‐mediated BSEP and MRP2, leading to impaired bile acid efflux and disruption of tight junctions between liver cells, resulting in liver damage. Activation of FXR and MRP2 mitigated OA‐induced liver injury, while inhibition of BSEP and MRP2 further aggravated OA‐induced liver injury. [ABSTRACT FROM AUTHOR]

Acute respiratory distress syndrome (ARDS) is a severe lung disease characterized by significant hypoxemia, which impairs the oxygen supply necessary for optimal lung function. This study aimed to investigate the effects of sodium propionate (SP), the primary end product of intestinal flora fermentation of dietary fiber, on lipopolysaccharide (LPS)-induced ARDS in rats. The rats were treated with SP, after which the lung wet/dry ratio, arterial partial oxygen pressure (PaO2), levels of pro- and anti-inflammatory cytokines, tight junction proteins ZO-1 and Occludin, as well as LC3 and phosphorylated PI3K (p-PI3K)/p-AKT/p-mTOR protein levels, were measured. Additionally, histopathological analysis was conducted. The results indicated that SP effectively alleviated arterial hypoxemia in rats and mitigated the pathological damage to both intestinal and lung tissues caused by LPS. Notably, SP significantly reduced the levels of inflammatory factors TNF-α and IL-6 in the blood and bronchoalveolar lavage fluid (BALF) of ARDS rats, while increasing the concentration of the anti-inflammatory factor IL-10. Furthermore, SP inhibited the activation of the PI3K/AKT/mTOR signaling pathway and enhanced the LC3II/LC3I ratio in lung tissue. Therefore, SP may improve LPS-induced ARDS in rats by inhibiting the activation of the PI3K/AKT/mTOR signaling pathway, promoting autophagy, decreasing the production and release of inflammatory markers, and reducing alveolar epithelial damage. [ABSTRACT FROM AUTHOR]

Background: Lactiplantibacillus species are extensively studied for their ability to regulate host immune responses and functional therapeutic potentials. Nevertheless, there is a lack of understanding on the mechanisms of interactions with the hosts during immunoregulatory activities. Methods: Two Lactiplantibacillus plantarum strains MKMB01 and MKMB02 were tested for probiotic potential following Indian Council of Medical Research (ICMR) guidelines. Human colorectal adenocarcinoma cells such as HT-29, caco-2, and human monocytic cell THP-1 were also used to study the potential of MKMB01 and MKMB02 in regulating the host immune response when challenged with enteric pathogen Salmonella enterica typhimurium. Cells were pre-treated with MKMB01 and MKMB02 for 4 h and then stimulated with Salmonella. qRT-PCR and ELISA were used to analyze the genes and protein expression. Confocal microscopy and field emission scanning electron microscopy (FESEM) were used to visualize the effects. An Agilent Seahorse XF analyzer was used to determine real-time mitochondrial functioning. Results: Both probiotic strains could defend against Salmonella by maintaining gut integrity via expressing tight junction proteins (TJPs), MUC-2, and toll-like receptors (TLRs) negative regulators such as single Ig IL-1-related receptor (SIGIRR), toll-interacting protein (Tollip), interleukin-1 receptor-associated kinase (IRAK)-M, A20, and anti-inflammatory transforming growth factor-β and interleukin-10. Both strains also downregulated the expression of pro-inflammatory cytokines/chemokines interleukin-1β, monocyte chemoattractant protein (MCP)-1, tumor necrosis factor-alpha (TNF-α), interleukin 6, and nitric oxide (NO). Moreover, TNF-α sheddase protein, a disintegrin and metalloproteinase domain 17 (ADAM17), and its regulator iRhom2 were downregulated by both strains. Moreover, the bacteria also ameliorated Salmonella-induced mitochondrial dysfunction by restoring bioenergetic profiles, such as non-mitochondrial respiration, spare respiratory capacity (SRC), basal respiration, adenosine triphosphate (ATP) production, and maximal respiration. Conclusions: MKMB01 and MKMB02 can reduce pathogen-induced gut-associated disorders and therefore should be further explored for their probiotic potential. [ABSTRACT FROM AUTHOR]

Contact inhibition of proliferation is a critical cell density control mechanism governed by the Hippo signalling pathway. The biochemical signalling underlying cell density‐dependent cues regulating Hippo signalling and its downstream effectors, YAP, remains poorly understood. Here, we reveal that the tight junction protein ZO‐2 is required for the contact‐mediated inhibition of proliferation. We additionally determined that the well‐established molecular players of this process, namely Hippo kinase LATS1 and YAP, are regulated by ZO‐2 and that the scaffolding function of ZO‐2 promotes the interaction with and phosphorylation of YAP by LATS1. Mechanistically, YAP is phosphorylated when ZO‐2 brings LATS1 and YAP together via its SH3 and PDZ domains, respectively, subsequently leading to the cytoplasmic retention and inactivation of YAP. In conclusion, we demonstrate that ZO‐2 maintains Hippo signalling pathway activation by promoting the stability of LATS1 to inactivate YAP. [ABSTRACT FROM AUTHOR]

Ciliopathies are a diverse group of disorders resulting from abnormalities in the development or function of multiple organs. While significant research has clarified the role of the primary cilium in transducing numerous signalling pathways, elucidating causes of neuronal and skeletal development disorders, the origins of other ciliopathy-related conditions, such as hepatic fibrocystic diseases, remain elusive. Additionally, attempts to correlate specific ciliary proteins with distinct phenotypes have been largely unsuccessful due to the variable and overlapping symptoms of ciliopathies. This study aims to elucidate the extraciliary roles of the protein B9D2 in the development of biliary dysgenesis, a condition present in Meckel-Gruber and Joubert syndromes caused by mutations in this protein. Traditionally, B9D2 is known for its role at the transition zone of the primary cilium in the transduction of signalling pathways notably Wingless and Hedgehog. Our work demonstrates that before ciliogenesis occurs, B9D2 is crucial for the maturation and maintenance of tight junctions ensuring epithelial barrier tightness and appropriate biliary lumen formation. This study provides new insights into the mechanisms underlying biliary dysgenesis in hepatic ciliopathies, suggesting that further exploration of the non-ciliary functions of proteins involved in ciliopathies could lead to a better understanding and treatment of these complex disorders. [ABSTRACT FROM AUTHOR]

In patients with rheumatoid arthritis (RA), intestinal flora imbalance and butyrate metabolism disorders precede clinical arthritis and are associated with the pathogenesis of RA. This imbalance can alter the immunology and intestinal permeability of the intestinal mucosa, leading to damage to the intestinal barrier. In this context, bacteria and their metabolites can enter the bloodstream and reach the distant target tissues of the host, resulting in local inflammation and aggravating arthritis. Additionally, arthritis is also exacerbated by bone destruction and immune tolerance due to disturbed differentiation of osteoclasts and adaptive immune cells. Of note, butyrate is a metabolite of intestinal flora, which not only locally inhibits intestinal immunity and targets zonulin and tight junction proteins to alleviate intestinal barrier-mediated arthritis but also inhibits osteoclasts and autoantibodies and balances the immune responses of T and B lymphocytes throughout the body to repress bone erosion and inflammation. Therefore, butyrate is a key intermediate linking intestinal flora to the host. As a result, restoring the butyrate-producing capacity of intestinal flora and using exogenous butyrate are potential therapeutic strategies for RA in the future. [ABSTRACT FROM AUTHOR]

Ulcerative colitis (UC) is a debilitating inflammatory bowel disease characterized by intestinal inflammation, barrier dysfunction, and dysbiosis, with limited treatment options available. This study systematically investigates the therapeutic potential of a synbiotic composed of galactooligosaccharides (GOS) and Limosilactobacillus reuteri in a murine model of colitis, revealing that GOS and L. reuteri synergistically protect against intestinal inflammation and barrier dysfunction by promoting the synthesis of pentadecanoic acid, an odd-chain fatty acid, from Bacteroides acidifaciens. Notably, the synbiotic, B. acidifaciens, and pentadecanoic acid are each capable of suppressing intestinal inflammation and enhancing tight junction by inhibiting NF-κB activation. Furthermore, similar reduction in B. acidifaciens and pentadecanoic acid levels are also observed in the feces from both human UC patients and lipopolysaccharide-induced intestinal inflammation in pigs. Our findings elucidate the protective mechanism of the synbiotic and highlight its therapeutic potential, along with B. acidifaciens and pentadecanoic acid, for UC and other intestinal inflammatory disorders. Here, Yujun Wu and colleagues report that a synbiotic composed of galactooligosaccharides and Limosilactobacillus reuteri alleviates gut inflammation in animals by enriching pentadecanoic acid synthesis from Bacteroides acidifaciens, showing potential for treating ulcerative colitis. [ABSTRACT FROM AUTHOR]

Sepsis is a global health challenge marked by limited clinical options and high mortality rates. AMP-activated protein kinase (AMPK) is a cellular energy sensor that mediates multiple crucial metabolic pathways that may be an attractive therapeutic target in sepsis. Pre-clinical experimental studies have demonstrated that pharmacological activation of AMPK can offer multiple potential benefits during sepsis, including anti-inflammatory effects, induction of autophagy, promotion of mitochondrial biogenesis, enhanced phagocytosis, antimicrobial properties, and regulation of tight junction assembly. This review aims to discuss the existing evidence supporting the therapeutic potential of AMPK activation in sepsis management. [ABSTRACT FROM AUTHOR]

Introduction: The resolution of the skin's inflammatory response is only possible if its barrier function is restored. TRPV1 channel activation plays an important role during inflammation but the effect of this activation on the skin barrier under inflammatory conditions has not been clarified. We hypothesize that it could potentially aid the keratinocyte barrier by reducing inflammatory cytokine release and promoting tight junction development. Methods: To explore the role of TRPV1 activation in inflammation, we designed and optimized an in vitro model of keratinocytes with basal epidermal layer characteristics using HaCaT cells and TNFα to induce inflammation. Results: TNFα increased the gene expression of tight junction protein claudin 1 (CLDN1) by at least 2.60 ± 0.16-fold, in a concentration-dependent manner, over a 48 h period. The administration of a capsaicin pre-treatment reduced the CLDN1 expression to 1.51 ± 0.16-fold during the first 6 h after TNFα induction, whereas IL-8 cytokine release was reduced 0.64 ± 0.17-fold. After 48 h, CLDN1 protein levels increased by a factor of 6.57 ± 1.39 compared to cells only treated with TNFα. Discussion: These results suggest that activation of TRPV1 by capsaicin can potentiate the increase in CLDN1 expression and CLDN1 protein synthesis induced by TNFα in cultured keratinocytes, while reducing the release of IL-8. [ABSTRACT FROM AUTHOR]

Background: Brain microglia and infiltrating monocyte-derived macrophages are vital in preserving blood vessel integrity after stroke. Understanding mechanisms that induce immune cells to adopt vascular-protective phenotypes may hasten the development of stroke treatments. IL-33 is a potent chemokine released from damaged cells, such as CNS glia after stroke. The activation of IL-33/ST2 signaling has been shown to promote neuronal viability and white matter integrity after ischemic stroke. The impact of IL-33/ST2 on blood-brain barrier (BBB) integrity, however, remains unknown. The current study fills this gap and reveals a critical role of IL-33/ST2 signaling in macrophage-mediated BBB protection after stroke. Methods: Transient middle cerebral artery occlusion (tMCAO) was performed to induce ischemic stroke in wildtype (WT) versus ST2 knockout (KO) male mice. IL-33 was applied intranasally to tMCAO mice with or without dietary PLX5622 to deplete microglia/macrophages. ST2 KO versus WT bone marrow or macrophage cell transplantations were used to test the involvement of ST2+ macrophages in BBB integrity. Macrophages were cocultured in transwells with brain endothelial cells (ECs) after oxygen-glucose deprivation (OGD) to test potential direct effects of IL33-treated macrophages on the BBB in vitro. Results: The ST2 receptor was expressed in brain ECs, microglia, and infiltrating macrophages. Global KO of ST2 led to more IgG extravasation and loss of ZO-1 in cerebral microvessels 3 days post-tMCAO. Intranasal IL-33 administration reduced BBB leakage and infarct severity in microglia/macrophage competent mice, but not in microglia/macrophage depleted mice. Worse BBB injury was observed after tMCAO in chimeric WT mice reconstituted with ST2 KO bone marrow, and in WT mice whose monocytes were replaced by ST2 KO monocytes. Macrophages treated with IL-33 reduced in vitro barrier leakage and maintained tight junction integrity after OGD. In contrast, IL-33 exerted minimal direct effects on the endothelial barrier in the absence of macrophages. IL-33-treated macrophages demonstrated transcriptional upregulation of an array of protective factors, suggesting a shift towards favorable phenotypes. Conclusion: Our results demonstrate that early-stage IL-33/ST2 signaling in infiltrating macrophages reduces the extent of acute BBB disruption after stroke. Intranasal IL-33 administration may represent a new strategy to reduce BBB leakage and infarct severity. [ABSTRACT FROM AUTHOR]

Giardia duodenalis, an important zoonotic protozoan parasite, adheres to host intestinal epithelial cells (IECs) via the ventral disc and causes giardiasis characterized mainly by diarrhea. To date, it remains elusive how excretory-secretory products (ESPs) of Giardia enter IECs and how the cells respond to the entry. Herein, we initially demonstrated that ESPs evoked IEC endocytosis in vitro. We indicated that ESPs contributed vitally in triggering intrinsic apoptosis, pro-inflammatory responses, tight junction (TJ) protein expressional changes, and autophagy in IECs. Endocytosis was further proven to be implicated in those ESPs-triggered IEC responses. Ten predicted virulent excretory-secretory proteins of G. duodenalis were investigated for their capability to activate clathrin/caveolin-mediated endocytosis (CME/CavME) in IECs. Pyridoxamine 5'-phosphate oxidase (PNPO) was confirmed to be an important contributor. PNPO was subsequently verified as a vital promoter in the induction of giardiasis-related IEC apoptosis, inflammation, and TJ protein downregulation. Most importantly, this process seemed to be involved majorly in PNPO-evoked CME pathway, rather than CavME. Collectively, this study identified Giardia ESPs, notably PNPO, as potentially important pathogenic factors during noninvasive infection. It was also noteworthy that ESPs-evoked endocytosis might play a role in triggering giardiasis-inducing cellular regulation. These findings would deepen our understanding about the role of ESPs, notably PNPO, in the pathogenesis of giardiasis and the potential attributed endocytosis mechanism. [ABSTRACT FROM AUTHOR]

Apical cell-cell junctions, including adherens junctions and tight junctions, adhere epithelial cells to one another and regulate selective permeability at both bicellular junctions and tricellular junctions (TCJs). Although several specialized proteins are known to localize at TCJs, it remains unclear how actomyosin-mediated tension transmission at TCJs contributes to the maintenance of junction integrity and barrier function at these sites. Here, utilizing the embryonic epithelium of gastrula-stage Xenopus laevis embryos, we define a mechanism by which the mechanosensitive protein Vinculin helps anchor the actomyosin network at TCJs, thus maintaining TCJ integrity and barrier function. Using an optogenetic approach to acutely increase junctional tension, we find that Vinculin is mechanosensitively recruited to apical junctions immediately surrounding TCJs. In Vinculin knockdown (KD) embryos, junctional actomyosin intensity is decreased and becomes disorganized at TCJs. Using fluorescence recovery after photobleaching (FRAP), we show that Vinculin KD reduces actin stability at TCJs and destabilizes Angulin-1, a key tricellular tight junction protein involved in regulating barrier function at TCJs. When Vinculin KD embryos are subjected to increased tension, TCJ integrity is not maintained, filamentous actin (F-actin) morphology at TCJs is disrupted, and breaks in the signal of the tight junction protein ZO-1 signal are detected. Finally, using a live imaging barrier assay, we detect increased barrier leaks at TCJs in Vinculin KD embryos. Together, our findings show that Vinculin-mediated actomyosin organization is required to maintain junction integrity and barrier function at TCJs and reveal new information about the interplay between adhesion and barrier function at TCJs. [Display omitted] • Vinculin is mechanosensitively recruited to tricellular junctions • Vinculin's actin-binding function is needed for tricellular actomyosin organization • Vinculin reinforces tricellular junction integrity in response to increased tension • Vinculin is required to maintain barrier function at tricellular junctions Epithelial tricellular junctions are vulnerable to disruption. Here, van den Goor et al. reveal that Vinculin is mechanosensitively recruited to vertebrate tricellular junctions, where it regulates actomyosin organization and barrier function. Vinculin reinforces tricellular junction integrity when epithelia experience increased mechanical force. [ABSTRACT FROM AUTHOR]

Background: RNA editing, especially A-to-I editing sites, is a common RNA modification critical for stem cell differentiation, muscle development, and disease occurrence. Unveiling comprehensive RNA A-to-I editing events associated with myogenesis of the skeletal muscle satellite cells (MuSCs) is essential for extending our knowledge of the mechanism underpinning muscle development. Results: A total of 9,632 RNA editing sites (RESs) were screened in the myoblasts (GM), myocytes (DM1), and myotubes (DM5) samples. Among these sites, 4,559 A-to-I edits were classified and further analyzed. There were 3,266 A-to-I sites in the protein-coding region, out of which 113 missense sites recoded protein. Notably, five A-to-I sites in the 3' UTR of four genes (TRAF6, NALF1, SLC38A1, ENSCHIG00000019092) altered their targeted miRNAs. Furthermore, a total of 370 A-to-I sites with different editing levels were detected, including FBN1, MYH10, GSK3B, CSNK1D, and PRKACB genes. These genes were predominantly enriched in the cytoskeleton in muscle cells, the hippo signaling pathway, and the tight junction. Furthermore, we identified 14 hub genes (TUFM, GSK3B, JAK2, RPSA, YARS1, CDH2, PRKACB, RUNX1, NOTCH2, CDC23, VCP, FBN1, RARS1, MEF2C) that potentially related to muscle development. Additionally, 123 stage-specific A-to-I editing sites were identified, with 43 sites in GM, 25 in DM1, and 55 in DM5 samples. These stage-specific edited genes significantly enriched essential biological pathways, including the cell cycle, oocyte meiosis, motor proteins, and hedgehog signaling pathway. Conclusion: We systematically identified the RNA editing events in proliferating and differentiating goat MuSCs, which was crucial for expanding our understanding of the regulatory mechanisms of muscle development. [ABSTRACT FROM AUTHOR]

Background: Sepsis is defined as a life‐threatening disease. Autophagy and the microbiome are increasingly connected with sepsis. The aim of this study was to investigate the protective effect of autophagy and the possible mechanisms. Methods: The septic rat model was established by cecal ligation perforation (CLP). Rapamycin (Rap), 3‐methyladenine (3‐MA), and chloroquine (CQ) were administered to interfere autophagy. Western blot (WB) was used to detect the expression of key proteins in autophagy. Hematoxylin and eosin (H&E) staining and enzyme‐linked immunosorbent assays (ELISAs) were used to identify the effect of autophagy on various organs. 16S ribosomal RNA gene sequencing was used to analyze the changes of the gut microbiota. Results: Rap significantly upregulated the expression of key autophagy proteins, and 3‐MA reduced the relative expression compared to the CLP group. The autophagic flux showed a corresponding trend. Interestingly, the autophagy inducer significantly decreased the mortality and the lipopolysaccharide (LPS) level in serum compared with the CLP group. Autophagy activation significantly improves the inflammatory response in sepsis. Histopathological sections showed that CLP destroyed the tight junctions between ileal epithelial cells, while autophagy induction reversed the damage. The sequencing results showed that autophagy activation increased the alpha diversity and alterted the composition and structure of gut microbiota. The abundance of Proteobacteria was markedly decreased in the Rap group, whereas Bacteroidetes was notably increased compared with the CLP group. Additionally, the protective effect of autophagy further changed the biomarkers in the microbial community. The top 35 functions in each sample were analyzed to obtain 18 genes including RNA synthesis, ATP binding and transport, chromosome assignment, osmotic polysaccharide transport, transcytosis, and methylation. Conclusion: Autophagy is able to improve inflammation and may directly or indirectly regulate the microbiota of septic rats. Autophagy may be an important target for future clinical interventions in the treatment of sepsis. [ABSTRACT FROM AUTHOR]

The blood-brain barrier (BBB) is a critical interface that maintains the central nervous system homeostasis by controlling the exchange of substances between the blood and the brain. Disruption of the BBB plays a vital role in the development of neuroinflammation and neurological dysfunction in sepsis, but the mechanisms by which the BBB becomes disrupted during sepsis are not well understood. Here, we induced endotoxemia, a major type of sepsis, in mice by intraperitoneal injection of lipopolysaccharide (LPS). LPS acutely increased BBB permeability, activated microglia, and heightened inflammatory responses in brain endothelium and parenchyma. Concurrently, LPS or proinflammatory cytokines activated the NF-κB pathway, inhibiting Wnt/β-catenin signaling in brain endothelial cells in vitro and in vivo. Cell culture study revealed that NF-κB p65 directly interacted with β-catenin to suppress Wnt/β-catenin signaling. Pharmacological NF-κB pathway inhibition restored brain endothelial Wnt/β-catenin signaling activity and mitigated BBB disruption and neuroinflammation in septic mice. Furthermore, genetic or pharmacological activation of brain endothelial Wnt/β-catenin signaling substantially alleviated LPS-induced BBB leakage and neuroinflammation, while endothelial conditional ablation of the Wnt7a/7b co-receptor Gpr124 exacerbated the BBB leakage caused by LPS. Mechanistically, Wnt/β-catenin signaling activation rectified the reduced expression levels of tight junction protein ZO-1 and transcytosis suppressor Mfsd2a in brain endothelial cells of mice with endotoxemia, inhibiting both paracellular and transcellular permeability of the BBB. Our findings demonstrate that endotoxemia-associated systemic inflammation decreases endothelial Wnt/β-catenin signaling through activating NF-κB pathway, resulting in acute BBB disruption and neuroinflammation. Targeting the endothelial Wnt/β-catenin signaling may offer a promising therapeutic strategy for preserving BBB integrity and treating neurological dysfunction in sepsis. [ABSTRACT FROM AUTHOR]

Background and Purpose Experimental Approach Key Results Conclusions and Implications Intracranial atherosclerosis is one of the most common causes of ischaemic stroke. However, there is a substantial knowledge gap on the development of intracranial atherosclerosis. Intracranial arteries are characterized by an upregulation of tight junctions between endothelial cells, which control endothelial permeability. We investigated the role of N6‐methyladenosine (m6A), a common RNA modification, on endothelial integrity, focusing on the pro‐atherogenic microRNA miR‐494‐3p and tight junction proteins TJP1 and PECAM1.We assessed the m6A landscape, along with the expression of miR‐494‐3p, TJP1 and PECAM1 in postmortem human vertebral arteries (VA), internal carotid arteries (ICA), and middle cerebral arteries (MCA) with various stages of intimal thickening and plaque formation. The interactions between m6A‐modified miR‐494‐3p mimics, TJP1 and PECAM1, were investigated in vitro using primary human (brain) endothelial cells.Increased m6A expression was observed in the luminal lining of atherosclerosis‐affected VAs, accompanied by reduced TJP1 and PECAM1, but not VE‐cadherin, expression. Colocalization of m6A and miR‐494‐3p in the luminal lining of VA plaques was confirmed, indicating m6A methylation of miR‐494‐3p in intracranial atherosclerosis. Moreover, site‐specific m6A‐modification of miR‐494‐3p led to repression specifically of TJP1 protein expression at cell–cell junctions of brain microvascular endothelial cells, while unmodified miR‐494‐3p showed no effect.This study highlights increasing m6A levels during intracranial atherogenesis. Increases in m6A‐miR‐494‐3p contribute to the observed decreased TJP1 expression in endothelial cell–cell junctions. This is likely to have a negative effect on endothelial integrity and may thus accelerate intracranial atherosclerosis progression. [ABSTRACT FROM AUTHOR]

Neurogenic bladder often occurs after pelvic ganglia injury. Its symptoms, like severe urinary retention and incontinence, have a significant impact on individuals' quality of life. Unfortunately, there are currently no effective treatments available for this type of injury. Here, we designed a fiber-enhanced tissue bandage for injured pelvic ganglia. Tight junctions formed in tissue bandages create a mini tissue structure that enhances resistance in an in vivo environment and delivers growth factors to support the healing of ganglia. Strength fibers are similar to clinical bandages and guarantee ease of handling. Furthermore, tissue bandages can be stored at low temperatures over 5 months without compromising cell viability, meeting the requirements for clinical products. A tissue bandage was applied to a male rat with a bilateral major pelvic ganglia crush injury. Compared to the severe neurogenic bladder symptoms observed in the injury and scaffold groups, tissue bandages significantly improved bladder function. We found that tissue bandage increases resistance to mechanical injury by boosting the expression of cytoskeletal proteins within the major pelvic ganglia. Overall, tissue bandages show promise as a practical therapeutic approach for ganglia repair, offering hope for developing more effective treatments for this thorny condition. Neurogenic bladder often occurs after pelvic ganglia injury. Here, the authors designed a fiber-enhanced tissue bandage for injured pelvic ganglia that facilitates the delivery of growth factors essential for ganglia healing. [ABSTRACT FROM AUTHOR]

The nematode Caenorhabditis elegans is widely employed as a model organism to study basic biological mechanisms. However, transgenic C. elegans are generated by manual injection, which remains low-throughput and labor-intensive, limiting the scope of approaches benefitting from large-scale transgenesis. Here, we report a robotic microinjection system, integrating a microfluidic device capable of reliable worm immobilization, transfer, and rotation, for high-speed injection of C. elegans. The robotic system provides an injection speed 2-3 times faster than that of experts with 7–22 years of experience while maintaining comparable injection quality and only limited trials needed by users to become proficient. We further employ our system in a large-scale reverse genetic screen using multiplexed alternative splicing reporters, and find that the TDP-1 RNA-binding protein regulates alternative splicing of zoo-1 mRNA, which encodes variants of the zonula occludens tight junction proteins. With its high speed, high accuracy, and high efficiency in worm injection, this robotic system shows great potential for high-throughput transgenic studies of C. elegans. Manual injection, which remains low-throughput and labor-intensive, is a technical bottleneck for large-scale genetic studies of C. elegans. Here, the authors report a robotic microinjection system which facilitates injection speed while maintaining injection quality which is comparable to experienced experts. [ABSTRACT FROM AUTHOR]

Simple Summary: Juncao-substrate Ganoderma lucidum Residue (HWE-JGLR) is a dark brown powder. It is obtained by cultivating G. lucidum using mycorrhizae as culture material and extracting it from the remaining culture substrate after G. lucidum harvesting. The application of HWE-JGLR in animal feeds not only reduces the waste of resources but also promotes the reuse of agricultural resources and sustainable development. In our study, HWE-JGLR not only optimised the growth performance of broilers but also enhanced their immune responses. Specifically, dietary supplementation with HWE-JGLR was associated with increased production and gene expression of the immunomodulatory cytokine interleukin-4 (IL-4) and IL-10. Meanwhile, levels and gene expression of the pro-inflammatory cytokine IL-1β were significantly reduced. In addition, HWE-JGLR enhanced the intestinal barrier aspect in poultry. The addition of HWE-JGLR upregulated the gene expression of the broiler jejunal mucosal tight junction proteins Claudin-1 and ZO-1. The purpose of this experiment was to investigate the effects of Hot Water Extract of Juncao-substrate Ganoderma lucidum Residue (HWE-JGLR) on the immune function and intestinal health of yellow-feather broilers. In an animal feeding experiment, 288 male yellow-feather broilers (1 day old) were randomly allocated to four treatment groups with six replicates of 12 birds each. The control (CON) group was fed a basal diet. HJ-1, HJ-2, and HJ-3 were fed a basal diet supplemented with 0.25%, 0.50%, and 1.00% HWE-JGLR, respectively. The feeding trial lasted for 63 d. The results showed increased ADFI (p = 0.033) and ADG (p = 0.045) of broilers in HJ-3, compared with the CON group. Moreover, higher contents of serum IL-4 and IL-10 and gene expression of IL-4 and IL-10 in jejunum mucosa and lower contents of serum IL-1β and gene expression of IL-1β in jejunum mucosa in HJ-3 were observed (p < 0.05). Additionally, the jejunal mucosal gene expression of Claudin-1 and ZO-1 in HJ-2 and HJ-3 was higher than that in the CON group (p < 0.05). As for the microbial community, compared with the CON group, the ACE index, Shannon index, and Shannoneven index of cecal microorganisms in HJ-2 and HJ-3 were elevated (p < 0.05). PCoA analysis showed that the cecal microbial structure of broilers in HJ-2 and HJ-3 was different from the CON group (p < 0.05). In contrast with the CON group, the broilers in HJ-2 and HJ-3 possessed more abundant Desulfobacterota at the phylum level and unclassified Lachnospiraceae, norank Clostridia vadinBB60 group and Blautia spp. at the genus level, while Turicibacter spp. and Romboutsia spp. were less (p < 0.05). In conclusion, dietary supplementation with HWE-JGLR can improve growth performance, enhance body immunity and intestinal development, and maintain the cecum microflora balance of yellow-feather broilers. [ABSTRACT FROM AUTHOR]

Obstructive sleep apnea syndrome (OSAS) affects a large part of the aging population. It is characterized by chronic intermittent hypoxia and associated with neurocognitive dysfunction. One hypothesis is that the blood–brain barrier (BBB) functions could be altered by exosomes. Exosomes are nanovesicles found in biological fluids. Through the study of exosomes and their content in tau and amyloid beta (Aβ), the aim of this study was to show how exosomes could be used as biomarkers of OSAS and of their cognitive disorders. Two groups of 15 volunteers from the PROOF cohort were selected: severe apnea (AHI > 30) and control (AHI < 5). After exosome isolation from blood serum, we characterized and quantified them (CD81, CD9, CD63) by western blot and ELISAs and put them 5 h in contact with an in vitro BBB model. The apparent permeability of the BBB was measured using sodium-fluorescein and TEER. Cell ELISAs were performed on tight junctions (ZO-1, claudin-5, occludin). The amount of tau and Aβ proteins found in the exosomes was quantified using ELISAs. Compared to controls, OSAS patients had a greater quantity of exosomes, tau, and Aβ proteins in their blood sera, which induced an increase in BBB permeability in the model and was reflected by a loss of tight junction' expression. Elderly patients suffering severe OSAS released more exosomes in serum from the brain compartment than controls. Such exosomes increased BBB permeability. The impact of such alterations on the risk of developing cognitive dysfunction and/or neurodegenerative diseases is questioned. [ABSTRACT FROM AUTHOR]

Cadmium is a contributing factor to cardiovascular diseases and highly toxic to vascular endothelial cells. It has a distinct mode of injury, causing the de-endothelialization of regions in the monolayer structure of endothelial cells in a concentration-dependent manner. However, the specific molecules involved in the cadmium toxicity of endothelial cells remain unclear. The purpose of this study was to identify the specific molecular mechanisms through which cadmium affects endothelial detachment. Cadmium inhibited the expression of claudin-5 and zonula occludens (ZO)-1, which are components of tight junctions (strongest contributors to intercellular adhesion), in a concentration- and time-dependent manner. Compared to arsenite, zinc, and manganese, only cadmium suppressed the expression of both claudin-5 and ZO-1 molecules. Moreover, the knockdown of claudin-5 and ZO-1 exacerbated cadmium-induced endothelial cell injury and expansion of the detachment area, whereas their overexpression reversed these effects. CRE-binding protein inhibition reduced cadmium toxicity, suggesting that CRE-binding protein activation is involved in the cadmium-induced inhibition of claudin-5 and ZO-1 expression and endothelial detachment. These findings provide new insights into the toxicological mechanisms of cadmium-induced endothelial injury and risk of cardiovascular disease. [ABSTRACT FROM AUTHOR]

Bovine mastitis is an infectious disease that causes substantial economic losses to the dairy industry worldwide. Current antibiotic therapy faces issues of antibiotic misuse and antimicrobial resistance, which has aroused concerns for both veterinary and human medicine. Thus, this study explored the potential of Colo EVs (bovine colostrum-derived extracellular vesicles) to address mastitis. Using LPS-induced murine mammary epithelial cells (HC11), mouse monocyte macrophages (RAW 264.7), and a murine mastitis model with BALB/C mice, we evaluated the safety and efficacy of Colo EVs, in vivo and in vitro. Colo EVs had favorable biosafety profiles, promoting cell proliferation and migration without inducing pathological changes after injection into murine mammary glands. In LPS-induced murine mastitis, Colo EVs significantly reduced inflammation, improved inflammatory scores, and preserved tight junction proteins while protecting milk production. Additionally, in vitro experiments demonstrated that Colo EVs downregulated inflammatory cytokine expression, reduced inflammatory markers, and attenuated NF-κB pathway activation. In summary, we inferred that Colo EVs have promise as a therapeutic approach for mastitis treatment, owing to their anti-inflammatory properties, potentially mediated through the NF-κB signaling pathway modulation. [ABSTRACT FROM AUTHOR]

The blood-retina barrier (BRB), which is disrupted in diabetic retinopathy (DR) and uveitis, is an important anatomical characteristic of the retina, regulating nutrient, waste, water, protein, and immune cell flux. The BRB is composed of endothelial cell tight junctions, pericytes, astrocyte end feet, a collagen basement membrane, and perivascular macrophages. Despite the importance of the BRB, retinal perivascular macrophage function remains unknown. We found that retinal perivascular macrophages resided on postcapillary venules in the superficial vascular plexus and expressed MHC class II. Using single-cell RNA-Seq, we found that perivascular macrophages expressed a prochemotactic transcriptome and identified platelet factor 4 (Pf4, also known as CXCL4) as a perivascular macrophage marker. We used Pf4Cre mice to specifically deplete perivascular macrophages. To model retinal inflammation, we performed intraocular CCL2 injections. Ly6C+ monocytes crossed the BRB proximal to perivascular macrophages. Depletion of perivascular macrophages severely hampered Ly6C+ monocyte infiltration. These data suggest that retinal perivascular macrophages orchestrate immune cell migration across the BRB, with implications for inflammatory ocular diseases including DR and uveitis. [ABSTRACT FROM AUTHOR]

Mitochondrial dysfunction may precipitate intestinal dysfunction, while inflammatory bowel disease manifests as a chronic inflammatory ailment affecting the gastrointestinal tract. This condition disrupts the barrier function of the intestinal epithelium and alters metabolic products. Increasing mitochondrial adenosine triphosphate (ATP) synthesis in intestinal epithelial cells presents a promising avenue for colitis treatments. Nevertheless, the impact of cedrol on ATP and the intestinal barrier remains unexplored. Hence, this study is dedicated to examining the cedrol's protective effect on an inflammatory cocktail (IC)–induced intestinal epithelial barrier dysfunction in Caco‐2 cells. The finding reveals that cedrol enhances ATP content and the transepithelial electrical resistance value in the intestinal epithelial barrier. Moreover, cedrol mitigates the IC‐induced decrease in the messenger ribonucleic acid (mRNA) expression of tight junction proteins (ZO‐1, Occludin, and Claudin‐1), thereby ameliorating intestinal epithelial barrier dysfunction. Furthermore, nuclear magnetic resonance (NMR)–based metabolomic analysis indicated that IC‐exposed Caco‐2 cells are restored by cedrol treatments. Notably, cedrol elevates metabolites such as amino acids, thereby enhancing the intestinal barrier. In conclusion, cedrol alleviates IC‐induced intestinal epithelial barrier dysfunction by promoting ATP‐dependent proliferation of Caco‐2 cells and bolstering amino acid levels to sustain tight junction messenger ribonucleic acid expression. [ABSTRACT FROM AUTHOR]

Chronic pancreatitis (CP) is a progressive inflammatory disorder that impairs endocrine and exocrine function. Our previous work showed that mesenchymal stem/stromal cells (MSCs) and MSCs overexpressing alpha‐1 antitrypsin (AAT‐MSCs) could be therapeutic tools for CP. However, primary MSCs are predisposed to undergo senescence during culture expansion, which limits their therapeutic applications. We generated and characterized immortalized human MSCs (iMSCs) and AAT‐MSCs (iAAT‐MSCs) and tested their protective effect on 2,4,6‐Trinitrobenzenesulfonic acid (TNBS)‐induced acinar cell death in an in vitro cell culture system. Primary MSCs were immortalized by transduction with simian virus 40 large T antigen (SV40LT), and the resulting iMSC and iAAT‐MSC lines were analysed for proliferation, senescence, phenotype and multi‐differentiation potential. Subsequently, apoptosis and ferroptosis pathways were investigated by assessing changes before and after TNBS treatment. Coculture of iMSCs and iAAT‐MSCs with acinar cell lines inhibited early cell death induced by TNBS, reduced ER stress and reversed TNBS‐induced protein reduction at tight junctions. Additionally, iMSCs and iAAT‐MSCs exerted such protection by regulating mitochondrial respiration, ATP content and ROS production in TNBS‐induced acinar cells. Furthermore, iMSCs and iAAT‐MSCs ameliorated TNBS‐induced ferroptosis by modulating iron generation and ROS production and regulating the ferritin heavy chain 1 (FTH1)/protein disulfide isomerase (PDI)/glutathione peroxide 4 (GPX4) signalling pathways in acinar cells. These findings identify ferroptosis as an unrecognized mechanism that leads to TNBS‐induced cell death and offer mechanistic insights relevant to using stem cell therapy to treat acinar cell death associated with CP. [ABSTRACT FROM AUTHOR]

Background/Objectives: Our objective was to review published studies of the intestinal barrier and permeability, the deleterious effects of dietary components (particularly fat), the impact of altered intestinal permeability in disease models and human diseases, the role of the microbiome and epigenomics in control of barrier function, and the opportunities to restore normal barrier function with dietary interventions and products of the microbiota. Methods: We conducted a literature review including the following keywords alone or in combination: intestinal barrier, permeability, microbiome, epigenomics, diet, irritable bowel syndrome, inflammatory bowel disease, probiotics. Results: Intestinal permeability is modified by a diet including fat, which increases permeability, and nutrients such as fiber, glutamine, zinc, vitamin D, polyphenols, emulsifiers, and anthocyanins, which decrease permeability. There is significant interaction of the microbiome and barrier function, including the inflammatory of luminal/bacterial antigens, and anti-inflammatory effects of commensals or probiotics and their products, including short-chain fatty acids. Epigenomic modification of barrier functions are best illustrated by effects on junction proteins or inflammation. Detailed documentation of the protective effects of diet, probiotics, prebiotics, and microbiota is provided. Conclusion: intestinal permeability is a critical factor in protection against gastrointestinal diseases and is impacted by nutrients that preserve or heal and repair the barrier and nurture anti-inflammatory effects. [ABSTRACT FROM AUTHOR]

Background: Recirculating aquaculture systems can cause chronic stress in fish when stocking density is too high. However, this study tested whether low stocking density can cause fish stress. Adult rainbow trout, with an average weight of 1.517 kg (± 0.39), were subjected to low (12 kg/m3 ± 0.94) and moderate (43 kg/m3 ± 2.03) stocking densities for 24 days in a recirculating system maintained at 15 °C. At the end of the experiment, fecal microbiome analysis was carried out using a 16S rRNA amplicon sequencing. Additionally, an untargeted plasma metabolomics analysis was conducted. Results: The moderate stocking density group harboured greater numbers of commensals, such as C. somerae, R. lituseburensis, and L. plantarum. In contrast, detrimental species such as S. putrifacens and P. putida were abundant in the low-stocking density fish. Functional microbiome profiling revealed vitamin B12 salvage and synthesis in moderate stocking densities, which may support intestinal tight junction function. Additionally, vitamin B1 biosynthesis pathways were more abundant in the moderate stocking density group, which may function towards oxidative energy metabolism and protect against oxidative stress. A complementary plasma metabolomics study, although done at slightly different stocking densities and duration, confirmed the presence of blood metabolic stress markers. Elevated levels of L-lactic acid and L-Norvaline, L-Valine, and L-glutamine, indicate low stocking density fish were under stress. Furthermore, a P4HA2 stress gene biomarker confirmed the occurrence of stress in low-density fish. This study suggests that low stocking density can induce stress in fish. Moreover, moderate stocking density leads to a distinct and beneficial fecal microbiome profile. Conclusion: Our study highlights the potential benefits of optimizing the stocking density of fish in recirculating aquaculture systems. This can improve fish health and welfare, promoting a more resilient fecal microbiome. [ABSTRACT FROM AUTHOR]

ShenZhu TiaoPi granule (STG) is a compound prescription that is used in Chinese medicine for the treatment of type 2 diabetes mellitus (T2DM). Previous studies have indicated a hypoglycaemic effect, but the underlying mechanism remains unclear. Goto-Kakizaki (GK) rats were used to establish an in vivo T2DM model (Mod). The metformin (Met) and STG treatment time was 12 weeks. Fasting blood glucose (FBG) and insulin levels and the area under the glucose curve (GAUC) were measured. Intestinal pathology and permeability were observed. Microbial diversity analysis and metabolomics were used to investigate the underlying mechanisms. Compared with the Con group, the T2DM Mod group presented significant differences in weight, FBG, GAUC, and homeostasis model assessment-insulin resistance (HOMA-IR) indices (p < 0.01). Met and STG improved these indicators (p < 0.01). The pathological morphology and zonula occludens 1 protein levels in the intestines of the Mod group of rats were altered, leading to increases in the lipopolysaccharide (LPS) and interleukin-1ß (IL-1ß) levels. In the Met and STG groups, the intestinal conditions improved, and the LPS and IL-1ß levels significantly decreased (p < 0.01). Changes in the gut microbiota and metabolites occurred in the Mod group. In the STG group, the abundance of Intestinimonas increased, and the abundance of Eubacterium coprostanoligenes decreased significantly (p < 0.05). Moreover, STG also altered 2-deoxyglucose, beta-muricholic acid and dioxolithocholic acid production. In addition, the main metabolic pathways affected by STG were bile acid biosynthesis and cholesterol metabolism. Intestinimonas, D-maltose_and_ alpha-lactose may be potential biomarkers for the effects of STG. STG alleviates hyperglycaemia via the gut microbiota and metabolites in GK rats. [ABSTRACT FROM AUTHOR]

Spermatogenesis constitutes a complex and intricate cascade of differentiation, indispensable for the male reproductive competence. The intercellular communication conduits of Sertoli cells (SCs) are pivotal in orchestrating this cascade ensuring sustenance and development of germ cells. Single cells and bioinformatics recently demonstrated articles are used for the regulatory modalities through which SCs modulate spermatogenesis, specifically via androgen receptors (ARs), the transforming growth factor-beta/Smad axis, mitogen-activated protein kinases, cAMP/protein kinase A (PKA), phosphatidylinositol 4,5-bisphosphate 3-kinase (PI3k)/AKT serine threonine kinase (Akt), AMP-activated protein kinase, and AR pathways. Within this framework, homeostasis of gap junction dynamics, cryptic sites and the activities at tight junctions and adherens junctions, with the integrity of the testicular barrier, glucose assimilation, lactate distribution, being governed also along with SC maturation. Disruptions in activities or abnormal concentration in derangements in AR, cAMP/PKA, and PI3k/Akt pathways, and as well as the molecules that comprise them, would present male infertility. [ABSTRACT FROM AUTHOR]

The epidermis, the most superficial layer of the human skin, serves a critical barrier function, protecting the body from external pathogens and allergens. Dysregulation of epidermal differentiation contributes to barrier dysfunction and has been implicated in the pathology of various dermatological diseases, including atopic dermatitis (AD). Mucopolysaccharide polysulphate (MPS) is a moisturising agent used to treat xerosis in patients with AD. However, its mechanism of action on keratinocytes, the main constituents of the epidermis, remains unclear. In this study, we investigated the effect of MPS on keratinocytes by subjecting adult human epidermal and three‐dimensional cultured keratinocytes to MPS treatment, followed by transcriptome analysis. The analysis revealed that MPS treatment enhances keratinocyte differentiation and suppresses proliferation. We focused on amphiregulin (AREG), a membrane protein that belongs to the epidermal growth factor (EGF) family and possesses a heparin‐binding domain, as a significant target among the genes altered by MPS. MPS exerted an inhibitory effect directly on AREG, rather than on EGF receptors or other members of the EGF family. Furthermore, AREG leads to a reduction in epidermal barrier function, whereas MPS contributes to barrier enhancement via AREG inhibition. Collectively, these findings suggest that MPS modulates barrier function through AREG inhibition, offering insights into potential therapeutic strategies for skin barrier restoration. [ABSTRACT FROM AUTHOR]

Simple Summary: This paper presents a meta-analysis of currently available single-cell RNA sequencing (scRNAseq) datasets from zebrafish kidneys. Our work aims to identify key marker genes for various immune cell types and transcription factors (TFs) involved in fish myelopoiesis. Our results validate previous studies and expand them; newly discovered markers and TFs have experimental confirmation of their roles in complementary cell types of mammals and fish species. Reconstructed gene networks of macrophage- and neutrophil-specific genes revealed the potential interactions of identified TFs and marker genes. Obtained results could be taken into account during the design of immunogenetics experiments (e.g., creating zebrafish lines with TF knockouts) and serve as a basis for novel applications for zebrafish. The revealed cell-specific markers could help in the accurate determination of fish cell types in the future. Our methodology of data integration showed its reliability and could be further adapted to meta-analyses of various scRNAseq datasets. The innate immune system (IIS) is an ancient and essential defense mechanism that protects animals against a wide range of pathogens and diseases. Although extensively studied in mammals, our understanding of the IIS in other taxa remains limited. The zebrafish (Danio rerio) serves as a promising model organism for investigating IIS-related processes, yet the immunogenetics of fish are not fully elucidated. To address this gap, we conducted a meta-analysis of single-cell RNA sequencing (scRNA-seq) datasets from zebrafish kidney marrow, encompassing approximately 250,000 immune cells. Our analysis confirms the presence of key genetic pathways in zebrafish innate immune cells that are similar to those identified in mammals. Zebrafish macrophages specifically express genes encoding cathepsins, major histocompatibility complex class II proteins, integral membrane proteins, and the V-ATPase complex and demonstrate the enrichment of oxidative phosphorylation ferroptosis processes. Neutrophils are characterized by the significant expression of genes encoding actins, cytoskeleton organizing proteins, the Arp2/3 complex, and glycolysis enzymes and have demonstrated their involvement in GnRH and CLR signaling pathways, adherents, and tight junctions. Both macrophages and neutrophils highly express genes of NOD-like receptors, phagosomes, and lysosome pathways and genes involved in apoptosis. Our findings reinforce the idea about the existence of a wide spectrum of immune cell phenotypes in fish since we found only a small number of cells with clear pro- or anti-inflammatory signatures. [ABSTRACT FROM AUTHOR]

Hemolysis is associated with pulmonary hypertension (PH), but the direct contribution of circulating free heme to the PH pathogenesis remains unclear. Here, we show that the elevated levels of circulating free heme are sufficient to induce PH and inflammatory response in mice and confirm the critical role of mitogen-activated protein kinase kinase-3 (MKK3)-mediated pathway in free heme signaling. Following the continuous infusion of heme for 2 wk, wild-type (WT) but not MKK3 knockout (KO) mice develop PH, as evidenced by a significantly elevated right ventricular (RV) systolic pressure, RV hypertrophy, and pulmonary vascular remodeling. The MKK3/p38 axis, markedly activated by heme infusion in WTs, results in upregulated proliferative/cytokine signaling targets Akt, ERK1/2, and STAT3, which were abrogated in MKK3 KO mice. Moreover, the MKK3 KOs were protected against heme-mediated endothelial barrier dysfunction by restoring the tight junction protein zonula occludens-1 expression and diminishing the inflammatory cell infiltration in the lungs. Plasma cytokine multiplex analysis revealed a severe cytokine storm already 24 h after initiation of heme infusion, with a significant increase of 19 cytokines, including IL-1b, IL-2, IL-6, IL-9, and TNF-a, in WT animals and complete attenuation of cytokine production in MKK3 KO mice. Together, these findings reveal a causative role of circulating free heme in PH through activating inflammatory and proliferative responses. The central role of MKK3 in orchestrating the heme-mediated pathogenic response supports MKK3 as an attractive therapeutic target for PH and other lung inflammatory diseases linked to hemolytic anemia. NEW & NOTEWORTHY: This study demonstrates that elevated levels of circulating free heme can induce pulmonary hypertension (PH) and inflammation in mice. Continuous heme infusion activated the MKK3/p38 pathway, leading to increased right ventricular pressure, right ventricular hypertrophy, and vascular remodeling. This activation upregulated signaling cascades such as Akt, ERK1/2, and STAT3, whereas MKK3 knockout mice were protected against these changes and had reduced inflammatory responses, highlighting MKK3's potential as a therapeutic target for PH. [ABSTRACT FROM AUTHOR]

Confluent populations of the epithelial cell line, MDCK II, develop circumferential tight junctions joining adjacent cells to create a barrier to the paracellular movement of solutes and water. Treatment of MDCK II cell populations from the apical surface with 1 mM Na-caprate increased permeability to macromolecules (Leak Pathway) without increasing monolayer disruption or cell death. Graphical analysis of the apparent permeability versus solute Stokes radius for a size range of fluorescein-dextran species indicates apical 1 mM Na-caprate enhances Leak Pathway permeability by increasing the number of Leak Pathway openings without significantly affecting opening size. Na-caprate treatment did not alter the content of any tight junction protein examined. Treatment of MDCK II cell populations with apical 1 mM Na-caprate disrupted basal F-actin stress fibers and decreased the tortuosity of the tight junctions. Treatment of MDCK II cell populations with blebbistatin, a myosin ATPase inhibitor, alone had little effect on Leak Pathway permeability but synergistically increased Leak Pathway permeability when added with 1 mM Na-caprate. Na-caprate exhibited a similar ability to increase Leak Pathway permeability in wild-type MDCK II cell monolayers and ZO-1 knockdown MDCK II cell monolayers but an enhanced ability to increase Leak Pathway permeability in monolayers of TOCA-1 knockout MDCK II cells. These results demonstrate that Na-caprate increases MDCK II cell population Leak Pathway permeability by increasing the number of Leak Pathway openings. This action is likely mediated by alterations in F-actin organization, primarily involving disruption of basal F-actin stress fibers. [ABSTRACT FROM AUTHOR]

Cells depend on precisely regulating barrier function within the vasculature to maintain physiological stability and facilitate essential substance transport. Endothelial cells achieve this through specialized adherens and tight junction protein complexes, which govern paracellular permeability across vascular beds. Adherens junctions, anchored by vascular endothelial (VE)-cadherin and associated catenins to the actin cytoskeleton, mediate homophilic adhesion crucial for barrier integrity. In contrast, tight junctions composed of occludin, claudin, and junctional adhesion molecule A interact with Zonula Occludens proteins, reinforcing intercellular connections essential for barrier selectivity. Endothelial cell-cell junctions exhibit dynamic conformations during development, maturation, and remodeling, regulated by local biochemical and mechanical cues. These structural adaptations play pivotal roles in disease contexts such as chronic inflammation, where junctional remodeling contributes to increased vascular permeability observed in conditions from cancer to cardiovascular diseases. Conversely, the brain microvasculature's specialized junctional arrangements pose challenges for therapeutic drug delivery due to their unique molecular compositions and tight organization. This commentary explores the molecular mechanisms underlying endothelial cell-cell junction conformations and their implications for vascular permeability. By highlighting recent advances in quantifying junctional changes and understanding mechanotransduction pathways, we elucidate how physical forces from cellular contacts and hemodynamic flow influence junctional dynamics. [ABSTRACT FROM AUTHOR]

Background: Helichrysum italicum is a Mediterranean plant with well-known anti-inflammatory activity, but our previous whole transcriptome analysis has found that H. italicum infusion (HII) can also affect cytoskeletal rearrangement and tight junctions. The goal of the present study was to determine if HII improves the intestinal barrier (IB) dysfunction and by what mechanism. Methods: Caco-2 cells on Transwell inserts were used as a model of IB permeability. Heat-killed (HKB) or live Salmonella Infantis bacteria were used to induce IB integrity disruption upon three different testing conditions: pre-, co-, and post-treatment with 0.2 v/v% HII. Transepithelial electrical resistance values were used as an indicator of monolayer integrity before and after all treatments, and RT-PCR was used to assess the expression of tight junction proteins (TJPs) and inflammatory cytokines known to regulate intestinal permeability. Results: We found that all three treatments with HII improved the HKB-induced integrity disruption and decreased the down-regulation of TJP1, OCLN, and CLDN1, with the greatest effect observed in the pre-treated cells. Treatment with HII also decreased the up-regulation of CLDN2, TNF-α, IL-1β, and IL-6. In addition, pre-treatment of Caco-2 cells with HII prevented translocation of S. Infantis but did not prevent adhesion and invasion. Conclusion: This study showed that HII can improve inflammation-disrupted IB function by indirect modulation of mRNA expression of TJPs, especially in a preventive manner. [ABSTRACT FROM AUTHOR]

Purpose: Claudins are tight junction proteins partaking in epithelial-mesenchymal transition and cancer progression. In this study, we investigated the expression patterns of claudin-1 and claudin-4 in thyroid pathologies, discussed their links with the pathogenesis of thyroid cancers, and reviewed the therapeutic potential of targeting claudins in cancers. Methods: The research group 162 cores of thyroid samples from patients (70 female and 11 male) diagnosed with thyroid adenoma, goiter, papillary, medullary, and anaplastic thyroid cancers. All samples were stained for the expression of claudin-1 and claudin-4, and the analysis of IHC was performed. Results: Goiter samples showed negative claudin-1 and mostly positive expression of claudin-4. Papillary thyroid cancer and thyroid adenoma showed positive expression of claudin-1, while claudin-4 was positive in papillary thyroid cancers, goiters, and adenomas. In The Cancer Genome Atlas cohort, claudin-1 and claudin-4 were overexpressed in papillary thyroid cancer compared to normal thyroid tissues. Patients with high claudin-1 expression had significantly lower 5-year overall survival than patients with low claudin-1 levels (86.75% vs. 98.65, respectively). In multivariate analysis, high claudin-1 expression (HR 7.91, CI 95% 1.79–35, p = 0.006) and advanced clinical stage remained statistically significant prognostic factors of poor prognosis in papillary thyroid cancer. Conclusions: The pattern of claudin-1 staining was pathology-specific and changed between cancers of different histology. This phenomenon may be associated with the different pathogenesis of thyroid cancers and early metastasis. The loss of claudin-1 and claudin-4 characterized more aggressive cancers. Several studies have shown the benefits of targeting claudins in cancers, but their implementation into clinical practice requires further trials. [ABSTRACT FROM AUTHOR]

Lateolabrax maculatus iridovirus (LMIV) is a variant strain of red sea bream iridovirus (RSIV), causing serious economic losses in aquaculture. Claudins (CLDNs) are major components of tight junctions (TJs) forming an important line of defense against pathogens. Our pilot miRNA-mRNA joint analysis indicated the degradation of CLDN3, as well as its interaction with miR-181a during LMIV infection. To elucidate the miR-181a/CLDN3/LMIV interactions, in vitro assays were carried out on LMB-L cells. We first confirmed that LMIV infection could decrease the expression of CLDN3, accompanied by the enhancement of permeability, suggesting the dysfunction of TJs. Contrary to the inhibition of CLDN3, the activation of miR-181a was proved, presenting a negative correlation between miR-181a and CLDN3 (Pearson r = −0.773 and p < 0.01). In addition, the influence of CLDN3 on LMIV replication was analyzed by knockdown and over-expression of CLDN3. When CLDN3 was silenced in LMB-L cells with siCLDN3-623 at 9 days post transfection (dpt), LMIV copies and titers were significantly up-regulated by 1.59-fold and 13.87-fold, respectively. By contrast, LMIV replication in LMB-L cells was reduced by 60% and 71%, post transfection with pcDNA3.1-CLDN3 over-expressed plasmid at 6 dpt and 9 dpt, respectively. Ultimately, the regulatory relationship between miR-181a and CLDN3 was further validated by dual luciferase reporter assays. Taking into account the above-described results, we proposed a "miR-181a/CLDN3/LMIV" regulatory relationship. This study provides a new insight for understanding the mechanism of LMIV replication. [ABSTRACT FROM AUTHOR]

Allergens and Th2 cytokines affect the homeostatic environment in the airways, leading to increased mucus production by goblet cells associated with altered adherens junctional complex (AJC) and tight junction (TJ) proteins responsible for maintaining epithelial barrier function. Circadian clock-dependent regulatory mechanisms such as inflammation and epithelial barrier function are gaining more attention due to their therapeutic potential against allergic inflammatory lung diseases. Currently, there are no studies to support whether REV-ERBa activation can attenuate Th2 cytokine-induced epithelial barrier dysfunction in human bronchial epithelial cells. We hypothesized that Th2 cytokine-induced epithelial barrier dysfunction may be protected by activating REV-ERBa. Treatment with Th2 cytokines or HDM significantly reduced the cell impedance, as confirmed by transepithelial electrical resistance (TEER). However, pre-treatment with SR10067 attenuated Th2 cytokine-induced barrier dysfunction, such as decreased permeability, improved TEER, localization of AJC and TJ proteins, and mRNA and protein levels of selected epithelial barrier and circadian clock targets. Overall, we showed for the first time that REV-ERBa activation regulates altered epithelial barrier function that may have direct implications for the treatment of asthma and other allergic diseases. [ABSTRACT FROM AUTHOR]