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S Schaefer

Publications and source records attributed to S Schaefer.

At least 91 records · Page 5Linked to original sources

31P MRS of myocardial inorganic phosphate using radiofrequency gradient echoes.

Determination of the chemical shift and integral of the myocardial intracellular inorganic phosphate (Pi) resonance by 31P magnetic resonance spectroscopy (MRS) is often precluded due to a large overlapping signal from 2,3-diphosphoglycerate (2,3-DPG) from chamber and myocardial blood. This report demonstrates the use of radiofrequency (RF) magnetic field gradient echoes (RFGE) to eliminate signals from 2,3-DPG in flowing blood, while retaining signals from intracellular myocardial Pi, ATP, and phosphocreatine (PCr). The ECG-triggered 31P spectra were acquired from the myocardium of open chest pigs using a Philips Gyroscan 2-T magnetic resonance spectrometer. A 2.5-cm-diameter surface coil attached to the myocardium was used to provide the RF gradient as well as for excitation and detection of signals. Optimal performance of the RFGE pulse sequence was obtained when the RF gradient pulses were centered at peak diastole or peak systole. Under these conditions, 2,3-DPG signals were completely suppressed, and sensitivity was usually sufficient to allow detection of a well-resolved Pi signal. Myocardial pH determined from RFGE experiments was 7.16 +/- 0.10, and the ratio of the integrals of the Pi and ATP resonances (Pi/ATP) was 0.24. The mean signal-to-noise ratio (S/N) for PCr in control spectra acquired in 4 min was 19/1, while the mean S/N for PCr in RFGE-edited spectra acquired in 15 min was 11/1, demonstrating that the present implementation of the RFGE method results in significant loss in sensitivity. These experiments demonstrate that RFGE-editing allows accurate determination of the chemical shift and integral of the Pi resonance in blood-perfused myocardium in situ.

2,3-Diphosphoglycerate↗

DNA replication of wheat dwarf virus, a geminivirus, requires two cis-acting signals.

Replication of the single-stranded DNA genome of wheat dwarf virus (WDV) leads to the accumulation of covalently closed double-stranded DNA of genome length in infected cells. By studying the replication properties of a naturally occurring deletion mutant of WDV isolated from infected plants and of deletion mutants constructed in vitro, we have defined cis-acting regions required for viral DNA replication. The results show that two distinct regions are required in cis to yield the normal replicative forms of WDV-DNA.

Blotting, Southern↗

In vitro tumorigenicity of hepatitis B virus DNA and HBx protein.

Persistent infection by hepatitis B virus (HBV) is strongly associated with the development of hepatocellular carcinoma. This linkage may be caused by oncogenic HBV gene products. Our initial in vitro studies have revealed that a non-tumorigenic, fetal mouse hepatocyte line (FMH202-1), harboring simian virus 40 large tumor antigen (SV40TAg) as transgene, can be converted into a full-malignant phenotype by transfection with dimeric HBV-DNA Höhne, M., Schaefer, S., Seifer, M., Feitelson, M.A., Paul, D. and Gerlich, W.H. (1990) EMBO J 9, 1137-1145. The oncogenic effect was neither dependent on simultaneous expression of SV40TAg nor on the cell type, since HBV-transfected permanent mouse fibroblasts (LTK-) also displayed enhanced colony formation in soft agar. Transfection of FMH202-1 with the X region of HBV generated clones that also formed colonies in soft agar and were tumorigenic in nude mice. Growth in soft agar and induction of nude mice tumors both depended on high expression of HBx protein. Although HBx expression was stronger in X-transfected than HBV-transfected clones, the former did not grow well in soft agar, and the X-derived tumors developed more slowly. In the tumors, expression of HBx was almost shut off, but again high in the tumor-derived cell lines. Constitutive expression of c-fos was strongly enhanced in the X-transfected cell lines and tumors. Transfection of FMH202-1 with an isogenic HBx-deficient mutant fragment generated several clones, which expressed normal levels of HBx transcripts, but did not grow in soft agar.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

Contrast-enhanced magnetic resonance imaging of hypoperfused myocardium.

Contrast-enhanced magnetic resonance (MR) imaging can define myocardial perfusion defects due to acute coronary occlusion. However, since most clinically important diagnostic examinations involve coronary arteries with subtotal stenoses, we investigated the ability of MR imaging with a manganese contrast agent to detect perfusion abnormalities in a canine model of partial coronary artery stenosis. The contrast agent was administered after the creation of a partial coronary artery stenosis with the addition of the coronary vasodilator dipyridamole in six of 12 animals. The hearts were imaged ex situ using gradient reversal and spin-echo sequences, and images were analyzed to determine differences in signal intensity between hypoperfused and normally perfused myocardium. Comparison of MR images with regional blood flow and thallium-201 measurements showed good concordance of hypoperfused segments in those animals given dipyridamole, with 75% of the abnormal segments correctly identified. In those animals not given dipyridamole, 48% of segments were correctly identified. Thus, ex vivo MR imaging with a paramagnetic contrast enhancement can be used to detect acute regional myocardial perfusion abnormalities due to severe partial coronary artery stenoses.

Animals↗

Wheat dwarf virus vectors replicate and express foreign genes in cells of monocotyledonous plants.

Wheat dwarf virus (WDV) is a geminivirus that infects monocotyledonous plants. To exploit the potential of WDV as a replicative gene vector, we developed a transient replication and expression system based on the transfection of protoplasts derived from Triticum monococcum suspension culture cells. Cloned genomic copies of various WDV isolates as well as mutants constructed in vitro were introduced into the protoplasts and assayed for their ability to replicate. As a result, regions of the WDV genome necessary or dispensable for the viral DNA replication could be defined. In addition, the gene encoding the viral capsid protein was replaced by three different bacterial marker genes, neomycin phosphotransferase, chloramphenicol acetyltransferase, and beta-galactosidase. The beta-galactosidase gene doubled the size of the WDV genome. The replication of the recombinant WDV genomes and the expression of these genes were monitored in suspension culture cells of T. monococcum. The potential of replicative expression vectors based on the WDV genome is discussed.

Base Sequence↗

Metabolic and functional consequences of blunted myocardial reactive hyperemia.

This study determined whether the rapidity of myocardial metabolic and contractile recovery after brief coronary occlusion depends upon the intensity of reactive hyperemia. We also tested the hypothesis that coronary flow rate modulates contractility after brief myocardial ischemia, independent of changes in phosphorus metabolites. Eight open-chest pigs were studied with phosphorus-31 nuclear magnetic resonance (NMR) spectroscopy with 14 s time resolution. After a 29-s anterior descending coronary occlusion, peak Doppler coronary flow velocity was alternately unrestricted (normal hyperemia, 443 +/- 40% of control) or limited to 159 +/- 9% of control. During 29 s coronary occlusion, phosphocreatine-to-inorganic phosphate ratio (PCr/Pi) and systolic segment shortening in the ischemic region fell to 28 +/- 4 and 7 +/- 7% of control, respectively. With normal hyperemia, PCr/Pi and segment shortening recovered within 29 s. With blunted hyperemia, recovery of both parameters was delayed an additional 29-43 s, associated with reduced subendocardial blood flow (measured with radioactive microspheres) and persistent intracellular acidosis. However, the relationship between segment shortening and PCr/Pi was unaffected by the intensity of reactive hyperemia. Thus blunted reactive hyperemia significantly delays metabolic and contractile recovery from brief ischemia, probably via transient maldistribution of transmural perfusion. However, coronary blood flow rate does not independently modulate contractility after brief reversible ischemia.

Adenosine Triphosphate↗

Clinical nuclear magnetic resonance spectroscopy: insight into metabolism.

Nuclear magnetic resonance (NMR) spectroscopy can nondestructively evaluate changes in metabolites with different disease states, as well as with therapeutic interventions. Animal studies have provided the basis for understanding changes in high-energy phosphates with myocardial ischemia. Studies of graded ischemia due to partial coronary stenosis have shown the sensitivity of the ratio of phosphocreatinine to inorganic phosphate to small reductions in myocardial blood flow and its relation to myocardial function. The application of NMR spectroscopy to humans requires precise localization techniques to avoid acquiring contaminating information from structures around the heart, such as the chest wall and diaphragm. With these localization techniques, metabolic evidence of ischemia has been demonstrated in patients with myocardial infarction and patients with known coronary disease, although the sensitivity of this technique for the diagnosis of inducible ischemia is unknown. At rest, patients with dilated and hypertrophic cardiomyopathies often have an elevated phosphodiester resonance, possibly signifying abnormal breakdown of membrane phospholipids. Increasing oxygen demand in these patients does not usually alter high-energy phosphates, suggesting that oxidative energy metabolism is preserved under these conditions. NMR spectroscopy is a powerful tool to increase understanding of metabolic changes in a variety of pathologic conditions.

Animals↗

In vivo phosphorus-31 spectroscopic imaging in patients with global myocardial disease.

The goals of this study were to determine whether abnormalities in phosphorus metabolism could be noninvasively detected using phosphorus-31 nuclear magnetic resonance spectroscopy in patients with dilated cardiomyopathy and left ventricular hypertrophy, and whether these patient groups could be distinguished from each other based on parameters obtained using this technique. Seventeen patients and 14 control subjects were studied using nuclear magnetic resonance spectroscopy. Spectra were obtained from the human heart at rest using 3-dimensional spectroscopic imaging as a localization technique. Data were acquired over an average volume of 48 cc in 26.3 minutes using a 2 tesla imaging and spectroscopy unit. The ratio of phosphocreatine to adenosine triphosphate was 0.89 +/- 0.88 (mean +/- standard error) in normal subjects and did not differ significantly in patients with dilated cardiomyopathy or left ventricular hypertrophy. A prominent peak in the phosphodiester region was seen much more frequently in patients with dilated cardiomyopathy, resulting in significantly higher ratios of phosphodiester to phosphocreatine (1.28 +/- 0.35) and phosphodiester to adenosine triphosphate (0.79 +/- 0.18) in this group compared to normal subjects (0.33 +/- 0.08 and 0.29 +/- 0.08, respectively). However, the various patient groups could not be reliably distinguished from each other based on spectral patterns. These studies demonstrate the feasibility of performing phosphorus-31 nuclear magnetic resonance spectroscopic imaging in patients with myocardial disease. The initial results indicate that, under resting conditions, the ratio of phosphocreatine to adenosine triphosphate is not consistently altered in patients with severe global cardiomyopathies or hypertrophy. Phosphodiesters are elevated in some patients with dilated cardiomyopathy, a finding that may signify abnormal phospholipid metabolism in this condition.

Adenosine Triphosphate↗

The post-occlusive hyperemic response in patients with systemic sclerosis.

We investigated post-ischemic hyperreactive cutaneous blood flow in patients with primary Raynaud's phenomenon and Raynaud's phenomenon secondary to systemic sclerosis (SSc). Reactive hyperemia was measured over a locally warmed area of skin, using a laser Doppler flowmeter, following 5 minutes of suprasystolic occlusion of blood flow. We found that patients with primary Raynaud's phenomenon had normal post-ischemic blood flow compared with normal controls. In contrast, patients with SSc had reduced levels of baseline and peak blood flow compared with either the primary Raynaud's phenomenon patients or the normal subjects. Infusion of carbaprostacyclin, a potent prostacyclin analog vasodilator, did not increase blood flow in the SSc patients, nor did it restore the reactive hyperemic response. These findings are consistent with the hypothesis that patients with the nonvasoconstricted condition of SSc have fixed structural defects that limit cutaneous microvascular blood flow.

Adult↗

Malignant transformation of immortalized transgenic hepatocytes after transfection with hepatitis B virus DNA.

Persistent infection by hepatitis B virus (HBV) is epidemiologically correlated with the prevalence of hepatocellular carcinoma, but its role in tumor development is not yet understood. To study the putative oncogenic potential of HBV, a non-malignant immortal mouse hepatocyte line FMH202 harboring metallothionein promoter-driven simian virus 40 large tumor antigen was transfected with HBV DNA. All stably transfected clones which replicated HBV displayed malignant growth characteristics in soft agar and were tumorigenic upon inoculation in nude mice. The nude mice tumors were histologically classified as differentiated or anaplastic hepatocellular carcinomas. As with human liver carcinomas, rearrangements of in vitro integrated HBV sequences were observed in the nude mouse tumors, and in tumor-derived cell lines. In one case, expression of viral core and surface antigens was blocked in the tumors, correlating with hypermethylation of the HBV genome. However, the expression of X gene was maintained in most tumors and tumor-derived cell lines. X protein was detected in nuclei by immune fluorescence and by immune blot. These results provide the first demonstration that HBV displays oncogenic potential in an experimental system. This system could be useful to functionally identify HBV genes which convey a tumorigenic phenotype.

Animals↗

Epicardial and endocardial localized 31P magnetic resonance spectroscopy: evidence for metabolic heterogeneity during regional ischemia.

Previous studies have noted that myocardial blood flow and high energy phosphates are heterogeneous across the myocardial wall during ischemia. In order to determine whether differences in metabolites between the subendocardium and subepicardium could be detected using 31P magnetic resonance spectroscopy, the Fourier series window (FSW) experiment was implemented on a porcine model of graded regional ischemia. FSW experiments using a planar phantom showed a 46% improvement in localization to the subendocardium compared to a one-pulse experiment. Animal studies of graded ischemia demonstrated a gradient in the phosphocreatine to inorganic phosphate ratio in the myocardium that paralleled the gradient in blood flow. These studies demonstrate the ability of spatially localized 31P magnetic resonance spectroscopy to detect regional changes in myocardial high energy phosphates localized to the subepicardium and subendocardium.

Animals↗

Wheat dwarf virus Ac/Ds vectors: expression and excision of transposable elements introduced into various cereals by a viral replicon.

The maize transposon Activator (Ac) and in vitro-generated nonautonomous derivatives thereof [Ac delta or Dissociation (Ds) elements] were inserted into the genome of a geminivirus of graminaceous plants, wheat dwarf virus, at a site that does not interfere with viral replication. These recombinant viral genomes were introduced into wheat, maize, and rice protoplasts, where rapid and efficient excision of Ac was observed. Excision was detected only in vectors in which, after transfection, the virus could replicate. This result is not restricted to the autonomous Ac; excision of Ds elements was also induced by transposase activity provided in trans by plasmids expressing the cDNA of Ac. The potential of this combination of a transposon with a viral replicon for plant molecular genetic engineering is discussed.

Base Sequence↗

Myocardial high-energy phosphates in reactive hyperemia.

This study used 31P nuclear magnetic resonance (NMR) spectroscopy to determine whether the magnitude and duration of myocardial reactive hyperemia (RH) reflect a commensurate derangement of myocardial high-energy phosphate (HEP) metabolism, or if coronary blood flow (CBF) and HEP metabolism are dissociated during RH. Twelve open-chest anesthetized pigs were studied during and after 24 s occlusion of the anterior descending coronary artery. CBF velocity was measured with a Doppler probe. NMR time resolution (4.8 s) was obtained by summing corresponding short blocks of data from multiple occlusions. During occlusion, phosphocreatine (PCr) declined to 65 +/- 5% (mean +/- SE) of control accompanied by increased spectral intensity of the Pi + phosphomonester region. By 20 s of reflow, HEPs had returned to control levels, but CBF was still elevated at 282 +/- 18% of control and remained elevated for an additional 53 +/- 7 s, during which 44 +/- 6% of total RH flow occurred. Therefore, the control of CBF is not closely coupled to the levels of myocardial HEPs during RH and the duration of RH does not reflect prolonged depletion of myocardial HEPs.

Animals↗

Dynamic relation between myocardial contractility and energy metabolism during and following brief coronary occlusion in the pig.

Changes in high-energy phosphate metabolism may be important in the regulation of myocardial contractile function during ischemia. This study sought to determine the dynamic relation between myocardial contractile function and high-energy phosphate metabolism during and following brief (24-second) coronary occlusion, when large and rapid changes in both parameters occur. Eight anesthetized, open-chest pigs were instrumented with a Doppler flow probe and occluder on the anterior descending coronary artery, segment length crystals in the anterior left ventricular wall, and a surface coil for phosphorus-31 nuclear magnetic resonance spectroscopy. Phosphorus-31 spectra were reconstructed with a 4.8-second time resolution by summing corresponding short blocks of data from multiple occlusions. Metabolic and functional parameters were unchanged during the first 4.8 seconds of occlusion. During the remainder of occlusion, phosphocreatine progressively declined to 66 +/- 3% of control, inorganic phosphate rose to 170 +/- 8% of control, and segment shortening fell to 25 +/- 9% of control. A strong linear correlation was found between dynamic changes in segment shortening and phosphocreatine (r2 = 0.97), inorganic phosphate (r2 = 0.96), and the ratio of phosphocreatine to inorganic phosphate (r2 = 0.98) during occlusion. At any level of the ratio between phosphocreatine and inorganic phosphate, segment shortening was greater during reflow than during occlusion. The close, dynamic relation between segment shortening and phosphorus metabolites supports the regulation of contractility by changes in energy metabolism or its by-products during ischemia. During reactive hyperemia, the high coronary flow rate may be an independent factor modulating contractility.

Adenosine Triphosphate↗

Relationship between myocardial metabolites and contractile abnormalities during graded regional ischemia. Phosphorus-31 nuclear magnetic resonance studies of porcine myocardium in vivo.

The mechanisms responsible for changes in myocardial contractility during regional ischemia are unknown. Since changes in high-energy phosphates during ischemia are sensitive to reductions in myocardial blood flow, it was hypothesized that myocardial function under steady-state conditions of graded regional ischemia is closely related to changes in myocardial high-energy phosphates. Therefore, phosphorus-31 nuclear magnetic resonance spectroscopy was employed in an in vivo porcine model of graded coronary stenosis. Simultaneous measurements of regional subendocardial blood flow, high-energy phosphates, pH, and myocardial segment shortening were made during various degrees of regional ischemia in which subendocardial blood flow was reduced by 16-94%. During mild reductions in myocardial blood flow (subendocardial blood flow = 83% of nonischemic myocardium), only the ratio of phosphocreatine to inorganic phosphate (PCr/Pi), Pi, and [H+] were significantly changed from control. PCr, ATP, and PCr/ATP were not significantly reduced from control with mild reductions in blood flow. Changes in myocardial segment shortening were most closely associated with changes in PCr/Pi (r = 0.94). Pi and [H+] were negatively correlated with segment shortening (r = -0.64 and -0.58, respectively) and increased over twofold when blood flow was reduced by 62%. Thus, these data demonstrate that PCr/Pi is sensitive to reductions in myocardial blood flow and closely correlates with changes in myocardial function. These data are also consistent with a role for Pi or H+ as inhibitors of myocardial contractility during ischemia.

Adenosine Triphosphate↗

Cardiovascular applications of nuclear magnetic resonance spectroscopy.

Nuclear magnetic resonance spectroscopy has great potential for defining noninvasively the metabolic status of the heart and skeletal muscle. This technique uses the spin properties of certain nuclei (such as phosphorus-31, hydrogen-1 and carbon-13) to measure high energy phosphates, intracellular pH, lactate and glycogen. Animal studies have formed the basis for human investigations and have demonstrated well-defined changes in high energy phosphates during myocardial ischemia and reperfusion, as well as in cardiomyopathies. Human studies have been limited by issues of sensitivity and localization, although techniques such as rotating frame, depth-resolved surface coil spectroscopy, image-selected in vivo spectroscopy and spectroscopic imaging have been used to acquire phosphorus-31 spectra from the human heart. The few human studies of patients with disease have demonstrated elevated inorganic phosphate peaks after myocardial infarction and abnormal phosphodiester peaks in patients with hypertrophic cardiomyopathy. Studies of patients with heart failure have shown that these patients acidify their peripheral muscles with exercise more easily than do control subjects. Clinical application of nuclear magnetic resonance spectroscopy will depend on technical advances and the demonstration of sensitivity of metabolic changes with disease.

Animals↗

Phosphorus-31 magnetic resonance spectroscopy in humans by spectroscopic imaging: localized spectroscopy and metabolite imaging.

In in vivo phosphorus magnetic resonance spectroscopy (MRS), spectroscopic imaging (SI) can be used as a flexible localization technique, producing spectra from multiple volumes in a single examination. Presented here are phosphorus SI studies of human organs in which a selective-volume SI reconstruction was used rather than the usual array-format SI reconstruction. A linear predictor technique was used to estimate the initial points of the free induction decay missing because of the delay needed for phase-encoding gradients, significantly reducing the baseline artifacts which commonly complicate interpretation of SI spectra. In studies of heart, brain, liver, and kidney, the performance of SI was found to compare favorably with that of ISIS. SI phosphorus metabolite intensity images from a brain tumor patient were obtained at 2 X 2-cm in-plane resolution (with "slice" thickness of roughly 16 cm, determined by coil sensitivity) in 34 min, demonstrating the feasibility of obtaining clinically useful metabolite images in clinically reasonable examination times.

Astrocytoma↗

Clinical magnetic resonance spectroscopy of brain, heart, liver, kidney, and cancer. A quantitative approach.

Clinical studies using 31P and 1H MRS with a whole body 2.0 T MRI/MRS system are described. In most cases, techniques to quantitate absolute molar concentrations of metabolites in various organs were used. In the brain, AIDS, chronic stroke, and white matter lesions were associated with alterations of brain 31P metabolites. Epilepsy was associated with increased pH in the seizure focus. In the heart, dilated cardiomyopathy was associated with increased PDE/ATP while PCr/ATP was unchanged. In the liver, alcoholic hepatitis and cirrhosis were associated with diminished hepatic ATP while alcoholic hepatitis had increased pH and cirrhosis had decreased pH. This allowed differentiation of normal liver, alcoholic hepatitis, and alcoholic cirrhosis without biopsy. In the prostate, malignancy was associated with increased PME/ATP and decreased PCr/ATP. The PME/PCr was greatly increased in malignant prostate with no overlap in normals. Other cancers outside the brain had increased PME and effective treatment was often associated with diminished PME. 1H MRS of the brain was performed using ISIS and outer volume suppression pulses for volume localization. Excellent high resolution 1H water-suppressed spectra were obtained at echo times as short as 30 ms, showing well resolved peaks for lactate, N-acetylaspartate, glutamate, choline, creatinine, and inositol. 1H MRS demonstrated that the uptake of ethanol by the brain was slower than the rise of ethanol in blood. 31P spectroscopic imaging of the brain with resolution of 2.25 x 2.25 x 2.5 cm produced metabolic images and high resolution spectra from desired regions of interest.(ABSTRACT TRUNCATED AT 250 WORDS)

Brain↗