Arterial Vascular Endothelium Research Articles

Diagnostic criteria of endothelial dysfunction, subclinical atherosclerosis and their relationship with NOS3 and HIF1A gene polymorphism in military personnel in extreme conditions of the arctic zone

The state of the arterial vascular endothelium against the background of subclinical atherosclerosis and its relationship with gene polymorphism in military personnel serving in the Arctic climatic zone, depending on the duration of service, was evaluated. In total, 251 male servicemen aged 28–40 years were examined. The group of servicemen of the Arctic climate zone included 99 people, who were divided into three groups according to the duration of military service in the Arctic: group 1 included 8 (8%) servicemen with up to 5 years of military service; group 2, 21 (21%) people with 5–10 years; group 3, 70 (71%) servicemen with 10 years. The control group included 152 military personnel aged 28–40 years from the temperate climate zone. All participants underwent a test with reactive hyperemia, elastography, measurement of the thickness of the intima–media complex on the brachial artery, ultrasound examination of the common carotid arteries, photoplethysmography, pulse oximetry, spirometry, anthropometry, biochemical blood analysis, and determination of gene polymorphism using polymerase chain reaction. The results revealed that endothelium-dependent vasodilation rats in groups 2 and 3 were 1. 7 and 2. 7 times less than that in group 1 (p 0. 05). In 11 (16%) military personnel of group 3, thickening of the intima–media complex was found in 50% of the adjacent areas of the common carotid arteries. In groups 2 and 3, the total blood cholesterol level exceeded the value by 10% and 14%, respectively, compared with group 1 (p 0. 05). The artery wall was rigid in carriers of the T/T genotypes of the HIF1A and NOS3 genes and was most elastic in the G/T genotypes of the NOS3 gene and C/C of the HIF1A gene. In general, military personnel serving in the Arctic climatic zone with a service duration of 10 years have endothelial dysfunction and peripheral vascular stiffness with subclinical atherosclerosis phenomena, particularly in military personnel with the T/T genotypes of HIF1A (rs11549465) and NOS3 (rs1799983). We believe that the leading pathogenetic link of endothelial dysfunction of arterial vessels is the resistance of myocytes to the effects of vasodilation factors expressed by the vascular endothelium, particularly nitric oxide. The longer the service experience of military personnel under conditions of polar hypoxia, the higher the probability of developing endothelial dysfunction.

Concurrent Ex Vivo Differentiation of a Vascular Niche and Hematopoietic Stem/Progenitor Cells from Human iPSCs

One of the most promising objectives of clinical hematology is to derive transplantable autologous hematopoietic stem/progenitor cells (HSPCs) ex vivo. Recent reports have described the generation of engraftable HSPCs via forced expression of key transcription factors in either endothelial cells, or hemogenic endothelium (HE) derived from induced pluripotent stem cells (iPSCs). While protocols for the generation of transgene-free engraftable HSPCs suitable for therapy remain unavailable, these studies, along with observations in the developing embryo, have shown the close association of HE populations with arterial vascular endothelium is of critical importance to HSPC production and maturation. To facilitate the development of a vascular niche supporting the emergence of transgene-free, functional HSPCs from human iPSCs, we developed a simple, monolayer-based, chemically-defined, and scalable differentiation protocol requiring no replating or embryoid body (EB) formation (commercially available as STEMdiffTM Hematopoietic Kit, Stem Cell Technologies). During the first 3 days, a mesodermal lineage specification was induced with morphogens (BMP4, bFGF and VEGF). For the subsequent 18 days, cells were further differentiated into HSPCs with the addition of hematopoietic cytokines (SCF, Flt3-L and TPO). Starting at day 5, an adherent monolayer composed in part of CD34+CD43- endothelial cells rapidly formed, followed by the initiation of endothelial-to-hematopoietic transition (EHT), as demonstrated by the emergence of round cells directly budding off the endothelial monolayer in time-lapse imaging studies (Panel A). Importantly, these cells were released within the culture supernatant and could be easily harvested by simple pipetting. Flow cytometric analysis of the supernatant cells indicated that the emerging cells initially had a predominant CD45-CD235a+ erythroid phenotype (maximum 69.9±12.4% at day 7), consistent with a primitive yolk-sac wave of hematopoiesis (PanelB). From day 7, CD45+CD34+ HSPCs increased gradually, with a maximum number (average 1x105 cells/cm2, n=10) observed at day 12, and a peak percentage at day 12 (47.4±12.4%) and 14 (48.8±9.9%) (Panel B). These cells could form colonies in clonogenic progenitor assays, albeit at reduced capacity compared to bona fide CD45+CD34+ cells. Strikingly, cells with a phenotype that enables the highest reported purity of human HSCs (CD34+ CD38- CD45RA- CD90+ CD49f+ Rholo) were highly enriched in the culture supernatant at day 10 through 14 (Panel C). However, these cells did not result in efficient, long-term engraftment in immuno-deficient (NSG) mice. To identify possible causes for the lack of durable repopulating potential of iPSC-derived HSPCs in this system, we further characterized the endothelial monolayer between day 5 and 12 of differentiation. We focused on recently described endothelial subsets within the CD34hiCD43- population, including the in vitro equivalent of definitive HE (CD34hi CD43- CD73- CD184- DLL4-), arterial vascular endothelium (CD34hi CD43- CD73mid CD184+), and venous vascular endothelium (CD34hi CD43- CD73hi CD184-) (Ditadi et al., Nat. Cell Biol 2015). At each timepoint analyzed, definitive HE comprised <1% of the total cell population (Panel D). Arterial vascular endothelium was also scarce, rising to a maximum of 6.6% at day 7 before declining to <1% by day 9 of culture (Panel D). Venous vascular endothelium accounted for approximately 2% of total cells (Panel D). The remaining cells of the monolayer expressed varying levels of CD34 and CD43, consistent with the recently demonstrated heterogeneity of cells undergoing EHT (Guibentif et al., Cell Rep 2017). Thus, our data suggest that the absence of a definitive HE population within a developing arterial niche, as occurs in the dorsal aorta (DA) during development, may explain in part the lack of engraftment potential of iPSC-derived HSPCs. The unique adaptable and scalable nature of this monolayer-based differentiation protocol will facilitate precise temporal activation or downregulation of signaling pathways essential for the generation of engraftable human HSPCs. Quantitative and temporal modulations of Wnt/Nodal pathways for mesodermal specification as well as VEGF/Notch pathways for arterial vascular endothelium are underway. [Display omitted] DisclosuresChen:Stem Cell Technologies: Patents & Royalties: StemDiff Hematopoietic Kit. Boehm:StemCell Technologies: Patents & Royalties: StemDiff Hematopoietic Kit. Larochelle:Novartis: Research Funding; StemCell Technologies: Patents & Royalties: StemDiff Hematopoietic Kit; Novartis: Research Funding.

Robust generation of erythroid and multilineage hematopoietic progenitors from human iPSCs using a scalable monolayer culture system.

One of the most promising objectives of clinical hematology is to derive engraftable autologous hematopoietic stem cells (HSCs) from human induced pluripotent stem cells (iPSCs). Progress in translating iPSC technologies to the clinic relies on the availability of scalable differentiation methodologies. In this study, human iPSCs were differentiated for 21 days using STEMdiff™, a monolayer-based approach for hematopoietic differentiation of human iPSCs that requires no replating, co-culture or embryoid body formation. Both hematopoietic and non-hematopoietic cells were functionally characterized throughout differentiation. In the hematopoietic fraction, an early transient population of primitive CD235a+ erythroid progenitor cells first emerged, followed by hematopoietic progenitors with multilineage differentiation activity in vitro but no long-term engraftment potential in vivo. In later stages of differentiation, a nearly exclusive production of definitive erythroid progenitors was observed. In the non-hematopoietic fraction, we identified a prevalent population of mesenchymal stromal cells and limited arterial vascular endothelium (VE), suggesting that the cellular constitution of the monolayer may be inadequate to support the generation of HSCs with durable repopulating potential. Quantitative modulation of WNT/β-catenin and activin/nodal/TGFβ signaling pathways with CHIR/SB molecules during differentiation enhanced formation of arterial VE, definitive multilineage and erythroid progenitors, but was insufficient to orchestrate the generation of engrafting HSCs. Overall, STEMdiff™ provides a clinically-relevant and readily adaptable platform for the generation of erythroid and multilineage hematopoietic progenitors from human pluripotent stem cells.

Effects of Porphyromonas gingivalis on interleukin-33 expression in rabbit vascular endothelium tissues

To investigate interleukin-33 (IL-33) in the arterial vascular endothelium of rabbits infected with Porphyromonas gingivalis (P. gingivalis), and to explore the relationship between P. gingivalis and atherosclerosis. A total of 24 rabbits were randomly divided into control and experimental groups. The experimental group received intravenous injection of P. gingivalis once a week for 12 weeks to establish a coronary atherosclerosis model. The rabbits in the control group were injected with equal volume of physiological saline. All the rabbits were killed after 13 weeks. The IL-33 expression levels in the arterial vascular endothelium of the rabbits were detected through immunohistochemistry, reverse transcription polymerase chain reaction, and Western blot analysis. The effects of P. gingivalis on the IL-33 expression in the arterial vascular endothelium of the rabbits were analyzed. The relative expression levels of IL-33 mRNA in the vascular endothelium cells were 58.244±2.407, and the relative expression levels of IL-33 protein were 1.863±0.171 in the experimental group. The relative expression levels of IL-33 mRNA were 3.143±0.805, and the relative expression levels of IL-33 protein were 0.537± 0.028 in the control group. The expression levels of IL-33 mRNA and protein of vascular endothelium cells in the experimental group were significantly higher than those of the control group (P<0.01). P. gingivalis infection promotes IL-33 expression levels in vascular endothelial cells and may regulate the occurrence and development of atherosclerosis. .

Human definitive haemogenic endothelium and arterial vascular endothelium represent distinct lineages

The generation of haematopoietic stem cells (HSCs) from human pluripotent stem cells (hPSCs) will depend on the accurate recapitulation of embryonic haematopoiesis. In the early embryo, HSCs develop from the haemogenic endothelium (HE) and are specified in a Notch-dependent manner through a process named endothelial-to-haematopoietic transition (EHT). As HE is associated with arteries, it is assumed that it represents a subpopulation of arterial vascular endothelium (VE). Here we demonstrate at a clonal level that hPSC-derived HE and VE represent separate lineages. HE is restricted to the CD34+CD73−CD184− fraction of day 8 embryoid bodies (EBs) and it undergoes a NOTCH-dependent EHT to generate RUNX1C+ cells with multilineage potential. Arterial and venous VE progenitors, by contrast, segregate to the CD34+CD73medCD184+ and CD34+CD73hiCD184− fractions, respectively. Together, these findings identify HE as distinct from VE and provide a platform for defining the signalling pathways that regulate their specification to functional HSCs.

Porcine Extrahepatic Vascular Endothelial Asialoglycoprotein Receptor 1 Mediates Xenogeneic Platelet Phagocytosis In Vitro and in Human-to-Pig Ex Vivo Xenoperfusion

Asialoglycoprotein receptor-1 (ASGR1) mediates capture and phagocytosis of platelets in pig-to-primate liver xenotransplantation. However, thrombocytopenia is also observed in xenotransplantation or xenoperfusion of other porcine organs than liver. We therefore assessed ASGR1 expression as well as ASGR1-mediated xenogeneic platelet phagocytosis in vitro and ex vivo on porcine aortic, femoral arterial, and liver sinusoidal endothelial cells (PAEC/PFAEC/PLSEC). Porcine forelimbs were perfused with whole, heparinized human or autologous pig blood. Platelets were counted at regular intervals. Pig limb muscle and liver, as well as PAEC/PFAEC/PLSEC, were characterized for ASGR1 expression. In vitro, PAEC cultured on microcarrier beads and incubated with non-anticoagulated human blood were used to study binding of human platelets and platelet-white blood cell aggregation. Carboxyfluorescein diacetate succinimidyl ester-labeled human platelets were exposed to PAEC/PFAEC/PLSEC and analyzed for ASGR1-mediated phagocytosis. Human platelet numbers decreased from 102 ± 33 at beginning to 13 ± 6 × 10/μL (P < 0.0001) after 10 minutes of perfusion, whereas no significant decrease of platelets was seen during autologous perfusions (171 ± 26 to 122 ± 95 × 10/μL). The PAEC, PFAEC, and PLSEC all showed similar ASGR1 expression. In vitro, no correlation was found between reduction in platelet count and platelet-white blood cell aggregation. Phagocytosis of human carboxyfluorescein diacetate succinimidyl ester-labeled platelets by PAEC/PFAEC/PLSEC peaked at 15 minutes and was inhibited (P < 0.05 to P < 0.0001) by rabbit anti-ASGR1 antibody and asialofetuin. The ASGR1 expressed on aortic and limb arterial pig vascular endothelium plays a role in binding and phagocytosis of human platelets. Therefore, ASGR1 may represent a novel therapeutic target to overcome thrombocytopenia associated with vascularized pig-to-primate xenotransplantation.

Declined Presentation: Human definitive hemogenic endothelium represents a distinct lineage from arterial vascular endothelium

The successful derivation of hematopoietic stem cells (HSCs) from human pluripotent stem cells (hPSCs) will be dependent on our ability to accurately recapitulate critical aspects of embryonic hematopoiesis in the differentiation cultures. In the embryo, HSCs arise from a progenitor population known as hemogenic endothelium (HE), through a Notch dependent process that involves an endothelial to hematopoietic transition (ETH). Given that HE expresses endothelial markers and is associated with the major arteries in the embryo, it is assumed that this progenitor population derives from the arterial vascular endothelium (VE).

Generation of definitive hematopoietic progenitors from human pluripotent stem cells

The hematopoietic system in the early embryo consists of two distinct programs, primitive hematopoiesis and definitive hematopoiesis that can be distinguished by site and timing of development and the spectrum of lineages generated. Primitive hematopoiesis, the first program to develop, is restricted to the yolk sac and produces primitive erythrocytes, macrophages and megakaryocytes, but does not give rise to lymphoid cells or hematopoietic stem cells. Lymphoid lineage cells and HSCs are generated during the definitive stage of hematopoiesis that emerges following the onset of primitive hematopoiesis. Definitive program develops from a progenitor population known as hemogenic endothelium that is specified at different sites within the vasculature of the embryo. To derive HSCs from human pluripotent stem cells, it is essential to be able to accurately recapitulate these development stages including the formation of HE and its specification to a hematopoietic fate in the differentiation cultures. Using T lymphoid potential and primitive erythropoiesis to monitor the emergence of the two hematopoietic programs from hPSCs, we have recently found that both primitive and definitive hematopoiesis develop from KDR progenitors that are specified within the first three days of differentiation. The primitive KDR progenitors can be distinguished from those of the definitive lineage by the expression of CD235a and by the requirement for Wnt/b-catenin signaling. Development of the primitive KDR+CD235a+ progenitor is Wnt independent, whereas specification of the KDR+CD235a- definitive progenitor is dependent on signaling through this pathway, specifically between days 2 and 3 of differentiation. Following an additional five days of differentiation, the Wnt-induced definitive progenitors generate a CD34+ population that expresses cell surface markers indicative of HE and displays the capacity to give rise to lymphoid, myeloid and erythroid progeny. Based on expression of the vascular markers, CD73 and CD184, this day 8 CD34+ population can be separated into three distinct fractions: CD73+CD184+, CD73+CD184- and CD73-CD184-. Functional analyses revealed that hematopoietic potential reflective of HE segregated to the CD73-CD184- fraction, whereas the CD73+CD184+ and CD73+CD184- fractions were found to contain progenitors of arterial and venous vascular endothelium, respectively. Taken together, these findings demonstrate that it is possible to model definitive hematopoietic development from hPSCs and to identify and isolate a population enriched in HE, the putative progenitor of the human HSC.

2011 Vascular Research Initiatives Conference: basic foundations of translational research in vascular disease

The Vascular Research Initiatives Conference (VRIC) is an annual conference organized by the Society for Vascular Surgery (SVS). The 2011 VRIC was held in Chicago (IL, USA) to precede and coincide with the first day of the meeting of the Council on Arteriosclerosis, Thrombosis and Vascular Biology (ATVB) of the American Heart Association. The event is designed to present world class vascular research results, encourage collaboration between vascular surgeons and basic scientists in related disciplines, as well as to stimulate interest in research among aspiring academic vascular surgeons. The 2011 VRIC featured plenary sessions addressing peripheral arterial disease, vascular endothelium and thrombosis, aneurysms, and stem cells and tissue engineering. Recipients of the SVS partner grants with the National Institutes of Health K08 awardees presented their progress reports, and keynote addresses were given by Linda Graham and Frank LoGerfo.

Pharmacodynamic Characterization of Nitric Oxide-Mediated Vasodilatory Activity in Isolated Perfused Rat Mesenteric Artery Bed

Vasodilation profiles following a short-term infusion of nitric oxide (NO), acetylcholine (ACh), and sodium nitroprusside (SNP) into an isolated perfused mesenteric artery bed were analyzed in rats to examine their vasodilatory efficacy under physiological conditions. These compounds commonly increase the intracellular NO concentration to exert vasodilatory activity. In an experiment with exogenous NO infusion where 100 µl of 1 : 300 diluted NO-saturated solution (approx. 53 pmol of NO) was applied, the infusion caused transient vasodilation in a dose-dependent manner, with the peak vasodilation value being 74.7% of the maximum relaxation value. In experiments with ACh, the peak vasodilation value was 81.5% of the maximum at a dose of 60 pmol. The vasodilation profile of ACh was similar to that of NO infusion, but the ACh-induced vasodilation reduced at a slower rate than that induced by NO infusion. The vasodilatory activity of SNP was less potent than that of ACh, and its peak value was 62.8% of the maximum at a dose of 2000 pmol. However, SNP activity was augmented by removing the vascular endothelia of the mesenteric artery bed, and the peak value reached 67.3% of the maximum at a dose of 60 pmol. Pharmacodynamic analysis indicated that NO and ACh are equivalent regarding their vasodilatory efficacy, while the efficacy of SNP was less than 1% of theirs, as the arterial vascular endothelium impeded intracellular SNP-related NO generation, by which 95% of SNP's vasodilatory efficacy was negated. These findings will be helpful to understand factors influencing the therapeutic efficacy of vasodilators.

Role of NFκB in age-related vascular endothelial dysfunction in humans

Role of NFκB in age-related vascular endothelial dysfunction in humans

Chlamydia pneumoniae stimulates the proliferation of HUVEC through the induction of VEGF by THP-1

Chlamydia pneumoniae, a gram-negative bacterium, is an important human intracellular pathogen; studies of C. pneumoniae pathogenesis have shown that the organism can infect many cell types associated with both respiratory and vascular sites, including arterial smooth muscle cells, macrophages and vascular endothelium. Recently, the recognition of atherosclerosis as an inflammatory disease in its genesis, progression and ultimate clinical manifestations has created an interesting area of vascular research. We assessed the hypothesis that growth factors from THP-1 macrophages infected with C. pneumoniae are involved in the regulation of cell proliferation in HUVEC.The induction of these factors were dependent on time of infection, as medium harvested 48 h after infection had a greater activity than media harvested at 12 or 24 h after infection. Heat-killed C. pneumoniae produced similar results to those of live bacteria. In addition, conditioned medium filtered sterile from infected macrophages induced the proliferation of HUVEC, thus demonstrating its angiogenic potential. Moreover, pretreatment of macrophages with cytochalasin D, a phagocytosis inhibitor, yielded almost comparable results, suggesting that bacterium cell-attachment is sufficient for VEGF, IL-1β and IL-8 induction.Further studies are necessary to elucidate the biological role of chlamydial involvement in the complex and mutifactored processes of angiogenesis and possibly contribute to the development of therapeutic strategies.

CCM2 Expression Parallels That of CCM1

Mutations in CCM2 (MGC4607 or malcavernin) cause familial cerebral cavernous malformation (CCM), an autosomal dominant neurovascular disease. Both the function of this molecule and the pathogenesis of the disease remain elusive. We analyzed the mRNA expression of Ccm1 and Ccm2 in the embryonic and postnatal mouse brain by in situ hybridization. Subsequently, we generated CCM2-specific polyclonal antibodies and tested their specificity using transient transfection experiments in various cell lines. We then investigated CCM2 protein expression in cerebral and extracerebral tissues by Western blot analysis as well as immunohistochemistry and compared these results with CCM1 (KRIT1) protein expression. In situ analysis shows similar temporal and spatial expression patterns for Ccm1 and Ccm2, although Ccm1 expression appears more widespread. Immunohistochemical analysis shows that CCM2 is expressed in various human organs, most noticeably in the arterial vascular endothelium. As is the case with CCM1, CCM2 is not expressed in other vascular wall elements such as smooth muscle cells or the venous circulation. Within cerebral tissue, it is also expressed in pyramidal neurons, astrocytes, and their foot processes. In extracerebral tissues, CCM2 is present in various epithelial cells necessary for blood-organ barrier formation. CCM1 and CCM2 have similar expression patterns during development and postnatally thereafter. Given the fact that the disease phenotypes caused by mutations in either gene are clinically and pathologically indistinguishable, the significant overlap in expression pattern supports the hypothesis that both molecules are involved in the same pathway important for central nervous system vascular development.

Chlamydia pneumoniae and atherosclerosis.

Exposure to Chlamydia pneumoniae is extremely common, and respiratory infections occur repeatedly among most people. Strong associations exist between C. pneumoniae infection and atherosclerosis as demonstrated by: (i) sero-epidemiological studies showing that patients with cardiovascular disease have higher titres of anti-C. pneumoniae antibodies compared with control patients; (ii) detection of the organism within atherosclerotic lesions, but not in adjacent normal tissue by immunohistochemistry, polymerase chain reaction and electron microscopy and by culturing the organism from lesions; and (iii) showing that C. pneumoniae can either initiate lesion development or cause exacerbation of lesions in rabbit and mouse animal models respectively. The association of this organism with atherosclerosis has also provided sufficient impetus to conduct a variety of human secondary prevention antibiotic treatment trials. The results of these studies have been mixed and, thus far, no clear long-lasting benefit has emerged from these types of investigations. Studies of C. pneumoniae pathogenesis have shown that the organism can infect many cell types associated with both respiratory and cardiovascular sites, including lung epithelium and resident alveolar macrophages, circulating monocytes, arterial smooth muscle cells and vascular endothelium. Infected cells have been shown to exhibit characteristics associated with the development of cardiovascular disease (e.g. secretion of proinflammatory cytokines and procoagulants by infected endothelial cells and foam cell formation by infected macrophages). More detailed analysis of C. pneumoniae pathogenesis has been aided by the availability of genomic sequence information. Genomic and proteomic analyses of C. pneumoniae infections in relevant cell types will help to define the pathogenic potential of the organism in both respiratory and cardiovascular disease.

Enhanced release of nitric oxide in response to changes in flow and shear stress in the superior mesenteric arteries of portal hypertensive rats.

Increased nitric oxide (NO) release has been implicated in the pathogenesis of the hyperdynamic circulation in portal hypertension. NOS 3 (eNOS) causes NO release from the endothelium in response to physical stimuli, such as increased blood flow and shear stress. We evaluated the functional activity of the endothelium in the superior mesenteric arterial bed of portal hypertensive rats through direct measurement of NO metabolites (NOx) during changes in flow and shear stress. The in vitro perfusion system (McGregor) was used in sham and portal vein-ligated (PVL) rats. Shear stress was applied gradually to superior mesenteric arterial beds by increasing the perfusion rate. Flow studies were performed serially before and after incubation with either Krebs solution alone or with the NO-inhibitor, NG-monomethyl-L-arginine (L-NMMA). NOx concentrations in the perfusate were measured using chemiluminescence. The slope of NOx release versus flow-induced shear stress was calculated. Before L-NMMA administration, NOx concentrations and release of NO in PVL rats were significantly elevated in comparison with sham rats at flow rates of 32, 40, and 48 mL/min. The slope of NOx production versus shear stress index was significantly higher in PVL than in sham rats. After L-NMMA incubation, the decrease in slope was significantly larger in PVL rats. This study provides direct evidences of an increased NO synthesis by the superior mesenteric arterial vascular endothelium of PVL animals in response to shear stress. The increased NO output in response to shear stress suggests an adaptative mechanism developed by the vascular endothelial cells in response to a chronic increase in flow-mediated shear stress.

Cold ischemia–induced damage to vascular endothelium results in permeability alterations in transplanted lungs

Despite suggestions of a connection between endothelial damage and permeability alterations after ischemia and reperfusion in pulmonary tissue undergoing transplantation, no direct correlation between vascular endothelial discontinuity and parenchymal edema has yet been shown. Methods: Forty-two rat lungs were harvested and stored for 48 or 72 hours under hypothermic and ischemic conditions. Stored pulmonary tissue was studied before transplantation and 5 minutes or 24 hours after transplantation by light microscopy and scanning electron microscopy of arterial vascular endothelium. Results: Stored lungs not subjected to revascularization showed moderate perivascular edema, with small intercellular gaps in endothelial monolayers. Five minutes after transplantation, pulmonary tissue appeared congested, with perivascular and alveolar edema. Examination of vascular endothelium by scanning electron microscopy showed detachment of endothelial cells. Twenty-four hours after transplantation, edema, hemorrhage, and vascular congestion were found in all specimens. Arterial vascular endothelium showed weak intercellular connections, numerous intercellular gaps, and widespread cell detachment. Bronchial epithelial cells appeared damaged after storage, with loss of cilia, blebbing of apical cytoplasm, and cellular rounding. These changes were maintained 5 minutes after transplantation but appeared totally reversed after 24 hours in specimens stored 48 hours, whereas bronchial denudation was observed in 72-hour stored lungs. Statistically significant positive correlations (Kendall p < 0.001) between revascularization time and alveolar edema and hemorrhage were found for both storage periods. Conclusion: The results from this study demonstrate correlation between loss of endothelial monolayer continuity and histologic evidence of vascular permeability increases in pulmonary tissue before and after lung transplantation. (J T HORAC C ARDIOVASC S URG 1996;112:1027-35)