ENDOTHELIUM

Endothelium-dependent responses are impaired in various models of hypertension. The effects of antihypertensive treatment on endothelium-dependent relaxations were studied in Dahl salt-sensitive (DS) and Dahl salt-resistant rats (DR) on a high or low sodium diet. The rats were given either a diet containing 8% NaCI or 0.1% NaCI for 8 weeks or a diet containing 8% NaCI and a combination of reserpine, hydrochlorothiazide, and hydralazine for 8 or 2 weeks. DS on the 8% NaCI diet developed hypertension, while the other rats did not. Antihypertensive therapy for 8 or 2 weeks prevented or reversed hypertension in DS and lowered blood pressure in DR on the 8% NaCI diet. Aortic rings with and without endothelium were suspended in organ chambers for isometric tension recording. In all groups, acetylcholine, adenosine 5'-diphosphate, and thrombin caused endotheliumdependent relaxations. The relaxations in response to all agonists were significantly decreased in DS on 8% NaCI compared to relaxations in the other rats. Antihypertensive treatment for 8 or 2 weeks prevented or reversed the decreased endothelium-dependent relaxations in response to all agonists tested, but not those to the endothelium-independent agonist, sodium nitroprusside. These results suggest that antihypertensive treatment normalizes endothelium-dependent relaxations. This effect of antihypertensive treatment might be important for the prevention of cardiovascular complications in patients with hypertension. (Hypertension 9 [Suppl III]: IIM93- III-197, 1987) KEY WORDS * acetylcholine • adenosine 5'-diphosphate • Dahl rats • norepinephrine sodium nitroprusside • thrombin • thoracic aorta From the Department of Physiology and Biophysics, Mayo Clinic and Foundation, Rochester, and the Renal Section, Veterans

A novel compact illumination device in variable-angle total internal reflection fluorescence microscopy (VA-TIRFM) is described. This device replaces the standard condensor of an upright microscope. Light from different laser sources is delivered via a monomode fibre and focused onto identical parts of a sample under variable angles of total internal reflection. Thus, fluorophores in close proximity to a cell-substrate interface are excited by an evanescent wave with variable penetration depth, and localized with high (nanometre) axial resolution. In addition to quantitative measurements in solution, fluorescence markers of the cytoplasm and the plasma membrane, i.e. calcein and laurdan, were examined using cultivated endothelial cells. Distances between the glass substrate and the plasma membrane were determined using the mathematical algorithm of a four-layer model, as well as a Gaussian-shaped intensity profile of the illumination spot on the samples. Distances between 0 and 30 nm in focal contacts and between 100 and 300 nm in other parts of the cell were thus determined. In addition to measurements of cell-substrate topology, the illumination device appears appropriate for numerous applications in which high axial resolution is required, e.g. experiments on endocytosis or exocytosis, as well as measurements of ion concentrations proximal to the plasma membrane. The compact illumination device is also suitable for combining TIRFM with further innovative techniques, e.g. time-resolved fluorescence spectroscopy, fluorescence lifetime imaging (FLIM) or fluorescence resonance energy transfer (FRET).

The effects of rabbit exposure to ozone (O 3) (0.4 ppm for 4 h) on two different cytochrome P-450 (CYP450)-dependent activities were investigated. In turn, the role of CYP450 in the inhibitory effect of O 3 on acetylcholine (ACh)-evoked increase in endothelial permeability was also assessed. Immediately after the period of exposure, rabbits of both sexes were sacrificed and their lungs were extracted. Some lungs were used for preparation of microsomes and measurement of 7-ethoxyresorufin O-deethylase (EROD) and parathion oxidase activities. Other rabbit lungs were isolated and recirculatingly blood-free perfused. Arterial, venous pressures, and lung weight were continuously recorded. Capillary pressure was measured by applying the double occlusion method. Capillary filtration coefficient (K f,c) was evaluated by measuring the amount of fluid filtering through the endothelium when vascular pressures were suddenly increased. Dose-response curves to ACh were constructed in air-or O 3-exposed rabbits. Some animals were pretreated with piperonyl butoxide (PBO), a well-known inhibitor of CYP450. O 3 significantly reduced both EROD and parathion oxidase lung microsomal 999

Chronic treatment with several antihypertensive agents, including calcium channel blockers, may interfere with remodeling of large arteries and increased arterial stiffness. We hypothesize that even a short, sevenday administration of calcium channel blockers might alter an aortic remodeling in spontaneously hypertensive rat (SHR). Male SHR and normotensive WKY rats (n = 14 each) were treated by either vehicle, vasculoselective calcium channel blocker nifedipine (1 mg/kg/day) or cardiac/vascular calcium channel blockers diltiazem (5 mg/kg/day) or verapamil (4 mg/kg/day, n = 6 for each treatment) subcutaneously twice daily for seven days. Additional SHR rats were randomized for termination 24, 72 or 120 h (n = 5 each) after the withdrawal of nifedipine. Systolic blood pressure was measured by tail cuff and thoracic aorta was collected for histomorphometric and functional analysis including acetylcholine-induced endotheliumdependent relaxation. Seven-day administration of diltiazem and nifedipine, but not verapamil decreased blood pressure in SHR. All drugs significantly attenuated abnormal aortic wall thickness, cross-sectional area and media-to-lumen ratio, but only nifedipine improved impaired endothelium-dependent relaxation. Following the withdrawal of nifedipine, all measured parameters returned back to control SHR values within 72 h. Seven-day treatment with distinct calcium channel blockers attenuates hypertensive remodeling of aorta, which might be, in case of nifedipine, reactivated even by a very short withdrawal of the drug. Therefore, vasculoprotection by calcium channel blockers is not restricted to a prolonged blood pressure modulation, but occurs rapidly. These findings could be relevant for an intervention in augmented vascular stiffness and related cardiovascular risk.

Background: Increased oxidative stress is associated with all cardiovascular risk factors and reactive oxygen species appear to be the principal mediators of cardiomyocite dysfunction in various carviovascular diseases. Carnitine has been shown to be effective in pathologic conditions characterized by increased oxidative stress and an antioxidant effect of l-carnitine and its derivatives has been described but the specific mechanism is unclear. Methods: We evaluated in human endothelial cells in culture the effect of l-carnitine (C), acetyl-l-carnitine (AC) and propionyl-l-carnitine (PC) on gene and protein expression (RT-PCR and Western blot) of oxidative stress related proteins heme oxygenase-1 (HO-1) and of endothelial NO synthase (ecNOS) in absence and presence of oxidative stress induced by H 2 O 2 . Results: HO-1 as well as ecNOS gene and protein expression significantly increased upon Carnitines incubation. Induction of oxidative stress increased HO-1 gene expression compared to basal condition (0.62 T 0.02 densitometric units vs. 0.48 T 0.05, p < 0.01) while decreased ecNOS gene expression (0.75 T 0.04 vs. 0.40 T 0.08, p < 0.001). These results were paralleled by similar results at protein level. Coincubation of C (0.5 -1.0 -2.0 mM), AC (0.1 -0.2 -0.4 mM) and PC (0.05 -0.1 -0.2 mM) with H 2 O 2 further increased HO-1 gene expression and not only normalized vs. H 2 O 2 but even increased vs. basal ecNOS mRNA. HO-1 and ecNOS gene expression was also paralleled at protein level by coincubation with C, AC and PC of cells exposed to oxidative stress. Conclusion: This is the first report that has utilized a molecular biological approach to demonstrate a direct stimulatory effect of Carnitines on gene and protein expression of the oxidative stress related markers HO-1 and ecNOS. As HO-1 and NO are known as antioxidant, antiproliferative and anti-inflammatory, their increased expression would be expected to protect from oxidative stress related cardiovascular risk factors and myocardial damage, therefore adding this effect to the multiple pathways involved in the effects of carnitines. D

Neutrophil collagenase (matrix metalloproteinase-8 or MMP-8) is regarded as being synthesized exclusively by polymorphonuclear neutrophils (PMN). However, in vivo MMP-8 expression was observed in mononuclear fibroblast-like cells in the rheumatoid synovial membrane. In addition, we detected MMP-8 mRNA expression in cultured rheumatoid synovial fibroblasts and human endothelial cells. Up-regulation of MMP-8 was observed after treatment of the cells with either tumor necrosis factor-alpha (10 ng/ml) or phorbol 12-myristate 13-acetate (10 nM). Western analysis showed a similar regulation at the protein level. The size of secreted MMP-8 was 50 kDa, which is about 30 kDa smaller than MMP-8 from PMN. Conditioned media from rheumatoid synovial fibroblasts contained both type I and II collagen degrading activity. However, degradation of type II collagen, but not that of type I collagen, was completely inhibited by 50 microM doxycycline, suggesting specific MMP-8 activity. In addition, doxycycline down-regulated MMP-8 induction, at both the mRNA and protein levels. Thus MMP-8 exerts markedly wider expression in human cells than had been thought previously, implying that PMN are not the only source of cartilage degrading activity at arthritic sites. The inhibition of both MMP-8 activity and synthesis by doxycycline provides an incentive for further studies on the clinical effects of doxycycline in the treatment of rheumatoid arthritis.

With the aim of contributing to a better understanding of the haemolytic function of the spleen, a morphologic and morphometric study of this organ fixed by arterial perfusion was performed in nine patients with hereditary spherocytosis (HS). three with autoimmune haemolytic anaemia (AHA) and six with Hodgkin's disease without splenic involvement (controls). The spleen weight in HS and AHA (6 2 1 1 4 2 9 g. mean 1 SD) was significantly increased with respect to controls (I 68 f 36 g) (P = 0.003 1. In HS the red cell retention in the cords of Billroth was significantly increased (203 1 6 8 per I O4 pmL) with respect to the cases with AHA (9 3 1 3 5 per 10* pmZ) and to the controls (5 7 1 2 8 per lo4 pmL) (P=0,004). In HS and AHA the number of macrophages per The role of the spleen in haemolytic anaemia has been extensively studied (

The endothelium is a highly dynamic cell layer that is involved in a multitude of physiological functions, including the control of vascular tone, the movement of cells and nutrients, the maintenance of blood fluidity and the growth of new vessels. During severe sepsis, the endothelium becomes proadhesive, procoagulant, antifibrinolytic and is characterized by alterations of vasomotor regulation. Most of these functions have been discovered using in vitro and animal models, but in vivo exploration of endothelium in patients remains difficult. New tools to analyze endothelial dysfunction at bedside have to be developed.

Background: Vitamin D is a pro-hormone that plays an essential role in the vasculature and in kidney function. Aims: To review the extra-skeletal effects of vitamin D on blood pressure, endothelial and renal function with emphasis on recent findings in postmenopausal women. Methods: Included in this review was a PubMed database search for English language articles through March 2013. This review discussed the physiology and definition of vitamin D deficiency, the recent evidence for the role vitamin D in blood pressure, vascular and renal function. Results: Experimental and epidemiological data suggest that vitamin D plays an important role in the vasculature and in kidney function. Low vitamin D concentrations appear to significantly associate with hypertension, endothelial and renal dysfunction. However, the results of clinical trials have generally been mixed. Studies specifically conducted among postmenopausal women are limited and findings are still inconsistent. Conclusions: Definitive studies are warranted to elucidate the effects of vitamin D supplementation on vascular and renal function and a more detailed work is needed to outline the route, duration and optimal dose of supplementation. It is premature to recommend vitamin D as a therapeutic option in the improvement of vascular and renal function at the current stage. OPEN ACCESS Nutrients 2013, 5 2591

Oxidized LDL is a key factor in the pathogenesis of atherosclerosis and its thrombotic complications such as stroke and myocardial infarction. 1 It activates endothelial cells and platelets through poorly defined mechanisms. Whereas heavily oxidized LDL has often toxic effects on cells, mildly oxidized LDL (mox-LDL) and minimally modified LDL (mm-LDL) alter the function of cells through mechanisms that involve the stimulation of receptor-mediated signal transduction pathways. 2 The present study summarizes our recent results concerning the identification of the components in mildly oxidized LDL that induce platelet and endothelial cell activation and describes the elucidation of the underlying signaling pathways.

Beyond the disorders recognized as mitochondrial diseases, abnormalities in function and/or ultrastructure of mitochondria have been reported in several unrelated pathologies. These encompass ageing, malformations, and a number of genetic or acquired diseases, as diabetes and cardiologic, haematologic, organ-specific (e.g., eye or liver), neurologic and psychiatric, autoimmune, and dermatologic disorders. The mechanistic grounds for mitochondrial dysfunction (MDF) along with the occurrence of oxidative stress (OS) have been investigated within the pathogenesis of individual disorders or in groups of interrelated disorders. We attempt to review broad-ranging pathologies that involve mitochondrial-specific deficiencies or rely on cytosol-derived prooxidant states or on autoimmune-induced mitochondrial damage. The established knowledge in these subjects warrants studies aimed at elucidating several open questions that are highlighted in the present review. The relevance of OS and MDF i...

Beyond the disorders recognized as mitochondrial diseases, abnormalities in function and/or ultrastructure of mitochondria have been reported in several unrelated pathologies. These encompass ageing, malformations, and a number of genetic or acquired diseases, as diabetes and cardiologic, haematologic, organ-specific (e.g., eye or liver), neurologic and psychiatric, autoimmune, and dermatologic disorders. The mechanistic grounds for mitochondrial dysfunction (MDF) along with the occurrence of oxidative stress (OS) have been investigated within the pathogenesis of individual disorders or in groups of interrelated disorders. We attempt to review broad-ranging pathologies that involve mitochondrial-specific deficiencies or rely on cytosol-derived prooxidant states or on autoimmune-induced mitochondrial damage. The established knowledge in these subjects warrants studies aimed at elucidating several open questions that are highlighted in the present review. The relevance of OS and MDF in different pathologies may establish the grounds for chemoprevention trials aimed at compensating OS/MDF by means of antioxidants and mitochondrial nutrients.

Polyphenols from Hibiscus sabdariffa calices were administered to patients with metabolic syndrome (125 mg/kg/day for 4 wk, n = 31) and spontaneously hypertensive rats (125 or 60 mg/kg in a single dose or daily for 1 wk, n = 8 for each experimental group). The H. sabdariffa extract improved metabolism, displayed potent anti-inflammatory and antioxidant activities, and significantly reduced blood pressure in both humans and rats. Diuresis and inhibition of the angiotensin I-converting enzyme were found to be less important mechanisms than those related to the antioxidant, anti-inflammatory, and endothelium-dependent effects to explain the beneficial actions. Notably, polyphenols induced a favorable endothelial response that should be considered in the management of metabolic cardiovascular risks.

L a symptomatologie de la drépanocytose, maladie génétique la plus fréquente des populations d'origine africaine, mais aussi présente dans le bassin méditerranéen, la péninsule arabique et le sous-continent indien, est dominée par une anémie hémolytique chronique, une susceptibilité accrue aux infections et la survenue de crises douloureuses, témoins d'une occlusion des petits vaisseaux. Malgré ces complications très invalidantes, l'amélioration de la prise en charge, avec notamment le dépistage néonatal, permet à la plupart des patients d'atteindre l'âge adulte et de dépasser maintenant la 5 e décennie. La physiopathologie de cette maladie et singulièrement les événements responsables de la vaso-occlusion sont encore imparfaitement connus ; ils ne peuvent se résumer à la simple polymérisation de l'hémoglobine S (HbS) en conditions hypoxiques, aboutissant à la falciformation du globule rouge (GR) drépanocytaire, puis à l'obstruction des microveinules postcapillaires par ces GR rigidifiés et moins déformables.

This study was designed to examine the effect of benzalkonium chloride/ethylenediaminetetraacetic acid (BAK/EDTA) on the ocular bioavailability (Focular) of ketorolac tromethamine after ocular instillation to normal and de-epithelialized corneas of rabbits both in vitro and in vivo. The in vivo Focular of the formulations was measured in flow-through perfusion chambers. For in vitro studies, a 35 µL dose of 0.5% ketorolac tromethamine with or without BAK/EDTA was instilled into rabbit eyes with intact or de-epithelialized corneas. At 0.5, 1, 2, 4, 6, and 8 h postdose, rabbits were euthanized, and the corneas and aqueous humor were collected from both eyes. The ketorolac concentrations from both in vitro and in vitro samples were quantified by reversed-phase high-performance liquid chromatography. The in vitro study results indicated that BAK/EDTA statistically significantly increased the Focular of ketorolac through de-epithelialized corneas but not through intact corneas. The in vitro study results showed that BAK/EDTA had no effect on the Focular of ketorolac in rabbits with intact corneas, based on the values of the area under the aqueous humor concentration versus time curves (AUC0−6h) of ketorolac. As expected, de-epithelialization of the corneas produced a faster and greater ocular absorption of ketorolac as evidenced by the smaller Tmax and larger AUC values compared to those for the intact corneas in vitro. However, BAK/EDTA decreased the ocular absorption of ketorolac in rabbits with de-epithelialized corneas. The half-lives (t1/2) of ketorolac in corneal tissue and aqueous humor were longer in rabbits with intact corneas than those in rabbits with de-epithelialized corneas. In conclusion, the in vitro Focular of ketorolac was not altered by BAK/EDTA in rabbits with intact corneas, but it was decreased by BAK/EDTA in rabbits with de-epithelialized corneas. Therefore, the formulation with ketorolac alone may be better as a postoperative ocular analgesic.

Although the endothelium is an extremely thin
single-cell layer, it performs exceedingly well in
preventing blood fluids from leaking into the surrounding
tissues. However, specific pathological conditions
can affect this cell layer, compromising the integrity of
the barrier. Vascular leakage is a hallmark of many
cardiovascular diseases and despite its medical importance,
no specialized therapies are available to prevent it
or reduce it. Small guanosine triphosphatases (GTPases)
of the Rho family are known to be key regulators of
various aspects of cell behavior and studies have shown
that they can exert both positive and negative effects on
endothelial barrier integrity. Moreover, extracellular matrix
stiffness has now been implicated in the regulation
of Rho-GTPase signaling, which has a direct impact on
the integrity of endothelial junctions. However, knowledge
about both the precise mechanism of this regulation
and the individual contribution of the specific regulatory
proteins remains fragmentary. In this review, we
discuss recent findings concerning the balanced activities
of Rho-GTPases and, in particular, aspects of the
regulation of the endothelial barrier. We highlight the
role of Rho-GTPases in the intimate relationships between
biomechanical forces, microenvironmental influences
and endothelial intercellular junctions, which are
all interwoven in a beautiful filigree-like fashion.

Adenoviral (Ad) vectors are promising gene therapy vehicles due to their in vivo stability and efficiency, but their potential utility is compromised by their restricted tropism. Targeting strategies have been devised to improve the efficacy of these agents, but specific targeting following in vivo systemic administration of vector has not previously been demonstrated. The distinct aim of the current study was to determine whether an Ad-targeting strategy could maintain fidelity upon systemic vascular administration. We used a bispecific antibody to target Ad infection specifically to angiotensin-converting enzyme (ACE), which is preferentially expressed on pulmonary capillary endothelium and which may thus enable gene therapy for pulmonary vascular disease.

High density lipoprotein (HDL) activates endothelial nitric oxide synthase (eNOS), leading to increased production of the antiatherogenic molecule NO. A variety of stimuli regulate eNOS activity through signaling pathways involving Akt kinase and/or MAP kinase. In the present study we investigated the role of kinase cascades in HDL-induced eNOS stimulation in cultured endothelial cells and COS M6 cells transfected with eNOS and the HDL receptor, scavenger receptor B-I (SR-BI). HDL (10-50 µg/ml, 20 min) caused eNOS phosphorylation at Ser-1179, and dominant negative Akt inhibited both HDL-mediated phosphorylation and activation of the enzyme. PI3 kinase inhibition or dominant negative PI3 kinase also blocked the phosphorylation and activation of eNOS by HDL. Studies with genistein and PP2 showed that the non-receptor tyrosine kinase, src, is an upstream stimulator of the PI3 kinase-Akt pathway in this paradigm. In addition, HDL activated MAP kinase through PI3 kinase, and MEK inhibition fully attenuated eNOS stimulation by HDL without affecting Akt or eNOS Ser-1179 phosphorylation. Conversely, dominant negative Akt did not alter HDL-induced MAP kinase activation. These results indicate that HDL stimulates eNOS through common upstream, src-mediated signaling which leads to parallel activation of Akt and MAP kinases and their resultant independent modulation of the enzyme. by guest on June 4, 2016 http://www.jbc.org/ Downloaded from Recently Li and colleagues also reported that HDL binding to SR-BI activates eNOS (5). The HDL-induced increase in NO production may be critical to the atheroprotective features of HDL since diminished bioavailablity of endotheliumderived NO has a key role in the early pathogenesis of hypercholesterolemiainduced vascular disease and atherosclerosis (6-8). However, the mechanisms by which HDL activates eNOS are yet to be clarified. eNOS is one of three isoenzymes which convert L-arginine to L-citrulline plus NO. The activity of eNOS is regulated by complex signal transduction pathways that involve various phosphorylation events and protein-protein interactions. Many stimuli modulate eNOS activity by activating kinases which alter the phosphorylation of the enzyme (9-15). Akt kinase (also known as PKB) by guest on June 4, 2016 http://www.jbc.org/ Downloaded from activates eNOS by directly phosphorylating the enzyme at serine-1179 (Ser-1179) (19). Akt itself is phosphorylated and activated by phosphoinositide 3kinase (PI3 kinase), which in turn is activated by a tyrosine kinase (TK). Both receptor TK and non-receptor TK are involved in PI3 kinase-Akt mediated eNOS activation by various agonists . In contrast to Ser-1179, phosphorylation of threonine-497 (Thr-497) of eNOS attenuates enzyme activity . eNOS is also modulated by MAP kinases (23,24) and unlike Akt, the effect of MAP kinases on eNOS activity can be either positive or negative (9,25-27). The role of kinase cascades in signaling by HDL from SR-BI to eNOS is entirely unknown.

We have demonstrated recently that peroxisome proliferatoractivated receptor (PPAR)␣ deficiency in the host leads to overt inflammation that suppresses angiogenesis via excess production of thrombospondin (TSP)-1 and prevents tumor growth. Hence, we speculated that pharmacologic activation of PPAR␣ would promote tumor growth. Surprisingly, the PPAR␣ agonist fenofibrate potently suppressed primary tumor growth in mice. This effect was not mediated by cancer-cell-autonomous antiproliferative mechanisms but by the inhibition of angiogenesis and inflammation in the host tissue. Although PPAR␣-deficient tumors were still susceptible to fenofibrate, absence of PPAR␣ in the host animal abrogated the potent antitumor effect of fenofibrate. In addition, fenofibrate suppressed endothelial cell proliferation and VEGF production, increased TSP-1 and endostatin, and inhibited corneal neovascularization. Thus, both genetic abrogation of PPAR␣ as well as its activation by ligands cause tumor suppression via overlapping antiangiogenic pathways. These findings reveal the potential utility of the well tolerated PPAR␣ agonists beyond their use as lipid-lowering drugs in anticancer therapy. Our results provide a mechanistic rationale for evaluating the clinical benefits of PPAR␣ agonists in cancer treatment, alone and in combination with other therapies. stroma ͉ inflammation ͉ fibrates ͉ microenvironment P eroxisome proliferator-activated receptors (PPARs) are a ‡ Present address

The female gender reduces the risk, but succumbs more to cardiovascular disease. The hypothesis that shortterm
(8 weeks) Streptozotocin-induced diabetes could produce greater female thanmale vascular tissue reactivity
and themechanistic basiswere explored. Aortic ring responses to Phenylephrine were examined in age- and
sex-matched normoglycaemic/diabetic rats. The normoglycaemic male tissue contracted significantly more than
the normoglycaemic female and the male/female diabetic tissues. Endothelial-denudation, L-NAME or MB
reversed these differences suggesting an EDNO-cGMP dependence. 17β-oestradiol exerted relaxant effect
on all endothelium-denuded (and normoglycaemic endothelium-intact male) tissues, but not endotheliumintact
normoglycaemic female. The greater male tissue contraction is attributable to absent 17β-oestradiolmodulated
relaxation. Indomethacin blockade of COX attenuatedmale normoglycaemic and female diabetic
tissue contraction (both reversed by L-NAME), but augmented diabetic male tissue contraction. These data
are consistent with the raised contractile TXA2 and PGE2 in normoglycaemic male and diabetic female tissues,
and the relaxant PGI2 in diabetic male (and female). The higher levels of PGI2 in the normoglycaemic
and diabetic female perhaps explain their greater relaxant response to Acetylcholine compared to the respective
male. In conclusion, there is an endothelium-dependent gender difference in the effect of short
term diabetes on vascular tissue reactivity which is COX mediated.

Inflammation is a fundamental response to tissue injury and invasion of pathogens, but it is detrimental in clinically important inflammatory disorders. Leukocytes are key players in the inflammatory response because of their antimicrobial, secretory and phagocytic activities. They are recruited to the inflamed tissue by sequential adhesive interactions between leukocytes and the endothelium that are mediated by cell-adhesion molecules (CAMs) on the surface of the interacting cells. The effects of many anti-inflammatory drugs can be ascribed, in part, to inhibition of the expression of CAMs. However, in the search for more selective and potent drugs for clinically important diseases such as multiple sclerosis, asthma, rheumatoid arthritis, inflammatory bowel disease, allergies and atherosclerosis, direct inhibition of the function of CAMs has attracted increasing interest. In recent years, the development of synthetic antagonists has provided better opportunities for drug targeting. Future advances in this field hold new prospects for therapeutic intervention in human inflammatory disorders.

Since the time a precise role of coenzyme Q 10 (CoQ 10 ) in myocardial bioenergetics was established, the involvement of CoQ in the pathophysiology of heart failure was hypothesized. This provided the rationale for numerous clinical trials of CoQ 10 as adjunctive treatment for heart failure. A mild hypotensive effect of CoQ was reported in the early years of clinical use of this compound. We review early human and animal studies on the vascular effects of CoQ. We then focus on endothelial dysfunction in type 2 diabetes and the possible impact on this condition of antioxidants and nutritional supplements, and in particular the therapeutic effects of CoQ. The effect of CoQ 10 on endothelial dysfunction in ischemic heart disease is also reviewed together with recent data highlighting that treatment with CoQ 10 increases extracellular SOD activity.

The aging process decreases tissue function and regenerative capacity, which has been associated with cellular senescence and a decline in adult or somatic stem cell numbers and self-renewal within multiple tissues. The potential therapeutic application of stem cells to reduce the burden of aging and stimulate tissue regeneration after trauma is very promising. Much research is currently ongoing to identify the factors and molecular mediators of stem cell self-renewal to reach these goals. Over the last two decades, fibroblast growth factors (FGFs) and their receptors (FGFRs) have stood up as major players in both embryonic development and tissue repair. Moreover, many studies point to somatic stem cells as major targets of FGF signaling in both tissue homeostasis and repair. FGFs appear to promote self-renewing proliferation and inhibit cellular senescence in nearly all tissues tested to date. Here we review the role of FGFs and FGFRs in stem cell self-renewal, cellular senescence,...

SUMMARY Carboxypeptidase E (CPE) is highly concentrated in neuroendocrine tissues and is the only carboxypeptidase detected in mature secretory vesicles. Carboxypeptidase D (CPD), a carboxypeptidase with CPE-like activity, is widely distributed in tissues and is present in the trans -Golgi network. Previous work had shown that both CPE and CPD are expressed in the human placenta and that CPD is

... E-mail hyoshi{at}cardiol.med.kyushu-u.ac.jp. ... General Procedures The study was conducted with the participants in a supine position and in a postabsorptive state in an air-conditioned room at 25°C to 26°C. Under local anesthesia with 2% procaine, the left brachial artery was ...

A fast and convenient method for silver staining of proteins on electroblotting membranes was developed based on Gallyas' histochemical intensifier and applied to human endothelial cell proteins separated by one-and two-dimensional electrophoresis and electroblotted to polyvinyl difluoride membranes. The method allowed detection of proteins on membranes with a sensitivity equal to the sensitivity of the most sensitive silver-staining protocols for electrophoresis gels. Also, the method was compatible with preceding immunostaining on the same membrane. Furthermore, an intensifying method for proteins in silverstained SDS-PAGE gels was developed based on Gallyas' histochemical intensifier. This method was applied to proteins separated by one-and two-dimensional gel electrophoresis and visualized by one of several silver-staining methods. Maximal intensification was achieved for the less sensitive but fast acidic silver-staining protocols, but even for the very sensitive alkaline protocols a significant increase in signal to noise ratio was obtained. In particular, negatively stained or invisible proteins on the silver-stained gels were found to be visualized by the Gallyas stain. Proteins from silver-stained and Gallyas-stained gels were identified by mass spectrometry, and the intensification procedure was fully compatible with mass spectrometry. © 2002 Elsevier Science (USA)

ollowing the discovery of endothelium-derived relaxing factor (EDRF), the chase was on to identify it. Its further characterization led gradually to a growing realization that it had much in common with nitric oxide (NO), including its very short half-life, the association of its relaxant action with elevation of cGMP levels, and inhibition of its action by methylene blue or hemoglobin. This paper showed experimentally that the effects of authentic NO and EDRF were indistinguishable. It stands as a point of focus in the important discovery of NO as a biological agent, a discovery, as is often the case, towards which a number of different groups were converging.

The additional health-promoting properties of functional virgin olive oil (FVOO) enriched with its own phenolic compounds (OOPC) versus the parental virgin olive oil (VOO) must be tested in appropriate human clinical trials. Our aim was to assess the effects of FVOO on endothelial function in hypertensive patients. Thirteen pre- and stage-1 hypertensive patients received a single dose of 30 mL of FVOO (OOPC=961 mg/kg) or VOO (OOPC=289 mg/kg) in a postprandial randomised, double blind, crossover trial. Endothelial function, measured as ischemic reactive hyperemia (IRH) and related biomarkers, were followed for 5h after consumption. Compared with VOO, FVOO increased IRH (P&lt;0.05) and plasma Cmax of hydroxytyrosol sulphate, a metabolite of OOPC 2h postprandial (P=0.05). After FVOO ingestion, oxidised LDL decreased (P=0.010) in an inverse relationship with IRH AUC values (P=0.01). FVOO provided more benefits on endothelial function than a standard natural virgin olive oil in pre- and ...

Vascular diseases supported by aberrant angiogenesis have increased incidence in HIV-1-infected patients. Several data suggest that endothelium dysfunction relies on action of HIV-1 proteins rather than on a direct effect of the virus itself. The HIV-1 matrix protein p17 is known to deregulate the biological activity of different immune cells. Recently, p17 was found to mimic IL-8 chemokine activity by binding to the IL-8 receptor CXCR1. Here we show that p17 binds with high affinity to CXCR2, a CXCR1-related receptor, and promotes the formation of capillary-like structures on human endothelial cells (ECs) by interacting with both CXCR1 and CXCR2 expressed on the EC surface. ERK signaling via Akt was defined as the pathway responsible for p17-induced tube formation. Ex vivo and in vivo experimental models confirmed the provasculogenic activity of p17, which was comparable to that induced by VEGF-A. The hypothesis of a major role for p17 in HIV-1-induced aberrant angiogenesis is enforced by the finding that p17 is detected, as a single protein, in blood vessels of HIV-1-patients and in particular in the nucleus of ECs. Localization of p17 in the nucleus of ECs was evidenced also in in vitro experiments, suggesting the internalization of exogenous p17 in ECs by mechanisms of receptormediated endocytosis. Recognizing p17 interaction with CXCR1 and CXCR2 as the key event in sustaining EC aberrant angiogenesis could help us to identify new treatment strategies in combating AIDSrelated vascular diseases. extracellular viral proteins | virokine | Akt-mediated ERK pathway | vasculogenic assays | surface plasmon resonance A ngiogenesis is a physiological process requiring growth of new blood vessels from preexisting vessels. This process involves coordinated endothelial cell (EC) proliferation, invasion, migration, and tube formation (1). When new vessels are required, proangiogenic factors are produced, whereas restoration of physiological conditions is achieved by producing inhibitors of angiogenesis and vessel stabilization factors. Breakdown of the tight regulated angiogenic balance leads to dysfunctional endothelium, abnormal angiogenesis, and vascular diseases.

Polycythemia vera (PV) and essential thrombocythemia (ET) are two Philadelphianegative myeloproliferative neoplasms (MPN) associated with an acquired mutation in the JAK2 tyrosine kinase gene. There is a rare incidence of progression to myelofibrosis and myeloid metaplasia in both disorders, which may or may not precede transformation to acute myeloid leukemia, but thrombosis is the main cause of morbidity and mortality. The pathophysiology of thrombosis in patients with MPN is complex. Traditionally, abnormalities of platelet number and function have been claimed as the main players, but increased dynamic interactions between platelets, leukocytes, and the endothelium do probably represent a fundamental interplay in generating a thrombophilic state. In addition, endothelial dysfunction, a well-known risk factor for vascular disease, may play a role in the thrombotic risk of patients with PV and ET. The identification of plasma markers translating the hemostatic imbalance in patients with PV and ET would be extremely helpful in order to define the subgroup of patients with a significant clinical risk of thrombosis.

In the liver, tissue-type plasminogen activator (t-PA) is endocytosed by hepatic parenchymal (PC), endothelial (EC) and Kupffer (KC) cells. Although the endocytosis is receptor-mediated, it remains a matter of discussion which receptors are involved in this catabolic process. To evaluate the role of a protein-specific receptor, as well as the possible involvement of the galactose receptor on PC and the mannose receptor on EC, we have employed different glycosylation variants of t-PA in biochemical and immunocytochemical studies. Partial or total removal of carbohydrate side-chains by endoglycosidases did not prevent clearance and hepatic endocytosis of t-PA by either of the liver cell types. Blockade of the galactose and mannose receptors by co-application of a large excess of the glycoprotein ovalbumin remained without effect on the binding and uptake of t-PA by hepatic cells. However, the contribution of different liver cell types to the hepatic clearance of t-PA was to a certain ...