Activation of AMP-activated protein kinase alleviates High-glucose-induced dysfunction of brain microvascular endothelial cell tight-junction dynamics

The blood-brain barrier, formed by specialized brain endothelial cells that are interconnected by tight junctions, strictly regulates paracellular permeability to maintain an optimal extracellular environment for brain homeostasis. Diabetes is known to compromise the blood-brain barrier, although the underlying mechanism remains unknown. The aim of this study was to elucidate the molecular mechanisms underlying disruption of the blood-brain barrier in diabetes and to determine whether activation of AMP-activated protein kinase prevents diabetes-induced blood-brain barrier dysfunction. Exposure of human brain microvascular endothelial cells to high glucose (25mmol/L d-glucose), but not to high osmotic conditions (20mmol/L l-glucose plus 5mmol/L d-glucose), for 2h to 1 week significantly increased the permeability of the blood-brain barrier in parallel with lowered expression levels of zonula occludens-1, occludin, and claudin-5, three proteins that are essential to maintaining endothelial cell tight junctions. In addition, high glucose significantly increased the generation of superoxide anions. Adenoviral overexpression of superoxide dismutase or catalase significantly attenuated the high-glucose-induced reduction of endothelial cell tight-junction proteins. Furthermore, administration of apocynin reversed the effects of high glucose on endothelial cell tight-junction proteins. Finally, activation of AMP-activated protein kinase with 5-amino-4-imidazole carboxamide riboside or adenoviral overexpression of constitutively active AMP-activated protein kinase mutants abolished both the induction of NAD(P)H oxidase-derived superoxide anions and the tight-junction protein degradation induced by high glucose. We conclude that high glucose increases blood-brain barrier dysfunction in diabetes through induction of superoxide anions and that the activation of AMP-activated protein kinase protects the integrity of the blood-brain barrier by suppressing the induction of NAD(P)H oxidase-derived superoxide anions.     

Identification of paxilin domains interacting with β-catenin

Barrier-protective agonists induce association of focal adhesions (FA) and adherens junctions (AJ) in endothelial cells. Here we identified specific domains of FA protein paxillin interacting with AJ protein and examined regulation of paxillin domain interactions with β-catenin by Rac GTPase. Co-expression of paxillin LD-1,2; LD-3,4; LIM-1,2; and LIM-3,4 domains with β-catenin showed exclusive interaction of LIM-1,2 and LIM-3,4 with β-catenin, which was enhanced by agonist-induced Rac activation or expression of activated Rac mutant. These results demonstrate a novel function of paxillin LIM domains in targeting β-catenin in a Rac-dependent manner, which may play a role in Rac-dependent control of FA-AJ interactions and monolayer integrity.     

Contractile activity and cell-cell contact regulate myofibrillar organization in cultured cardiac myocytes

Adult feline ventricular myocytes cultured on a laminin-coated substratum reestablish intercellular junctions, yet disassemble their myofibrils. Immunofluorescence microscopy reveals that these non- beating heart cells lack vinculin-positive focal adhesions; moreover, intercellular junctions are also devoid of vinculin. When these quiescent myocytes are stimulated to contract with the beta-adrenergic agonist, isoproterenol, extensive vinculin-positive focal adhesions and intercellular junctions emerge. If solitary myocytes are stimulated to beat, an elaborate series of vinculin-positive focal adhesions develop which appear to parallel the reassembly of myofibrils. In cultures where neighboring myocytes reestablish cell-cell contact, myofibrils appear to reassemble from the fascia adherens rather than focal contacts. Activation of beating is accompanied by a significant reduction in the rate of total and cytoskeletal protein synthesis; in fact, myofibrillar reassembly, redevelopment of focal adhesions and fascia adherens junctions require no protein synthesis for at least 24 h, implying the existence of an assembly competent pool of cytoskeletal proteins. Maturation of the fasciae adherens and the appearance of vinculin within Z-line/costameres, does require de novo synthesis of new cytoskeletal proteins. Changes in cytoskeletal protein turnover appear dependent on beta agonist-induced cAMP production, but myofibrillar reassembly is a cAMP-independent event. Such observations suggest that mechanical forces, in the guise of contractile activity, regulate vinculin distribution and myofibrillar order in cultured adult feline heart cells.     

Antibodies to kidney endothelial cells contribute to a "leaky" glomerular barrier in patients with chronic kidney diseases

Anti-endothelial cell antibodies (AECA) have been reported to cause endothelial dysfunction, but their clinical importance for tissue-specific endothelial cells is not clear. We hypothesized that AECA reactive with human kidney endothelial cells (HKEC) may cause renal endothelial dysfunction in patients with chronic kidney diseases. We report that a higher fraction (56%) of end-stage renal disease (ESRD) patients than healthy controls (5%) have AECA reactive against kidney endothelial cells (P <0.001). The presence of antibodies was associated with female gender (P < 0.001), systolic hypertension (P < 0.01), and elevated TNF-α (P < 0.05). These antibodies markedly decrease expression of both adherens and tight junction proteins VE-cadherin, claudin-1, and zonula occludens-1 and provoked a rapid increase in cytosolic free Ca(2+) and rearrangement of actin filaments in HKEC compared with controls. This was followed by an enhancement in protein flux and phosphorylation of VE-cadherin, events associated with augmented endothelial cell permeability. Additionally, kidney biopsies from ESRD patients with AECA but not controls demonstrated a marked decrease in adherens and tight junctions in glomerular endothelium, confirming our in vitro data. In summary, our data demonstrate a causal link between AECA and their capacity to induce alterations in glomerular vascular permeability.     

Redox regulation of 4-hydroxy-2-nonenal-mediated endothelial barrier dysfunction by focal adhesion, adherens, and tight junction proteins

4-Hydroxy-2-nonenal (4-HNE), one of the major biologically active aldehydes formed during inflammation and oxidative stress, has been implicated in a number of cardiovascular and pulmonary disorders. 4-HNE has been shown to increase vascular endothelial permeability; however, the underlying mechanisms are unclear. Hence, in the current study, we tested our hypothesis that 4-HNE-induced changes in cellular thiol redox status may contribute to modulation of cell signaling pathways that lead to endothelial barrier dysfunction. Exposure of bovine lung microvascular endothelial cells (BLMVECs) to 4-HNE induced reactive oxygen species generation, depleted intracellular glutathione, and altered cell-cell adhesion as measured by transendothelial electrical resistance. Pretreatment of BLM-VECs with thiol protectants, N-acetylcysteine and mercaptopropionyl glycine, attenuated 4-HNE-induced decrease in transendothelial electrical resistance, reactive oxygen species generation, Michael protein adduct formation, protein tyrosine phosphorylation, activation of ERK, JNK, and p38 MAPK, and actin cytoskeletal rearrangement. Treatment of BLMVECs with 4-HNE resulted in the redistribution of FAK, paxillin, VE-cadherin, beta-catenin, and ZO-1, and intercellular gap formation. Western blot analyses confirmed the formation of 4-HNE-derived Michael adducts with the focal adhesion and adherens junction proteins. Also, 4-HNE decreased tyrosine phosphorylation of FAK without affecting total cellular FAK contents, suggesting the modification of integrins, which are natural FAK receptors. 4-HNE caused a decrease in the surface integrin in a time-dependent manner without altering total alpha5 and beta3 integrins. These results, for the first time, revealed that 4-HNE in redox-dependent fashion affected endothelial cell permeability by modulating cell-cell adhesion through focal adhesion, adherens, and tight junction proteins as well as integrin signal transduction that may lead dramatic alteration in endothelial cell barrier dysfunction during heart infarction, brain stroke, and lung diseases.     

蛋白激酶在微血管通透性中的作用

微血管内皮细胞层是一层半选择通透性屏障,可以调节血液中的液体、溶质和血浆蛋白进入组织间隙。在炎症刺激作用下,可通过旁细胞途径和跨细胞途径引起内皮通透性上升。旁细胞通路主要由内皮细胞间的紧密连接、黏附连接和细胞与外基质的黏着斑组成。炎症介质,如脂多糖和肿瘤坏死因子α可激活多种蛋白激酶。活化的蛋白激酶主要包括Rho相关的卷曲蛋白激酶、肌球蛋白轻链激酶、蛋白激酶C、酪氨酸激酶和丝裂原活化蛋白激酶等,参与引发内皮屏障生化和结构改变,旁细胞通路开放,导致通透性上升。该文对上述蛋白激酶在微血管通透性中作用机制的研究进展进行综述。     

Localization of paxillin, a focal adhesion protein, to smooth muscle dense plaques, and the myotendinous and neuromuscular junctions of skeletal muscle

In this report we have demonstrated that paxillin, a cytoskeletal protein which is present in focal adhesions, localizes in vivo to regions of cell-extracellular matrix interaction which are believed to be analogous to focal adhesions. Specifically, it is enriched in the dense plaques of chicken gizzard smooth muscle tissue and in the myotendinous junctions formed in Xenopus laevis tadpole tail skeletal muscle. In addition, paxillin was identified at the rat diaphragm neuromuscular junction. The distribution of paxillin is thus comparable to that of other focal adhesion proteins, for example, talin and vinculin, in these structures.     

Actopaxin, a new focal adhesion protein that binds paxillin LD motifs and actin and regulates cell adhesion

Paxillin is a focal adhesion adapter protein involved in the integration of growth factor- and adhesion-mediated signal transduction pathways. Paxillin LD motifs have been demonstrated to bind to several proteins associated with remodeling of the actin cytoskeleton including the focal adhesion kinase, vinculin, and a complex of proteins comprising p95PKL, PIX, and PAK (Turner, C.E., M. C. Brown, J.A. Perrotta, M.C. Riedy, S.N. Nikolopoulos, A.R. McDonald, S. Bagrodia, S. Thomas, and P.S. Leventhal. 1999. J. Cell Biol. 145:851-863). In this study, we report the cloning and initial characterization of a new paxillin LD motif-binding protein, actopaxin. Analysis of the deduced amino acid sequence of actopaxin reveals a 42-kD protein with two calponin homology domains and a paxillin-binding subdomain (PBS). Western blotting identifies actopaxin as a widely expressed protein. Actopaxin binds directly to both F-actin and paxillin LD1 and LD4 motifs. It exhibits robust focal adhesion localization in several cultured cell types but is not found along the length of the associated actin-rich stress fibers. Similar to paxillin, it is absent from actin-rich cell-cell adherens junctions. Also, actopaxin colocalizes with paxillin to rudimentary focal complexes at the leading edge of migrating cells. An actopaxin PBS mutant incapable of binding paxillin in vitro cannot target to focal adhesions when expressed in fibroblasts. In addition, ectopic expression of the PBS mutant and/or the COOH terminus of actopaxin in HeLa cells resulted in substantial reduction in adhesion to collagen. Together, these results suggest an important role for actopaxin in integrin-dependent remodeling of the actin cytoskeleton during cell motility and cell adhesion.     

Cell confluence-dependent remodeling of endothelial membranes mediated by cholesterol

The plasma membranes of endothelial cells reaching confluence undergo profound structural and functional modifications, including the formation of adherens junctions, crucial for the regulation of vascular permeability and angiogenesis. Adherens junction formation is accompanied by the tyrosine dephosphorylation of adherens junctions proteins, which has been correlated with the strength and stability of adherens junctions. Here we show that cholesterol is a critical determinant of plasma membrane remodeling in cultures of growing cow pulmonary aortic endothelial cells. Membrane cholesterol increased dramatically at an early stage in the formation of confluent cow pulmonary aortic endothelial cell monolayers, prior to formation of intercellular junctions. This increase was accompanied by the redistribution of caveolin from a high density to a low density membrane compartment, previously shown to require cholesterol, and increased binding of the annexin II-p11 complex to membranes, consistent with other studies indicating cholesterol-dependent binding of annexin II to membranes. Furthermore, partial depletion of cholesterol from confluent cells with methyl-beta-cyclodextrin both induced tyrosine phosphorylation of multiple membrane proteins, including adherens junctions proteins, and disrupted adherens junctions. Both effects were dramatically reduced by prior complexing of methyl-beta-cyclodextrin with cholesterol. Our results reveal a novel physiological role for cholesterol regulating the formation of adherens junctions and other plasma membrane remodeling events as endothelial cells reach confluence.     

Paxillin: a new vinculin-binding protein present in focal adhesions

The 68-kD protein (paxillin) is a cytoskeletal component that localizes to the focal adhesions at the ends of actin stress fibers in chicken embryo fibroblasts. It is also present in the focal adhesions of Madin-Darby bovine kidney (MDBK) epithelial cells but is absent, like talin, from the cell-cell adherens junctions of these cells. Paxillin purified from chicken gizzard smooth muscle migrates as a diffuse band on SDS-PAGE gels with a molecular mass of 65-70 kD. It is a protein of multiple isoforms with pIs ranging from 6.31 to 6.85. Using purified paxillin, we have demonstrated a specific interaction in vitro with another focal adhesion protein, vinculin. Cleavage of vinculin with Staphylococcus aureus V8 protease results in the generation of two fragments of approximately 85 and 27 kD. Unlike talin, which binds to the large vinculin fragment, paxillin was found to bind to the small vinculin fragment, which represents the rod domain of the molecule. Together with the previous observation that paxillin is a major substrate of pp60src in Rous sarcoma virus-transformed cells (Glenney, J. R., and L. Zokas. 1989. J. Cell Biol. 108:2401-2408), this interaction with vinculin suggests paxillin may be a key component in the control of focal adhesion organization.     

Vinculin associates with endothelial VE-cadherin junctions to control force-dependent remodeling

To remodel endothelial cell-cell adhesion, inflammatory cytokine- and angiogenic growth factor-induced signals impinge on the vascular endothelial cadherin (VE-cadherin) complex, the central component of endothelial adherens junctions. This study demonstrates that junction remodeling takes place at a molecularly and phenotypically distinct subset of VE-cadherin adhesions, defined here as focal adherens junctions (FAJs). FAJs are attached to radial F-actin bundles and marked by the mechanosensory protein Vinculin. We show that endothelial hormones vascular endothelial growth factor, tumor necrosis factor α, and most prominently thrombin induced the transformation of stable junctions into FAJs. The actin cytoskeleton generated pulling forces specifically on FAJs, and inhibition of Rho-Rock-actomyosin contractility prevented the formation of FAJs and junction remodeling. FAJs formed normally in cells expressing a Vinculin binding-deficient mutant of α-catenin, showing that Vinculin recruitment is not required for adherens junction formation. Comparing Vinculin-devoid FAJs to wild-type FAJs revealed that Vinculin protects VE-cadherin junctions from opening during their force-dependent remodeling. These findings implicate Vinculin-dependent cadherin mechanosensing in endothelial processes such as leukocyte extravasation and angiogenesis.     

Lysophosphatidic Acid Increases Tight Junction Permeability in Cultured Brain Endothelial Cells

Abstract: Brain capillary endothelial cells are coupled by a continuous belt of complex high-electrical-resistance tight junctions that are largely responsible for the blood-brain barrier. We have investigated mechanisms regulating tight junction permeability in brain endothelial cells cultured to maintain high-resistance junctions. The phospholipid lysophosphatidic acid (LPA) was found to cause a rapid, reversible, and dose-dependent decrease in transcellular electrical resistance in brain endothelial cells. LPA also increased the paracellular flux of sucrose, which, together with the resistance decrease, indicated increased tight junction permeability. Activation of protein kinase C attenuated the effect of LPA, suggesting that it was mediated by activation of a signalling pathway. LPA did not cause any obvious relocalization of adherens junction- or tight junction-associated proteins. However, it did stimulate the formation of stress fibres, the recruitment of focal adhesion components, and the appearance of tyrosine phosphorylated protein at focal contacts. Our study shows that LPA is a modulator of tight junction permeability in brain endothelial cells in culture and raises the possibility that it triggers blood-brain barrier permeability changes under (patho)physiological conditions.     

Unique cell type-specific junctional complexes in vascular endothelium of human and rat liver sinusoids

Liver sinusoidal endothelium is strategically positioned to control access of fluids, macromolecules and cells to the liver parenchyma and to serve clearance functions upstream of the hepatocytes. While clearance of macromolecular debris from the peripheral blood is performed by liver sinusoidal endothelial cells (LSECs) using a delicate endocytic receptor system featuring stabilin-1 and -2, the mannose receptor and CD32b, vascular permeability and cell trafficking are controlled by transcellular pores, i.e. the fenestrae, and by intercellular junctional complexes. In contrast to blood vascular and lymphatic endothelial cells in other organs, the junctional complexes of LSECs have not yet been consistently characterized in molecular terms. In a comprehensive analysis, we here show that LSECs express the typical proteins found in endothelial adherens junctions (AJ), i.e. VE-cadherin as well as α-, β-, p120-catenin and plakoglobin. Tight junction (TJ) transmembrane proteins typical of endothelial cells, i.e. claudin-5 and occludin, were not expressed by rat LSECs while heterogenous immunreactivity for claudin-5 was detected in human LSECs. In contrast, junctional molecules preferentially associating with TJ such as JAM-A, B and C and zonula occludens proteins ZO-1 and ZO-2 were readily detected in LSECs. Remarkably, among the JAMs JAM-C was considerably over-expressed in LSECs as compared to lung microvascular endothelial cells. In conclusion, we show here that LSECs form a special kind of mixed-type intercellular junctions characterized by co-occurrence of endothelial AJ proteins, and of ZO-1 and -2, and JAMs. The distinct molecular architecture of the intercellular junctional complexes of LSECs corroborates previous ultrastructural findings and provides the molecular basis for further analyses of the endothelial barrier function of liver sinusoids under pathologic conditions ranging from hepatic inflammation to formation of liver metastasis.     

Ovarian hormones regulate expression of the focal adhesion proteins,talin and paxillin,in rat uterine luminal but not glandular epithelial cells

During early pregnancy in the rat, focal adhesions disassemble in uterine luminal epithelial cells at the time of implantation to facilitate their removal so that the implanting blastocyst can invade into the underlying endometrial decidual cells. This study investigated the effect of ovarian hormones on the distribution and protein expression of two focal adhesion proteins, talin and paxillin, in rat uterine luminal and glandular epithelial cells under various hormone regimes. Talin and paxillin showed a major distributional change between different hormone regimes. Talin and paxillin were highly concentrated along the basal cell surface of uterine luminal epithelial cells in response to oestrogen treatment. However, this prominent staining of talin and paxillin was absent and also a corresponding reduction of paxillin expression was demonstrated in response to progesterone alone or progesterone in combination with oestrogen, which is also observed at the time of implantation. In contrast, the distribution of talin and paxillin in uterine glandular epithelial cells was localised on the basal cell surface and remained unchanged in all hormone regimes. Thus, not all focal adhesions are hormonally dependent in the rat uterus; however, the dynamics of focal adhesion in uterine luminal epithelial cells is tightly regulated by ovarian hormones. In particular, focal adhesion disassembly in uterine luminal epithelial cells, a key component to establish successful implantation, is predominantly under the influence of progesterone.     

Activation of MMP-2, cleavage of matrix proteins,and adherens junctions during a snake venom metalloproteinase-induced endothelial cell apoptosis

Snake venom metalloproteinases (SVMPs) are structurally and functionally similar to matrix metalloproteinases (MMPs). We have previously demonstrated that a SVMP, named gaminelysin, can induce endothelial cell apoptosis [Biochem J. 357 (2001) 719]. In this study, the action mechanism of graminelysin in causing endothelial cell apoptosis was further investigated. We showed that the apoptosis was initiated with cell shape change and extracellular matrix degradation and occurred before cell detachment. Cleaved forms of MMP-2 might act in concert with graminelysin to cause apoptosis. During apoptosis, adherens junctions, including VE-cadherin and beta- and gamma-catenin were cleaved and alpha-catenin was decreased. VE-cadherin and beta-catenin at cell periphery were decreased and the discontinuity in alignment was found as observed with immunofluorescence microscopy. This was accompanied with a diffuse beta-catenin staining in the cytoplasm and a decreased F-actin stress fibers in some rounded cells. The decrease of VE-cadherin and beta-catenin in Triton-insoluble fractions confirmed that the association of adherens junctions with actin cytoskeleton was altered during apoptosis. Graminelysin-induced cleavage in adherens junctions was paralleled with the changes in paracellular permeability. We also detected the activation of caspase-3 and the decrease of Bcl-2/Bax ratio during apoptosis. However, caspase inhibitors showed differential effects in blocking the cleavage of PARP, adherens junctions, and DNA fragmentation. Taken together, the data presented suggest that metalloproteinase can control cell fates via the degradation of matrix proteins, the change of cell shape, and the cleavage of adherens junctions.     

Electron microscopic study of the intercellular junctions during the ciliary epithelial differentiation of the eye of the common frog]

Intercellular junctions in the eye ciliary epithelium were studied in Rana temporaria by means of transmission electron microscopy beginning from the stage of appearance of the light sensitivity in the larval eye and until the completion of metamorphosis. In the ciliary epithelium inner layer the apical parts of adjacent cells are tied by contacts forming the junction complex already at a very early developmental stage. The most apical position in this complex is occupied by the focal junction which appears to be an early stage of zonula occludens; this complex includes also zonula adherens and macula adherens which may follow in any succession focal junction or zonula occludens. No definite order was found in the localization of cell contacts between the outer and inner layers of ciliary epithelium, as well as between the side surfaces of the cells within each layer. Contacts were found everywhere termed as "lengthy" which may be considered as gap junctions. Regional differences were found in the ultrastructure of the ciliary epithelium cells with respect to unequal distribution of functional loads.     

Afadin controls p120-catenin-ZO-1 interactions leading to endothelial barrier enhancement by oxidized phospholipids

Afadin is a novel regulator of epithelial cell junctions assembly. However, its role in the formation of endothelial cell junctions and the regulation of vascular permeability remains obscure. We previously described protective effects of oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine (OxPAPC) in the in vitro and in vivo models of lung endothelial barrier dysfunction and acute lung injury, which were mediated by Rac GTPase. This study examined a role of afadin in the OxPAPC-induced enhancement of interactions between adherens junctions and tight junctions as a novel mechanism of endothelial cell (EC) barrier preservation. OxPAPC induced Rap1-dependent afadin accumulation at the cell periphery and Rap1-dependent afadin interaction with adherens junction and tight junction proteins p120-catenin and ZO-1, respectively. Afadin knockdown using siRNA or ectopic expression of afadin mutant lacking Rap1 GTPase binding domain suppressed OxPAPC-induced EC barrier enhancement and abolished barrier protective effects of OxPAPC against thrombin-induced EC permeability. Afadin knockdown also abolished protective effects of OxPAPC against ventilator-induced lung injury in vivo. These results demonstrate for the first time a critical role of afadin in the regulation of vascular barrier function in vitro and in vivo via coordination of adherens junction-tight junction interactions.     

A helping hand: How vinculin contributes to cell-matrix and cell-cell force transfer

Vinculin helps cells regulate and respond to mechanical forces. It is a scaffolding protein that tightly regulates its interactions with potential binding partners within adhesive structures—including focal adhesions that link the cell to the extracellular matrix and adherens junctions that link cells to each other—that physically connect the force-generating actin cytoskeleton (CSK) with the extracellular environment. This tight control of binding partner interaction—mediated by vinculin's autoinhibitory head–tail interaction—allows vinculin to rapidly interact and detach in response to changes in the dynamic forces applied through the cell. In doing so, vinculin modulates the structural composition of focal adhesions and the cell's ability to generate traction forces and adhesion strength. Recent evidence suggests that vinculin plays a similar role in regulating the fate and function of cell–cell junctions, further underscoring the importance of this protein. Using our lab's recent work as a starting point, this commentary explores several outstanding questions regarding the nature of vinculin activation and its function within focal adhesions and adherens junctions.     

Type XIII collagen: a novel cell adhesion component present in a range of cell-matrix adhesions and in the intercalated discs between cardiac muscle cells.

Recent analysis of type XIII collagen surprisingly showed that it is anchored to the plasma membranes of cultured cells via a transmembrane segment near its amino terminus. Here we demonstrate that type XIII collagen is concentrated in cultured skin fibroblasts and several other human mesenchymal cell lines in the focal adhesions at the ends of actin stress fibers, co-localizing with the known focal adhesion components talin and vinculin. This co-occurrence was also observed in rapidly forming adhesive structures of spreading and moving fibroblasts and in disrupting focal adhesions following microinjection of the Rho-inhibitor C3 transferase into the cells, suggesting that type XIII collagen is an integral focal adhesion component. Moreover, it appears to have an adhesion-related function since cell-surface expression of type XIII collagen in cells with weak basic adhesiveness resulted in improved cell adhesion on selected culture substrata. In tissues type XIII collagen was found in a range of integrin-mediated adherens junctions including the myotendinous junctions and costameres of skeletal muscle as well as many cell-basement membrane interfaces. Some cell-cell adhesions were found to contain type XIII collagen, most notably the intercalated discs in the heart. Taken together, the results strongly suggest that type XIII collagen has a cell adhesion-associated function in a wide array of cell-matrix junctions.