SC lipid model membranes designed for studying impact of ceramide species on drug diffusion and permeation, part III: influence of penetration enhancer on diffusion and permeation of model drugs.

The impact of the lipophilic penetration enhancer, oleic acid (OA), on the barrier properties of stratum corneum (SC) lipid model membranes was investigated based on diffusion and permeation studies of model drugs covering a broad range of lipophilicities. Diffusion and permeation experiments of urea, caffeine and diclofenac sodium were conducted using Franz-type diffusion cells. HPLC and capillary electrophoresis techniques were employed to analyze the amount of permeated drug. An incorporation of OA to the SC lipid model membranes did not change the relation between the diffusion and permeation behavior of model drugs presented previously for SC lipid model membranes without OA. The fastest rate of diffusion through SC lipid model membranes occurred in the case of the most hydrophilic drug, urea. In the case of permeation studies of caffeine and diclofenac sodium across SC lipid model systems, the permeability parameters were either equal or slightly larger in favor of the most lipophilic drug, diclofenac sodium. OA had a pronounced impact on the barrier properties of SC lipid model membranes. It caused the impairment of the barrier function of the SC lipid model membrane with Cer [AP] (phytosphingosine-based ceramide), however, surprisingly improved the barrier properties of the SC lipid model system with Cer [EOS] (sphingosine-based acylceramide).

SC lipid model membranes designed for studying impact of ceramide species on drug diffusion and permeation--part II: diffusion and permeation of model drugs.

The barrier function of two quaternary stratum corneum (SC) lipid model membranes, which were previously characterized with regard to the lipid organization, was investigated based on diffusion studies of model drugs with varying lipophilicities. Diffusion experiments of a hydrophilic drug, urea, and more lipophilic drugs than urea (i.e. caffeine, diclofenac sodium) were conducted using Franz-type diffusion cells. The amount of permeated drug was analyzed using either HPLC or CE technique. The subjects of interest in the present study were the investigation of the influence of physicochemical properties of model drugs on their diffusion and permeation through SC lipid model membranes, as well as the study of the impact of the constituents of these artificial systems (particularly ceramide species) on their barrier properties. The diffusion through both SC lipid model membranes and the human SC of the most hydrophilic model drug, urea, was faster than the permeation of the more lipophilic drugs. The slowest rate of permeation through SC lipid systems occurred in the case of caffeine. The composition of SC lipid model membranes has a significant impact on their barrier function. Model drugs diffused and permeated faster through Membrane II (presence of Cer [EOS]). In terms of the barrier properties, Membrane II is much more similar to the human SC than Membrane I.

Long-term outcome of patients with mild intermittent claudication under secondary prevention.

BACKGROUND: The known natural history of peripheral arterial disease (PAD) is determined by the generalization of atherosclerosis with resulting high cardiovascular and cerebrovascular morbidity and mortality. The aim of this prospective study was to record all vascular and non-vascular events in patients with mild intermittent claudication (IC) undergoing secondary preventive measures in a 10-year follow-up and to determine the time-points at which systemic localizations of atherosclerotic events develop. PATIENTS AND METHODS: Patients who originally had an isolated Fontaine stage IIa PAD were included in the follow-up which involved assessments carried out on an annual basis. The incidence and timepoints of vascular (i.e., myocardial infarction, stroke, critical limb ischemia, vascular death), and nonvascular events (i.e., cancer, non vascular death) were recorded and compared with the known natural history. RESULTS: 534 events (vascular: 433) concerning 109 claudicants (M/F: 88/21; 60.8 +/- 8.8 years; ABI 0.66 +/- 0.11) were recorded over an average follow-up period of 104 months. 25.7% of the claudicants died, 39% due to vascular events, 36% due to cancer disease and 25% due to other events. A deterioration of PAD (n = 108) was the most frequent event after 20 months, followed by angina pectoris (n = 41) and cancer diseases (n = 20) after 42 and 45 months, stroke (n = 19) after 58 months, myocardial infarction (n = 12) after 63 months, and finally critical limb ischemia (n = 27) and amputations (n = 10) after 80 and 114 months (median). 111 revascularizations were carried out. 62.3% of the claudicants developed a polyvascular disease, with 20% in a trivascular territory. CONCLUSIONS: A high vascular comorbidity also develops under secondary prevention as an expression of the continuing generalization of the atherosclerotic process in PAD. The causes for death are determined both by the vascular and the tumor related comorbidity.

Prevention of deep-vein thrombosis after total hip and knee replacement. Low-molecular-weight heparin in combination with intermittent pneumatic compression.

After total hip (THR) or knee replacement (TKR), there is still an appreciable risk of developing deep-vein thrombosis despite prophylaxis with low-molecular-weight heparin (LMWH). In a prospective, randomised study we examined the efficacy of LMWH in combination with intermittent pneumatic compression in patients undergoing primary unilateral THR or TKR. We administered 40 mg of enoxaparin daily to 131 patients combined with either the use of intermittent pneumatic compression or the wearing of graduated compression stockings. Compression ultrasonography showed no evidence of thrombosis after LMWH and intermittent pneumatic compression. In the group with LMWH and compression stockings the prevalence of thrombosis was 28.6% (40% after TKR, 14% after THR). This difference was significant (p < 0.0001). In the early post-operative phase after THR and TKR, combined prophylaxis with LMWH and intermittent pneumatic compression is more effective than LMWH used with graduated compression stockings.

Hyperhomocysteinemia: marker of systemic atherosclerosis in peripheral arterial disease.

AIM: Patients suffering from peripheral arterial disease (PAD) are increasingly described as having hyperhomocysteinemia more than in patients with coronary artery or cerebrovascular disease. Cases of symptomatic PAD usually present with associated coronary artery or cerebrovascular disease and renal artery disease. It can thus be postulated that multilocular atherosclerosis is linked to hyperhomocysteinemia and that the extent of atherosclerosis has a possible correlation with homocysteine concentrations. The aim of this study was to ascertain whether fasting total homocysteine concentrations in patients with PAD are associated with the extent and the localization of systemic atherosclerosis in cerebrovascular, coronary and/or renal vascular zones. METHODS: A total of 183 patients with PAD, Fontaine stages II-IV, were divided into 2 groups: Group A contained patients with isolated PAD (n=98) and Group B patients with systemic atherosclerosis in PAD (n=85). Characterization of vascular disease in various vascular zones was indication-adapted using non-invasive and/or invasive METHODS: Patients with renal insufficiency were excluded from the study. RESULTS: Homocysteine concentrations were significantly lower in patients with isolated PAD than in patients with additional systemic atherosclerosis (10.1+/-4.4 vs 16.7+/-7.04 micromol/l, p<0.0001). There were no differences in localization or extent of concomitant systemic atherosclerosis. Logistic regression analysis indicated that elevated plasma homocysteine and decreasing ABPI served independently as significant risk indicators for systemic atherosclerosis in patients with PAD (p<0.0001). CONCLUSION: Hyperhomocysteinemia is a precursoral marker of systemic atherosclerosis and thus a prognostic indicator of cardiovascular morbidity and mortality in PAD.

[Cerebrovascular insufficiency--1. How to reduce the risk of stroke]. / Zerebrovaskuläre Insuffizienz--Teil 1. Wie Sie das Schlaganfallrisiko minimieren können.

Extracranial stenosis of the carotid artery may lead to cerebral ischemic events and stroke. For this reason, such symptoms as transient ischemic attacks (TIA), unilateral syndrome or amaurosis fugax, the arteries feeding the brain should be investigated. In addition to bilateral palpation and auscultation of the arteries, modern ultrasonographic methods have an important role to play. The aim of treatment is secondary prevention of ischemic stroke, and possibly even to achieve regression of the vascular lesion. This necessitates the elimination of atherogenic risk factors. Particularly suitable as preventive medication, are platelet aggregation inhibitors, such as, acetylsalicylic acid and clopidogrel, which can also be applied postoperatively and postinterventionally to prevent recurrence.

Calcium waves in agarose gel with cell organelles: implications of the velocity curvature relationship.

Calcium oscillations and waves have been observed not only in several types of living cells but also in less complex systems of isolated cell organelles. Here we report the determination of apparent Ca2+ diffusion coefficients in a novel excitable medium of agarose gel with homogeneously distributed vesicles of skeletal sarcoplasmic reticulum. Spatiotemporal calcium patterns were visualized by confocal laser scanning fluorescence microscopy. To obtain characteristic parameters of the velocity curvature relationship, namely, apparent diffusion coefficient, velocity of plane calcium waves, and critical radius, positively and negatively curved wave fronts were analyzed. It is demonstrated that gel-immobilized cell organelles reveal features of an excitable medium. Apparent Ca2+ diffusion coefficients of the in vitro system, both in the absence or in the presence of mitochondria, were found to be higher than in cardiac myocytes and lower than in unbuffered agarose gel. Plane calcium waves propagated markedly slower in the in vitro system than in rat cardiac myocytes. Whereas mitochondria significantly reduced the apparent Ca2+ diffusion coefficient of the in vitro system, propagation velocity and critical size of calcium waves were found to be nearly unchanged. These results suggest that calcium wave propagation depends on the kinetics of calcium release rather than on diffusion.

Angiotensin-converting enzyme gene insertion/deletion polymorphism and carotid artery wall thickness in patients with peripheral arterial occlusive disease.

BACKGROUND: It has been suggested that the deletion polymorphism of the angiotensin converting enzyme (ACE) gene is linked to a high risk of cardiovascular disease. The relationship between the insertion/deletion (I/D) polymorphism of the ACE gene and the carotid intima-media thickness in patients with peripheral arterial occlusive disease is unknown. We tested the hypothesis that the early progression of atherosclerosis in the extracranial carotid arteries in patients with peripheral arterial disease is associated with a genetic predisposition. METHODS: This prospective trial included 98 patients who only had manifestations of arteriosclerotic disease in peripheral arterial vascular regions of the lower extremities (stable stage II PAOD). Maximal common carotid intima-media thickness (mIMT) was measured using high resolution B-mode ultrasonography. Determinations of ACE gene polymorphism were made using a polymerase chain reaction technique. Multivariate regression analysis was performed to assess the influence of ACE genotypes, ACE activity and vascular risk factors on intima-media thickness. RESULTS: There was no significant association between intima-media thickness and ACE gene polymorphism. History of symptomatic peripheral arterial disease without local or systemic progression exists in subjects with the II-genotype significantly longer than in subjects with the DD genotype (p=0.01). With the presence of an II-genotype, there was also a tendency towards a thinner intima-media thickness. We found significant correlations between intima-media thickness and age (p<0.0001), fasting serum insulin (p=0.001), and lipoprotein (a) (p=0.008). CONCLUSIONS: In the present study involving patients with stage II peripheral arterial occlusive disease, ACE gene polymorphism could not be identified as a determining marker for the development of intima-media thickening in the common carotid artery. However, it can be assumed that there is a reduced risk for the systemic progression of atherosclerosis in patients with the II genotype.

Sarcoplasmic reticulum vesicles embedded in agarose gel exhibit propagating calcium waves.

In different cell types, activation of signal transduction pathways leads to the generation of calcium oscillations and/or waves. Due to this important impact for cellular function, calcium waves are the subject of intensive investigations. To study interactions of cell organelles with no influence of the cell membrane, sarcoplasmic reticulum (SR) vesicles and well-coupled mitochondria were reconstituted. For the first time, we demonstrate the generation and propagation of calcium waves in a suspension of sarcoplasmic reticulum vesicles, embedded in an agarose gel. The propagation dynamics resemble those of calcium waves in living cells. Moreover, the addition of well-coupled mitochondria leads to more pronounced and significantly faster propagating waves, demonstrating the importance of the mitochondrial Ca(2+) transport. The experimental and simulation results indicate the resemblance of the in vitro system to an excitable medium.

[Ultrasound image of the hypothenar hammer syndrome]. / Das sonographische Bild des Hypothenar-Hammer-Syndroms.

UNLABELLED: A traumatic lesion of the ulnar artery (Hypothenar Hammer Syndrome, HHS) is a very rare cause of acral ischemia of the upper extremity. AIM: Evaluation of the efficacy of ultrasound techniques in the diagnosis of the HSS. METHOD: Based on the knowledge of the patient's history and clinical examination we investigated forearm, hand and digital arteries by continuous wave Doppler. By means of colour-coded duplex sonography (CCDS) we demonstrated the course of the radial and ulnar arteries to the palmar arch. For comparison we used the results of preintervention angiography. RESULTS: The diameter of the distal ulnary artery measured in healthy women was 1.8 +/- 0.32 and in men 2.2 +/- 0.46 mm. Among 268 patients with ischemia of the hands we diagnosed HHS in four cases. We identified an aneurysm of the ulnary artery in one case and a thrombotic occlusion in three cases. The thrombotic occlusion of the ulnar artery led to a dilatation of the vascular lumen. The occlusion length could be determined. Vessels diameter and echogenecity gave information about the age of the thrombosis. CONCLUSION: The continuous wave Doppler is a major diagnostic contribution because of its ability to register hemodynamic changes. Additionally, CCDS with its combination of sonomorphology and hemodynamics enables a distinction between HHS and other causes of digital ischemia. The CCDS more precisely refines the indications for preinterventional angiography in acute ischemia syndromes of the hand.

Angiotensin-converting enzyme gene insertion/deletion polymorphism and peripheral arterial occlusive disease.

BACKGROUND: The deletion polymorphism of the ACE gene is linked to a high risk of cardiovascular disease due to the permanent activation of the local and systemic renin-angiotensin systems (RAS). The aim of this prospective study was 1. to compare the ACE insertion/deletion polymorphism in individuals with a healthy vasculature with that of patients suffering from peripheral arterial occlusive disease (PAOD), and 2. to determine whether associations existed between specific clinical parameters and the ACE genotype which the PAOD patients expressed. PATIENTS AND METHODS: Determinations of ACE I/D gene polymorphism were made using a polymerase chain reaction (PCR) technique on 98 patients with clinical stage II PAOD according to Fontaine and 240 healthy individuals who served as controls. All patients and controls came from central Germany. Clinical variables which included duration of clinical symptoms, a familial history of the disease, arteriosclerosis score (ASF, providing an estimate of the extent of atheromatosis at femoral artery bifurcation) and plasma ACE activity were correlated with the genotypes taking the cardiovascular disease risk factors which were present into consideration. RESULTS: Differences in ACE genotypes between patients with PAOD (D/I: 0.57/0.43) and control group individuals (D/I: 0.59/0.41) were not observed. In comparison with the II genotype, the DD genotype was associated with a shorter duration of disease (p = 0.01), a positive family medical history (p = 0.022) and a higher plasma ACE activity (p = 0.026). The ASF did not correlate with the ACE I/D gene polymorphism. CONCLUSION: Evidence that the deletion allele is linked to the manifestation of PAOD could not be found in the patients studied. One can assume, however, that the deletion allele has a progression promoting effect on the disease.

Aspirin increases ferritin synthesis in endothelial cells: a novel antioxidant pathway.

Aspirin has recently been shown to increase endothelial resistance to oxidative damage. However, the mechanism underlying aspirin-induced cytoprotection is still unknown. Using cultured cells, the present study investigates the effect of aspirin on the expression of ferritin, a cytoprotective protein that sequesters free cytosolic iron, the main catalyst of oxygen radical formation. In bovine pulmonary artery endothelial cells, aspirin at low antithrombotic concentrations (0.03 to 0.3 mmol/L) induced the synthesis of ferritin protein in a time- and concentration-dependent fashion up to 5-fold over basal levels, whereas ferritin H (heavy chain) mRNA remained unaltered. Aspirin-induced cytoprotection from hydrogen peroxide toxicity was mimicked by exogenous iron-free apoferritin but not iron-loaded ferritin, demonstrating the antioxidant function of newly synthesized ferritin under these conditions. Ferritin induction by aspirin was specific in that other nonsteroidal anti-inflammatory drugs such as salicylic acid, indomethacin, or diclofenac failed to alter ferritin protein levels. Aspirin-induced ferritin synthesis was abrogated in the presence of the iron chelator desferrioxamine, pointing to an interaction of aspirin with iron-responsive activation of ferritin translation. Together, our results suggest induction of ferritin as a novel mechanism by which aspirin may prevent endothelial injury in cardiovascular disease, eg, during atherogenesis.

[Quantitative sonography of peripheral arteriovenous malformations]. / Quantitative Sonographie peripherer arteriovenöser Malformationen.

UNLABELLED: Congenital vascular malformations occur in 1.5% of the general population. Depending on their localization and morphology, with dysfunction of local and central hemodynamics as well as tissue metabolism, congenital peripheral arteriovenous malformations (AVM) are of particular importance within the larger, heterogeneous group of diseases which comprise vascular malformations. AIM: The aim of this study was to evaluate our ability to quantity peripheral congenital AVM using sonographic, functional-hemodynamic, and morphological parameters. METHOD: Apart from obtaining a medical history and applying clinical and venous occlusion plethysmographic diagnostics, sonographic examinations of peripheral arteries and veins using cw-Doppler sonography and color-coded duplex sonography were performed on 8 patients with peripheral congenital AVM. For comparative purposes, the results of pre-intervention angiography examinations were also considered. RESULTS: For a sonographic quantification of AVM at the time of diagnosis, during assessments of diseases progress, and postinterventionally, both the resistance index of Pourcelot and measurement of the vascular blood flow velocities appeared to be particularly appropriate as functional-hemodynamic parameters. The sonomorphological parameters reflect the individual vascular pathology, which is determined by hemodynamic changes. CONCLUSION: Functional-hemodynamic and sonomorphologic parameters allow a quantification of AVM. With knowledge of the clinical picture acquired at the time, they provide the basis for deciding upon a more extensive pre-interventional invasive radiological diagnosis.

Aspirin protects endothelial cells from oxidative stress--possible synergism with vitamin E.

A 24-h incubation with hydrogen peroxide (0.65 mM) markedly reduced viability of cultured endothelial cells. Preincubation with aspirin (3-30 microM) protected endothelial cells from hydrogen peroxide-induced toxicity and increased viability in a concentration-dependent fashion by up to 64% of control. A similar protection was observed with D-alpha-tocopherol acetate (vitamin E, 3-30 microM). The cytoprotective effects of aspirin and vitamin E against hydrogen peroxide were overadditive suggesting different mechanisms of antioxidant action. In agreement with this, cytotoxicity induced by iron, the main catalyst of oxygen radical formation, was substantially reduced by aspirin but not vitamin E. These results show that aspirin protects endothelial cells from oxidative stress possibly via binding or chelation of free cytosolic iron. Moreover, a combination of aspirin and vitamin E might be useful for the prevention of endothelial injury in cardiovascular disease, e.g. during atherogenesis.