Release Of Plasminogen Activator Research Articles

Tissue plasminogen activator (tPA) attenuates propofol-induced apoptosis in developing hippocampal neurons.

BackgroundWe investigated the effect of propofol on the tissue plasminogen activator (tPA) release in developing hippocampal neurons, and explored the effects of exogenous tPA on the propofol-induced neuron apoptosis.MethodsPrimary hippocampal neurons isolated from neonatal Sprague-Dawley rats were exposed to propofol (20, 50, and 100 μM) for 6 h either one time or three times. Finally, neurons were pretreated with exogenous tPA (5 µg/ml), followed by propofol exposure (100 μM, 6 h). The neuron apoptosis was detected by terminal transferase deoxyuridine triphosphate-biotin nick-end labeling (TUNEL) and the protein expression of cleaved caspase-3 (Cl-Csp3) was analyzed by western blot, the tPA in media was tested by enzyme-linked immunosorbent assay.ResultsPropofol exposure significantly increased the number of TUNEL-positive neurons and Cl-Csp3 expression in developing hippocampal neurons. Propofol decreased tPA level in the media of developing hippocampal neurons. The neuron appotosis induced by propofol was attenuated by pretreatment of tPA.ConclusionPropofol exposure decreased tPA release in developing hippocampal neurons. The addition of tPA could partially reverse the apoptotic effect of propofol.

Tissue-type plasminogen activator induces synaptic vesicle endocytosis in cerebral cortical neurons

The release of the serine proteinase tissue-type plasminogen activator (tPA) from the presynaptic terminal of cerebral cortical neurons plays a central role in the development of synaptic plasticity, adaptation to metabolic stress and neuronal survival. Our earlier studies indicate that by inducing the recruitment of the cytoskeletal protein βII-spectrin and voltage-gated calcium channels to the active zone, tPA promotes Ca2+-dependent translocation of synaptic vesicles (SVs) to the synaptic release site where they release their load of neurotransmitters into the synaptic cleft. Here we used a combination of in vivo and in vitro experiments to investigate whether this effect leads to depletion of SVs in the presynaptic terminal. Our data indicate that tPA promotes SV endocytosis via a mechanism that does not require the conversion of plasminogen into plasmin. Instead, we show that tPA induces calcineurin-mediated dynamin I dephosphorylation, which is followed by dynamin I-induced recruitment of the actin-binding protein profilin II to the presynaptic membrane, and profilin II-induced F-actin formation. We report that this tPA-induced sequence of events leads to the association of newly formed SVs with F-actin clusters in the endocytic zone. In summary, the data presented here indicate that following the exocytotic release of neurotransmitters tPA activates the mechanism whereby SVs are retrieved from the presynaptic membrane and endocytosed to replenish the pool of vesicles available for a new cycle of exocytosis. Together, these results indicate that in murine cerebral cortical neurons tPA plays a central role coupling SVs exocytosis and endocytosis.

Magnetically controlled release of recombinant tissue plasminogen activator from chitosan nanocomposites for targeted thrombolysis.

Ionic cross-linking of water-soluble chitosan with sodium tripolyphosphate in the presence of recombinant tissue plasminogen activator (rtPA) and magnetite (Fe3O4) nanoparticles could produce rtPA-encapsulated magnetic chitosan nanoparticles (MCNPs-rtPA). MCNPs do not elicit cytotoxicity and hemolysis in vitro. MCNPs-rtPA showed a negligible release of the rtPA protein when stored in phosphate buffer for 28 days. In contrast, the burst release of rtPA from MCNPs-rtPA was found in the serum with 60% of the original activity released in 30 min. The drug release into the serum is also magnet-sensitive; the release could be turned down with a magnetic field when MCNPs-rtPA was pelleted and reversibly turned on after removing the magnetic field when MCNPs-rtPA was dispersed. An in vitro thrombolytic study by thromboelastometry indicated a controlled release of rtPA from MCNPs-rtPA. In a rat embolic model where a preformed blood clot lodged in the left iliac artery upstream of the pudic epigastric branch, MCNPs-rtPA (0.2 mg kg-1 rtPA) was administered and guided magnetically to the clot, followed by mobile magnetic guidance for 60 min. Iliac blood flow increased immediately in response to the treatment, and reached a stable level ∼50 min after drug administration and the hind limb perfusion rate was restored from 53% to 75% of the basal level. Effective thrombolysis was therefore successfully demonstrated at an rtPA dose equivalent to 20% of the regular dose when the MCNPs-rtPA pellet was magnet-guided to the blood clot, followed by a triggered release of rtPA when switched to mobile magnetic guidance.

Dependence of Proximal GC Boxes and Binding Transcription Factors in the Regulation of Basal and Valproic Acid-Induced Expression of t-PA.

Objective. Endothelial tissue-type plasminogen activator (t-PA) release is a pivotal response to protect the circulation from occluding thrombosis. We have shown that the t-PA gene is epigenetically regulated and greatly induced by the histone deacetylase (HDAC) inhibitor valproic acid (VPA). We now investigated involvement of known t-PA promoter regulatory elements and evaluated dependence of potential interacting transcription factors/cofactors. Methods. A reporter vector with an insert, separately mutated at either the t-PA promoter CRE or GC box II or GC box III elements, was transfected into HT-1080 and HUVECs and challenged with VPA. HUVECs were targeted with siRNA against histone acetyl transferases (HAT) and selected transcription factors from the Sp/KLF family. Results. An intact VPA-response was observed with CRE mutated constructs, whereas mutation of GC boxes II and III reduced the magnitude of the induction by 54 and 79% in HT-1080 and 49 and 50% in HUVECs, respectively. An attenuated induction of t-PA mRNA was observed after Sp2, Sp4, and KLF5 depletion. KLF2 and p300 (HAT) were identified as positive regulators of basal t-PA expression and Sp4 and KLF9 as repressors. Conclusion. VPA-induced t-PA expression is dependent on the proximal GC boxes in the t-PA promoter and may involve interactions with Sp2, Sp4, and KLF5.

Promotion of Wound Healing by an Agonist of Adenosine A2A Receptor Is Dependent on Tissue Plasminogen Activator.

Impaired wound healing, as it occurs in diabetes mellitus or long-term corticoid treatment, is commonly associated with disability, diminished quality of life, and high economic costs. Selective agonists of the A2A receptor subtype of adenosine, an endogenous regulator of inflammation, promote tissue repair in animal models, both healthy and with impaired healing. Plasmin-mediated proteolysis of fibrin and other matrix proteins is essential for cell migration at sites of injury. Since adenosine A2A receptor activation increases plasminogen activator release from macrophages and mast cells, we studied the effect of a selective agonist, CGS-21680, on full-thickness excisional wound closure in wild-type, urokinase plasminogen activator (uPA)-deficient, and tissue plasminogen activator (tPA)-deficient mice. Wound closure was impaired in tPA- and uPA-deficient mice as compared with wild-type mice, and topical application of CGS-21680 significantly increased the rate at which wounds closed in wild-type mice and uPA-deficient mice, but not in tPA-deficient mice. Immunostaining of tissue sections showed that tPA was present in endothelial cells and histiocytes by day 3 post-wound and also by day 6. In contrast, uPA was more prominent in these cell types only by day 6 post-wound. Our results confirm that plasminogen activation contributes to wound repair and are consistent with the hypothesis that adenosine A2A receptor activation promotes wound closure by a mechanism that depends upon tPA, but not uPA. Moreover, our results suggest that topical adenosine A2A receptor agonists may be useful in promotion of wound closure in patients with impaired wound healing.

Endogenous plasminogen activators mediate progressive intracerebral hemorrhage after traumatic brain injury in mice

AbstractPersistent intracerebral hemorrhage (ICH) is a major cause of death and disability after traumatic brain injury (TBI) for which no medical treatment is available. Delayed bleeding is often ascribed to consumptive coagulopathy initiated by exposed brain tissue factor. We examined an alternative hypothesis, namely, that marked release of tissue-type plasminogen activator (tPA) followed by delayed synthesis and release of urokinase plasminogen activator (uPA) from injured brain leads to posttraumatic bleeding by causing premature clot lysis. Using a murine model of severe TBI, we found that ICH is reduced in tPA−/− and uPA−/− mice but increased in PAI-1−/− mice compared with wild-type (WT) mice. tPA−/−, but not uPA−/−, mice developed a systemic coagulopathy post-TBI. Tranexamic acid inhibited ICH expansion in uPA−/−mice but not in tPA−/− mice. Catalytically inactive tPA-S481A inhibited plasminogen activation by tPA and uPA, attenuated ICH, lowered plasma d-dimers, lessened thrombocytopenia, and improved neurologic outcome in WT, tPA−/−, and uPA−/− mice. ICH expansion was also inhibited by tPA-S481A in WT mice anticoagulated with warfarin. These data demonstrate that protracted endogenous fibrinolysis induced by TBI is primarily responsible for persistent ICH and post-TBI coagulopathy in this model and offer a novel approach to interrupt bleeding.

Limitations of component therapy for massive haemorrhage: is whole blood the whole solution?

Limitations of component therapy for massive haemorrhage: is whole blood the whole solution?

Effect of ascorbate on plasminogen activator inhibitor-1 expression and release from platelets and endothelial cells in an in-vitro model of sepsis

The microcirculation during sepsis fails due to capillary plugging involving microthrombosis. We demonstrated that intravenous injection of ascorbate reduces this plugging, but the mechanism of this beneficial effect remains unclear. We hypothesize that ascorbate inhibits the release of the antifibrinolytic plasminogen activator inhibitor-1 (PAI-1) from endothelial cells and platelets during sepsis. Microvascular endothelial cells and platelets were isolated from mice. Cells were cultured and stimulated with lipopolysaccharide (LPS), tumor necrosis factor alpha (TNFα), or thrombin (agents of sepsis), with/without ascorbate for 1-24 h. PAI-1 mRNA was determined by quantitative PCR. PAI-1 protein release into the culture medium was measured by ELISA. In platelets, PAI-1 release was measured after LPS, TNFα, or thrombin stimulation, with/without ascorbate. In endothelial cells, LPS and TNFα increased PAI-1 mRNA after 6-24 h, but no increase in PAI-1 release was observed; ascorbate did not affect these responses. In platelets, thrombin, but not LPS or TNFα, increased PAI-1 release; ascorbate inhibited this increase at low extracellular pH. In unstimulated endothelial cells and platelets, PAI-1 is released into the extracellular space. Thrombin increases this release from platelets; ascorbate inhibits it pH-dependently. The data suggest that ascorbate promotes fibrinolysis in the microvasculature under acidotic conditions in sepsis.

Impaired endogenous fibrinolytic capacity in prehypertensive men.

Prehypertension (BP 120–139/80–89 mmHg) is associated with an increased risk for future atherothrombotic events. Although the mechanisms underlying this elevated risk are not completely understood, one possibility is that prehypertension is associated with impaired endothelial fibrinolytic capacity. We tested the hypothesis that vascular endothelial release of t-PA is impaired in prehypertensive men. Net endothelial release of t-PA was determined, in vivo, in response to intrabrachial infusions of bradykinin (12.5, 25, 50 ng/100 mL tissue/min) and sodium nitroprusside at (1.0, 2.0, 4.0 µg/100 mL tissue/min) in 42 middle-age and older men: 16 normotensive (BP range: 100–119/57–79 mmHg); 16 prehypertensive (BP range: 120–139/76–89 mmHg); and 10 hypertensive (BP range: 140–150/74–100 mmHg). Net release of t-PA antigen was ~25% lower (P < 0.05) in the prehypertensive (−0.9 ± 0.8 to 42.4 ± 5.3 ng/100 mL tissue/min) compared with the normotensive (0.5 ± 1.0 to 53.9 ± 6.5 ng/100 mL tissue/min) men. There was no significant difference in t-PA release between the hypertensive (−1.8±1.6 to 40.8±6.6 ng/100 mL tissue/min) and prehypertensive groups. Sodium nitroprusside did not significantly alter t-PA release in any group. These data indicate that endothelial t-PA release is diminished in prehypertensive men. Further, the level of impairment in t-PA release seen with clinical hypertension is already apparent in the prehypertensive state. Impaired endothelial fibrinolytic function may underlie the increased atherothrombotic risk associated with blood pressure in the prehypertensive range.

Influence of Dietary Saturated Fat Intake on Endothelial Fibrinolytic Capacity in Adults

Approximately 50% of middle-aged and older adults in the United States regularly consume a diet high in saturated fat. High dietary saturated fat intake has been linked to promote atherothrombotic vascular disease. We tested the hypothesis that endothelial fibrinolytic function is diminished in middle-aged and older adults who habitually consume a diet high in saturated fat. Twenty-four healthy, sedentary middle-aged, and older adults (54 to 71years) were studied: 10 (8 men and 2 women) with a dietary saturated fat intake <10% (lower saturated fat) of total calories and 14 (9 men and 5 women) with a dietary saturated fat intake ≥10% of total calories (high saturated fat). Net endothelial release of tissue-type plasminogen activator (t-PA), the primary activator of fibrinolysis, was determined, invivo, in response to intrabrachial infusions of bradykinin (12.5 to 50.0ng/100ml tissue/min) and sodium nitroprusside (1.0 to 4.0μg/100ml tissue/min). Capacity of the endothelium to release t-PA in response to bradykinin was ∼30% less (p <0.05) in the high (from-0.7 ± 0.6 to 36.9 ± 3.3ng/100ml tissue/min) compared with the lower (from-0.3 ± 0.3 to 53.4 ± 7.8ng/100ml tissue/min) dietary saturated fat group. Moreover, total amount of t-PA released was significantly less (∼30%) (201 ± 22 vs 274 ± 29ng/100ml tissue) in the adults who reported consuming a diet high in saturated fat. These results indicate that the capacity of the endothelium to release t-PA is lower in middle-aged and older adults who habitually consume a diet high in saturated fat. In conclusion, endothelial fibrinolytic dysfunction may underlie the increased atherothrombotic disease risk with a diet high in saturated fat.

Endothelium-Derived Hyperpolarizing Factor Mediates Bradykinin-Stimulated Tissue Plasminogen Activator Release in Humans

Aims: Bradykinin (BK) stimulates tissue plasminogen activator (t-PA) release from human endothelium. Although BK stimulates both nitric oxide and endothelium-derived hyperpolarizing factor (EDHF) release, the role of EDHF in t-PA release remains unexplored. This study sought to determine the mechanisms of BK-stimulated t-PA release in the forearm vasculature of healthy human subjects. Methods: In 33 healthy subjects (age 40.3 ± 1.9 years), forearm blood flow (FBF) and t-PA release were measured at rest and after intra-arterial infusions of BK (400 ng/min) and sodium nitroprusside (3.2 mg/min). Measurements were repeated after intra-arterial infusion of tetraethylammonium chloride (TEA; 1 µmol/min), fluconazole (0.4 µmol·min^-1·l^-1), and N^G-monomethyl-<smlcap>L</smlcap>-arginine (<smlcap>L</smlcap>-NMMA, 8 µmol/min) to block nitric oxide, and their combination in separate studies. Results: BK significantly increased net t-PA release across the forearm (p < 0.0001). Fluconazole attenuated both BK-mediated vasodilation (-23.3 ± 2.7% FBF, p < 0.0001) and t-PA release (from 50.9 ± 9.0 to 21.3 ± 8.9 ng/min/100 ml, p = 0.02). TEA attenuated FBF (-14.7 ± 3.2%, p = 0.002) and abolished BK-stimulated t-PA release (from 22.9 ± 5.7 to -0.8 ± 3.6 ng/min/100 ml, p = 0.0002). <smlcap>L</smlcap>-NMMA attenuated FBF (p < 0.0001), but did not inhibit BK-induced t-PA release (nonsignificant). Conclusion: BK-stimulated t-PA release is partly due to cytochrome P₄₅₀-derived epoxides and is inhibited by K⁺_Ca channel blockade. Thus, BK stimulates both EDHF-dependent vasodilation and t-PA release.

Histone Deacetylase Inhibitor Treatment Increases Coronary t-PA Release in a Porcine Ischemia Model

BackgroundThe expression of the tissue plasminogen activator gene can be affected by histone deacetylation inhibition and thus appears to be under epigenetic control.ObjectivesThe study aimed to test if in vivo pharmacological intervention by valproic acid treatment would lead to increase in tissue plasminogen activator release capacity.MethodsIn an anaesthetized pig model, a controlled transient coronary occlusion was used to stimulate coronary tissue plasminogen activator release in a valproic acid treated (one week) and a non-treated group. Coronary venous blood samples from the ischemic region were collected, great cardiac vein thermodilution flow measurements were performed, and trans-coronary tissue plasminogen activator fluxes were calculated. Plasminogen activator inhibitor-1 was also measured.ResultsAdequate sampling from the affected area after the 10 minute ischemic period was confirmed by lactate measurements. Fluxes for tissue plasminogen activator at minutes 1, 3, 5, 7 and 10 were measured and then used to present cumulative net tissue plasminogen activator release for the whole measurement period for both groups. Area under the curve was higher for the valproic acid treated group at 10 minutes; 932±173 nanograms (n = 12) compared to the non-treated group, 451±78 nanograms (n = 10, p = 0.023). There was no difference in levels of plasminogen activator inhibitor-1 between groups.ConclusionsThese findings support a proof of concept for histone deacetylation inhibition positive effect on tissue plasminogen activator expression in an in vivo setting. Further studies are needed to find an optimal way to implement histone deacetylation inhibition to achieve desired clinical changes in tissue plasminogen activator expression.

Cortical‐layer‐specific effects of PACAP and tPA on interneuron migration during post‐natal development of the cerebellum

During early post-natal development of the cerebellum, granule neurons (GN) execute a centripetal migration toward the internal granular layer, whereas basket and stellate cells (B/SC) migrate centrifugally to reach their final position in the molecular layer (ML). We have previously shown that pituitary adenylate cyclase-activating polypeptide (PACAP) stimulates in vitro the expression and release of the serine protease tissue-type plasminogen activator (tPA) from GN, but the coordinated role of PACAP and tPA during interneuron migration has not yet been investigated. Here, we show that endogenous PACAP is responsible for the transient arrest phase of GN at the level of the Purkinje cell layer (PCL) but has no effect on B/SC. tPA is devoid of direct effect on GN motility in vitro, although it is widely distributed along interneuron migratory routes in the ML, PCL, and internal granular layer. Interestingly, plasminogen activator inhibitor 1 reduces the migration speed of GN in the ML and PCL, and that of B/SC in the ML. Taken together, these results reveal for the first time that tPA facilitates the migration of both GN and fast B/SC at the level of their intersection in the ML through degradation of the extracellular matrix. Crucial role of tissue plasminogen activator (tPA) in interneuron migration. Interneuron migration is a critical step for normal establishment of neuronal network. This study indicates that, in the post-natal cerebellum, tPA facilitates the opposite migration of immature excitatory granule neurons (GN) and immature inhibitory basket/stellate cells (B/SC) along the same migratory route. These data show that tPA exerts a pivotal role in neurodevelopment.

Clinical relevance and cellular source of elevated soluble urokinase plasminogen activator receptor (suPAR) in acute liver failure

Acute liver failure (ALF) is a life-threatening condition with a high mortality rate. The expression of urokinase plasminogen activator receptor (uPAR, CD87) and release of its shedded receptor into serum as soluble uPAR (suPAR) have been closely related to immune activation and prognosis in systemic inflammation and cirrhosis. We now aimed at investigating the clinical relevance and cellular source of uPAR and circulating suPAR in ALF. Serum suPAR concentrations were measured in 48 ALF patients and 62 healthy controls from a German liver transplantation centre. Hepatic immune cell subsets and uPAR expression were studied by FACS, qPCR and immunohistochemistry. Circulating suPAR levels were significantly increased in ALF patients, independent from the underlying aetiology, in comparison to controls. Serum suPAR concentrations were closely correlated with parameters reflecting liver cell injury, decreased liver function and the model of end-stage liver disease (MELD) score in ALF patients. By immunohistochemistry from explanted livers, ALF was associated with distinct immune cell accumulation and strong up-regulation of intrahepatic uPAR mRNA expression. CD87 (uPAR) expression was specifically detected on intrahepatic 'non-classical' monocytes (CD14(+) CD16(+) ), NKT and CD56(dim) NK cells isolated from human liver, but not on parenchymal or other non-parenchymal hepatic cell types. Membrane-bound uPAR was rapidly cleaved from monocytes upon inflammatory stimulation by lipopolysaccharide (LPS) and partially by co-cultured lymphocytes. Similar to its prognostic properties in patients with sepsis or cirrhosis, intrahepatic uPAR activation and serum suPAR concentrations might serve as an interesting biomarker in ALF.

Substance P Increases Sympathetic Activity During Combined Angiotensin-Converting Enzyme and Dipeptidyl Peptidase-4 Inhibition

Dipeptidyl peptidase-4 inhibitors prevent the degradation of incretin hormones and reduce postprandial hyperglycemia in patients with type 2 diabetes mellitus. Dipeptidyl peptidase-4 degrades other peptides with a penultimate proline or alanine, including bradykinin and substance P, which are also substrates of angiotensin-converting enzyme (ACE). During ACE inhibition, substance P is inactivated primarily by dipeptidyl peptidase-4, whereas bradykinin is first inactivated by aminopeptidase P. This study tested the hypothesis that dipeptidyl peptidase-4 inhibition potentiates vasodilator and fibrinolytic responses to substance P when ACE is inhibited. Twelve healthy subjects participated in this randomized, double-blinded, placebo-controlled crossover study. On each study day, subjects received sitagliptin 200 mg by mouth or placebo. Substance P and bradykinin were infused via brachial artery before and during intra-arterial enalaprilat. Sitagliptin and enalaprilat each reduced forearm vascular resistance and increased forearm blood flow without affecting mean arterial pressure, but there was no interactive effect of the inhibitors. Enalaprilat increased bradykinin-stimulated vasodilation and tissue plasminogen activator release; sitagliptin did not affect these responses to bradykinin. The vasodilator response to substance P was unaffected by sitagliptin and enalaprilat; however, substance P increased heart rate and vascular release of norepinephrine during combined ACE and dipeptidyl peptidase-4 inhibition. In women, sitagliptin diminished tissue plasminogen activator release in response to substance P both alone and during enalaprilat. Substance P increases sympathetic activity during combined ACE and dipeptidyl peptidase-4 inhibition. - URL: http://www.clinicaltrials.gov. Unique identifier: NCT01413542.

High-density lipoprotein from patients with coronary heart disease loses anti-thrombotic effects on endothelial cells: impact on arterial thrombus formation.

Thrombus formation is determined by the balance between pro- thrombotic mediators and anti-thrombotic factors.High-density lipoprotein (HDL) from healthy subjects exerts anti-thrombotic properties. Whether this is also the case for HDL from patients with stable coronary heart disease (CHD) or acute coronary syndrome (ACS) is unknown.In human aortic endothelial cells in culture,HDL (50 µg/ml) from healthy subjects (HS) inhibited thrombin-induced tissue factor (TF) expression and activity, while HDL (50 µg/ml) from CHD and ACS patients did not. Similarly, only healthy HDL increased endothelial tissue factor pathway inhibitor (TFPI) expression and tissue plasminogen activator (tPA) release, while HDL from CHD and ACS patients had no effect. Healthy HDL inhibited thrombin-induced plasminogen activator inhibitor type 1 (PAI-1) expression, while HDL from ACS patients enhanced endothelial PAI-1 expression. Inhibition of nitric oxide (NO) formation with L-NAME (100 µmol/l) abolished the anti-thrombotic effects of healthy HDL on TF, TFPI, and tPA expression. The exogenous nitric oxide donor, DETANO, mimicked the effects of healthy HDL and counterbalanced the loss of anti-thrombotic effects of HDL from CHD and ACS patients in endothelial cells. In line with this observation, healthy HDL, in contrast to HDL from CHD and ACS patients, increased endothelial NO production. In the laser-injured carotid artery of the mouse, thrombus formation was delayed in animals treated with healthy HDL compared with mice treated with vehicle or HDL from patients with CHD or ACS. In conclusion, HDL from CHD and ACS patients loses the ability of healthy HDL to suppress TF and to increase TFPI and t-PA and instead enhances PAI-1 and arterial thrombus formation.

Histamine‐induced vasodilatation in the human forearm vasculature

To investigate the mechanism of action of intra-arterial histamine in the human forearm vasculature. Three studies were conducted to assess changes in forearm blood flow (FBF) using venous occlusion plethysmography in response to intra-brachial histamine. First, the dose-response was investigated by assessing FBF throughout a dose-escalating histamine infusion. Next, histamine was infused at a constant dose to assess acute tolerance. Finally, a four way, double-blind, randomized, placebo-controlled crossover study was conducted to assess FBF response to histamine in the presence of H1 - and H2 -receptor antagonists. Flare and itch were assessed in all studies. Histamine caused a dose-dependent increase in FBF, greatest with the highest dose (30 nmol min(-1) ) infused [mean (SEM) infused arm vs. control: 26.8 (5.3) vs. 2.6 ml min(-1) 100 ml(-1) ; P < 0.0001]. Dose-dependent flare and itch were demonstrated. Acute tolerance was not observed, with an increased FBF persisting throughout the infusion period. H2 -receptor antagonism significantly reduced FBF (mean (95% CI) difference from placebo at 30 nmol min(-1) histamine: -11.9 ml min(-1) 100 ml(-1) (-4.0, -19.8), P < 0.0001) and flare (mean (95% CI) difference from placebo: -403.7 cm(2) (-231.4, 576.0), P < 0.0001). No reduction in FBF or flare was observed in response to the H1 -receptor antagonist. Itch was unaffected by the treatments. Histamine did not stimulate vascular release of tissue plasminogen activator or von Willebrand factor. Histamine causes dose-dependent vasodilatation, flare and itch in the human forearm. H2 -receptors are important in this process. Our results support further exploration of combined H1 - and H2 -receptor antagonist therapy in acute allergic syndromes.

Growth hormone replacement normalizes impaired fibrinolysis: New insights into endothelial dysfunction in patients with hypopituitarism and growth hormone deficiency

BackgroundCardiovascular morbidity in adult patients with growth hormone deficiency (GHD) and hypopituitarism is increased. Clustering of cardiovascular risk factors leading to endothelial dysfunction and impaired fibrinolysis has also been reported and may account for progression to overt vascular changes in these patients. However, effect of long lasting GH replacement therapy on fibrinolytic capacity in GH deficient patients has not been investigated so far. ObjectiveTo investigate fibrinolysis before and after challenge with venous occlusion in GHD patients with hypopituitarism before and during one year of growth hormone replacement. DesignHospital based, interventional, prospective study. Investigated subjectsTwenty one patient with GHD and fourteen healthy control subjects matched for age, sex and body mass index (BMI). MethodsAnthropometric, metabolic and fibrinolytic parameters were measured at the start and after three, six and twelve months of treatment with human recombinant GH. ResultsAt baseline GHD patients had significantly impaired fibrinolysis compared to healthy persons. During treatment with GH, significant changes were observed in insulin like growth factor 1(IGF-1) [from baseline 6.9(2.4–13.5) to 22.0(9.0–33.0) nmol/l after one month of treatment; p<0.01] and fibrinolysis. Improvement in fibrinolysis was mostly attributed to improvement of stimulated endothelial tissue plasminogen activator (t-PA) release in response to venous occlusion [from baseline 1.1(0.4–2.6) to 1.9(0.5–8.8) after one year of treatment; p<0.01]. ConclusionGrowth hormone replacement therapy has favorable effects on t-PA release from endothelium and net fibrinolytic capacity in GHD adults, which may contribute to decrease their risk of vascular complications.

Genomics of corneal wound healing: a review of the literature

Corneal wound healing is a complex process: its mechanisms and the underlying genetic control are not fully understood. It involves the integrated actions of multiple growth factors, cytokines and proteases produced by epithelial cells, stromal keratocytes, inflammatory cells and lacrimal gland cells. Following an epithelial insult, multiple cytokines are released triggering a cascade of events that leads to repair the epithelial defect and remodelling of the stroma to minimize the loss of transparency and function. In this review, we examine the literature surrounding the genomics of corneal wound healing with respect to the following topics: epithelial and stromal wound healing (including inhibition); corneal neovascularisation; the role of corneal nerves in wound healing; the endothelium; the role of aquaporins and aptamers. We also examine the effect of ectasia on corneal wound healing with regard to keratoconus and following corneal surgery. A better understanding of the cellular and molecular changes that occur during repair of corneal wounds will provide the opportunity to design treatments that selectively modulate key phases of the healing process resulting in scars that more closely resemble normal corneal architecture.

Coagulopathy, catecholamines, and biomarkers of endothelial damage in experimental human endotoxemia and in patients with severe sepsis: A prospective study

PurposeThe aim of this study was to investigate associations between circulating catecholamines, endothelial damage, and coagulopathy in experimental human endotoxemia and septic patients. Materials and MethodsNine healthy male volunteers undergoing endotoxemia (4-hour 0.5 ng/kg/hour infusion of E. coli lipopolysaccharide, blood sampling at 0, 4, and 6 hours) and 20 patients with severe sepsis. Analysis of plasma biomarkers (adrenaline, noradrenaline, thrombomodulin, syndecan-1, soluble vascular endothelial cadherin, histone-complexed DNA fragments, soluble CD40 ligand [sCD40L], protein C, tissue-type plasminogen activator, plasminogen activator inhibitor 1) and routine coagulation tests. ResultsEndotoxemia increased heart rate, temperature, white blood cell count, C-reactive protein and procalcitonin, decreased blood pressure and induced a hemostatic response with platelet consumption, reduced protein C and sCD40L levels and enhanced tissue-type plasminogen activator release (all P < .05). Septic patients had increased levels of noradrenaline, syndecan-1, thrombomodulin, histone-complexed DNA and sCD40L but reduced soluble vascular endothelial cadherin and plasminogen activator inhibitor 1 (all P < .05) and plasma catecholamines correlated positively with syndecan-1 (adrenaline and noradrenaline) and sTM (only noradrenaline) (all P < .05), biomarkers reflecting endothelial damage. Furthermore, noradrenaline, syndecan-1 and thrombomodulin levels correlated with INR and disease severity scores (noradrenaline and thrombomodulin) (all P < .05). ConclusionsExperimental endotoxemia induced a discrete hemostatic response without sympathoadrenal activation or endothelial damage. Septic patients had high levels of catecholamines and endothelial damage biomarkers that correlated with each other and with markers of hypocoagulability and disease severity.