Comparison between Bioimpedance Analysis and Dual-Energy X-ray Absorptiometry in assessment of body composition in a cohort of elderly patients aged 65-90 years.

We compare bioimpedance analysis (BIA) with dual-energy X-ray absorptiometry (DXA) in the assessment of free fat mass (FFM), fat mass (FM) and percentage of body fat under different conditions in relation to age categories, hydration parameters, body mass index (BMI) and sarcopenia. A cross-sectional analysis of body composition was estimated by BIA and DXA in 379 hospitalized elderly patients. In addition, estimates of FFM, FM and percentage of body fat were investigated across different conditions. Paired t-tests, Bland-Altman plot and intraclass correlation coefficient analysis were used to compare methods. Data showed an underestimation of means (BIA versus DXA) of FFM (women: 0,97 kg, p<0,01; men: 1,99 kg; p<0,01), and an overestimation of both the FM (women: +1,11 kg; p<0,01; men: +1,67 kg; p<0,01) and percentage of body fat (women: +2,07 %, p<0,01; men: +2,82 %, p<0,01). BIA underestimated FFM and overestimated FM and percentage of body fat in patients from the age group of 75 to 85 years, in patients with a total body water content <60%, in underweight and normal weight patients and in patients with sarcopenia (p<0,01). The intraclass coefficient results were indicative of poor reproducibility between BIA and DXA for FFM (women: +0,197; men: +0,250) and FM (women: +0,141; men +0,144). BIA is a good alternative for estimation of FFM and FM only in overweight or obese patients or in patients with good hydration status. BIA, on the other hand, is not an accurate method for assessing FFM in sarcopenic patients.

Berberine phospholipid exerts a positive effect on the glycemic profile of overweight subjects with impaired fasting blood glucose (IFG): a randomized double-blind placebo-controlled clinical trial.

OBJECTIVE: Berberine is a plant alkaloid known to exert positive metabolic effects. Human studies have confirmed its ability to improve the lipid and glycemic profile. This study aimed to evaluate the potential benefit of oral supplementation of Berberine PhytosomeTM (2 tablets/day, 550 mg/tablet) on the metabolic profile of subjects with impaired fasting blood glucose (IFG). PATIENTS AND METHODS: A total of 49 overweight subjects, 28 females and 21 males, were randomly assigned to either the supplemented group (n=24) or placebo (n=25). We considered glycemia as the primary endpoint and total cholesterol, high-density lipoprotein (HDL), total cholesterol/HLD, low-density lipoprotein (LDL), LDL/HDL, triglycerides, insulin, glycated hemoglobin, Homeostasis Model Assessment (HOMA), ApoA, ApoB, ApoB/ApoA, androgen suppression treatment (AST), alternative lengthening of telomeres (ALT), gamma-glutamyl transferase (GGT), creatinine, and body composition by dual-energy X-ray absorptiometry (DXA) as secondary endpoints. These parameters have been assessed at baseline, after 30 days, and after 60 days. RESULTS: After two months of treatment, through the use of linear mixed effect models, a statistically significant difference between supplemented and placebo groups was observed for glycemia [ß=-0.2495% C.I. (-0.47; -0.06), p=0.004], total cholesterol [ß=-0.25, 95% C.I. (-0.45; -0.04), p=0.05], total cholesterol/HDL [ß=-0.25, 95% C.I. (-0.43; -0.06), p=0.04], triglycerides [ß=-0.14, 95% C.I. (-0.25; -0.02), p=0.05], insulin [ß=-1.78, 95% C.I. (-2.87; -0.66), p=0.009], ApoB/ApoA [ß=-0.08, 95% C.I. (-0.13; -03), p=0.004], Visceral adipose tissue (VAT) [ß=-91.50, 95% C.I. (-132.60; -48.19), p<0.0001] and fat mass [ß=-945.56, 95% C.I. (-1,424.42; -441.57), p=0.004]. CONCLUSIONS: The use of berberine had no adverse events, supporting its use as a natural alternative to pharmacological therapies in the case of IFG.

Transgenic activation of Ras in neurons promotes hypertrophy and protects from lesion-induced degeneration.

Ras is a universal eukaryotic intracellular protein integrating extracellular signals from multiple receptor types. To investigate its role in the adult central nervous system, constitutively activated V12-Ha-Ras was expressed selectively in neurons of transgenic mice via a synapsin promoter. Ras-transgene protein expression increased postnatally, reaching a four- to fivefold elevation at day 40 and persisting at this level, thereafter. Neuronal Ras was constitutively active and a corresponding activating phosphorylation of mitogen-activated kinase was observed, but there were no changes in the activity of phosphoinositide 3-kinase, the phosphorylation of its target kinase Akt/PKB, or expression of the anti-apoptotic proteins Bcl-2 or Bcl-X(L). Neuronal Ras activation did not alter the total number of neurons, but induced cell soma hypertrophy, which resulted in a 14.5% increase of total brain volume. Choline acetyltransferase and tyrosine hydroxylase activities were increased, as well as neuropeptide Y expression. Degeneration of motorneurons was completely prevented after facial nerve lesion in Ras-transgenic mice. Furthermore, neurotoxin-induced degeneration of dopaminergic substantia nigra neurons and their striatal projections was greatly attenuated. Thus, the Ras signaling pathway mimics neurotrophic effects and triggers neuroprotective mechanisms in adult mice. Neuronal Ras activation might become a tool to stabilize donor neurons for neural transplantation and to protect neuronal populations in neurodegenerative diseases.

Natural Killer Response and Lipo-Metabolic Profile in Adults with Low HDL-Cholesterol and Mild Hypercholesterolemia: Beneficial Effects of Artichoke Leaf Extract Supplementation.

The aim of the present study is to evaluate the effects of 60-day artichoke leaf extract (ALE) supplementation (250mg, twice daily) on cytokines levels, natural killer cell (NK) response, and lipo-metabolic profile (HDL, LDL, and total-cholesterol, triglycerides (TG), ApoB, ApoA, lipid accumulation product (LAP), glucose, insulin, and homeostasis model assessment of insulin resistance (HOMA-IR)) in twenty adults (9/11 males/females, age=49.10 ± 13.74 years, and BMI=33.12 ± 5.14 kg/m2) with low HDL-C and mild hypercholesterolemia. Hierarchical generalized linear model, adjusted for sex, BMI, and age, has been used to evaluate pre-post treatment changes. A significant increase for HDL-C (ß=0.14, p=0.0008) and MCP-1 (ß=144.77, p=0.004) and a significant decrease for ApoB/ApoA (ß=-0.07, p=0.03), total-C/HDL-C ratio (ß=-0.58, p<0.001), and NK response at stimulus low (ß=0.43, p=0.04), medium (ß=0.40, p<0.001), and high (ß=0.42, p=0.001) have been found. These results support the benefits of ALE supplementation on metabolic profile.

A novel boswellic acids delivery form (Casperome®) in the management of musculoskeletal disorders: a review.

Standard pharmacological treatment of musculoskeletal conditions is often associated with relevant side effects. Botanical preparations endowed with a good tolerability profile, therefore, could have a role in the management of these disorders. Among different natural products, Boswellia serrata extracts have long been used for the treatment of musculoskeletal disorders, given their marked anti-inflammatory activity and their ability to promote tissue regeneration. However, standard preparations of Boswellia serrata show overall modest pharmacokinetic properties, a limitation which may ultimately lead to reduced efficacy. In an effort to improve the pharmacokinetic properties, Casperome®, a lecithin-based formulation of Boswellia serrata extract representing the whole natural bouquet, has been developed. This formulation was effective in the treatment of Achilles tendonitis, epicondylitis, radiculopathies, ankle sprains and sport injuries as shown in several clinical studies, the majority of which with a randomized design and all evaluating a number of well-recognized parameters of efficacy for the therapy of musculoskeletal disorder. All studies were consistent in showing a prompt decrease of pain and improvement of functionality of the affected area after supplementation with Casperome®, without any relevant adverse effect. Remarkably, these symptomatic improvements were paralleled by reduced plasmatic levels of inflammatory markers and by a diminished need for rescue analgesics. On these bases, Casperome® may have a role in the treatment of musculoskeletal disorders. Clinical studies in other similar conditions (e.g., osteoarthritis) appear warranted to further investigate the efficacy of this botanical product in more specific settings.

Partitioning of local anesthetics into membranes: surface charge effects monitored by the phospholipid head-group.

The binding of the charged form of two local anesthetics, dibucaine and etidocaine, to bilayers composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) was measured simultaneously with ultraviolet spectroscopy and deuterium magnetic resonance. Because of their amphiphilic molecular structure, both drugs intercalate between the lipid molecules, increasing the surface area and imparting a positive electric charge onto the membrane. The ultraviolet (UV) binding isotherms were therefore analyzed in terms of a model which specifically took into account the bilayer expansion as well as the charge-induced concentration variations near the membrane surface. By formulating a quantitative expression for the change in surface area upon drug intercalation and combining it with the Gouy-Chapman theory, the binding of charged dibucaine and etidocaine to the lipid membrane was best described by a partition equilibrium, with surface partition coefficients of 660 +/- 80 M-1 and 11 +/- 2 M-1 for dibucaine and etidocaine, respectively (pH 5.5, 0.1 M NaCl/50 mM buffer). Deuterium magnetic resonance demonstrated further that the binding of drug changed the head-group conformation of the lipid molecules. Invoking the intercalation model, a linear variation of the deuterium quadrupole splittings of the choline segments with the surface charge density was observed, suggesting that the phosphocholine head-group may act as a 'molecular electrometer' with respect to surface charges.

Group-directed modification of bacteriorhodopsin by arylisothiocyanates. Labeling, identification of the binding site and topology.

Group-directed hydrophobic modification of membrane-integrated protein segments by arylisothiocyanates is applied to bacteriorhodopsin. Labeling of purple membrane with phenylisothiocyanate and 4-N,N'-dimethylamino-azobenzene-4'-isothiocyanate results in covalent modification of a unique lysine epsilon-amino group of bacteriorhodopsin. Lysine residue 41, located in the amino-terminal chymotryptic fragment, has been identified as the arylisothiocyanate binding site by established sequencing techniques. The phenylisothiocyanate binding site is not accessible for the aqueously soluble analog p-sulfophenylisothiocyanate. Furthermore, the acid-induced bathochromic shift of the bound chromophore reagent is not observed following acidification of 4-N,N'-dimethylamino-azobenzene-4'-isothiocyanate-labeled purple membrane. The modification thus occurs in the hydrophobic membrane domain, providing further evidence for intramembraneous disposition of the modified protein segment. Light-induced proton translocation is preserved in reconstituted vesicles containing either phenylisothiocyanate-modified or 4-N,N'-dimethylamino-azobenzene-4'-isothiocyanate-modified bacteriorhodopsin.

Bradykinin improves postischaemic recovery in the rat heart: role of high energy phosphates, nitric oxide, and prostacyclin.

OBJECTIVE: The aim was to define: (1) whether bradykinin administration during reperfusion improves postischaemic myocardial recovery; (2) whether high energy phosphate compounds are involved in the protective effects of bradykinin; and (3) whether bradykinin-induced release of prostacyclin and nitric oxide mediate the protective effects of bradykinin. METHODS: In the Langendorff rat heart preparation, coronary flow, left ventricular developed pressure, and, using 31P magnetic resonance spectroscopy, the high energy phosphate compounds phosphocreatine and beta-ATP were assessed during 15 min of global ischaemia and 30 min of reperfusion. Administration of 10(-7) M bradykinin was started before ischaemia and maintained throughout the experiment (BK-pre). This was compared to 10(-7) M bradykinin given exclusively with reperfusion (BK-post). Then 10(-7) M bradykinin was given simultaneously with 10(-4) M N omega-nitro-L-arginine-methyl ester (BK-LNAME) or 10(-5) M indomethacin (BK-indo). RESULTS: In comparison to control hearts, BK-pre exerted a significant protective effect on the postischaemic recovery of coronary flow [71(5)% v 43(4)%, P < 0.05], left ventricular pressure [81(8)% v 42(5)%, P < 0.05], phosphocreatine [105(4)% v 67(8)%, P < 0.05], and beta-ATP [78(9)% v 48(7)%, P < 0.05]. With BK-post, recovery of coronary flow [71(4)% v 43(4)%, P < 0.05] and left ventricular pressure [78(4)% v 42(5)%, P < 0.05] significantly improved; however the recovery of phosphocreatine [70(4)% v 67(8)%, NS] and beta-ATP [58(2)% v 48(7)%, NS] was not different from control. When bradykinin and L-NAME or indomethacin was given the beneficial effects of bradykinin on ischaemic hearts were abolished. CONCLUSIONS: (1) Bradykinin improved postischaemic myocardial recovery when given before ischaemia or starting exclusively with reperfusion; (2) this was only partially related to a protective action on the high energy phosphate compounds during ischaemia; (3) the beneficial effects of bradykinin on ischaemic hearts are dependent from an unrestrained action of prostacyclin and nitric oxide.

Left ventricular wall stress and sarcoplasmic reticulum Ca(2+)-ATPase gene expression in renal hypertensive rats: dose-dependent effects of ACE inhibition and AT1-receptor blockade.

BACKGROUND: Cardiac hypertrophy is associated with altered Ca2+ handling and may predispose to the development of LV dysfunction and cardiac failure. At the cellular level, the re-expression of ANF represents a well-established marker of myocyte hypertrophy while the decreased expression of the sarcoplasmatic reticulum (SR) Ca(2+)-ATPase is thought o play a crucial role in the alterations of Ca2+ handling and LV function. We assessed the dose-dependent effect of chronic ACE inhibition or AT1 receptor blockade on cardiac function in relation to the cardiac expression of the SR Ca(2+)-ATPase and ANF. METHODS AND RESULTS: Renal hypertensive rats (2K-1C) were treated for 12 weeks with three different doses of the ACE inhibitor benazepril, the AT1-receptor antagonist valsartan (each drug 0.3, 3, and 10 mg/kg per day i.p.) or placebo. LV dimensions, hypertrophy and wall stress were determined in vivo by magnetic resonance imaging and the gene expressions of ANF and SR Ca(2+)-ATPase were quantified by Northern blot. Low doses of both drugs did not affect blood pressure, hypertrophy, systolic wall stress and the ANF and SR Ca(2+)-ATPase gene expression. High doses of each drug reduced systolic blood pressure, wall stress, and LV hypertrophy to a similar extent and to values comparable to normotensive, age-matched rats. In addition, high dose treatment reduced LV end-systolic and end-diastolic volume as compared to untreated 2K-1C animals and normalized the mRNA levels of both ANF and SR Ca(2+)-ATPase (as compared to normotensive animals). CONCLUSIONS: We conclude that in this model, high doses of ACE inhibition and AT1-receptor blockade are necessary to normalize systolic blood pressure, LV hypertrophy and systolic LV wall stress which, in turn, is associated with restoration of a normal cardiac phenotype with respect to SR Ca(2+)-ATPase and ANF and normalization of cardiac function.

Cell type specific upregulation of vascular endothelial growth factor in an MCA-occlusion model of cerebral infarct.

Vascular endothelial growth factor (VEGF) is an endothelial cell specific mitogen that has been implicated in hypoxia-mediated angiogenesis under physiological and pathological conditions. We used the middle cerebral artery occlusion model (MCAO) in the rat to investigate VEGF mRNA and protein localization, and VEGFR-1 mRNA and VEGFR-2 mRNA expression in cerebral ischemia. By nonradioactive in situ hybridization we observed upregulation of VEGF mRNA and VEGFR-1 mRNA, but not of VEGFR-2 mRNA in the hemisphere ipsilateral to MCA occlusion. VEGF mRNA was upregulated in the periphery of the ischemic area commencing 3 hours (h) after onset of MCAO, reached a peak after 24 h, and remained expressed at lower levels until 7 days (d) after MCAO. Double labelling experiments revealed that the majority of VEGF expressing cells in the penumbra and within the infarct were immunoreactive for Ox-42, Iba-1, and Ed1, but not for GFAP and neurofilament proteins, suggesting that microglial cells/macrophages are the major cell type expressing VEGE Since VEGF was also expressed in Ox-42 immunoreactive cells distant from the infarct (e.g. in the corpus callosum and hippocampus), activated microglial cells expressing VEGF may migrate towards the ischemic stimulus. VEGF protein was also detected on capillaries within the peri-ischemic area, suggesting that VEGF produced and secreted by microglial cells/macrophages binds to its receptors on nearby vascular endothelial cells and initiates an angiogenic response which counterbalances tissue hypoxia. Accordingly, apoptosis of neuroectodermal cells in the penumbra was highly depressed after the onset of angiogenesis. The spatial and temporal correlation between the induction of angiogenesis with VEGF and VEGFR-1 expression suggests that the ischemic upregulation of VEGF represents a physiological response of the brain to counterbalance hypoxia/ischemia in order to protect neuroectodermal tissue.

Characterization of the cerebroprotective efficacy of the competitive NMDA receptor antagonist CGP40116 in a rat model of focal cerebral ischemia: an in vivo magnetic resonance imaging study.

The cerebroprotective properties of the competitive NMDA antagonist D-(E)-2-amino-4-methyl-5-phosphono-3-pentenoic acid (CGP 40116) were evaluated in a rat model of focal cerebral ischemia. CGP 40116 (5-40 mg/kg i.v.) was injected immediately following permanent occlusion of the left middle cerebral artery (MCA). MK 801 (1 or 3 mg/kg i.v.), D-CPPene (20 mg/kg i.v.), and CGS 19755 (40 mg/kg i.v.) were used for comparison. Lesion volume was assessed using in vivo magnetic resonance imaging, which in initial experiments with parallel histological determinations proved to be an accurate method for the measurement of brain infarction and the determination of a cerebroprotective drug effect. CGP 40116 dose-dependently reduced the volume of cortical infarction, with an ED50 of 11 mg/kg i.v. and a maximal effect equivalent to a 62% reduction in cortical edema volume. Its cerebroprotective efficacy was thus comparable to that of MK 801. The rank order of potency for the NMDA antagonists was MK 801 > CGP 40116 approximately D-CPPene > CGS 19755. Neuroprotection by CGP 40116 was still apparent when treatment was started 30 min after MCA occlusion. It is concluded that CGP 40116 is an effective cerebroprotectant with potential clinical utility for amelioration of focal cerebral ischemic damage.

Increased cerebral infarct volumes in polyglobulic mice overexpressing erythropoietin.

There is increasing evidence that erythropoietin (Epo) has a protective function in cerebral ischemia. When used for treatment, high Epo plasma levels associated with increases in blood viscosity, however, may counteract beneficial effects of Epo in brain ischemia. The authors generated two transgenic mouse lines that overexpress human Epo preferentially, but not exclusively, in neuronal cells. In mouse line tg21, a fourfold increase of Epo protein level was found in brain only, whereas line tg6 showed a dramatic increase of cerebral and systemic transgene expression resulting in hematocrit levels of 80%. Cerebral blood flow (CBF), as determined by bolus tracking magnetic resonance imaging, was not altered in the tg6 line. The time-to-peak interval for the tracer, however, increased approximately threefold in polyglobulic tg6 mice. Immunohistochemical analysis revealed an increase in dilated vessels in tg6 mice, providing an explanation for unaltered CBF in polyglobulic animals. Permanent occlusion of the middle cerebral artery (pMCAO) led to similar perfusion deficits in wild-type, tg6, and tg21 mice. Compared with wild-type controls, infarct volumes were not significantly smaller (22%) in tg21 animals 24 hours after pMCAO, but were 49% enlarged (P < 0.05) in polyglobulic tg6 mice. In the latter animals, elevated numbers of Mac-1 immunoreactive cells in infarcted tissue suggested that leukocyte infiltration contributed to enlarged infarct volume. The current results indicate that moderately increased brain levels of Epo in tg21 transgenic mice were not sufficient to provide significant tissue protection after pMCAO. The results with tg6 mice indicate that systemic chronic treatment with Epo associated with elevated hematocrit might deteriorate outcome after stroke either because of the elevated hematocrit or other chronic effects.

The competitive NMDA antagonist CGP 40116 permanently reduces brain damage after middle cerebral artery occlusion in rats.

In this study we evaluated the effect of the competitive N-methyl-D-aspartate (NMDA) antagonist D-(E)-4-(3-phosphonoprop-2-enyl)piperazine-2-carboxylic acid (CGP 40116) on both early (2 days) and late (28 days) ischemic brain damage in a rodent model of focal cerebral ischemia by means of magnetic resonance imaging (MRI) and conventional histology. Immediately after occlusion of the left middle cerebral artery (MCA), rats received either CGP 40116 (20 mg/kg i.p.) or isotonic saline. Two MRI scans were performed in each animal 2 and 28 days after MCA occlusion. After the second scan, rats were perfusion fixed for histological evaluation. The volume of lesioned brain tissue as determined by MRI or histology was calculated from the damaged area in single sections and the distance between them. CGP 40116 reduced acute infarct volume as measured by MRI 2 days after MCA occlusion by 44% (p < 0.05, analysis of variance). After 28 days the lesion detected by MRI was still significantly smaller in the drug-treated animals. This finding was confirmed by the histological analysis showing a 64% reduction in the volume of brain atrophy in the CGP 40116 group (p < 0.05, analysis of variance). There was a good correlation between the MRI data and the results of the histological evaluation (r = 0.9). Our results indicate that (a) the competitive NMDA antagonist CGP 40116 permanently protects brain tissue from the consequences of cerebral ischemia in a rat model for human stroke and (b) early and late pathological changes can be accurately measured by MRI.

Reperfusion after thrombolytic therapy of embolic stroke in the rat: magnetic resonance and biochemical imaging.

The effect of thrombolytic therapy was studied in rats submitted to thromboembolic stroke by intracarotid injection of autologous blood clots. Thrombolysis was initiated after 15 minutes with an intracarotid infusion of recombinant tissue-type activator (10 mg/kg body weight). Reperfusion was monitored for 3 hours using serial perfusion- and diffusion magnetic resonance imaging, and the outcome of treatment was quantified by pictorial measurements of ATP, tissue pH, and blood flow. In untreated animals, clot embolism resulted in an immediate decrease in blood flow and a sharp decrease in the apparent diffusion coefficient (ADC) that persisted throughout the observation period. Thrombolysis successfully recanalized the embolized middle cerebral artery origin and led to gradual improvement of blood flow and a slowly progressing reversal of ADC changes in the periphery of the ischemic territory, but only to transient and partial improvement in the center. Three hours after initiation of thrombolysis, the tissue volume with ADC values less than 80% of control was 39 +/- 22% as compared to 61 +/- 20% of ipsilateral hemisphere in untreated animals (means +/- SD, P = .03) and the volume of ATP-depleted brain tissue was 25 +/- 31% as compared to 46 +/- 29% in untreated animals. Recovery of ischemic brain injury after thromboembolism is incomplete even when therapy is started as early as 15 minutes after clot embolism. Possible explanations for our findings include downstream displacement of clot material, microembolism of the vascular periphery, and events associated with reperfusion injury.

Exploration of P-type Ca2+ channels as drug targets for the treatment of epilepsy or ischemic stroke.

We investigated the neuroprotective efficacy of the P-type Ca2+ channel antagonist daurisoline against electroshock-induced convulsions in rats and mice, hypoxic/hypoglycemic-induced damage in rat hippocampal slices and brain damage induced by occlusion of the middle cerebral artery (MCA) in rats. Daurisoline applied intravenously (i.v.) (bolus of 1-60 mg/kg) reduced the spontaneous activity of rat cerebellar Purkinje cells in a dose-dependent manner, a result demonstrating activity in the brain with systemic administration of the compound. While this effect reversed rapidly in about 10-20 min following bolus-application of the drug at doses of up to 30 mg/kg, a dose of 60 mg/kg consistently induced a depression of respiration followed by death of the animals. Daurisoline administered at 10-30 mg/kg did not prevent electroshock-induced convulsions in mice or rats, nor did it reduce the neuronal damage in hippocampal slices induced by a hypoxic/hypoglycemic insult in vitro by MCA occlusion in vivo. These observations do not support the hypothesis that P-type Ca2+ channels are promising drug targets for the acute treatment of epileptic convulsions and/or ischemic stroke.

Combined blockade of endothelin-1 and thromboxane A(2) receptors against postischaemic contractile dysfunction in rat hearts.

1. Endothelin-1 (ET-1) may play a role in myocardial ischaemia/reperfusion injury because both the release and vasoconstrictor effect of ET-1 are increased after ischaemia. Since the increased vasoconstrictor effect of ET-1 can be mediated by ET-1-induced release of thromboxane A(2) (TXA(2)), the aim of this study was to test whether combined blockade of ET and TXA(2) receptors protects the coronary flow, contractile performance, and cardiac energy metabolism during ischaemia and reperfusion. 2. Bosentan (antagonist for ET(A) and ET(B) receptors, 1 microM based on concentration-response curves of ET-1), SQ 30,741 (antagonist of TXA(2) receptors, 0.1 microM), or the combination thereof was administered to isolated perfused rat hearts undergoing 15 min of global ischaemia and 60 min of reperfusion. 3. Neither bosentan or SQ 30,741 alone, nor the combination thereof, improved the incomplete postischaemic recovery of coronary flow, left ventricular developed pressure, phosphocreatine, or ATP. However, they attenuated ischaemia-induced acidosis but this did not translate into a measurable effect on haemodynamic or metabolic variables. 4. Thus, combined blockade of ET and TXA(2) receptors does not protect the coronary flow, contractile performance, and cardiac energy metabolism during ischaemia and reperfusion in isolated perfused rat hearts. This finding suggests that neither ET-1 nor ET-1-induced release of TXA(2) play a major role in the postischaemic recovery of the cardiac contractile function and energy metabolism.

CGP 53153: a new potent inhibitor of 5alpha-reductase.

CGP 53153 (N-2-(cyano-2-propyl)-3-oxo-4-aza-5alpha-androst-1-ene-17beta-carb oxamide) is a steroidal inhibitor of 5alpha-reductase, the enzyme which effects the conversion of testosterone (T) to 5alpha-dihydrotestosterone (DHT). In vitro, CGP 53153 competitively inhibited rat microsomal 5alpha-reductase from prostate by 50% (IC50) at a concentration of 36nM compared to the reference compound finasteride which inhibited 5alpha-reductase with an IC50 of 11 nM in the same system. In vivo, inhibition of 5alpha-reductase activity was characterized in three different test systems. Inhibition of 5alpha-reductase activity was first assessed in a standard test designed to compare directly the potency of different 5alpha-reductase inhibitors. This test assesses potency through the inhibition of prostate growth in juvenile castrate male rats treated with a standard dose of T-propionate (1 mg/kg, s.c.) and a 5alpha-reductase inhibitor administered orally at various doses for 4 days. CGP 53153 and finasteride significantly reduced T-propionate-mediated prostate growth by about 25% (ED25) compared to T-propionate-treated controls at oral doses of 0.01 and 0.1 mg/kg, respectively. Second, the effects on prostate weight were studied in normal adult male rats treated orally once daily for 14 days with 1, 3 and 10 mg/kg CGP 53153 and with 10 mg/kg finasteride. CGP 53153 significantly reduced prostate weight at 3 and 10 mg/kg by 31% and 37%, respectively, compared to vehicle-treated controls, whereas the dose of 10 mg/kg finasteride did not significantly reduce prostate weight. Third, the effects on prostate volume were studied in normal 6-9-year-old male dogs treated orally once daily with 5 mg/kg CGP 53153 and with 5 mg/kg finasteride for 12 weeks. Prostate volume was monitored with magnetic resonance imaging every 2 weeks beginning 6 weeks before start of the treatment with 5alpha-reductase inhibitors and ending after a recovery period of 8 weeks after termination of treatment. Treatment for 12 weeks with both CGP 53153 and finasteride was equally effective in reducing prostate volume by more than 70% in individual dogs. Anti-androgenic potency of CGP 53153 and finasteride was assessed in juvenile castrate male rats treated with DHT-propionate (1 mg/kg, s.c.) and a 5alpha-reductase inhibitor (p.o.) for 4 days. Neither CGP 53153 nor finasteride given at a dose of 10 mg/kg had any significant effect on DHT-propionate-mediated prostate growth, whereas the reference anti-androgen flutamide given at a dose of 10 mg/kg reduced prostate weight to levels comparable to those seen in untreated castrate animals. For CGP 53153, the dose of 10 mg/kg is 1000-fold higher than the ED25 for 5alpha-reductase inhibition in vivo. In conclusion, both CGP 53153 and finasteride are potent inhibitors of the rat 5alpha-reductase enzyme system in vitro without showing any anti-androgenic effects in vivo. Both CGP 53153 and finasteride were equally potent in reducing prostate volume in aged male dogs, whereas in rats, CGP 53153 is up to 10 times more potent than finasteride in reducing prostate weight as shown in two different rat models.

Evaluation of quinolinic acid induced excitotoxic neurodegeneration in rat striatum by quantitative magnetic resonance imaging in vivo.

Excitotoxic neurodegeneration in the rat striatum was induced by direct injection of quinolinic acid. The degree of damage was evaluated in vivo 1 day later by quantitative magnetic resonance imaging (MRI) and 7 days later in the same animals by measuring the activities of the neuronal marker enzymes choline acetyltransferase and glutamic acid decarboxylase. Striatal damage assessed using the two approaches was highly correlated. Moreover the cerebroprotective efficacy of the N-methyl-D-aspartate receptor antagonist CGP 40116 was indistinguishable based on all analytical parameters. MRI, however, was more reproducible than the enzymatic methods and was faster and simpler for routine analyses of excitotoxic damage and cerebroprotection in vivo.

Plasminogen activation in experimental permanent focal cerebral ischemia.

BACKGROUND: Previous experimental work using in situ zymography has shown very early increased plasminogen activation in ischemic regions after 3 h of ischemia with and without reperfusion. The objective of the present study was to evaluate the time course and extent of plasminogen activation in long-term permanent focal cerebral ischemia. MATERIAL AND METHODS: The middle cerebral artery in male Fisher rats was irreversibly occluded by electrocoagulation. Duration of ischemia was 48, 72, and 168 h. Occlusion was controlled in vivo by MRI at day 2. Plasminogen activation was detected by in situ zymography of 10 microm cryosections with an overlay containing plasminogen and the plasmin substrate caseine. Areas of plasminogen activation were compared to structural lesions (immunohistochemical loss of microtubule-associated protein 2; MAP 2). RESULTS: Compared to controls, increased plasminogen activation was observed in the basal ganglia and the cortex of the ischemic hemisphere after 48, 72, and 168 h (affected area of basal ganglia: 44.5+/-21.9, 70.1+/-2.3 and 66.6+/-2.8%, respectively; affected area of cortex: 63.4+/-9.8, 67.7+/-0.7 and 64.0+/-3.7%, respectively). The duration of ischemia had no significant influence on the extent of plasminogen activation. Areas of increased plasminogen activation significantly overlapped with and exceeded areas of MAP 2 loss (P<0.005). DISCUSSION: Permanent focal cerebral ischemia leads to increased plasminogen activation in ischemic regions. This plasminogen activation remains elevated at persistent levels over days. It may contribute to extracellular matrix (ECM) disruption, secondary hemorrhage, and brain edema in subacute stages of ischemic stroke.

Reduction of excitotoxicity-induced brain damage by the competitive NMDA antagonist CGP 40116: a longitudinal study using diffusion-weighted imaging.

The cerebroprotective properties of the competitive N-methyl-D-aspartate (NMDA) antagonist CGP 40116 were evaluated in a rat model of excitotoxicity-induced brain damage using direct intrastriatal injection of quinolinic acid and subsequent (5 or 45 min later) i.p. administration of the drug. Diffusion-weighted magnetic resonance imaging (DWI) was used to follow the temporal lesion growth during the acute phase (4 h) and T2-weighted MRI (T2WI) to quantify vasogenic edema extent 2 days later. For control animals, we found a rapid increase in lesion volume during the first hour followed by a moderate growth over the following hours. The DWI-visible hyperintensity was partially reversible after treatment with CGP 40116. The onset of action of CGP 40116 was immediate. The final outcome (63% reduction of lesion volume within 2-4 h post-surgery) was independent of the time of drug administration. DWI data after 4 h correlated well with those obtained by T2WI 2 days later. DWI is a valuable method for early prediction of the outcome of therapeutic interventions of excitotoxic insults.