Clinical predictors of complications in patients with left-sided infective endocarditis: A retrospective study of 206 episodes.

PURPOSE: Early identification of specific patient subgroups at high risk of developing life-threatening infective endocarditis (IE) complications is of paramount importance. Better stratification may allow more intensive treatment of these patients and positively influences clinical outcomes. METHODS: We carried out a retrospective survey of consecutive left-sided IE adult patients, admitted over a 15-year period to two main tertiary care centres in the Czech Republic. RESULTS: Among a group of 196 patients (155 males; median age 64 years), a total of 206 left-sided IE episodes were identified. Perivalvular extension of infection was most frequently seen in prosthetic aortic valve endocarditis (OR 6.706, p<0.0001). Valve prolapse/perforation during IE episodes was significantly associated with mitral valve IE (OR 2.136, p=0.026) and vegetation length (OR 1.055, p=0.009). Septic shock was significantly related to two main risk factors: S. aureus infection (OR 8.459, p=<0.0001) and smoking (OR 8.403, p=0.001). Mitral valve IE with a vegetation length ≥13 mm was the strongest risk factor for this complication (OR 3.24, p=0.001), followed by S. aureus infection (OR 3.59, p=0.002). Finally, septic shock (OR 6.000, p=0.001) represented the most important risk factor of in-hospital mortality. CONCLUSIONS: This study provides the most detailed profile of complication predictors related to left-sided IE in Central Europe. Early individual stratification of IE related occurrence of complications might help to decrease extremely high morbidity and mortality of this disease (Tab. 5, Ref. 37).

The role of VEGF in the diabetic patients undergoing endovascular therapy of symptomatic aortic valve stenosis.

The aim of this study was to explore changes in plasma vascular endothelial growth factor (VEGF) in aged patients who undergone transcatheter aortic valve implantation or balloon angioplasty for the treatment of aortic stenosis. Plasma VEGF was measured in subjects with diabetes mellitus type 2 (DM) (n=21, age 79.2+/-1.6 years) and in non-diabetic subjects (non-DM) (n=23, age 84.4+/-0.7 years), using an ELISA kit. Before the procedure plasma levels of VEGF were significantly lower in DM than in non-DM patients (P<0.05). Plasma VEGF significantly increased in both groups (DM and non-DM) 24 h (387+/-64 vs. 440+/-30 pg/ml, P<0.05) and 72 h (323+/-69 vs. 489+/-47 pg/ml, P<0.05) after the endovascular procedure. However, the VEGF in DM patients was significantly lower compared to non-DM subjects up to one month after the endovascular procedure (283+/-47 vs. 386+/-38 pg/ml, P<0.05). We conclude that increased plasma VEGF in aged patients associates with atherosclerotic aortic valve stenosis. In spite of that plasma VEGF in DM was constantly significantly lower than in non diabetic patients, both before and after the endovascular procedure, possibly reflecting a disturbance of angiogenic/anti-angiogenic balance in diabetes.

[Warning about risk of invasive infections in splenectomized patients. Experiences from University Hospital Brno, Czech Republic, in 2011]. / Upozornení na nebezpecí invazivních infekcí u splenektomovaných pacientu. Zkusenosti z FN Brno 2011.

Syndrome of fulminant sepsis in splenectomized (overwhelming postsplenectomy infection - OPSI) is feared and often fatal infectious complication in patients after splenectomy. The risk of syndrome of fulminant sepsis in splenectomized in these persons persists lifelong and doesn't diminish during the time. Etiologically, encapsulated bacterias like Streptococcus pneumoniae, Haemophilus influenzae group b and Neisseria meningitidis are involved. As the mortality of syndrome of fulminant sepsis in splenectomized is very high, it is indispensable to educate and vaccinate persons in risk. The authors present case reports of three splenectomized patients who were hospitalized for invasive pneumococcal infection in the University Hospital Brno, Czech Republic, in 2011.

Patient skin dosimetry in interventional cardiology in the Czech Republic.

In this study, skin dosimetry of patients undergoing interventional cardiology procedures is presented. Three hospitals were included. Two methods were used for skin dosimetry--radiochromic dosimetry films and reconstruction of skin dose distribution based on examination protocol. Maximum skin doses (MSD) obtained from both methods were compared for 175 patients. For patients for whom the film MSD was >1 Gy, the reconstruction MSD differed from the film MSD in the range of ± 50 % for 83 % of patients. For remaining patients, the difference was higher and it was caused by longer fluoroscopy time. For 59 patients for whom the cumulative dose was known, the cumulative dose was compared with the film MSD. Skin dosimetry with radiochromic films is more accurate than the reconstruction method, but films do not include X-ray fields from lateral projections whilest reconstructions do.

[Percutaneous coronary interventions in diabetic patients]. / Perkutánní intervencní výkony na vencitých tepnách u diabetiku.

Diabetes mellitus (DM) is a serious medical problem. The patients with diabetes have higher risk of coronary artery diseases. Diabetes mellitus also increases the risk of complication resulting from revascularization treatment in compare with non-diabetic patients. The first studies, which compare the surgical and percutaneous revascularization proved, that the major difference between these two revascularization techniques is in patients with diabetes. The difference is inflected by necessity of repeated revascularization after PCI, which is caused by higher incidence of a restenosis in patients with DM. Mortality, myocardial infarction and stroke are more frequent in diabetic patients, but the difference between PCI and surgical treated groups is not significant. Though the restenosis had significantly decreased due to of drug-eluted stents, the surgical revascularization is preferred method of treatment in patients with insulin-dependent diabetes, poorly compensated diabetes and patients with multiple complex coronary disease.

[Myocardial dysfunction in sepsis--diagnostics and therapy]. / Myokardiální dysfunkce v sepsi--diagnostika a terapie.

Incidence of myocardial dysfunction in studies with severe sepsis patients is up to two thirds of patients. On the other side, patients with normal echocardiography have some type of myocardial injury, which can be detected by elevated serum levels of troponins and natriuretic peptides. Strong prognostic value of these markers regarding morbidity and mortality of septic patients indicates an important role of this "occult" myocardial injury. Therapeutical interventions should take place only in situation in that low cardiac output is not capable to ensure metabolic demands of tissues. Nowadays, because of detrimental effects of classical inotropes, new strategies are under investigation. Namely levosimendan is promising alternative, not only related to its inotropic effects. Early diagnostics, assessment of prognosis and therapeutic strategy in patients with SMD are challenging for continuing research and for clinicians of different specialities.

Time-course of tissue factor plasma level in patients with acute coronary syndrome.

Enhanced expression of tissue factor (TF) may result in thrombosis contributing to acute clinical consequences of coronary artery disease. Several studies demonstrated elevated plasma levels of TF in patients with acute coronary syndrome (ACS). The aim of our study was to compare the concentrations of TF in coronary sinus (CS), proximal part of the left coronary artery (LCA) and peripheral vein (PV) of patients with ACS and stable coronary artery disease (SCAD). Time course of the TF plasma levels in PV was followed on day 1 and day 7 after index event of ACS presentation and was compared to day 0 values. No heparin was given prior to the blood sampling. Twenty-nine patients in the ACS group (age 63.6+/-10.8 years, 20 males, 9 females) and 24 patients with SCAD (age 62.3+/-8.1 years, 21 males, 3 females) were examined. TF plasma level was significantly higher in patients with ACS than in those with SCAD (239.0+/-99.3 ng/ml vs. 164.3+/-114.2 ng/ml; p=0.016). There was no difference in TF plasma levels in PV, CS and LCA (239.0+/-99.3 ng/ml vs. 253.7+/-131.5 ng/ml vs. 250.6+/-116.4 ng/ml, respectively). TF plasma levels tended to decrease only non-significantly on the day 7 (224.4+/-109.8 ng/ml). Significant linear correlation between TF and high sensitivity CRP (hs-CRP) levels on day 0 was found. In conclusion, TF plasma levels are elevated in patients with ACS not only locally in CS but also in systematic circulation. Our data support the relationship between TF production and proinflammatory mediators.

Neuroprotective use of mild hypothermia in patients with severe vasospasms after subarachnoid haemorrhage.

BACKGROUND: The authors describe two cases of patients with a severe subarachnoid haemorrhage, where mild hypothermia was successfully applied as a part of comprehensive neuroprotective therapy. PATIENTS: A 56-year-old patient was admitted to an intensive care unit with the diagnosis of subarachnoid haemorrhage, with a consciousness dysfunction requiring artificial ventilation. Angiography failed to establish the cause of the haemorrhage, even after repeated examinations. Furthermore, the authors describe the case of a 28-year-old woman with negative anamnesis and without long-term pharmacological medication, who was admitted to the hospital with a severe headache and a qualitative consciousness dysfunction. Angiography showed an aneurysm appearing closely above the left internal carotid artery bifurcation. After detecting vasospasms, mild hypothermia was repeatedly used in both patients, keeping the temperature of the body core between 34-34.5 degrees C. RESULTS: The total length of the introduced therapeutic hypothermia was 12 days in the first case and 6 days in the second case. The method used was non-invasive all-body cooling by means of blankets with circulating cooling liquid (Blanketrol II, Cinncinnati Sub Zero). In both cases the computed tomography findings and the clinical conditions gradually improved and the patients were released from the intensive care unit on the 22nd and 30th day, respectively, following the disorder detection. DISCUSSION: Mild hypothermia is a clinically attainable neuroprotective method, which--in combination with other therapeutic measures--led to minimising the neurological deficit in patients with severe subarachnoid haemorrhage (Ref. 11).

Plasma tissue factor in coronary artery disease: further step to the understanding of the basic mechanisms of coronary artery thrombosis.

Tissue factor is a cell surface protein that is expressed constitutively by monocytes, macrophages and fibroblasts, but also by some other cells in response to a variety of stimuli. The main function of the tissue factor is to form a complex with factor VII/VIIa that converts factors IX and X to their active forms. Tissue factor is also involved in the pathophysiology of systemic inflammatory disorders, coagulopathies, atherosclerotic disease, tumor angiogenesis and metastasis. Increased tissue factor expression either locally in the coronary plaques or systematically on circulating blood elements of patients with acute coronary syndromes may be responsible for increased thrombin generation, thus leading to platelet activation and fibrin formation. Tissue factor therefore plays a pivotal role in the initiation of thrombotic complications in patients with coronary artery disease.

Prevalence of normal coronary angiography in the acute phase of suspected ST-elevation myocardial infarction: experience from the PRAGUE studies.

BACKGROUND: Acute ST-elevation myocardial infarction in patients with normal coronary arteries has previously been described, but coronary angiography in these patients was performed after the acute phase of the infarction. It is possible that these patients did not have normal angiograms during the acute phase (transient coronary thrombosis or spasm were usually suspected to be the cause). Information on the prevalence of truly normal coronary angiograms during the acute phase of a suspected ST-elevation myocardial infarction is lacking. PATIENTS AND METHODS: The Primary Angioplasty in patients transferred from General community hospitals to specialized PTCA Units with or without Emergency thrombolysis-1 (PRAGUE-1) and PRAGUE-2 studies enrolled 1150 patients with ST-elevation acute myocardial infarction, in whom 625 coronary angiograms were performed within 2 h of the initial electrocardiogram. A simultaneous registry included an additional 379 coronary angiograms performed during the ST-elevation phase of a suspected myocardial infarction. Thus, a total of 1004 angiograms were retrospectively analyzed. A normal coronary angiogram was defined as one with the absence of any visible angiographic signs of atherosclerosis, thrombosis or spontaneous spasm. RESULTS: Normal coronary angiograms were obtained for 26 patients (2.6%). Among these, the diagnosis at discharge was a small myocardial infarction in seven patients (0.7%), acute (peri)myocarditis in five patients, dilated cardiomyopathy in four patients, hypertension with left ventricular hypertrophy in three patients, pulmonary embolism in two patients and misinterpretation of the electrocardiogram (ie, no cardiac disease) in five patients. Seven patients with small infarctions underwent angiography within 30 min to 90 min of complete relief of the signs of acute ischemia, and thus, angiograms during pain were not taken. None of the 898 patients catheterized during ongoing symptoms of ischemia had a normal coronary angiogram. Spontaneous coronary spasm as the only cause (without underlying coronary atherosclerosis) for the evolving infarction was not seen among these 898 patients. Thus, the causes of the seven small infarcts in patients with normal angiograms remain uncertain. CONCLUSIONS: The observed prevalence of normal coronary angiography in patients presenting with acute chest pain and ST elevations was 2.6%. Most of these cases were misdiagnoses, not infarctions. A normal angiogram during a biochemically confirmed infarction is extremely rare (0.7%) and was not seen during the ongoing symptoms of ischemia.

[Ischemia-modified albumin: new marker of myocardial ischemia?]. / Ischémií modifikovaný albumin: nový marker ischémie myokardu.

The Albumin Cobalt Binding Test is a quantitative in vitro diagnostic test used on human serum that detects ischemia-modified albumin by measuring the cobalt binding capacity of albumin in human serum. Ischemia modified albumin is intended for use in conjunction with ECG and cardiac troponin as an aid to short term risk stratification of patients presenting with chest pain suggestive of cardiac origin. Thus, in patients with chest pain or equivalent symptoms suggestive of cardiac origin, with non-diagnostic ECG and normal troponin, a negative IMA can be used as an aid to rule out acute coronary syndrome (ACS) in low risk patients.

Safety and dose relationship of recombinant human activated protein C for coagulopathy in severe sepsis.

OBJECTIVES: To assess the safety and effect on coagulopathy of a range of doses of recombinant human activated protein C (rhAPC). To determine an effective dose and duration of rhAPC for use in future clinical trials. DESIGN: Double-blind, randomized, placebo-controlled, multicenter, dose-ranging (sequential), phase II clinical trial. SETTING: Forty community or academic medical institutions in United States and Canada. PATIENTS: One hundred thirty-one adult patients with severe sepsis. INTERVENTIONS: Intravenous infusion of rhAPC (12, 18, 24, or 30 microg/kg/hr) or placebo for 48 or 96 hrs. MEASUREMENTS AND MAIN RESULTS: No significant differences in incidence of serious bleeding events (4% rhAPC, 5% placebo, p >.999) or incidence of serious adverse events (39% rhAPC, 46% placebo, p = 0.422) between rhAPC- and placebo-treated patients were observed. One of 53 rhAPC-treated patients with suitable immunogenicity samples had a low level, transient, non-neutralizing anti-APC antibody response not associated with any clinical adverse event. Significant dose-dependent decreases in both D-dimer (p <0.001) and end of infusion interleukin 6 levels (p =.021) were demonstrated. No statistically significant effects on fibrinogen or platelet counts were observed. A nonstatistically significant 15% relative risk reduction in 28-day all-cause mortality was observed between rhAPC- and placebo-treated patients. CONCLUSIONS: rhAPC was safe and well-tolerated and demonstrated a dose-dependent reduction in D-dimer and interleukin 6 levels relative to placebo. The dose of 24 microg/kg/hr for 96 hrs was selected for use in future clinical studies.

[Ventricular fibrillation in chronic heart disease]. / Komorová fibrilace u chronických onemocnení srdce.

The objective of the work was to describe in subjects with spontaneous ventricular fibrillation, after elimination of acute cardiac disease, the strategy of antiarrhythmic treatment and to evaluate, based on prospective follow-up, the effectiveness of this treatment. The authors included in the group 36 patients (30 men and 6 women) within the range from 34 to 78 years (mean age 58 +/- 11 years) with spontaneous ventricular fibrillation. They divided the group into a subgroup (15 subjects) without revascularization of the heart muscle, into a subgroup (17 subjects) with revascularization of the myocardium (coronary angioplasty and bypasses) and a subgroup (4 subjects) where ischaemic heart disease was ruled out (mostly cardiomyopathies). In all subgroups they used programmed ventricular stimulation (apparatuses of Quinton Co. USA, Biotronik Co. GFR), in the subgroup with revascularization within 3 months. During the diagnostic procedure of ventricular stimulation they tested antiarrhythmic drugs most frequently amiodarone per os (for 4 weeks). An implantable cardioverter--defibrillator was implanted in 17 patients (8 subjects without revascularization, 6 subjects with revascularization, 3 subjects without ischaemic heart disease). All patients were followed up till death, maximum 24 months. The authors evaluated the rate of cardiac deaths (death on cardiac grounds, incl. sudden arrhythmic death) and sudden arrhythmic deaths (within one hour after the onset of symptoms or the first malignant ventricular tachyarrhythmia recorded after implantation of the defibrillator). In the subgroup without revascularization with electric instability of the ventricles according to programmed stimulation 66.7% they described seven cardiac deaths (46.7%) and 6 sudden "arrhythmic" deaths (40%) incl. 5 subjects with ineffective testing of antiarrhythmic drugs. Conversely in the subgroup with revascularization and with diagnostic programmed stimulation in 47.1% they found 3 cardiac deaths (17.7%), one sudden "arrhythmic" death (5.9%)--a subject with ineffective testing. In the subgroup without ischaemic heart disease they recorded cardiac and sudden "arrhythmic" deaths in half the subjects, in all instances in subjects without inducible ventricular tachyarrhythmia. The authors found in the course of a two-year investigation a relapse of cardiac arrest in 25% of subjects after spontaneous ventricular fibrillation. A third of these subjects (all without a cardioverter-defibrillator) died. They confirm the benefit of implantation of a defibrillator for all subjects regardless of the basic diagnosis and revascularization of the heart muscle.

Regulation of interleukin-1-stimulated GMCSF mRNA levels in human endothelium.

The regulation of interleukin-1 (IL-1)-mediated increases in GMCSF mRNA levels in human endothelium was examined and determined to occur in a time- and protein kinase C (PKC)-dependent manner. IL-1beta induced the early activation and translocation of PKC isotypes alpha and beta2 to the nucleus and PKC inhibition attenuated the IL-1-mediated increase in GMCSF mRNA levels. PKC activation by PMA alone, in the absence of IL-1beta activation, however, was insufficient to allow GMCSF mRNA detection. Increasing cyclic adenosine nucleotide (cAMP) levels suppressed IL-1beta-induced increases in GMCSF mRNA levels. In contrast, botulinum toxin C, which mediates the ADP ribosylation of a 21 kD ras-related G protein, augmented IL-1beta-induced GMCSF mRNA expression. Inhibition of protein synthesis (with cycloheximide) raised basal GMCSF mRNA transcripts to detectable levels, augmented IL-1-induced increases in GMCSF mRNA levels, and exhibited negative regulation by cAMP. Finally, disruption of either microtubules (with colchicine) or microfilaments (with cytochalasin B) resulted in reduced GMCSF mRNA expression in response to IL-1beta. These results are compatible with a model wherein IL-1-mediated increases in human endothelial cell GMCSF mRNA may be linked to both nuclear protein kinase C activation and activation of a low molecular weight G-protein, although neither activity alone is sufficient to increase the levels of GMCSF mRNA.

[Changes in platelet activity in invasive cardiologic procedures]. / Zmeny aktivity desticek pri invazivních kardiologických výkonech.

The authors revealed some changes in the platelet activity in patients with invasive cardiological procedures. Changes of the platelet function were manifested by an enhanced aggregation of platelets in vivo, an increased secretion from alpha granules and increased release of prostaglandin metabolites from platelets and from the vascular wall. Acetylsalicylic acid (ASA) suppressed the formation of circulating platelet aggregates in vivo, but the platelet activity was manifested by another mechanism, independent on ASA. The authors recorded therefore an increase of prostaglandin metabolites and PF4 even in patients who were treated with ASA before the invasive examination.

Mechanisms of pertussis toxin-induced barrier dysfunction in bovine pulmonary artery endothelial cell monolayers.

We have previously characterized several G proteins in endothelial cells (EC) as substrates for the ADP-ribosyltransferase activity of both pertussis (PT) and cholera toxin and described the modulation of key EC physiological responses, including gap formation and barrier function, by these toxins. In this study, we investigated the mechanisms involved in PT-mediated regulation of bovine pulmonary artery endothelial cells barrier function. PT caused a dose-dependent increase in albumin transfer, dependent upon action of the holotoxin, since neither the heat-inactivated PT, the isolated oligomer, nor the protomer induced EC permeability. PT-induced gap formation and barrier dysfunction were additive to either thrombin- or thrombin receptor-activating peptide-induced permeability, suggesting that thrombin and PT utilize distinct mechanisms. PT did not result in Ca2+ mobilization or alter either basal or thrombin-induced myosin light chain phosphorylation. However, PT stimulated protein kinase C (PKC) activation, and both PKC downregulation and PKC inhibition attenuated PT-induced permeability, indicating that PKC activity is involved in PT-induced barrier dysfunction. Like thrombin-induced permeability, the PT effect was blocked by prior increases in adenosine 3',5'-cyclic monophosphate. Thus PT-catalyzed ADP-ribosylation of a G protein (possibly other than Gi) may regulate cytoskeletal protein interactions, leading to EC barrier dysfunction.

Phosphatidic acid directly activates endothelial cell protein kinase C.

The metabolism of phosphatidic acid (PA) yields diacylglycerol (DAG), a known activator of protein kinase C (PKC). To examine potential direct effects of PA on PKC activation, PKC purified from bovine pulmonary artery endothelial cells (BPAEC) was utilized in an in vitro assay examining gamma-[32P]ATP phosphorylation of H1 histone. In the presence of Ca2+ and phosphatidylserine (PS), DAG (80 microM) produced maximal PKC activity (6.4 pmol gamma-[32P]ATP incorporated/microgram/min). Dioleoyl-PA (80 microM) and 1-stearoyl,2-arachidonyl-PA (80 microM) activated PKC in a concentration-dependent manner (maximal activity of 2.01 +/- 0.1 pmol/microgram/min). Unlike unlabelled phorbol esters or DAG, dioleoyl-PA did not significantly alter the binding of [3H]-phorbol dibutyrate to PKC, suggesting that PA directly activates endothelial cell PKC in a manner distinct from DAG-mediated PKC activation.

Hypoxic pulmonary hypertension: haemodynamic changes after oxygen and various vasodilators in the same patient at different time periods.

The haemodynamic response to oxygen breathing and various vasodilatory drugs (Nifedipine, Ketanserin, Dihydralazine, Nitroglycerin, Iso-Mack, Nit-Ret) was repeatedly measured at different time intervals (4-24 months) in 10 males with hypoxic pulmonary hypertension due to chronic obstructive lung disease. It was found that the pulmonary artery mean pressure and pulmonary vascular resistance changes are stable in most of the patients, discordant reactions being exceptional and of small degree. The authors consider these findings important in the therapeutical decision making for longterm oxygen therapy and or vasodilatory drug administration.

Interleukin 1-stimulated prostacyclin synthesis in endothelium: lack of phospholipase C, phospholipase D, or protein kinase C involvement in early signal transduction.

The cascade of transmembrane signaling events that follow the occupancy of the interleukin 1 receptor remain poorly defined. We examined potential postreceptor transduction systems involved in human recombinant interleukin 1-beta-stimulated prostacyclin synthesis in human umbilical vein endothelium. Challenge of human umbilical vein endothelium monolayers with recombinant interleukin 1-beta resulted in dose- and time-dependent tritiated arachidonate release and prostacyclin synthesis consistent with phospholipase A2 activation. Prostacyclin synthesis after interleukin 1-beta (10 ng/ml) was detected 4 hours after stimulation and peaked at 16 to 24 hours. To examine whether interleukin 1-beta produced early activation of a phosphoinositide-specific phospholipase C, human umbilical vein endothelium monolayers were labeled with tritiated-2-myoinositol and inositol polyphosphates recovered after interleukin 1-beta stimulation. In contrast to the potent agonist, alpha-thrombin, interleukin 1-beta failed to significantly increase inositol phosphate production when examined for up to 4 hours. The absence of a significant increase in the Cai++ secretagogue, IP3, was confirmed in human umbilical vein endothelium monolayers loaded with the Ca++ photoprotein probe aequorin. Basal aequorin luminescence was unaltered after interleukin 1-beta (0 to 2 hours), whereas both alpha-thrombin and Ca++ ionophore A23187 produced rapid rises in Cai++. The intracellular Ca++ antagonist BAPTA and the extracellular Ca++ chelator EGTA produced significant inhibition of interleukin 1-beta-stimulated prostacyclin generation at 4 to 8 hours, suggesting either an indirect inhibitory effect of these agents on phospholipase A2 activity or that an increase in Ca++ may be a late event in the transduction scheme after interleukin 1 stimulation. Interleukin 1-beta-stimulated protein kinase C, phospholipase D, and adenylyl cyclase activities (0 to 4 hours) were unchanged from controls. Despite the absence of increased plasma membrane protein kinase C activity up to 4 hours after interleukin 1, pretreatment of human umbilical vein endothelium monolayers with staurosporine or phorbol myristate acetate (18 hours) to reduce protein kinase C activities, significantly attenuated the interleukin 1-stimulated prostanoid responses at 16 hours but not at 4 hours. Furthermore, short (5 minute) pretreatment with phorbol myristate acetate dramatically augmented interleukin 1-mediated prostacyclin responses in synergistic fashion, suggesting that protein kinase C may modulate interleukin 1 signal transducing pathways. In summary, these studies suggest that interleukin 1-beta-mediated endothelial cell phospholipase A2 activity and prostacyclin synthesis occur via a novel transducing pathway that does not involve early activation of phospholipase C, phospholipase D, or adenylate cyclase.(ABSTRACT TRUNCATED AT 400 WORDS)

Protein kinase C phosphorylates caldesmon77 and vimentin and enhances albumin permeability across cultured bovine pulmonary artery endothelial cell monolayers.

Cytoskeletal protein (CSP) interactions are critical to the contractile response in muscle and non-muscle cells. Current concepts suggest that activation of the contractile apparatus occurs through selective phosphorylation by specific cellular kinase systems. Because the Ca(2+)-phospholipid-dependent protein kinase C (PKC) is involved in the regulation of a number of key endothelial cell responses, the hypothesis that PKC modulates endothelial cell contraction and monolayer permeability was tested. Phorbol myristate acetate (PMA), a direct PKC activator, and alpha-thrombin, a receptor-mediated agonist known to increase endothelial cell permeability, both induced rapid, dose-dependent activation and translocation of PKC in bovine pulmonary artery endothelial cells (BPAEC), as assessed by gamma-[32P]ATP phosphorylation of H1 histone in cellular fractions. This activation was temporally associated with evidence of agonist-mediated endothelial cell contraction as demonstrated by characteristic changes in cellular morphology. Agonist-induced activation of the contractile apparatus was associated with increases in BPAEC monolayer permeability to albumin (approximately 200% increase with 10(-6) MPMA, approximately 400% increase with 10(-8) M alpha-thrombin). To more closely examine the role of PKC in activation of the contractile apparatus, PKC-mediated phosphorylation of two specific CSPs, the actin- and calmodulin-binding protein, caldesmon77, and the intermediate filament protein, vimentin, was assessed. In vitro phosphorylation of both caldesmon and vimentin was demonstrated by addition of exogenous, purified BPAEC PKC to unstimulated BPAEC homogenates, to purified bovine platelet caldesmon77, or to purified smooth muscle caldesmon150. Caldesmon77 and vimentin phosphorylation were observed in intact [32P]-labeled BPAEC monolayers stimulated with either PMA or alpha-thrombin, as detected by immunoprecipitation. In addition, BPAEC pretreatment with the PKC inhibitor, staurosporine, prevented alpha-thrombin- and PMA-induced phosphorylation of both cytoskeletal proteins, attenuated morphologic evidence of contraction, and abolished agonist-induced barrier dysfunction. These results demonstrate that agonist-stimulated PKC activity results in cytoskeletal protein phosphorylation in BPAEC monolayer, an event which occurs in concert with agonist-mediated endothelial cell contraction and resultant barrier dysfunction.