Building Biomaterials to Mimic 3D Cell-Cell Junctions.

Cell-cell interactions typically occur in a 3D context that is distinct from conventional 2D cell-substrate interactions in a Petri dish. Here, we describe a benchtop method to combine a 2D extracellular matrix surface with a 3D, vertical boundary functionalized with the extracellular domain of E-cadherin. The methodology is suitable for any biology laboratory without requiring advanced microfabrication equipment or training. Overall, this cell-mimetic interface uniquely recapitulates key aspects of cell-cell adhesion and can serve as a versatile, reductionist technique to study general cell-cell interactions in a 3D context.

Phospholipid scramblase 1: an essential component of the nephrocyte slit diaphragm.

Blood ultrafiltration in nephrons critically depends on specialized intercellular junctions between podocytes, named slit diaphragms (SDs). Here, by studying a homologous structure found in Drosophila nephrocytes, we identify the phospholipid scramblase Scramb1 as an essential component of the SD, uncovering a novel link between membrane dynamics and SD formation. In scramb1 mutants, SDs fail to form. Instead, the SD components Sticks and stones/nephrin, Polychaetoid/ZO-1, and the Src-kinase Src64B/Fyn associate in cortical foci lacking the key SD protein Dumbfounded/NEPH1. Scramb1 interaction with Polychaetoid/ZO-1 and Flotillin2, the presence of essential putative palmitoylation sites and its capacity to oligomerize, suggest a function in promoting SD assembly within lipid raft microdomains. Furthermore, Scramb1 interactors as well as its functional sensitivity to temperature, suggest an active involvement in membrane remodeling processes during SD assembly. Remarkably, putative Ca2+-binding sites in Scramb1 are essential for its activity raising the possibility that Ca2+ signaling may control the assembly of SDs by impacting on Scramb1 activity.

Regulation of vascular endothelial integrity by mesenchymal stem cell extracellular vesicles after hemorrhagic shock and trauma.

BACKGROUND: Patients with hemorrhagic shock and trauma (HS/T) are vulnerable to the endotheliopathy of trauma (EOT), characterized by vascular barrier dysfunction, inflammation, and coagulopathy. Cellular therapies such as mesenchymal stem cells (MSCs) and MSC extracellular vesicles (EVs) have been proposed as potential therapies targeting the EOT. In this study we investigated the effects of MSCs and MSC EVs on endothelial and epithelial barrier integrity in vitro and in vivo in a mouse model of HS/T. This study addresses the systemic effects of HS/T on multiorgan EOT. METHODS: In vitro, pulmonary endothelial cell (PEC) and Caco-2 intestinal epithelial cell monolayers were treated with control media, MSC conditioned media (CM), or MSC EVs in varying doses and subjected to a thrombin or hydrogen peroxide (H2O2) challenge, respectively. Monolayer permeability was evaluated with a cell impedance assay, and intercellular junction integrity was evaluated with immunofluorescent staining. In vivo, a mouse model of HS/T was used to evaluate the effects of lactated Ringer's (LR), MSCs, and MSC EVs on endothelial and epithelial intercellular junctions in the lung and small intestine as well as on plasma inflammatory biomarkers. RESULTS: MSC EVs and MSC CM attenuated permeability and preserved intercellular junctions of the PEC monolayer in vitro, whereas only MSC CM was protective of the Caco-2 epithelial monolayer. In vivo, both MSC EVs and MSCs mitigated the loss of endothelial adherens junctions in the lung and small intestine, though only MSCs had a protective effect on epithelial tight junctions in the lung. Several plasma biomarkers including MMP8 and VEGF were elevated in LR- and EV-treated but not MSC-treated mice. CONCLUSIONS: In conclusion, MSC EVs could be a potential cell-free therapy targeting endotheliopathy after HS/T via preservation of the vascular endothelial barrier in multiple organs early after injury. Further research is needed to better understand the immunomodulatory effects of these products following HS/T and to move toward translating these therapies into clinical studies.

The TRIP6/LATS1 complex constitutes the tension sensor of α-catenin/vinculin at both bicellular and tricellular junctions.

Cell-cell mechanotransduction regulates tissue development and homeostasis. α-catenin, the core component of adherens junctions, functions as a tension sensor and transducer by recruiting vinculin and transducing signals that influence cell behaviors. α-catenin/vinculin complex-mediated mechanotransduction regulates multiple pathways, such as Hippo pathway. However, their associations with the α-catenin-based tension sensors at cell junctions are still not fully addressed. Here, we uncovered the TRIP6/LATS1 complex co-localizes with α-catenin/vinculin at both bicellular junctions (BCJs) and tricellular junctions (TCJs). The localization of TRIP6/LATS1 complex to both TCJs and BCJs requires ROCK1 and α-catenin. Treatment by cytochalasin B, Y-27632 and blebbistatin all impaired the BCJ and TCJ junctional localization of TRIP6/LATS1, indicating that the junctional localization of TRIP6/LATS1 is mechanosensitive. The α-catenin/vinculin/TRIP6/LATS1 complex strongly localized to TCJs and exhibited a discontinuous button-like pattern on BCJs. Additionally, we developed and validated an α-catenin/vinculin BiFC-based mechanosensor that co-localizes with TRIP6/LATS1 at BCJs and TCJs. The mechanosensor exhibited a discontinuous distribution and motile signals at BCJs. Overall, our study revealed that TRIP6 and LATS1 are novel compositions of the tension sensor, together with the core complex of α-catenin/vinculin, at both the BCJs and TCJs.

Blood-testis barrier: a review on regulators in maintaining cell junction integrity between Sertoli cells.

The blood-testis barrier (BTB) is formed adjacent to the seminiferous basement membrane. It is a distinct ultrastructure, partitioning testicular seminiferous epithelium into apical (adluminal) and basal compartments. It plays a vital role in developing and maturing spermatocytes into spermatozoa via reorganizing its structure. This enables the transportation of preleptotene spermatocytes across the BTB, from basal to adluminal compartments in the seminiferous tubules. Several bioactive peptides and biomolecules secreted by testicular cells regulate the BTB function and support spermatogenesis. These peptides activate various downstream signaling proteins and can also be the target themself, which could improve the diffusion of drugs across the BTB. The gap junction (GJ) and its coexisting junctions at the BTB maintain the immunological barrier integrity and can be the "gateway" during spermatocyte transition. These junctions are the possible route for toxicant entry, causing male reproductive dysfunction. Herein, we summarize the detailed mechanism of all the regulators playing an essential role in the maintenance of the BTB, which will help researchers to understand and find targets for drug delivery inside the testis.

Conditional deficiency of Rho-associated kinases disrupts endothelial cell junctions and impairs respiratory function in adult mice.

The Ras homology (Rho) family of GTPases serves various functions, including promotion of cell migration, adhesion, and transcription, through activation of effector molecule targets. One such pair of effectors, the Rho-associated coiled-coil kinases (ROCK1 and ROCK2), induce reorganization of actin cytoskeleton and focal adhesion through substrate phosphorylation. Studies on ROCK knockout mice have confirmed that ROCK proteins are essential for embryonic development, but their physiological functions in adult mice remain unknown. In this study, we aimed to examine the roles of ROCK1 and ROCK2 proteins in normal adult mice. Tamoxifen (TAM)-inducible ROCK1 and ROCK2 single and double knockout mice (ROCK1flox/flox and/or ROCK2flox/flox;Ubc-CreERT2) were generated and administered a 5-day course of TAM. No deaths occurred in either of the single knockout strains, whereas all of the ROCK1/ROCK2 double conditional knockout mice (DcKO) had died by Day 11 following the TAM course. DcKO mice exhibited increased lung tissue vascular permeability, thickening of alveolar walls, and a decrease in percutaneous oxygen saturation compared with noninducible ROCK1/ROCK2 double-floxed control mice. On Day 3 post-TAM, there was a decrease in phalloidin staining in the lungs in DcKO mice. On Day 5 post-TAM, immunohistochemical analysis also revealed reduced staining for vascular endothelial (VE)-cadherin, ß-catenin, and p120-catenin at cell-cell contact sites in vascular endothelial cells in DcKO mice. Additionally, VE-cadherin/ß-catenin complexes were decreased in DcKO mice, indicating that ROCK proteins play a crucial role in maintaining lung function by regulating cell-cell adhesion.

Transcriptional Levels of Intercellular Junction Proteins in an Alveolar Epithelial Cell Line Exposed to Irradiation or Bleomycin.

We performed a comparative study of the effects of X-ray irradiation and bleomycin on the mRNA levels of E-cadherin and tight junction proteins (claudin-3, claudin-4, claudin-18, ZO-2, and occludin) in an alveolar epithelial cell line L2. Irradiation decreased claudin-4 levels and increased occludin levels, while the levels of other mRNAs remained unchanged. Bleomycin increased the expression levels of all proteins examined except claudin-3. Irradiation and bleomycin have different effects on the expression level of intercellular junction proteins, indicating different reactions triggered in alveolar epithelial cells and a great prospects of further comparative studies.

Regulation of Epithelial and Endothelial Barriers by Molecular Chaperones.

The integrity and permeability of epithelial and endothelial barriers depend on the formation of tight junctions, adherens junctions, and a junction-associated cytoskeleton. The establishment of this junction-cytoskeletal module relies on the correct folding and oligomerization of its protein components. Molecular chaperones are known regulators of protein folding and complex formation in different cellular compartments. Mammalian cells possess an elaborate chaperone network consisting of several hundred chaperones and co-chaperones. Only a small part of this network has been linked, however, to the regulation of intercellular adhesions, and the systematic analysis of chaperone functions at epithelial and endothelial barriers is lacking. This review describes the functions and mechanisms of the chaperone-assisted regulation of intercellular junctions. The major focus of this review is on heat shock protein chaperones, their co-chaperones, and chaperonins since these molecules are the focus of the majority of the articles published on the chaperone-mediated control of tissue barriers. This review discusses the roles of chaperones in the regulation of the steady-state integrity of epithelial and vascular barriers as well as the disruption of these barriers by pathogenic factors and extracellular stressors. Since cytoskeletal coupling is essential for junctional integrity and remodeling, chaperone-assisted assembly of the actomyosin cytoskeleton is also discussed.

Tight junction membrane proteins regulate the mechanical resistance of the apical junctional complex.

Epithelia must be able to resist mechanical force to preserve tissue integrity. While intercellular junctions are known to be important for the mechanical resistance of epithelia, the roles of tight junctions (TJs) remain to be established. We previously demonstrated that epithelial cells devoid of the TJ membrane proteins claudins and JAM-A completely lack TJs and exhibit focal breakages of their apical junctions. Here, we demonstrate that apical junctions fracture when claudin/JAM-A-deficient cells undergo spontaneous cell stretching. The junction fracture was accompanied by actin disorganization, and actin polymerization was required for apical junction integrity in the claudin/JAM-A-deficient cells. Further deletion of CAR resulted in the disruption of ZO-1 molecule ordering at cell junctions, accompanied by severe defects in apical junction integrity. These results demonstrate that TJ membrane proteins regulate the mechanical resistance of the apical junctional complex in epithelial cells.

Functional analysis reveals calcium-sensing receptor gene regulating cell-cell junction in renal tubular epithelial cells.

PURPOSE: Calcium-sensing receptor (CASR) influences the expression pattern of multiple genes in renal tubular epithelial cells. The objective of this inquiry was to explore the molecular mechanisms of CASR in renal tubular epithelial cells and nephrolithiasis. METHODS: HK-2 cells were transfected with lentiviruses carrying either CASR (named CASR) or an empty vector negative control (named NC), as well as shRNA intended to target CASR (named shCASR) or its corresponding negative control (named shNC). CCK-8 assay was used to detect the effect of CASR on the proliferation of HK-2 cells. RNA-Sequencing was applied to explore potential pathways regulated by CASR in HK-2 cells. RESULTS: PCR and western blot results showed that CASR expression was significantly increased in CASR cells and was decreased in shCASR cells when compared to their corresponding negative control, respectively. CCK-8 assay revealed that CASR inhibited the proliferation of HK-2 cells. RNA-Sequencing results suggested that the shCASR HK-2 cells exhibited a significant up-regulation of 345 genes and a down-regulation of 366 genes. These differentially expressed genes (DEGs) were related to cell apoptosis and cell development. In CASR HK-2 cells, 1103 DEGs primarily functioned in mitochondrial energy metabolism, and amino acid metabolism. With the Venn diagram, 4 DEGs (Clorf116, ENPP3, IL20RB, and CLDN2) were selected as the hub genes regulated by CASR. Enrichment analysis revealed that these hub genes were involved in cell-cell junction, and epithelial cell development. CONCLUSIONS: In summary, our investigation has the potential to offer novel perspectives on CASR regulating cell-cell junction in HK-2 cells.

Campylobacter jejuni Surface-Bound Protease HtrA, but Not the Secreted Protease nor Protease in Shed Membrane Vesicles, Disrupts Epithelial Cell-to-Cell Junctions.

Fundamental functions of the intestinal epithelium include the digestion of food, absorption of nutrients, and its ability to act as the first barrier against intruding microbes. Campylobacter jejuni is a major zoonotic pathogen accounting for a substantial portion of bacterial foodborne illnesses. The germ colonizes the intestines of birds and is mainly transmitted to humans through the consumption of contaminated poultry meat. In the human gastrointestinal tract, the bacterium triggers campylobacteriosis that can progress to serious secondary disorders, including reactive arthritis, inflammatory bowel disease and Guillain-Barré syndrome. We recently discovered that C. jejuni serine protease HtrA disrupts intestinal epithelial barrier functions via cleavage of the tight and adherens junction components occludin, claudin-8 and E-cadherin. However, it is unknown whether epithelial damage is mediated by the secreted soluble enzyme, by HtrA contained in shed outer-membrane vesicles (OMVs) or by another mechanism that has yet to be identified. In the present study, we investigated whether soluble recombinant HtrA and/or purified OMVs induce junctional damage to polarized intestinal epithelial cells compared to live C. jejuni bacteria. By using electron and confocal immunofluorescence microscopy, we show that HtrA-expressing C. jejuni bacteria trigger efficient junctional cell damage, but not soluble purified HtrA or HtrA-containing OMVs, not even at high concentrations far exceeding physiological levels. Instead, we found that only bacteria with active protein biosynthesis effectively cleave junctional proteins, which is followed by paracellular transmigration of C. jejuni through the epithelial cell layer. These findings shed new light on the pathogenic activities of HtrA and virulence strategies of C. jejuni.

The zonula adherens matura redefines the apical junction of intestinal epithelia.

Cell-cell apical junctions of epithelia consist of multiprotein complexes that organize as belts regulating cell-cell adhesion, permeability, and mechanical tension: the tight junction (zonula occludens), the zonula adherens (ZA), and the macula adherens. The prevailing dogma is that at the ZA, E-cadherin and catenins are lined with F-actin bundles that support and transmit mechanical tension between cells. Using super-resolution microscopy on human intestinal biopsies and Caco-2 cells, we show that two distinct multiprotein belts are basal of the tight junctions as the intestinal epithelia mature. The most apical is populated with nectins/afadin and lined with F-actin; the second is populated with E-cad/catenins. We name this dual-belt architecture the zonula adherens matura. We find that the apical contraction apparatus and the dual-belt organization rely on afadin expression. Our study provides a revised description of epithelial cell-cell junctions and identifies a module regulating the mechanics of epithelia.

Bicellular Localization of Tricellular Junctional Protein Angulin-3/ILDR2 Allows Detection of Podocyte Injury.

Podocytes serve as part of the renal filtration unit with slit diaphragms. Although the structure of slit diaphragms between two cells is well characterized, how the tricellular contact of podocytes is organized and how it changes in injured podocytes remains unknown. This study focused on a tricellular junction protein, angulin-3, and its localization in healthy podocytes, in developmental stages, and in pathologic conditions, using a newly established monoclonal antibody. Angulin-3 was confined at tricellular junctions of primordial podocytes, then transiently localized at bicellular junctions as foot process interdigitation developed and the intercellular junctions rearranged into slit diaphragm, and eventually distributed in a sparse punctate pattern on the foot processes of adult podocytes. In the rodent podocyte injury models, angulin-3 showed bicellular localization between the foot processes, and the localization turned from punctate to dashed linear pattern along the effaced foot processes with the progression of podocyte injury. Angulin-3 also accumulated between foot processes in a linear pattern in kidney biopsy samples of human nephrotic syndrome. Additionally, the line length of angulin-3 staining signal correlated with risk of relapse under glucocorticoid therapy in patients with minimal change nephrotic syndrome. This study proposes an image program to score the linearity of the accumulation pattern of angulin-3 to evaluate the relapse risk of patients with minimal change nephrotic syndrome.

The Dilute domain in Canoe is not essential for linking cell junctions to the cytoskeleton but supports morphogenesis robustness.

Robust linkage between adherens junctions and the actomyosin cytoskeleton allows cells to change shape and move during morphogenesis without tearing tissues apart. The Drosophila multidomain protein Canoe and its mammalian homolog afadin are crucial for this, as in their absence many events of morphogenesis fail. To define the mechanism of action for Canoe, we are taking it apart. Canoe has five folded protein domains and a long intrinsically disordered region. The largest is the Dilute domain, which is shared by Canoe and myosin V. To define the roles of this domain in Canoe, we combined biochemical, genetic and cell biological assays. AlphaFold was used to predict its structure, providing similarities and contrasts with Myosin V. Biochemical data suggested one potential shared function - the ability to dimerize. We generated Canoe mutants with the Dilute domain deleted (CnoΔDIL). Surprisingly, they were viable and fertile. CnoΔDIL localized to adherens junctions and was enriched at junctions under tension. However, when its dose was reduced, CnoΔDIL did not provide fully wild-type function. Furthermore, canoeΔDIL mutants had defects in the orchestrated cell rearrangements of eye development. This reveals the robustness of junction-cytoskeletal connections during morphogenesis and highlights the power of natural selection to maintain protein structure.

Palmitoylation of proteolipid protein M6 promotes tricellular junction assembly in epithelia of Drosophila.

Tricellular junctions (TCJs) seal epithelial cell vertices and are essential for tissue integrity and physiology, but how TCJs are assembled and maintained is poorly understood. In Drosophila, the transmembrane proteins Anakonda (Aka, also known as Bark), Gliotactin (Gli) and M6 organize occluding TCJs. Aka and M6 localize in an interdependent manner to vertices and act jointly to localize Gli, but how these proteins interact to assemble TCJs was not previously known. Here, we show that the proteolipid protein M6 physically interacts with Aka and with itself, and that M6 is palmitoylated on conserved juxta-membrane cysteine residues. This modification promotes vertex localization of M6 and binding to Aka, but not to itself, and becomes essential when TCJ protein levels are reduced. Abolishing M6 palmitoylation leads to delayed localization of M6 and Aka but does not affect the rate of TCJ growth or mobility of M6 or Aka. Our findings suggest that palmitoylation-dependent recruitment of Aka by M6 promotes initiation of TCJ assembly, whereas subsequent TCJ growth relies on different mechanisms that are independent of M6 palmitoylation.

ESCRT-III-dependent adhesive and mechanical changes are triggered by a mechanism detecting alteration of septate junction integrity in Drosophila epithelial cells.

Barrier functions of proliferative epithelia are constantly challenged by mechanical and chemical constraints. How epithelia respond to and cope with disturbances of barrier functions to allow tissue integrity maintenance is poorly characterised. Cellular junctions play an important role in this process and intracellular traffic contribute to their homeostasis. Here, we reveal that, in Drosophila pupal notum, alteration of the bi- or tricellular septate junctions (SJs) triggers a mechanism with two prominent outcomes. On one hand, there is an increase in the levels of E-cadherin, F-actin, and non-muscle myosin II in the plane of adherens junctions. On the other hand, ß-integrin/Vinculin-positive cell contacts are reinforced along the lateral and basal membranes. We found that the weakening of SJ integrity, caused by the depletion of bi- or tricellular SJ components, alters ESCRT-III/Vps32/Shrub distribution, reduces degradation and instead favours recycling of SJ components, an effect that extends to other recycled transmembrane protein cargoes including Crumbs, its effector ß-Heavy Spectrin Karst, and ß-integrin. We propose a mechanism by which epithelial cells, upon sensing alterations of the SJ, reroute the function of Shrub to adjust the balance of degradation/recycling of junctional cargoes and thereby compensate for barrier junction defects to maintain epithelial integrity.

Calcium influx promotes PLEKHG4B localization to cell-cell junctions and regulates the integrity of junctional actin filaments.

PLEKHG4B is a Cdc42-targeting guanine-nucleotide exchange factor implicated in forming epithelial cell-cell junctions. Here we explored the mechanism regulating PLEKHG4B localization. PLEKHG4B localized to the basal membrane in normal Ca2+ medium but accumulated at cell-cell junctions upon ionomycin treatment. Ionomycin-induced junctional localization of PLEKHG4B was suppressed upon disrupting its annexin-A2 (ANXA2)-binding ability. Thus, Ca2+ influx and ANXA2 binding are crucial for PLEKHG4B localization to cell-cell junctions. Treatments with low Ca2+ or BAPTA-AM (an intracellular Ca2+ chelator) suppressed PLEKHG4B localization to the basal membrane. Mutations of the phosphoinositide-binding motif in the pleckstrin homology (PH) domain of PLEKHG4B or masking of membrane phosphatidylinositol-4,5-biphosphate [PI(4,5)P2] suppressed PLEKHG4B localization to the basal membrane, indicating that basal membrane localization of PLEKHG4B requires suitable intracellular Ca2+ levels and PI(4,5)P2 binding of the PH domain. Activation of mechanosensitive ion channels (MSCs) promoted PLEKHG4B localization to cell-cell junctions, and their inhibition suppressed it. Moreover, similar to the PLEKHG4B knockdown phenotypes, inhibition of MSCs or treatment with BAPTA-AM disturbed the integrity of actin filaments at cell-cell junctions. Taken together, our results suggest that Ca2+ influx plays crucial roles in PLEKHG4B localization to cell-cell junctions and the integrity of junctional actin organization, with MSCs contributing to this process.

Integrin-based adhesions promote cell-cell junction and cytoskeletal remodelling to drive embryonic wound healing.

Embryos repair wounds rapidly, with no inflammation or scarring. Embryonic wound healing is driven by the collective movement of the cells around the lesion. The cells adjacent to the wound polarize the cytoskeletal protein actin and the molecular motor non-muscle myosin II, which accumulate at the wound edge forming a supracellular cable around the wound. Adherens junction proteins, including E-cadherin, are internalized from the wound edge and localize to former tricellular junctions at the wound margin, in a process necessary for cytoskeletal polarity. We found that the cells adjacent to wounds in the Drosophila embryonic epidermis polarized Talin, a core component of cell-extracellular matrix (ECM) adhesions, which preferentially accumulated at the wound edge. Integrin knockdown and inhibition of integrin binding delayed wound closure and reduced actin polarization and dynamics around the wound. Additionally, disrupting integrins caused a defect in E-cadherin reinforcement at tricellular junctions along the wound edge, suggesting crosstalk between integrin-based and cadherin-based adhesions. Our results show that cell-ECM adhesion contributes to embryonic wound repair and reveal an interplay between cell-cell and cell-ECM adhesion in the collective cell movements that drive rapid wound healing.

Oat ß-glucan repairs the epidermal barrier by upregulating the levels of epidermal differentiation, cell-cell junctions and lipids via Dectin-1.

BACKGROUND AND PURPOSE: Oat ß-glucan could ameliorate epidermal hyperplasia and accelerate epidermal barrier repair. Dectin-1 is one of the receptors of ß-glucan and many biological functions of ß-glucan are mediated by Dectin-1. Dectin-1 promotes wound healing through regulating the proliferation and migration of skin cells. Thus, this study aimed to investigate the role of oat ß-glucan and Dectin-1 in epidermal barrier repair. EXPERIMENTAL APPROACH: To investigate the role of Dectin-1 in the epidermal barrier, indicators associated with the recovery of a damaged epidermal barrier, including histopathological changes, keratinization, proliferation, apoptosis, differentiation, cell-cell junctions and lipid content were compared between WT and Dectin-1-/- mice. Further, the effect of oat ß-glucan on the disruption of the epidermal barrier was also compared between WT and Dectin-1-/- mice. KEY RESULTS: Dectin-1 deficiency resulted in delayed recovery and marked keratinization, as well as abnormal levels of keratinocyte differentiation, cell-cell junctions and lipid synthesis during the restoration of the epidermal barrier. Oat ß-glucan significantly reduces epidermal hyperplasia, promotes epidermal differentiation, increases cell-cell junction expression, promotes lipid synthesis and ultimately accelerates the recovery of damaged epidermal barriers via Dectin-1. Oat ß-glucan could promote CaS receptor expression and activate the PPAR-γ signalling pathway via Dectin-1. CONCLUSION AND IMPLICATIONS: Oat ß-glucan promote the recovery of damaged epidermal barriers through promoting epidermal differentiation, increasing the expression of cell-cell junctions and lipid synthesis through Dectin-1. Dectin-1 deficiency delay the recovery of epidermal barriers, which indicated that Dectin-1 may be a potential target in epidermal barrier repair.

Tight junctions.

Various functions within our bodies require the generation and maintenance of compartments with distinct compositions, which in turn necessitate the formation of semipermeable cellular diffusion barriers. For example, the blood-brain barrier protects the brain by allowing only specific molecules to pass through. Another instance is the intestinal barrier, which allows the uptake of essential nutrients, while restricting the passage of pathogenic molecules and bacteria. Breakdown of such barriers causes various pathologies, such as brain or retinal edema, or diarrhoea. Epithelia and endothelia are the most common barrier-forming cells. Individual cells in such barriers are held together by cell-cell adhesion structures - also known as intercellular junctions - that are essential for barrier formation and maintenance. Here, we will focus on the structure and assembly of tight junctions (TJs) and their functions as barriers, but will refer to other adhesive structures crucial for barrier regulation such as adherens junctions (AJs) and focal adhesions to the extracellular matrix (ECM) (Figure 1A,B). We will also discuss additional functions of TJs in cell surface polarity and the regulation of gene expression, cell function, and cell behaviour.