Tissue Vasculature Research Articles

Fluorescence imaging in colorectal surgery

Fluorescent imaging is an emerging technological tool that can guide surgeons during surgery by highlighting anatomical structures and pathology, and help with intraoperative decision making. A comprehensive review of published literature was performed using the search terms "fluorescence", "imaging" and "colorectal surgery" in PubMed. Only clinical trials that were published in English were included in this review. Ex vivo and animal studies were excluded. This review demonstrates the use of fluorescence imaging in colorectal surgery in four areas: (1) assessment of tissue perfusion and vasculature; (2) assessment of tumour; (3) lymphatic drainage and (4) identification of the urinary tract. The most commonly used fluorescent dyes are nonspecific, such as indocyanine green and methylene blue, but there is increasing interest in the development of specific fluorescently labelled molecular markers. Fluorescence imaging is a potentially useful tool for colorectal surgery. Early studies on fluorescence imaging have been promising but larger scale randomised controlled trials are warranted to demonstrate the effectiveness and benefits of using fluorescence imaging routinely. The development of molecular dyes that are specific to targets could significantly increase the potential use of fluorescence imaging during surgery.

Pancreas Optical Clearing and 3-D Microscopy in Health and Diabetes

Although first described over a hundred years ago, tissue optical clearing is undergoing renewed interest due to numerous advances in optical clearing methods, microscopy systems, and three-dimensional (3-D) image analysis programs. These advances are advantageous for intact mouse tissues or pieces of human tissues because samples sized several millimeters can be studied. Optical clearing methods are particularly useful for studies of the neuroanatomy of the central and peripheral nervous systems and tissue vasculature or lymphatic system. Using examples from solvent- and aqueous-based optical clearing methods, the mouse and human pancreatic structures and networks will be reviewed in 3-D for neuro-insular complexes, parasympathetic ganglia, and adipocyte infiltration as well as lymphatics in diabetes. Optical clearing with multiplex immunofluorescence microscopy provides new opportunities to examine the role of the nervous and circulatory systems in pancreatic and islet functions by defining their neurovascular anatomy in health and diabetes.

Optoacoustic imaging in endocrinology and metabolism.

Imaging is an essential tool in research, diagnostics and the management of endocrine disorders. Ultrasonography, nuclear medicine techniques, MRI, CT and optical methods are already used for applications in endocrinology. Optoacoustic imaging, also termed photoacoustic imaging, is emerging as a method for visualizing endocrine physiology and disease at different scales of detail: microscopic, mesoscopic and macroscopic. Optoacoustic contrast arises from endogenous light absorbers, such as oxygenated and deoxygenated haemoglobin, lipids and water, or exogenous contrast agents, and reveals tissue vasculature, perfusion, oxygenation, metabolic activity and inflammation. The development of high-performance optoacoustic scanners for use in humans has given rise to a variety of clinical investigations, which complement the use of the technology in preclinical research. Here, we review key progress with optoacoustic imaging technology as it relates to applications in endocrinology; for example, to visualize thyroid morphology and function, and the microvasculature in diabetes mellitus or adipose tissue metabolism, with particular focus on multispectral optoacoustic tomography and raster-scan optoacoustic mesoscopy. We explain the merits of optoacoustic microscopy and focus on mid-infrared optoacoustic microscopy, which enables label-free imaging of metabolites in cells and tissues. We showcase current optoacoustic applications within endocrinology and discuss the potential of these technologies to advance research and clinical practice.

Modular mucopolysaccharide gelatin naturapolyceutics hydrocolloid biomatrix with cobalt nano-additives for high density vascular network assembly

The present study demonstrates the development of polysaccharide gelatin naturapolyceutics hydrocolloidal biomatrix with cobalt nano-additives for restructuring native tissue vasculature for tissue regenerative applications. The engineered Gelatin/Aloevera mucilage polysaccharide/nanoscaled Cobalt (GAC) hydrocolloids resulted from the intermolecular interactions between the aloevera mucilage, cobalt nano-therapeutic and gelatin. GAC hydrocolloid showed enhanced thermal stability in comparison with control Gelatin/Aloevera mucilage (GA) hydrocolloid. FTIR analysis validated that the reinforcement of aloevera mucilage and cobalt nano-therapeutic did not affect the structural integrity of the gelatin molecule. 3-Dimensional sponge-like orientation of GAC hydrocolloid facilitates perfusable biomatrix for access to nutrients and gaseous exchange for high cell adhesion and proliferation. The combined therapeutic efficacy of mucilage polysaccharides, biodegradable nanoscaled cobalt and bio-polymer enhanced the pro-angiogenic capability of the hydrocolloids by stimulating Vascular Endothelial Growth Factor (VEGF) response at wounded tissue for faster healing. The experimental outcomes on in vivo angiogenesis profiling further confirmed the development of micro vessel in chick embryonic model and regeneration of blood vessels in zebra fish model. This study opens up the potential of mucilage polysaccharides in stimulating high density angiogenesis and conveys the progress of a biocompatible, biodegradable mucilaginous hydrocolloid as an effective bio-adhesive for vascular development in soft tissue regeneration.

CTP Synthase 2 From Arabidopsis thaliana Is Required for Complete Embryo Development.

Pyrimidine de novo synthesis is an essential pathway in all organisms. The final and rate-limiting step in the synthesis of the nucleotide cytidine triphosphate (CTP) is catalyzed by CTP synthase (CTPS), and Arabidopsis harbors five isoforms. Single mutant lines defective in each one of the four isoforms do not show apparent phenotypical alterations in comparison to wild-type plants. However, Arabidopsis lines that contain T-DNA insertions in the CTPS2 gene were unable to produce homozygous offspring. Here, we show that CTPS2 exhibits a distinct expression pattern throughout embryo development, and loss-of-function mutants are embryo lethal, as siliques from +/ctps2 plants contained nearly 25% aborted seeds. This phenotype was rescued by complementation with CTPS2 under control of its endogenous promoter. CTPS2::GFP lines revealed expression only in the tip of columella cells in embryo root tips of the heart and later stages. Furthermore, CTPS2 expression in mature roots, most pronounced in the columella cells, shoots, and vasculature tissue of young seedlings, was observed. Filial generations of +/ctps2 plants did not germinate properly, even under external cytidine supply. During embryo development, the CTPS2 expression pattern resembled the established auxin reporter DR5::GFP. Indeed, the cloned promoter region we used in this study possesses a repeat of an auxin response element, and auxin supply increased CTPS2 expression in a cell-type-specific manner. Thus, we conclude that CTPS2 is essential for CTP supply in developing embryos, and loss-of-function mutants in CTPS2 are embryo lethal.

Heterogeneity and Dynamics of Vasculature in the Endocrine System During Aging and Disease.

The endocrine system consists of several highly vascularized glands that produce and secrete hormones to maintain body homeostasis and regulate a range of bodily functions and processes, including growth, metabolism and development. The dense and highly vascularized capillary network functions as the main transport system for hormones and regulatory factors to enable efficient endocrine function. The specialized capillary types provide the microenvironments to support stem and progenitor cells, by regulating their survival, maintenance and differentiation. Moreover, the vasculature interacts with endocrine cells supporting their endocrine function. However, the structure and niche function of vasculature in endocrine tissues remain poorly understood. Aging and endocrine disorders are associated with vascular perturbations. Understanding the cellular and molecular cues driving the disease, and age-related vascular perturbations hold potential to manage or even treat endocrine disorders and comorbidities associated with aging. This review aims to describe the structure and niche functions of the vasculature in various endocrine glands and define the vascular changes in aging and endocrine disorders.

Quantification of microvasculature parameters based on optoacoustic angiography data

Optoacoustic (OA) angiography allows high-contrast three-dimensional (3D) visualization of hemoglobin-containing structures ranging from micrometers to millimeters. However, due to the large amount of 3D data acquired by modern high-throughput OA systems the resulting OA vasculature images might be difficult to analyze visually. This problem is especially relevant for monitoring of angiogenesis of experimental tumors, which blood vessels tend to be smaller and more tortuous compared to vasculature of healthy tissue. In this paper a novel algorithm for OA image processing is proposed to quantify vessel structure parameters automatically. The algorithm is based on creation of vasculature graphs which parameters (lengths of branches, number of branches, etc) can serve as a numerical characterization of vasculature: vessel density, vessel length, etc. The results of testing the developed algorithm on numerical simulation phantoms and in vivo OA images of tumor models in a mouse demonstrate a statistically significant difference of all the extracted parameters for tumor and normal tissue. The results show a high potential of the proposed approach for OA angiography in different applications including clinical and experimental oncology.

Tissue Clearing and 3-D Visualization of Vasculature with the PEGASOS Method.

Tissue clearing techniques turn tissue transparent through a series of chemical and physical treatments. They have provided a useful tool for three-dimensional (3-D) imaging to study tissue spatial organization and interactions. Many tissue clearing methods have been developed in recent years. Each method has its own application range depending on the purposes of the study. Three criteria for selecting an appropriate clearing method include clearing transparency, fluorescence preservation, and broad tissue applicability. PEG-associated solvent system (PEGASOS) emerged recently as a solvent-based tissue clearing method capable of rendering diverse tissues highly transparent while preserving fluorescence. Combined with vascular labeling techniques, PEGASOS method enables 3-D visualization of vasculature in whole tissues at subcellular resolution. Here, we describe the standard PEGASOS passive immersion protocol and several compatible vascular labeling techniques. Methods of 3-D imaging, data processing, and annotations are also briefly introduced.

Uncertainty propagation and sensitivity analysis: results from the Ocular Mathematical Virtual Simulator.

We propose an uncertainty propagation study and a sensitivity analysis with the Ocular Mathematical Virtual Simulator, a computational and mathematical model that predicts the hemodynamics and biomechanics within the human eye. In this contribution, we focus on the effect of intraocular pressure, retrolaminar tissue pressure and systemic blood pressure on the ocular posterior tissue vasculature. The combination of a physically-based model with experiments-based stochastic input allows us to gain a better understanding of the physiological system, accounting both for the driving mechanisms and the data variability.

Injection of Evans blue dye to fluorescently label and image intact vasculature.

Blood vessels perform critical functions in both health and disease. Understanding how vessels form, pattern and respond to damage is essential. However, labeling and imaging the vasculature to ascertain these properties can be difficult and time-consuming. Here, the authors present a novel methodology for rapidly and efficiently labeling whole vascular networks in vivo by exploiting the fluorescent properties of Evans blue. By combining the labeling with fluorescence microscopy, this method enables visualization of whole tissue vasculature for a fraction of the time and cost compared with traditional methods.

Harnessing Single-Cell RNA Sequencing to Better Understand How Diseased Cells Behave the Way They Do in Cardiovascular Disease.

The transition of healthy arteries and cardiac valves into dense, cell-rich, calcified, and fibrotic tissues is driven by a complex interplay of both cellular and molecular mechanisms. Specific cell types in these cardiovascular tissues become activated following the exposure to systemic stimuli including circulating lipoproteins or inflammatory mediators. This activation induces multiple cascades of events where changes in cell phenotypes and activation of certain receptors may trigger multiple pathways and specific alterations to the transcriptome. Modifications to the transcriptome and proteome can give rise to pathological cell phenotypes and trigger mechanisms that exacerbate inflammation, proliferation, calcification, and recruitment of resident or distant cells. Accumulating evidence suggests that each cell type involved in vascular and valvular diseases is heterogeneous. Single-cell RNA sequencing is a transforming medical research tool that enables the profiling of the unique fingerprints at single-cell levels. Its applications have allowed the construction of cell atlases including the mammalian heart and tissue vasculature and the discovery of new cell types implicated in cardiovascular disease. Recent advances in single-cell RNA sequencing have facilitated the identification of novel resident cell populations that become activated during disease and has allowed tracing the transition of healthy cells into pathological phenotypes. Furthermore, single-cell RNA sequencing has permitted the characterization of heterogeneous cell subpopulations with unique genetic profiles in healthy and pathological cardiovascular tissues. In this review, we highlight the latest groundbreaking research that has improved our understanding of the pathological mechanisms of atherosclerosis and future directions for calcific aortic valve disease.

Laminin N-terminus α31 protein distribution in adult human tissues.

Laminin N-terminus α31 (LaNt α31) is a netrin-like protein derived from alternative splicing of the laminin α3 gene. Although LaNt α31 has been demonstrated to influence corneal and skin epithelial cell function, its expression has not been investigated beyond these tissues. In this study, we used immunohistochemistry to characterise the distribution of this protein in a wide-array of human tissue sections in comparison to laminin α3. The data revealed widespread LaNt α31 expression. In epithelial tissue, LaNt α31 was present in the basal layer of the epidermis, throughout the epithelium of the digestive tract, and in much of the epithelium of the reproductive system. LaNt α31 was also found throughout the vasculature of most tissues, with enrichment in reticular-like fibres in the extracellular matrix surrounding large vessels. A similar matrix pattern was observed around the terminal ducts in the breast and around the alveolar epithelium in the lung, where basement membrane staining was also evident. Specific enrichment of LaNt α31 was identified in sub-populations of cells of the kidney, liver, pancreas, and spleen, with variations in intensity between different cell types in the collecting ducts and glomeruli of the kidney. Intriguingly, LaNt α31 immunoreactivity was also evident in neurons of the central nervous system, in the cerebellum, cerebral cortex, and spinal cord. Together these findings suggest that LaNt α31 may be functionally relevant in a wider range of tissue contexts than previously anticipated, and the data provides a valuable basis for investigation into this interesting protein.

Osseous lesions in the distal extremities of dogs with strangulating hair mats.

Strangulating hair mats in dogs are characterized as soft tissue injury secondary to chronically matted hair, which can encircle the extremities and constrict lymphatic tissue and local vasculature. As the tissue swells, the hair constriction tightens and may result in tissue necrosis and osseous changes secondary to ischemic necrosis. The objectives of this retrospective case series were to describe the radiographic characteristics of persistent hair mat strangulation in dogs, to identify clinical features that would distinguish between dogs with soft tissue lesions only and dogs with osseous changes and to describe the clinical outcome in these dogs. All dogs with clinically identified strangulating hair matting had radiographically identified soft tissues defects and over half of the limbs evaluated had osseous lesions including focal periostitis, bone remodeling, bone atrophy, lysis of bone, and occasionally subluxation of joints. Chronically matted hair and strangulating hair mats in dogs are recognized by veterinarians as a medical finding in animal neglect. The osseous features of strangulating hair mats have not been described in the veterinary medical literature. This study describes the clinical features and radiographic characteristics of strangulating hair mats.

A multitarget angiogenesis inhibitor, CTT peptide-endostatin mimic-kringle 5, prevents diet-induced obesity.

Adipose tissue vasculature has been considered an attractive target for prevention and treatment of obesity. AARP (CTT peptide-endostatin mimic-kringle 5) is a novel multitarget fusion protein against tumor angiogenesis. This study aimed to examine the effects of AARP on diet-induced obesity and its possible molecular mechanism. Treatment with AARP markedly prevented weight gains, improved metabolic disturbances, and decreased adipose tissue angiogenesis in diet-induced obese mice without noticeable toxicities. In addition to its potent antiangiogenic and MMP-2/9 inhibitory activities, AARP administration also significantly increased energy expenditure, influenced the metabolic and angiogenic gene expression profiles, and attenuated obesity-induced inflammation, demonstrating its systemic beneficial effects. Importantly, AARP exhibited no effect on mice fed with standard normal mouse diet. Furthermore, the AARP-treated HFD-fed mice experienced a significant increase in lifespan during the posttreatment observation period, compared with untreated HFD-fed mice. Our results suggest that AARP might be pharmacologically useful for treatment of obesity or obesity-related metabolic disorders in humans. KEY MESSAGES: What is already known • More effective and safe therapies for obesity are in urgent need. • AARP is a novel multitarget fusion protein against tumor angiogenesis. What this study adds • AARP prevents obesity, improves metabolic disorders in mice fed high-fat diet. • AARP increases energy expenditure, decreases adipose tissue angiogenesis, and increases lifespan. • AARP is well tolerated and exhibits no observable toxicity. Clinical significance • AARP may be a promising therapeutic agent against obesity or obesity-related metabolic disturbances.

Unique blood vasculature and innervation in the cavernous tissue of murine vomeronasal organs.

The vomeronasal organ (VNO) is an accessory olfactory device related to reproductive behavior. The soft tissue of the tubular organ is composed of sensory/non-sensory epithelia and a highly developed vasculature, which in the latter the dilation and contraction of blood vessels are thought to contribute to pumping in and out luminal fluid or air, like penile erectile tissue. The present histological observation of the murine VNO revealed a more complicated vasculature than previously evaluated ones with large differences along the rostro-caudal axis. An immunohistochemical study for vasoactive substances displayed extremely dense innervation by cholinergic nerves containing nitric oxide synthase and VIP/PHI in the thick smooth muscle layer surrounding venous sinuses at light and electron microscopic levels. Furthermore, the differential distribution of cholinergic nerves and adrenergic nerves may provide a novel insight into the pumping mechanism of VNO.

Tissue Engineering of Lymphatic Vasculature in the Arteriovenous Loop Model of the Rat.

Various therapeutic approaches, for example, in case of trauma or cancer require the transplantation of autologous tissue. Depending on the size and the origin of the harvested tissue, these therapies can lead to iatrogenic complications and donor-site morbidities. In future, these side effects could be avoided by transplanting artificially generated tissue consisting of different cell types and matrix components derived from the host body. Tissue that is grown in the patient could be advantageous compared with the more simply structured in vitro-grown alternatives. To overcome the limitations of graft vascularization, the arteriovenous (AV) loop technique has been established for different tissues in the last years and was adapted for lymphatic tissue engineering in the present study. We utilized the AV loop technique to grow human lymphatic vasculature in vivo in the Rowett nude (RNU) rat. A combination of human lymphatic endothelial cells (LECs) and bone marrow-derived mesenchymal stem cells was implanted in a fibrin matrix surrounding the AV loop. After 2 or 4 weeks of implantation, the animals were perfused and the tissue was harvested. It could be demonstrated by immunohistochemistry for human LYVE1, human CD31, and murine podoplanin that the implanted cells formed human lymphatic vasculature in the AV loop chamber. Beside development of murine podoplanin-positive vasculature in the AV loop tissue, vasculature positive for human marker proteins developed in comparable numbers. This suggests that implanted LECs are able to improve the lymphatic vascularization of the newly engineered tissue. Thus, we were able to establish an in vivo tissue engineering method to generate lymphatic vascularized soft tissue. An axially vascularized transplantable lymphatic vessel network was engineered without requiring advanced cell culture equipment, rendering the lymphatic AV loop highly suitable for applied regenerative medicine. Impact statement Various surgical procedures require the transplantation of autologous harvested tissue, for example, the vascularized lymph node transfer for the treatment of lymphedema. Tissue-engineered transplants could be used instead of autologous transplants and thereby help to reduce the side effects of those therapies. However, in vitro tissue engineering of large constructs requires a lot of know-how as well as advanced cell culture equipment, which might not be accessible in every hospital. In vivo tissue engineering approaches like the presented technique for the generation of transplantable networks of lymphatic vasculature could serve as an alternative for in vitro tissue engineering approaches in clinical settings.

Abstract 26: Intraductal procedure with refined ethanol-containing ablative solution for primary prevention of breast cancer

Abstract Breast cancer (BC) is the second leading cause of cancer-related deaths in women. One in every eight women will develop BC during her lifetime. For moderate and low risk women, there are few options available for prevention. For high-risk patients, prophylactic mastectomy is the most effective intervention for primary prevention of BC. Prophylactic mastectomy consists of the complete removal of mammary epithelial cells, from which the majority of BC arises, and the surrounding tissue which can have great physical, emotional, and psychological tolls on the individual. Therefore, fewer than 30% of high-risk individuals choose to undergo this aggressive procedure as their primary intervention. We seek to develop an alternative, minimally invasive procedure that intraductally delivers a cell-killing solution throughout the ductal tree to ablate the mammary epithelial cells before they become malignant. Existing methods such as ductography for clinical diagnostic imaging support the technical feasibility of this approach. We recently published a proof-of-concept study with ethanol as an intraductal (ID) ablative solution in non-transgenic FVB mice and in a C(3)1-tag FVB BC model. Ethanol is a safe compound with multiple clinical applications as an ablative and sclerosing agent. ID injections of 70% ethanol showed maximal and most consistent epithelial cell ablation than other tested concentrations with limited and tolerable collateral damage in the surrounding tissue. ID injections of 70% ethanol in cancer prone C(3)1-tag mice significantly delayed tumor formation and reduced tumor incidence. We followed up 10 ethanol-injected FVB animals until their natural death (median survival > 630 days), with no long-term adverse effects observed. Here, we tested different formulations to further minimize collateral tissue damage of 70% ethanol. We added ethyl cellulose, a gelling agent used to limit diffusion of ethanol in clinical treatments of venous malformation and pre-clinical liver cancer, to the 70% ethanol solution. Non-transgenic FVB mice and Sprague-Dawley rats ID injected with this refined ablative formulation achieved the same ablative rates as 70% ethanol solution but further minimized collateral damage to surrounding vasculature and adipose tissue. Introduction of tantalum oxide nanocrystals as high-resolution radiographic contrast agent into the ablative solution enabled unprecedented in vivo visualization of ductal tree filling and monitoring of 70% ethanol ablative effects in rodent models. Future studies will test the efficacy of this refined formulation to prevent BC in genetically engineered mouse models and chemical-induced rat models. The pre-clinical validation of this ID ablative procedure in aforementioned cancer-prone rodent models will further support the translatability of this approach to first-in-human clinical trials for high-risk individuals. Citation Format: Erin Kay Zaluzec, Elizabeth Kenyon, Maximilian Volk, Brooke L. Jackson, Jeremy Hix, Shatadru Chakravarty, Erik Shapiro, Anna Moore, Lorenzo F. Sempere. Intraductal procedure with refined ethanol-containing ablative solution for primary prevention of breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 26.

Exercise training improves adipose tissue metabolism and vasculature regardless of baseline glucose tolerance and sex

IntroductionWe investigated the effects of a supervised progressive sprint interval training (SIT) and moderate-intensity continuous training (MICT) on adipocyte morphology and adipose tissue metabolism and function; we also tested whether...

NADPH-quinone oxidoreductase-1 mediates Benzo-[a]-pyrene-1,6-quinone-induced cytotoxicity and reactive oxygen species production in human EA.hy926 endothelial cells

Numerous studies conducted in the past have reported deaths in the human population due to cardiovascular diseases (CVD) on exposure to air particulate matter (APM). BP-1,6-quinone (BP-1,6-Q) is one of the significant components of APM. However, the mechanism(s) by which it can exert its toxicity in endothelial cells is not yet completely understood. NAD(P)H: quinone oxidoreductase-1 (NQO1) is expressed highly in myocardium and vasculature tissues of the heart and plays a vital role in maintaining vascular homeostasis. This study, demonstrated that BP-1,6-Q diminishes NQO1 enzyme activity in a dose-dependent manner in human EA.hy926 endothelial cells. The decrease in the NQO1 enzyme causes potentiation in BP-1,6-Q-mediated toxicity in EA.hy926 endothelial cells. The enhancement of NQO1 in endothelial cells showed cytoprotection against BP-1,6-Q-induced cellular toxicity, lipid, and protein damage suggesting an essential role of NQO1 in cytoprotection against BP-1,6-Q toxicity. Using various biochemical assays and genetic approaches, results from this study further demonstrated that NQO1 also plays a crucial role in BP-1,6-Q-induced production of reactive oxygen species (ROS). These findings will contribute to elucidating BP-1,6-Q mediated toxicity and its role in the development of atherosclerosis.

JAM-A Acts via C/EBP-α to Promote Claudin-5 Expression and Enhance Endothelial Barrier Function.

Intercellular tight junctions are crucial for correct regulation of the endothelial barrier. Their composition and integrity are affected in pathological contexts, such as inflammation and tumor growth. JAM-A (junctional adhesion molecule A) is a transmembrane component of tight junctions with a role in maintenance of endothelial barrier function, although how this is accomplished remains elusive. We aimed to understand the molecular mechanisms through which JAM-A expression regulates tight junction organization to control endothelial permeability, with potential implications under pathological conditions. Genetic deletion of JAM-A in mice significantly increased vascular permeability. This was associated with significantly decreased expression of claudin-5 in the vasculature of various tissues, including brain and lung. We observed that C/EBP-α (CCAAT/enhancer-binding protein-α) can act as a transcription factor to trigger the expression of claudin-5 downstream of JAM-A, to thus enhance vascular barrier function. Accordingly, gain-of-function for C/EBP-α increased claudin-5 expression and decreased endothelial permeability, as measured by the passage of fluorescein isothiocyanate (FITC)-dextran through endothelial monolayers. Conversely, C/EBP-α loss-of-function showed the opposite effects of decreased claudin-5 levels and increased endothelial permeability. Mechanistically, JAM-A promoted C/EBP-α expression through suppression of β-catenin transcriptional activity, and also through activation of EPAC (exchange protein directly activated by cAMP). C/EBP-α then directly binds the promoter of claudin-5 to thereby promote its transcription. Finally, JAM-A-C/EBP-α-mediated regulation of claudin-5 was lost in blood vessels from tissue biopsies from patients with glioblastoma and ovarian cancer. We describe here a novel role for the transcription factor C/EBP-α that is positively modulated by JAM-A, a component of tight junctions that acts through EPAC to up-regulate the expression of claudin-5, to thus decrease endothelial permeability. Overall, these data unravel a regulatory molecular pathway through which tight junctions limit vascular permeability. This will help in the identification of further therapeutic targets for diseases associated with endothelial barrier dysfunction. Graphic Abstract: An graphic abstract is available for this article.