Leukocyte-endothelial Cell Adhesion Molecule Research Articles

MP07-13 THERAPEUTIC EFFECT OF RIPK1 INHIBITOR IN TESTICULAR ISCHEMIA-REPERFUSION

You have accessJournal of UrologyInfertility: Basic Research & Pathophysiology I1 Apr 2017MP07-13 THERAPEUTIC EFFECT OF RIPK1 INHIBITOR IN TESTICULAR ISCHEMIA-REPERFUSION Shin Ohira, Ryoei Hara, Shigenobu Tone, Seitetsu Kin, Shinjiro Shimizu, Tomohiro Fujii, Yoshiyuki Miyaji, and Atsushi Nagai Shin OhiraShin Ohira More articles by this author , Ryoei HaraRyoei Hara More articles by this author , Shigenobu ToneShigenobu Tone More articles by this author , Seitetsu KinSeitetsu Kin More articles by this author , Shinjiro ShimizuShinjiro Shimizu More articles by this author , Tomohiro FujiiTomohiro Fujii More articles by this author , Yoshiyuki MiyajiYoshiyuki Miyaji More articles by this author , and Atsushi NagaiAtsushi Nagai More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.279AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Testicular torsion makes testis ischemic. Surgery is immediately needed to reestablish blood flow. Even if the surgery succeed, testicular atrophy is often appear and lead to spermatic dysfunction. However, the etiology is still controversial in terms of pathophysiological changes. Recently it was reported that loss of the formation in any tissue after ischemia-reperfusion (IR) was involved in necroptosis, which is one of programmed cell death series. Necrostatin-1 (Nec-1) blocks both necroptosis and indoleamine 2,3-dioxygenase (IDO). In our previous research, we elucidated that IDO inhibitor decrease inflammation in testis and epididymis. First, we investigated pathophysiological change of testicular IR researching histological and biochemical phase in this study. Subsequently to the analysis, we tried to inhibit Receptor-interacting protein kinase 1 (RIPK1) and clarify a function of necroptosis in testicular IR. METHODS Twelve weeks old ICR male mice were used in this study. Their unilateral testicular artery was clamped under general anesthesia. Declamping three hours later, their testicular blood flow were resumed. After the procedure, bilateral testes were removed in time dependent manner (at day 1, 3, 5 and 7). Histological and biochemical change were evaluated by immunostaining and ELISA methods. Spermatic analysis from the epididymal cauda were evaluated by computer aided sperm analysis (CASA). In the following research, Nec-1 4 μg/g was administrated (iv) after declamping testicular blood flow. After the treatment, bilateral testes were removed in time dependent manner. Then, histological, biochemical and semen analysis were evaluated. Sham surgery was performed as control. Their experiments were duplicated at least. RESULTS Regarding histological change, invasion of lymphocyte-predominant inflammatory cells accumulated at day 3, 5 and destruction of seminiferous structure were observed at day 5, 7. Necroptosis cell using RIPK staining was abundantly expressed. In semen analysis, significant decreased spermatic concentration was observed at day 5 and 7 compared to control (p<0.05). Significant decreased spermatic motility was observed at day 1, 3, 5 and 7 compared to control (p<0.05). Interestingly, in contralateral (unaffected side) testes, significant decreased spermatic motility was observed at day 5 and 7 compared to control (p<0.05). Some candidates were picked up as molecular marker. Significant increased E-selectin expression, which is a marker of leukocyte-endothelial cell adhesion molecule, was observed at day 1 compared to control (p<0.05). Significant increased IL-6 expression, which is a marker of inflammation, was observed at day 3 compared to control (p<0.05). Significant increased 8-OHdG expression, which is a biomarker of oxidative stress, was observed at day 7 compared to control (p<0.05). Interestingly, significant increased the highest expression of E-selectin, IL-6 and 8-OHdG were observed in contralateral testes. Same results were introduced in immunohistochemical staining. After treatment of Nec-1, testicular structure were maintained and little invasion of inflammatory cells were observed. Significant decreased germ cell death (necroptosis) in seminiferous tubules were statistically observed. Significant decreased E-selectin, IL-6 and 8-OHdG were also observed. Importantly, Spermatic motility also maintained in both treated testis and contralateral testes. CONCLUSIONS Oxidative stress via inflammation and necroptosis should induce spermatogenetic dysfunction in IR testis. First, prominent inflammation was occurred in testicular ischemia-reperfusion model and not only expression of cytokines were increased, but also in contralateral testes. Subsequently, oxidative stress highly expressed and cell death appeared. Therefore, their change of bilateral testis would be one of pathophysiology in patients with ischemic testis. To inhibit RIPK1 would be contribute to protection of testicular tissue and spermatogenesis in IR model. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e86-e87 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Shin Ohira More articles by this author Ryoei Hara More articles by this author Shigenobu Tone More articles by this author Seitetsu Kin More articles by this author Shinjiro Shimizu More articles by this author Tomohiro Fujii More articles by this author Yoshiyuki Miyaji More articles by this author Atsushi Nagai More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Signal transduction mechanism of the NZW type IL‑10R1 and the effect on B lymphocyte function.

The present study aimed to investigate the signal transduction mechanism of the NZW type interleukin(IL)‑10R1 and the effect on the function of Blymphocytes. Vectors for the expression of wild‑type, NZW and G1146A IL‑10R1 were constructed and transfected into Ba/F3 cells. The cell proliferation curve was determined using an MTT assay. CD32 and leukocyte-endothelial cell adhesion molecule1 (LECAM‑1) expression levels were assessed using flow cytometry. The phosphorylation of Janus kinase 1 (Jak1), tyrosine kinase2 (Tyk2) and signal transducer and activator of transcription3 (Stat3) were measured by western blot analysis. As compared with wild‑type IL‑10R1 cells, the proliferation ability of NZW IL‑10R1 cells was increased following stimulation with IL‑10, which was even greater for G1146A cells. CD32 levels in NZW IL‑10R1 cells were lower, while the expression of LECAM‑1 was higher compared with wild‑type IL‑10R1 cells, which was also observed for G1146A cells. The phosphorylation levels of Jak1 and Stat3 in NZW IL‑10R1 and G1146A IL‑10R1 cells were significantly lower compared with the wild‑type IL‑10R1 cells. However, the phosphorylation levels in the three cell types were not significantly different. In conclusion, the deficiency in Jak1‑Stat3 signal transduction in NZW IL‑10R1 cells induces a loss in the inhibition ability of proliferation and activation as well as a migration tendency of Blymphocytes, which is hypothesized to be associated with the occurrence and development of the autoimmune disease systemic lupus erythematosus (SLE).

Abstract 184: Irradiation Induces Gene Expression Changes In Cultured Murine Brain Endothelial Cells

Brain arteriovenous malformations (AVMs) are abnormal connections between arteries and veins and are the leading cause of hemorrhagic stroke in children and young adults. Treatment of large and deep AVMs remains challenging, therefore new and safe treatment methods are required. Our hypothesis is that stereotactic radiosurgery can be used to selectively alter AVM endothelial cell phenotype, allowing targeted molecular therapies that spare normal vessels. This study examined the post-radiation time course of gene expression changes of potential molecular targets. A murine brain endothelial cell culture (bEnd.3) was treated with 25 Gy using a linear accelerator; non-irradiated cells were used as controls. Quantitative real-time PCR was used to measure the relative gene expression at 1, 6, 12, 24, 48, 72, 96, 120, 144, and 168 h after radiation. Genes encoding for intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) were found to be significantly up-regulated post irradiation (p&lt;0.0001), with the maximum level of gene expression for ICAM-1 being evident at 144 h (9-fold) and for VCAM-1 at 120 h (23-fold). Additionally, leukocyte-endothelial cell adhesion molecule 2 (E-selectin) and leukocyte-endothelial cell adhesion molecule 3 (P-selectin) showed significantly increased gene expression at 24 h with 1.5-fold (p&lt;0.001) and 2-fold (p&lt;0.0001) increases respectively. These data support the hypothesis that stereotactic radiosurgery can significantly alter endothelial cell phenotype in AVMs, potentially enabling the use of ligand-based molecular targeting therapies such as the stimulation of thrombosis for AVM treatment.

Targeting TNF-α to Neoangiogenic Vessels Enhances Lymphocyte Infiltration in Tumors and Increases the Therapeutic Potential of Immunotherapy

Abnormal tumor vasculature impairs T lymphocyte adhesion to endothelial cells and lymphocyte extravasation into neoplastic tissues, limiting the therapeutic potential of both active and adoptive immunotherapies. We have found that treatment of tumor-bearing mice with NGR-TNF, a Cys-Asn-Gly-Arg-Cys peptide-TNF fusion product capable of altering the endothelial barrier function and improving drug penetration in tumors, associated with the intratumor upregulation of leukocyte-endothelial cell adhesion molecules, the release of proinflammatory cytokines and chemokines, and the infiltration of tumor-specific effector CD8(+) T cells. As a result, NGR-TNF enhanced the therapeutic activity of adoptive and active immunotherapy, delaying tumor growth and prolonging survival. Furthermore, we have found that therapeutic effects of these combinations can be further increased by the addition of chemotherapy. Thus, these findings might be relevant for the design of novel immunotherapeutic approaches for cancer patients.

The complement system plays a critical role in the development of experimental autoimmune anterior uveitis.

The role of complement in ocular autoimmunity was explored in a experimental autoimmune anterior uveitis (EAAU) animal model. EAAU was induced in Lewis rats by immunization with bovine melanin-associated antigen. Complement activation in the eye was monitored by Western blot for iC3b. The importance of complement to the development of EAAU was studied by comparing the course of intraocular inflammation in normal Lewis rats (complement-sufficient) with cobra venom factor-treated rats (complement-depleted). Eyes were harvested from both complement-sufficient and complement-depleted rats for mRNA and protein analysis for IFN-gamma, IL-10, and interferon-inducible protein (IP)-10. Intracellular adhesion molecule (ICAM)-1 and leukocyte-endothelial cell adhesion molecule (LECAM)-1 were detected by immunofluorescent staining. OX-42 was used to investigate the importance of iC3b and CR3 interaction in EAAU. There was a correlation between ocular complement activation and disease progression in EAAU. The incidence, duration, and severity of disease were dramatically reduced after active immunization in complement-depleted rats. Complement depletion also completely suppressed adoptive transfer EAAU. The presence of complement was critical for local production of cytokines (IFN-gamma and IL-10), chemokines (IP-10), and adhesion molecules (ICAM-1 and LECAM-1) during EAAU. Furthermore, intraocular complement activation, specifically iC3b production and engagement of complement receptor 3 (CR3), had a significant impact on disease activity in EAAU. The study provided the novel finding that complement activation plays a central role in the pathogenesis of ocular autoimmunity and may serve as a potential target for therapeutic intervention.

Synthesis of α(1,3) Fucosyltransferases IV- and VII-Dependent Eosinophil Selectin Ligand and Recruitment to the Skin

Selectins mediate the initial adhesion of leukocytes to endothelial cells in many contexts of inflammation-dependent leukocyte recruitment. The glycans that contribute to P- and E-selectin counterreceptor activity arise through glycosylation reactions in which the terminal steps are catalyzed by alpha(1,3) fucosyltransferases (FTs). We examined how selectin ligand activities are controlled in eosinophils by characterizing FT expression profiles and regulatory mechanisms in eosinophils isolated from human blood. We found that FT-IV and FT-VII mRNAs were up-regulated by transforming growth factor-beta1, but the FT-IV transcript consistently predominated in eosinophils. To further define the physiological role of FT-IV and FT-VII in expression of eosinophil selectin ligand, we characterized models of dermal eosinophilia in FT-IV- and/or FT-VII-deficient mice in vivo. FT-IV deficiency yielded a significant decrease in eosinophil recruitment to the skin. Likewise, deficiency of FT-VII also yielded a decrease in eosinophil recruitment. Eosinophil recruitment that remained in the absence of FT-VII was further inhibited by blocking P- or E-selectin and was essentially absent in mice deficient in both enzymes. These observations indicate that FT-IV and FT-VII are both important contributors to selectin-dependent eosinophil recruitment to the skin and may represent therapeutic targets for treating diseases in which eosinophil recruitment contributes to pathophysiology.

Dextran sulfate as a leukocyte-endothelium adhesion molecule inhibitor of lung injury in pediatric open-heart surgery

In spite of the progress in operative techniques and pre- and postoperative management for congenital heart disease, lung injury induced by the extracorporeal circulation is still a serious insult in pediatric open-heart operations. To prevent this injury, we used a leukocyte-endothelial cell adhesion molecule blocking agent, dextran sulfate, in a clinical setting. Sixty mg/kg of dextran sulfate (DS) was intravenously infused to the patients just before cardiopulmonary bypass was started and 600 mg was added to the bypass circuit prime. Thirty patients (DS group, 14 patients with atrial septal defect, and 16 patients with ventricular septal defect) were compared with age and body-weight matched control patients (control group, 14 patients with atrial septal defect, 11 patients with ventricular septal defect). Postoperative respiratory index, white blood cell counts, complement C3 and plasma granulocyte elastase levels during and after the operation were measured. Respiratory index just after the termination of cardiopulmonary bypass was better preserved in the DS group than in the control group (0.50 +/- 0.08 versus 0.81 +/- 0.12, p < 0.01). The sum total amount of measured granulocyte elastase across whole study period was significantly lower in the DS group (p < 0.05). The data suggeste the possible effects of dextran sulfate in ameliorating post-perfusion lung damage by interfering with leukocyte-endothelial cell adhesion in pediatric open-heart operations. Future application to patients with more complex anomalies is anticipated.

Human immunodeficiency virus type 1 protease inhibitors block toll-like receptor 2 (TLR2)- and TLR4-Induced NF-kappaB activation.

Coinfections with opportunistic and pathogenic bacteria induce human immunodeficiency virus (HIV) replication through microbial antigen activation of NF-kappaB. Here, we assessed whether HIV type 1 protease inhibitors (PI) block microbial antigen activation of NF-kappaB. Human microvessel endothelial cells were transiently transfected with either endothelial cell-leukocyte adhesion molecule NF-kappaB luciferase or interleukin 6 (IL-6) promoter luciferase constructs by using FuGENE 6, and they were treated with PI (nelfinavir, ritonavir, or saquinavir) prior to stimulation with the Toll-like receptor 4 (TLR4) and TLR2 ligands, with lipopolysaccharide (LPS), soluble Mycobacterium tuberculosis factor, or Staphylococcus epidermidis phenol-soluble modulin, respectively, or with tumor necrosis factor alpha (TNF-alpha). Luciferase activity was measured by using a Promega luciferase kit. TNF-alpha release from the supernatant was measured by enzyme-linked immunosorbent assay. Cell death was assessed by lactate dehydrogenase assay. We observed that PI pretreatment blocked the TLR2- and TLR4- as well as the TNF-alpha-mediated NF-kappaB activation, in a dose-dependent manner. PI pretreatment also blocked the LPS-induced IL-6 promoter transactivation and TNF-alpha secretion. These data suggest that PI block HIV replication not only by inhibiting the HIV protease but also by blocking the TLR- and TNF-alpha-mediated NF-kappaB activation and proinflammatory cytokine production. These findings may help explain the immunomodulatory effects of PI, and they suggest an advantage for PI-containing drug regimens in the treatment of HIV-infected patients who are coinfected with opportunistic and pathogenic bacteria.

Roles of CD14 in LPS-induced liver injury and lethality in mice pretreated with Propionibacterium acnes

The mechanism of the liver damage and lethaliy in Propionibacterium acnes ( P. acnes)-LPS system remains obscure. To examine the role of CD14 in the system, M14M mice, in which CD14 was expressed heterotopically under the control of the metallothionein promoter were used. The production of soluble CD14 (sCD14) was increased by both P. acnes – priming and LPS challenge (1 μg per mouse) in both nontransgenic and M14M mice, although the plasma level was much higher in M14M nontransgenic than mice. The size of granulomas induced by an intraperitoneal administration of P. acnes in M14M mice 7 days after priming was smaller than that in nontransgenic mice. An LPS challenge induced apoptotic and necrotic changes in hepatocytes in nontransgenic mice but not in M14M mice. The challenge dose resulted in almost 90% lethality in nontransgenic mice but not in M14M mice 24 h after challenge. TNF-α, IFN-γ, IL-12, IL-18 and inducible nitric oxide synthase (iNOS) mRNA expressions produced by LPS challenge in M14M mice were low compared with those in nontransgenic mice. IL-18 mRNA expression was upregulated in P. acnes-primed nontransgenic mice but not in M14M mice. These results suggest that the high sCD14 concentration may account for less marked liver damage in M14M mice. Increase in the challenge dose of LPS (2 μg per mouse) resulted in increased lethality of M14M mice without liver damage. The levels of endothelial cell leukocyte adhesion molecule (ELAM)-1 mRNA expression in several organs in M14M mice 1–3 h after LPS challenge were, however, lower than those in nontransgenic mice. The high sCD14 concentration may stimulate endothelial cell activation, which may account for lethality without liver damage in M14M mice. Thus, CD14 is involved in both the priming and induction phases as well as lethality in P. acnes-LPS system.

Upregulation of P-selectin and intercellular adhesion molecule-1 after retinal ischemia-reperfusion injury.

Vascular endothelial cells and leukocytes express several inflammatory adhesion receptors, such as selectins and cell adhesion molecules, after retinal ischemia. They mediate the transmigration process of leukocytes. For further understanding of the role of leukocytes after retinal ischemia-reperfusion injury, the responses of the leukocyte-endothelial cell adhesion molecules P-selectin and intercellular adhesion molecule (ICAM)-1 during retinal ischemia and reperfusion were examined. Male pigmented rats were subjected to retinal ischemia by a 1-hour ligation of the optic nerve followed by reperfusion. Gene and protein expression of P-selectin and ICAM-1 were studied at 0.5, 1, 6, 12, and 24 hours after the onset of reperfusion with semiquantitative polymerase chain reaction and Western blot analysis. Immunohistochemical methods were used to detect specific lesions expressing ICAM-1. Significant upregulation of P-selectin and ICAM-1 mRNA (at 6, 12, and 24 hours of reperfusion) were observed, with the expression peaks of both P-selectin and ICAM-1 mRNA occurring at 12 hours after reperfusion. P-selectin protein gradually increased and reached a maximum 6 hours after reperfusion, whereas ICAM-1 protein increased until 24 hours after reperfusion. Immunostaining with ICAM-1 antibodies was positive in endothelial cells after ischemia-reperfusion injury. Retinal ischemia-reperfusion stimulates P-selectin and ICAM-1 expression. Endothelial ICAM-1 expression in retinal vessels was observed. Upregulation of P-selectin and ICAM-1, which contribute to leukocyte rolling and adhesion, were observed after retinal ischemia.

Leukocyte–endothelial cell adhesion molecule 1 (LECAM-1) polymorphism is associated with diabetic nephropathy in type 2 diabetes mellitus

Background/aims: Glomerular infiltration with monocytes/macrophages has been implicated in the pathogenesis of diabetic nephropathy. In this study, we evaluated the relationship between the genetic polymorphism in leukocyte–endothelial adhesion molecule-1 (LECAM-1) and diabetic nephropathy in patients with type 2 diabetes mellitus. Methods: We determined the frequency of the LECAM-1 P213S genotype in 102 diabetic patients with diabetic nephropathy, 90 diabetic patients with no evidence of diabetic nephropathy, and 200 healthy control individuals. Results: The frequency of the LECAM-1 213PP genotype and P allele in patients with diabetic nephropathy was significantly higher than that in patients without nephropathy (genotype 68% vs. 53%, χ 2=6.78, P=.034; allele 83% vs. 72%, χ 2=6.26, P=.012). The LECAM-1 P213 genotype was associated with a 1.86-fold increased risk for nephropathy independently of other risk factors. Conclusion: The data suggest that the LECAM-1 213PP genotype is a genetic risk factor for the development of nephropathy in type 2 diabetes mellitus.

HSP70 as Endogenous Stimulus of the Toll/Interleukin-1 Receptor Signal Pathway

Human heat-shock protein (HSP)70 activates innate immune cells and hence requires no additional adjuvants to render bound peptides immunogenic. Here we tested the assumption that endogenous HSP70 activates the Toll/IL-1 receptor signal pathway similar to HSP60 and pathogen-derived molecular patterns. We show that HSP70 induces interleukin-12 (IL-12) and endothelial cell-leukocyte adhesion molecule-1 (ELAM-1) promoters in macrophages and that this is controlled by MyD88 and TRAF6. Furthermore, HSP70 causes MyD88 relocalization and MyD88-deficient dendritic cells do not respond to HSP70 with proinflammatory cytokine production. Using the system of genetic complementation with Toll-like receptors (TLR) we found that TLR2 and TLR4 confer responsiveness to HSP70 in 293T fibroblasts. The expanding list of endogenous ligands able to activate the ancient Toll/IL-1 receptor signal pathway is in line with the "danger hypothesis" proposing that the innate immune system senses danger signals even if they originate from self.

Leukocyte endothelial cell interactions in pig to human organ xenograft rejection

In cases where antibody- and complement-mediated hyperacute rejection (HAR) of vascularized organ xenografts has been prevented, acute vascular rejection (AVR) and acute T cell-mediated rejection (ACR) cause graft destruction. Infiltration of leukocytes (innate and graft-primed T cells) into the graft execute the latter two rejection modalities. The leukocyte extravasation process, which is a prerequisite for graft infiltration, is governed by adhesion molecules, including the selectin, integrin and immunoglobulin protein families, and the chemokine protein family. The compatibility between porcine endothelial cell and human leukocyte adhesion molecules was investigated in dynamic adhesion and static transendothelial migration assays. The effect of human anti-pig antibodies on human leukocyte adhesion to, and transendothelial migration across, porcine endothelium was assessed under dynamic and static conditions, respectively. In contrast to previously published results, no difference in the ability of neutrophils to adhere to pig and human endothelium was observed. Furthermore, no evident quantitative or qualitative differences in the capacity of human and porcine endothelium to support transendothelial migration of human leukocytes (T, B and natural killer (NK) cells, monocytes, and neutrophils) could be detected. The presence of human anti-pig antibodies (Abs) modulated the migration of leukocytes across porcine endothelium, as well as neutrophil adhesion to porcine endothelium under conditions of flow. Antibodies specific for pig endothelial adhesion molecules can potentially be used as species (graft)-specific immunosuppressive reagents in order to prevent cellular organ xenograft rejection.

Soluble ICAM-1 and E-selectin levels correlate with clinical and biological aspects of severe ovarian hyperstimulation syndome

Objective: To assess the potential involvement of the soluble endothelial cell-leukocyte adhesion molecules E-selectin and intercellular adhesion molecule-1 (ICAM-1) in the pathophysiology of capillary hyperpermeability in the ovarian hyperstimulation syndrome (OHSS). Design: Controlled clinical study. Setting: Women hospitalized for severe OHSS after ovulation induction for IVF at two tertiary medical centers. Patient(s): Eleven patients with severe OHSS undergoing IVF and 20 controls who had received a similar ovulation induction regimen and did not develop OHSS. Intervention(s): Serial serum samples were obtained from all patients with OHSS from admission until discharge. Ascitic fluid was obtained from all patients by therapeutic paracentesis. Serum was obtained from all controls 4–8 days after ET. Main Outcome Measure(s): Samples were assayed for soluble E-selectin and soluble ICAM-1 by using enzyme-linked immunosorbent assay and results were correlated with clinical and biological aspects of OHSS. Result(s): Compared with controls, patients with severe OHSS had higher levels of soluble ICAM-1 and lower levels of soluble E-selectin detected in serum and ascites. Serum levels of soluble ICAM-1 decreased, while soluble E-selectin levels increased along with clinical and biological improvement. Serum soluble ICAM-1 showed significant positive correlation and serum soluble E-selectin showed significant negative correlation with clinical and biological aspects of severe OHSS. Conclusion(s): Soluble ICAM-1 and soluble E-selectin seem to be involved in the pathophysiology of capillary hyperpermeability in severe OHSS.

Nuclear factor-κB activation is higher in peripheral blood mononuclear cells of male smokers

Nuclear factor-κB (NF-κB) is an oxidative stress sensitive transcription factor involved in the regulation of inflammatory genes. Activation of NF-κB results into increased expression of inflammatory genes such as for interleukins, endothelial cell leukocyte adhesion molecules and inducible NO synthase. Our study, with six smokers and 10 non-smokers, showed that NF-κB activity in peripheral blood mononuclear cells of smokers compared to non-smokers is significantly higher ( P<0.05). This indicates that NF-κB is involved in cigarette smoke induced inflammatory responses and that NF-κB activation can be used as a functional marker of oxidative stress.

Increased plasma level of soluble E-selectin in acute myocardial infarction

Background E-selectin, also known as endothelial cell leukocyte adhesion molecule-1, is a member of the selectin family of adhesion molecules and is expressed on vascular endothelial cells in inflammatory reactions. The induction of surface E-selectin expression by endothelial cells is considered a marker of activation. Methods and Results We examined the plasma soluble E-selectin (sE-selectin) level in 41 patients within 6 hours after the onset of acute myocardial infarction (AMI) and in 37 patients with stable exertional angina and 27 control patients. Blood samples were obtained on admission, after reperfusion therapy, and at 4 hours, 8 hours, 12 hours, 24 hours, 48 hours, 3 days, 5 days, 1 week, and 2 weeks after admission in the AMI group. In this group, 21 patients had a history of prodromal unstable angina before infarction and 20 had sudden onset of infarction. The plasma sE-selectin level (ng/mL) on admission was higher in the AMI group than in the stable exertional angina group and control group (38.5 ± 3.1 vs 28.5 ± 1.5, P <.01, 26.0 ± 1.8, P <.01, respectively). In addition, plasma sE-selectin levels were higher in the patients with AMI with prodromal unstable angina than in those with a sudden onset of infarction on admission (44.7 ± 5.4 vs 32.0 ± 2.1, P <.05). The plasma sE-selectin level decreased slowly during the chronic phase both in patients with AMI with prodromal unstable angina (from 44.7 ± 5.4 to 33.8 ± 3.4, P <.01) and those with a sudden onset of infarction (from 32.0 ± 2.1 to 24.9 ± 2.4, P <.01). Conclusions These results suggest that an increase of sE-selectin may reflect enhanced endothelial cell activation in patients with AMI. The higher sE-selectin level in patients with AMI with prodromal unstable angina may have been associated with repeated episodes of myocardial ischemia and reperfusion. (Am Heart J 2000;140:243-8.)

Proinflammatory cytokines increase in sepsis after anti-adhesion molecule therapy.

Cytokine mediators and leukocyte-endothelial cell adhesion molecules are critical and interdependent components of the acute inflammatory response in sepsis. We hypothesized that the administration of monoclonal antibodies to intercellular adhesion molecule-1 (CD54) or E- and L-selectin (CD62E/L) would decrease serum levels of the proinflammatory cytokines interleukin-1beta (IL-1), IL-6, and IL-8 and tumor necrosis factor receptor (TNFR-1) in baboons during sepsis. Adult male baboons received infusions of 1 x 10(9) colony forming units (CFU)/kg heat-killed Escherichia coli (E. coli) followed 12 h later by live E. coli (1 x 10(10) CFU/kg). At the time of live bacterial infusion, six septic animals were treated with a monoclonal antibody to CD54 and six with an antibody to CD62E and L (1 mg/kg). Eight untreated septic animals served as controls. Sequentially drawn serum samples were assayed for IL-1, IL-6, IL-8, and TNFR-1 using enzyme-linked immunoassay (ELISA). Data were compared using Mann-Whitney U tests and Chi-square analyses. Median survival was decreased in both treatment groups compared to controls (P < 0.05). Peak IL-1 level was higher than controls in septic animals treated with anti-CD54 but not anti-CD62E/L (P < 0.05, P = NS, respectively). Elevations in IL-6, IL-8, and TNFR-1 were increased and prolonged in both antibody treated groups compared to controls (P < 0.05). These results provide the first in vivo evidence that leukocyte-endothelial adhesion molecules CD54 and CD62E/L regulate cytokine production in sepsis.

Expression of alpha-1 proteinase inhibitor in human islet microvascular endothelial cells.

There is a microcirculation system within the islets of Langerhans. However, little is known about the phenotypic and functional characterization of islet microvascular endothelial cells (MVEC). In this study, we purified MVEC from human pancreatic islets by using Ulex europaeus (Sigma, St. Louis, MO) agglutinin-1 (UEA-1)-coated dynabeads (Dynal A.S., Oslo, Norway). These purified human islet MVEC (HI-MVEC) express von Willebrand factor, take up high levels of acetylated LDL, and upregulate endothelial cell leukocyte adhesion molecule 1 in response to tumor necrosis factor-alpha. Ultrastructure examination shows the presence of microvilli and fenestrations on the cell surface, Weibel-Palade bodies in the cytoplasm, and tight junctions between cells. Furthermore, we show that vascular endothelial cell growth factor contributes to the formation of surface fenestrations on cultured HI-MVEC. After purification, HI-MVEC exhibit a very low proliferation capacity and are strongly resistant to trypsin, compared with other original MVEC. We also demonstrate that alpha-1 proteinase inhibitor (Api) is expressed on HI-MVEC and specifically located at the area of cell-cell junctions. By reverse transcription-polymerase chain reaction, a significant messenger RNA band of Api was found only in HI-MVEC, but not in other organ-derived MVEC, indicating that expression of Api is islet MVEC specific. Antibodies to Api significantly reversed the resistance to trypsin and promoted proliferation of HI-MVEC, suggesting that these specific functional characteristics of HI-MVEC are related to the expression of Api. These results indicate that HI-MVEC exhibit some specific morphological and functional characteristics that differ from MVEC derived from other organs.

Inflammation and lipid-lowering treatment.

Leukocytes play a critical role in both the initial steps of atherogenesis and the late events of plaque instability and thrombosis. Leukocyte-endothelial cell adhesion molecules (CAMs) modulate the interaction between circulating leukocytes and the vessel wall. Levels of soluble CAMs are increased in patients with dyslipidemia or other risk factors, and levels can be reduced by lipid-modifying therapy. Novel markers of inflammation and atherosclerosis may be useful to identify high-risk individuals and to assess the efficacy of antiatherosclerotic therapy.

Recent progress of the research on novel microbial metabolites

Abstract