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J Schrader

Publications and source records attributed to J Schrader.

At least 37 records · Page 2Linked to original sources
PubMed2004

A minimally invasive approach for efficient gene delivery to rodent hearts.

Transcoronary gene delivery represents a desirable option to achieve global myocardial transgene expression but still requires aggressive surgical preparation in rodents. We therefore developed a catheter-based approach for cardiac gene transfer in the closed chest rat. A double-lumen balloon catheter was used to create aortic occlusion for specific infusion of adenoviral vectors carrying a beta-galactosidase transgene (1 x 10(11) PFU) into the coronaries. Simultaneously, venous return was obstructed by a second balloon catheter in the right atrium. To prolong viral incubation time, we induced a transient cardiac arrest (2 and 5 min) by a combination of acetylcholine and the beta-receptor antagonist, esmolol. At 72 h after transfection, the hearts showed a homogeneous and widespread beta-galactosidase expression, and the transduction efficiency increased and up to about 43% of cardiac myocytes (histochemistry) with a 400-fold increase of beta-galactosidase activity (luminescence assay) compared to sham-operated hearts. Pharmacological treatment aimed at increasing vascular permeability (SNAP and histamine) did not bring about synergistic effects on transfection efficiency. In addition, the method using high intracoronary pressure delivery (>300 mmHg) in a single-pass manner resulted in rather sparse beta-galactosidase expression in the myocardium (3-5% of cardiac myocytes). Therefore, the percutaneous gene delivery system described here provides a simple and minimally invasive procedure that represents a novel strategy for a homogeneous and highly efficient in vivo gene transfer to rodent hearts. Our results also suggest that prolongation of viral incubation time is an effective means for achieving highly efficient myocardial gene transduction.

Adenoviridae↗
PubMed2003

Polar auxin transport in the wood-forming tissues of hybrid aspen is under simultaneous control of developmental and environmental signals.

Recent research has highlighted the importance of auxin concentration gradients during plant development. Establishment of these gradients is believed to involve polar auxin transport through specialized carrier proteins. We have used an experimental system, the wood-forming tissue of hybrid aspen, which allows tissue-specific expression analysis of auxin carrier genes and quantification of endogenous concentrations of the hormone. As part of this study, we isolated the putative polar auxin transport genes, PttLAX1-PttLAX3 and PttPIN1-PttPIN3, belonging to the AUX1-like family of influx and PIN1-like efflux carriers, respectively. Analysis of PttLAX and PttPIN expression suggests that specific positions in a concentration gradient of the hormone are associated with different stages of vascular cambium development and expression of specific members of the auxin transport gene families. We were also able demonstrate positive feedback of auxin on polar auxin transport genes. Entry into dormancy at the end of a growing season leads to a loss of auxin transport capacity, paralleled by reduced expression of PttLAX and PttPIN genes. Furthermore, data from field experiments show that production of the molecular components of the auxin transport machinery is governed by environmental controls. Our findings collectively demonstrate that trees have developed mechanisms to modulate auxin transport in the vascular meristem in response to developmental and environmental cues.

Base Sequence↗
PubMed2003

Coxsackie-and-adenovirus receptor mRNA expression in human heart failure.

BACKGROUND: Adenoviral vectors are widely used as gene-transfer vehicles in experimental and clinical studies. Since virus incorporation and transfection efficacy depend to a large extent on the concentration of the coxsackie-and-adenovirus (CAR) receptor on target cells the aim of this study was to quantify the CAR-receptor concentration in various human cardiomyopathies. METHODS: After RNA isolation from myocardial biopsies obtained during surgical procedures, cDNA was generated by reverse transcription. The relative RNA content was analyzed by quantitative PCR using glyceraldehydes-3-phosphate dehydrogenase (GAPDH) as a standard reference. The cardiomyopathies (CM) analyzed were categorized according to their etiology in dilated CM (DCM, n = 28), ischemic CM (ICM, n = 52), CM in mitral valve disease (MVCM, n = 32) and aortic valve disease (AVCM, n = 32). Data were related to non-cardiomyopathic tissue from donor hearts (non-CM, n = 64). RESULTS: Compared with non-CM hearts DCM showed a 34-fold (+/-5.4) increase in CAR mRNA concentration, in ICM CAR mRNA was elevated by a factor of 12 (+/-4.3), in MVCM by 27 (+/-7) and AVCM by factor 47 (+/-9.3) (ANOVA p < 0.001). Compared with the expression in rat hearts CAR levels were found to be similar to those in human ICM. CONCLUSIONS: These results show that cardiomyopathies associated with heart failure transcribe substantially higher levels - on average by a factor of 30 - of CAR-mRNA than non-failing control hearts. Myocardial gene transfer using adenoviral vectors should therefore be facilitated in human cardiomyopathies and may present a promising approach for therapeutic interventions.

Adenoviridae↗
PubMed2003

[Sequelae of hypertension on end-organs of the brain].

Hypertension is the most important risk factor for stroke and vascular dementia. Antihypertensive treatment reduces stroke risk by 40%. Most probably all antihypertensive drugs are equally effective with the exception of alpha-blockers. Blood pressure is increased in many patients with acute stroke. In this phase sudden drops in blood pressure should be avoided. The combination of an ACE-inhibitor and a diuretic reduced strokes by 28% after TIA or a first stroke. Whether this is a drug specific effect or due to lowering blood pressure per se is investigated at the moment.

Antihypertensive Agents↗
PubMed2003

Screening of yeasts for the production of the aroma compound 2-phenylethanol in a molasses-based medium.

Fourteen yeast strains were screened for production of 2-phenylethanol from L-phenylalanine with molasses as carbon source. Up to 1 g 2-phenylethanol l-1 was obtained. Using oleyl alcohol as a second phase for in situ product removal to enhance the production of 2-phenylethanol increased the yield to about 3 g 2-phenylethanol l-1 at 35 degrees C. The most productive strains were Kluyveromyces marxianus CBS 600 and CBS 397.

Cell Division↗
PubMed2002

Biotechnological production of 2-phenylethanol.

2-Phenylethanol (2-PE) is an important flavour and fragrance compound with a rose-like odour. Most of the world's annual production of several thousand tons is synthesised by chemical means but, due to increasing demand for natural flavours, alternative production methods are being sought. Harnessing the Ehrlich pathway of yeasts by bioconversion of L-phenylalanine to 2-PE could be an option, but in situ product removal is necessary due to product inhibition. This review describes the microbial production of 2-PE, and also summarizes the chemical syntheses and the market situation.

Bacteria↗
PubMed2002

Effects of nitric oxide donors on cardiac contractility in wild-type and myoglobin-deficient mice.

1. The effects of the nitric oxide (NO) donors S-nitroso-N-acetylpenicillamine (SNAP), sodium(Z)-1-(N,N-diethylamino)diazen-1-ium-1,2-diolate (DEA-NONOate), and (Z)-1-[N-(2-Aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate (DETA-NONOate) on force of contraction (F(c)) were studied in atrial and ventricular muscle strips obtained from wild-type (WT) and myoglobin-deficient (myo(-/-)) mice. 2. SNAP slightly reduced F(c) in preparations from WT mice at concentrations above 100 microM; this effect was more pronounced in myo(-/-) mice. 3. DEA-NONOate reduced F(c) in preparations from myo(-/-) mice to a larger extent than those from WT mice. 4. DETA-NONOate reduced F(c) in preparations from myo(-/-) but not from WT mice. 5. Pre-incubation with an inhibitor of the soluble guanylyl cyclase (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one; 100 microM) prevented the effects of SNAP, DEA-NONOate and DETA-NONOate on F(c) in myo(-/-) mice. 6. It is suggested that, in physiological conditions, myoglobin acts as intracellular scavenger preventing NO from reaching its intracellular receptors in cardiomyocytes, whereas, in myoglobin-deficient conditions, NO is able to reduce contractility via activation of the soluble guanylyl cyclase/cyclic GMP pathway.

Animals↗
PubMed2001

A transcriptional roadmap to wood formation.

The large vascular meristem of poplar trees with its highly organized secondary xylem enables the boundaries between different developmental zones to be easily distinguished. This property of wood-forming tissues allowed us to determine a unique tissue-specific transcript profile for a well defined developmental gradient. RNA was prepared from different developmental stages of xylogenesis for DNA microarray analysis by using a hybrid aspen unigene set consisting of 2,995 expressed sequence tags. The analysis revealed that the genes encoding lignin and cellulose biosynthetic enzymes, as well as a number of transcription factors and other potential regulators of xylogenesis, are under strict developmental stage-specific transcriptional regulation.

Cell Wall↗
PubMed2001

Defective hippocampal mossy fiber long-term potentiation in endothelial nitric oxide synthase knockout mice.

Mossy fiber long-term potentiation (mfLTP) was compared in hippocampal slices prepared from wild-type mice and mice lacking functional endothelial nitric oxide synthase (eNOS(-/-) mice) using field potential recording. In the presence of D-2-amino-5-phosphonovaleric acid (AP5, 50 microM), the mfLTP induced by tetanic stimulation (100 Hz, 1 sec) was significantly reduced in knockouts (n = 8) in comparison with wild-type (n = 8). Similarly, potentiation induced by forskolin (30 microM) or 8-bromo-cyclic adenosine monophosphate (8-Br-cAMP, 100 microM) was less pronounced in knockouts. However, in wild-types the mfLTP-induced in the presence of the nonselective pharmacological inhibitor of NOS (N-nitro-L-Arginine, 100 microM, n = 6) was not significantly different from control (n = 8). Thus, eNOS is not directly involved in mfLTP, but lack of eNOS during development leads to a deficit downstream of adenylyl cyclase.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed2001

The nitric oxide-induced reduction in cardiac energy supply is not due to inhibition of creatine kinase.

OBJECTIVES: While nitric oxide (NO) is a potent vasodilator already in the nM range, a cGMP-independent negative inotropic effect is observed at higher concentrations. Since inhibition of creatine kinase (CK) by NO-induced nitrosylation has been proposed as a possible mechanism of action, we measured the flux through CK in the intact heart. METHODS: In saline perfused, paced guinea pig hearts 31P NMR spectroscopy was employed to directly assess the cardiac energy status, i.e. free energy of ATP hydrolysis (DeltaG(ATP)) and flux through CK using magnetization transfer in absence and presence of NO. RESULTS: NO (50 microM) doubled coronary flow and induced a rapid drop in left ventricular developed pressure (39+/-10 vs. 81+/-10 mmHg) and MVO(2) (1.3+/-0.8 vs. 3.7+/-0.5 micromol/min/g) (n=7). This effect was associated with an immediate decrease in phosphocreatine (PCr) (-69%) and DeltaG(ATP). During the subsequent 35 min of NO infusion cardiac function and MVO(2) remained depressed, while PCr partially recovered. NO had no effect on the unidirectional forward flux through CK (98 +/- 21 vs. 99 +/- 20 micromol/min/g, n=7) which was 5- to 10-fold greater than the rate of ATP turnover. Upon cessation of NO infusion both cardiac function and PCr rapidly returned to baseline values. The NO-induced fall in the myocardial energy status was associated with an increase in mitochondrial NADH (n=7) as assessed by surface fluorescence. The observed change in fluorescence was similar to that observed with short term ischemia. CONCLUSION: The NO-mediated depression of myocardial function, MVO(2) and energy status is not mediated by changes in CK flux. Most likely a partial blockade of mitochondrial oxidative phosphorylation at the level of cytochrome c oxidase is responsible for this effect.

Animals↗
PubMed2001

Inotropic response to beta-adrenergic receptor stimulation and anti-adrenergic effect of ACh in endothelial NO synthase-deficient mouse hearts.

1. The functional consequences of a lack of endothelial nitric oxide synthase (eNOS) on left ventricular force development and the anti-adrenergic effect of acetylcholine (ACh) were investigated in isolated hearts and cardiomyocytes from wild type (WT) and eNOS knockout (eNOS-/-) mice. 2.eNOS expression in cardiac myocytes accounted for 20 % of total cardiac eNOS (Western blot analysis). These results were confirmed by RT-PCR analysis. 3. In the unstimulated perfused heart, the left ventricular pressure (LVP) and maximal rate of left ventricular force development (dP/dtmax) of eNOS-/- hearts were not significantly different from those of WT hearts (LVP: 97 +/- 11 mmHg WT vs. 111 +/- 11 mmHg eNOS-/-; dP/dtmax: 3700 +/- 712 mmHg s(-1) WT vs. 4493 +/- 320 mmHg s)-1) eNOS-/-). 4. The dobutamine (10-300 nM)-induced increase in LVP was enhanced in eNOS-/- hearts. In contrast, L-type Ca2+ currents (ICa,L) in isolated cardiomyocytes of WT and eNOS-/- hearts showed no differences after beta-adrenergic stimulation. Dibutyryl-cGMP (50 microM) reduced basal ICa,L in WT cells to 72 +/- 12 % while eNOS-/- ICa,L was insensitive to the drug. The pre-stimulated ICa,L (30 nM isoproterenol) was attenuated by dibutyryl-cGMP in WT and eNOS-/- cells to the same extent. 5. The Ca2+ (1.5-4.5 mM)-induced increase in inotropy was not different between the two experimental groups and beta-adrenergic receptor density was increased by 50% in eNOS-/- hearts. 6. The contractile effects of dobutamine could be inhibited almost completely by ACh or adenosine. The extent of the anti-adrenergic effect of both compounds was identical in WT and eNOS-/- hearts. Measurement of ICa,L in isolated cardiac myocytes yielded similar results. 7. These data demonstrate that in the adult mouse (1) lack of eNOS is associated with increased cardiac contractile force in response to beta-adrenergic stimulation and with elevated -adrenergic receptor density, (2) the unaltered response of ICa,L in eNOS-/- cardiac myocytes to beta-adrenergic stimulation suggests that endothelium-derived NO is important in mediating the whole-organ effects and (3) eNOS is unimportant for the anti-adrenergic effect of ACh and adenosine.

Acetylcholine↗
PubMed2001

Myoglobin: A scavenger of bioactive NO.

The present study explored the role of myoglobin (Mb) in cardiac NO homeostasis and its functional relevance by employing isolated hearts of wild-type (WT) and myoglobin knockout mice. (1)H NMR spectroscopy was used to measure directly the conversion of oxygenated Mb (MbO(2)) to metmyoglobin (metMb) by reaction with NO. NO was applied intracoronarily (5 nM to 25 microM), or its endogenous production was stimulated with bradykinin (Bk; 10 nM to 2 microM). We found that infusion of authentic NO solutions dose-dependently (>/= 2.5 microM NO) increased metMb formation in WT hearts that was rapidly reversible on cessation of NO infusion. Likewise, Bk-induced release of NO was associated with significant metMb formation in the WT (>/=1 microM Bk). Hearts lacking Mb reacted more sensitively to infused NO in that vasodilatation and the cardiodepressant actions of NO were more pronounced. Similar results were obtained with Bk. The lower sensitivity of WT hearts to changes in NO concentration fits well with the hypothesis that in the presence of Mb, a continuous degradation of NO takes place by reaction of MbO(2) + NO to metMb + NO(3)(-), thereby effectively reducing cytosolic NO concentration. This breakdown protects myocytic cytochromes against transient rises in cytosolic NO. Regeneration of metMb by metMb reductase to Mb and subsequent association with O(2) leads to reformation of MbO(2) available for another NO degradation cycle. Our data indicate that this cycle is crucial in the breakdown of NO and substantially determines the dose-response curve of the NO effects on coronary blood flow and cardiac contractility.

Animals↗
PubMed2001

[Spatial heterogeneity of myocardial circulation and energy metabolism].

Within the left ventricular myocardium, substantial differences can be observed in terms of both perfusion and energy turnover. In addition to the small transmural gradient from the subepi--to the subendocardium (1:1.2), more recent high-resolution studies reveal a major patchwork-pattern, e.g., in terms of flow. Adjacent 200 microliters areas can differ more than 3-fold in local perfusion. Low flow and high flow areas (< 50% or > 150% of mean flow, respectively) represent up to 1/5 of the left ventricular myocardium. This local flow pattern is temporally stable for at least days and possibly weeks. Low and high flow areas also differ in local energy metabolism. High flow areas are characterized by enhanced glucose phosphorylation and fatty acid permeability, resulting in increased uptake of these substrates. This is the basis for the recent finding that high flow areas are characterized by an enhanced turnover of the citric acid cycle and thus of local O2 consumption. Since local O2 supply and consumption are closely coupled, low flow areas display no biochemical signs of ischemia. Reducing local flow by 50% results in a similar rise of adenosine or lactate in low and high flow areas. Following complete cessation of perfusion, high flow areas display a greater risk of infarction, indicating enhanced energy demand. Further studies are needed to elucidate the molecular basis of this spatial heterogeneity and to test whether the 3-fold differences in local energy turnover within the myocardial wall also translate into comparable variations of local contractility.

Coronary Circulation↗
PubMed2001

Spatial heterogeneity of energy turnover in the heart.

Local myocardial blood flow varies substantially in spite of a rather homogeneous morphology. To further elucidate this paradox, the spatial heterogeneity of tricarboxylic acid cycle turnover (J(TCA), micromol min(-1) g(-1)) and coronary flow was assessed at a high spatial resolution (6x6x6 mm3) in the open chest dog. Local flow differed more than 2.5-fold between individual samples in each heart (n=7). Out of 1,500 myocardial samples, 1/10 received less than 60% and another 1/10 more than 138% of the normalized mean. In low- and high-flow samples, pyruvate uptake and metabolism were analyzed by 13C NMR spectroscopy. Following [3-13C]pyruvate infusion (2 mM, 12 min), glutamate [4-13C]/[3-13C] was significantly greater in low-flow (2.21+/-0.75, 40 samples) than in high-flow (1.64+/-0.49, 39 samples) areas. This suggests that there are major differences in J(TCA). Glutamate, citrate and lactate content positively correlated with flow. Anaplerotic pathways contributed a fraction similar to J(TCA) in low- and high-flow areas, as demonstrated by isotopomer analysis after 60 min of [3-13C]pyruvate application. Mathematical model analysis of NMR data and relevant pool sizes revealed that J(TCA) and thus myocardial oxygen consumption (MVO2) in high-flow areas exceed values in low-flow areas at least threefold. Thus low and high metabolic states normally coexist within the well perfused heart, suggesting that there is considerable spatial heterogeneity of cardiac energy generation and work.

Alanine↗
PubMed2001

Unaltered radial maze performance and brain acetylcholine of the endothelial nitric oxide synthase knockout mouse.

Proceeding from previous findings of a beneficial effect of endothelial nitric oxide synthase (eNOS) gene inactivation on negatively reinforced water maze performance, we asked whether this improvement in place learning capacities also holds for a positively reinforced radial maze task. Unlike its beneficial effects on the water maze task, eNOS gene inactivation did not facilitate radial maze performance. The acquisition performance over the days of place learning did not differ between eNOS knockout (eNOS-/-) and wild-type mice (eNOS+/+). eNOS-/- mice displayed a slight and eNOS+/+ mice a more severe working memory deficit in the place learning version of the radial maze compared to the genetic background C57BL/6 strain. Possible differential effects of eNOS inactivation, related to differences in reinforcement contingencies between the Morris water maze and radial maze tasks, behavioral strategy requirements, or to different emotional and physiological concomitants inherent in the two tasks are discussed. These task-unique characteristics might be differentially affected by the reported anxiogenic and hypertensional effects of eNOS gene inactivation. Post-mortem determination of acetylcholine concentrations in diverse brain structures revealed that acetylcholine and choline contents were not different between eNOS-/- and eNOS+/+ mice, but were increased in eNOS+/+ mice compared to C57BL/6 mice in the frontal cortex. Our findings demonstrate that phenotyping of learning and memory capacities should not rely on one learning task only, but should include tasks employing both negative and positive reinforcement contingencies in order to allow valid statements regarding differences in learning capacities between rodent strains.

Acetylcholine↗