Forms of a self associating autoantibody complex between a monoclonal human IgG1 and human serum albumin.

The mode of association of an unusual human autoantibody complex, composed of a monoclonal immunoglobulin, Tu IgG, and human serum albumin was investigated. A crystalline complex forms from these components in the cold and we have shown that it consists of IgG and albumin in a 1:2 molar ratio [Jentoft et al., Biochemistry 21, 289-294 (1982)]. The crystalline complex was analyzed by electron microscopy and the soluble natural complexes (formed by dissolving the crystals at 20 degrees C) were studied by sedimentation velocity. The sedimentation studies demonstrated that the soluble Tu IgG-albumin complexes are in equilibrium with free Tu IgG and albumin molecules and that the major soluble sedimenting species has a S20,w value of 12.5S. At a constant concn of complex, the size of the sedimenting complex can be reduced by lowering the pH, increasing the ionic strength, or adding CaCl2, citrate, ascorbate or urea. These intermediate, soluble forms have S20,w values that are consistent with 1:1 and 1:2 Tu IgG-albumin complexes. Parameters of repeat distances and angles that were obtained from electron micrographs of the crystalline form of the Tu IgG-albumin complex were used to propose a model for the 12.5S species and were also incorporated into a three-dimensional model for the complex. The 12.5S complex is proposed to form by dimerization of the 1:2 Tu IgG-albumin complex via interactions of albumin with the Fc region of the antibody. The 12.5S dimer may be the nucleating species for subsequent rapid associations that lead to spontaneous formation of crystals. In the proposed model for the Tu IgG-albumin crystals, the angle between the Fab arms of each Tu IgG molecule is 90 degrees, the antigenic determinant on the albumin is located near one end of the long axis of the cylindrical molecule, the site of interaction with Fc is located at the other end of the cylinder, and the CH3 domain of the IgG contains the binding site for albumin that is responsible for the formation of the dimeric 12.5S species. A series of sedimentation velocity experiments suggest that the association between the CH3 domain of IgG and albumin requires the prior formation of the antibody-antigen complex.

Phosphorylethanolamine--the major constituent of the phosphomonoester peak observed by 31P-NMR on developing dog brain.

31P-NMR spectra of newborn dog brains exhibit a prominent phosphomonoester (PME) peak (6.78 +/- SD 0.05 ppm from phosphocreatine peak), similar to those of human neonates. Studies were undertaken to identify the chemical constituents of this peak. Brains of puppies were funnel frozen for methanol-HCl-perchloric acid extraction after in vivo 31P-NMR spectra were taken. The pK of the major component of the PME region in the NMR spectrum of extract was 5.4, corresponding to that of phosphorylethanolamine (PEt). Addition of PEt increased the major peak on the PME region over a wide range of pH, while addition of phosphorylcholine or ribose 5-phosphate yielded distinct peaks. We suggest that the major constituent of phosphomonoester peak of 31P-NMR spectra of newborn dog brain is phosphorylethanolamine. Biochemical mechanisms relevant to changes of phosphorylethanolamine during brain development are discussed.

Structural determinants of Cys2His2 zinc fingers.

Two mutants of the zinc finger peptide Xfin-31 (Ac-YKCGLCERSFVEKSALSRHQRVHKN-CONH2) containing alterations to the conserved hydrophobic core have been constructed and their zinc-bound structures investigated by 1H NMR techniques. In the first (Xfin-31B) a double mutation R8F/F10G places the conserved core aromatic residue at position 8 rather than position 10. In the second (Xfin-31C), Phe-10 is replaced by Leu. A qualitative analysis of 1H chemical shifts, NOE connectivities and coupling constants indicates that the global folds of both mutants are similar to that of the wild-type protein. However, amide exchange rates suggest that the F10L mutant is much less stable than either the wild-type or the R8F/F10G mutant.

Validation of near-infrared spectroscopy in humans.

Near-infrared (NIR) spectroscopy is a noninvasive technique that uses the differential absorption properties of hemoglobin to evaluate skeletal muscle oxygenation. Oxygenated and deoxygenated hemoglobin absorb light equally at 800 nm, whereas at 760 nm absorption is primarily from deoxygenated hemoglobin. Therefore, monitoring these two wavelengths provides an index of deoxygenation. To investigate whether venous oxygen saturation and absorption between 760 and 800 nm (760-800 nm absorption) are correlated, both were measured during forearm exercise. Significant correlations were observed in all subjects (r = 0.92 +/- 0.07; P < 0.05). The contribution of skin flow to the changes in 760-800 nm absorption was investigated by simultaneous measurement of skin flow by laser flow Doppler and NIR recordings during hot water immersion. Changes in skin flow but not 760-800 nm absorption were noted. Intra-arterial infusions of nitroprusside and norepinephrine were performed to study the effect of alteration of muscle perfusion on 760-800 nm absorption. Limb flow was measured with venous plethysmography. Percent oxygenation increased with nitroprusside and decreased with norepinephrine. Finally, the contribution of myoglobin to the 760-800 nm absorption was assessed by using 1H-magnetic resonance spectroscopy. At peak exercise, percent NIR deoxygenation during exercise was 80 +/- 7%, but only one subject exhibited a small deoxygenated myoglobin signal. In conclusion, 760-800 nm absorption is 1) closely correlated with venous oxygen saturation, 2) minimally affected by skin blood flow, 3) altered by changes in limb perfusion, and 4) primarily derived from deoxygenated hemoglobin and not myoglobin.

Uncoupling effects of 2,4-dinitrophenol on electron transfer reactions and cell bioenergetics in rat brain in situ.

We have examined the effects of uncoupling on electron transfer reactions and cell bioenergetics in rat brain in situ. The redox reactions of cyt aa3, cyt c and cyt b as well as the levels of phosphocreatine/inorganic phosphate ratio (PCr/Pi) were monitored in isolated perfused rat head, using optical and [31P]NMR techniques, respectively. In the first series of experiments anesthetized (pentobarbital) and mechanically ventilated rats underwent bilateral carotid arterial cannulation. The head (skull intact, muscle removed) was perfused with a perfluorochemical solution (FC-43), and the animal was then decapitated. By means of reflectance spectrophotometry, the redox reactions of cyt aa3, c and b were monitored before and after perfusing the head with uncoupler 2,4-dinitrophenol (DNP) as well as after complete anoxia. The second series of experiments was performed using [31P]NMR spectroscopy (24.33 MHz; 10.25 in. magnet). PCr/Pi ratios were first determined for living rats (in vivo) before and after exchange transfusion with FC-43 (Hct less than 0.5%), and then in isolated perfused head (in situ) before and after 2,4-DNP addition. Reduction of cyt aa3 (32 +/- 1.4%, mean +/- S.E.M.), cyt c (29 +/- 6%), and cyt b (19 +/- 2%) was induced by addition of 2,4-DNP. Compared to in vivo aerobic metabolism with a PCr/Pi ratio of 3.0-5.0, the value in the uncoupled state in situ was less than 0.1. We conclude that in contrast to in vitro, uncoupling of oxidative phosphorylation induces reduction of intramitochondrial cytochromes in situ.(ABSTRACT TRUNCATED AT 250 WORDS)

MR identification of white matter abnormalities in multiple sclerosis: a comparison between 1.5 T and 4 T.

BACKGROUND AND PURPOSE: Although MR spectroscopy and functional MR imaging of the brain have been successful at 4 T, conventional fast spin-echo imaging of the brain at 4 T has not been adequately evaluated. The purpose of this study was to compare the detection of white matter abnormalities in multiple sclerosis (MS) at 1.5 T and 4 T. METHODS: Fifteen patients with clinically definite MS were imaged at both 1.5 T and 4 T within a 1-week period. Comparison was made between fast spin-echo long-TR images at both field strengths. Pulse sequences were tailored to maximize resolution and signal-to-noise ratio in clinically relevant imaging times (< 7 min). Four interpreters independently reviewed the images obtained at both field strengths in separate sessions and evaluated them for lesion identification, size, characterization, and subjective resolution. Differences in interpretations at 1.5 T and 4 T were subsequently recorded. RESULTS: Images obtained at 4 T showed a mean of 88 more lesions as compared with images obtained at 1.5 T. All the lesions measured less than 5 mm and were typically aligned along perivascular spaces. Twenty-five consensually identified lesions on 4-T images were not seen at all on 1.5-T images. Moreover, 4-T images showed 56 additional consensually identified lesions, which were indistinct and seen only in retrospect on 1.5-T images. These lesions were frequently (n = 48) identified in large confluent areas of white matter signal intensity abnormality at 1.5 T. All observers also agreed that 4-T images subjectively enhanced the perception of normal perivascular spaces and small perivascular lesions. CONCLUSION: MR imaging at 4 T can depict white matter abnormalities in MS patients not detectable at 1.5 T through higher resolution with comparable signal-to-noise ratio and imaging times.

Undecagold clusters for site-specific labeling of biological macromolecules: simplified preparation and model applications.

We report simple and rapid procedures for the synthesis of a variety of stable, water-soluble undecagold cluster, and model applications of a thiol-reactive gold cluster for the specific labeling of cysteine residues in proteins.

High-resolution imaging using Hadamard encoding.

High-resolution imaging techniques using noninvasive modalities such as magnetic resonance (MR) imaging are being pursued as in vivo cancer screening techniques in an attempt to eliminate the invasive nature of surgical biopsy. When acquiring high-resolution MR images for tissue screening, image fields of view have in the past been limited by the matrix sizes available in conventional MR scanners. We present here a technique that uses aliasing to produce high resolution images with larger matrix sizes than are currently available. The image is allowed to alias in both the frequency encoding and phase encoding dimensions, and the individual, aliased fields of view are recovered by Hadamard encoding methods. These fields may then be tiled to obtain a composite image with high spatial resolution and a large field of view. The technique is demonstrated using two-dimensional and three-dimensional in vivo imaging of the human brain and breast.

Relaxation and imaging of lithium in vivo.

It has recently been demonstrated that NMR imaging can be used to record the distribution of lithium. We now report on the parameters pertinent to this imaging. Specifically, the relaxation of Li in aqueous solution, in agarose gel, and in vivo has been investigated. In the latter case, both the longitudinal and transverse relaxations were biexponential, consistent with the behavior expected for a spin 3/2 quadrupole relaxed nucleus. The overall relaxation rate was quite slow in vivo with T'1 = 3.5 sec and T''1 = 6.6 sec.

Discrimination of MR images of breast masses with fractal-interpolation function models.

RATIONALE AND OBJECTIVES: The authors evaluated the feasibility of using statistical fractal-dimension features to improve discrimination between benign and malignant breast masses at magnetic resonance (MR) imaging. MATERIALS AND METHODS: The study evaluated MR images of 32 malignant and 20 benign breast masses from archived data at the University of Pennsylvania Medical Center. The test set included four cases that were difficult to evaluate on the basis of border characteristics. All diagnoses had been confirmed at excisional biopsy. The fractal-dimension feature was computed as the mean of a sample space of fractal-dimension estimates derived from fractal interpolation function models. To evaluate the performance of the fractal-dimension feature, the classification effectiveness of five expert-observer architectural features was compared with that of the fractal dimension combined with four expert-observer features. Feature sets were evaluated with receiver operating characteristic analysis. Discrimination analysis used artificial neural networks and logistic regression. Robustness of the fractal-dimension feature was evaluated by determining changes in discrimination when the algorithm parameters were perturbed. RESULTS: The combination of fractal-dimension and expert-observer features provided a statistically significant improvement in discrimination over that achieved with expert-observer features alone. Perturbing selected parameters in the fractal-dimension algorithm had little effect on discrimination. CONCLUSION: A statistical fractal-dimension feature appears to be useful in distinguishing MR images of benign and malignant breast masses in cases where expert radiologists may have difficulty. The statistical approach to estimating the fractal dimension appears to be more robust than other fractal measurements on data-limited medical images.

Analysis of speech-related variance in rapid event-related fMRI using a time-aware acquisition system.

Speech production introduces signal changes in fMRI data that can mimic or mask the task-induced BOLD response. Rapid event-related designs with variable ISIs address these concerns by minimizing the correlation of task and speech-related signal changes without sacrificing efficiency; however, the increase in residual variance due to speech still decreases statistical power and must be explicitly addressed primarily through post-processing techniques. We investigated the timing, magnitude, and location of speech-related variance in an overt picture naming fMRI study with a rapid event-related design, using a data acquisition system that time-stamped image acquisitions, speech, and a pneumatic belt signal on the same clock. Using a spectral subtraction algorithm to remove scanner gradient noise from recorded speech, we related the timing of speech, stimulus presentation, chest wall movement, and image acquisition. We explored the relationship of an extended speech event time course and respiration on signal variance by performing a series of voxelwise regression analyses. Our results demonstrate that these effects are spatially heterogeneous, but their anatomic locations converge across subjects. Affected locations included basal areas (orbitofrontal, mesial temporal, brainstem), areas adjacent to CSF spaces, and lateral frontal areas. If left unmodeled, speech-related variance can result in regional detection bias that affects some areas critically implicated in language function. The results establish the feasibility of detecting and mitigating speech-related variance in rapid event-related fMRI experiments with single word utterances. They further demonstrate the utility of precise timing information about speech and respiration for this purpose.

Aortic aneurysm following subclavian flap repair: diagnosis by magnetic resonance imaging.

Aortic aneurysm following subclavian flap repair of coarctation of the aorta is thought to occur infrequently. We present a case in which aneurysm of the subclavian flap was found 10 years after the original surgery. The location and extent of the aneurysm was clearly defined using cardiac magnetic resonance imaging (MRI). This report adds further support to recent recommendations to make cardiac MRI a routine imaging study for all postoperative patients from coarctation of the aorta repair.

The use of finite impulse response filters in pulse design.

We develop a new way of synthesizing pulse sequences with desirable frequency characteristics. By combining our previous results with techniques from the theory of finite impulse response filters, we can specify (1) the total duration of the pulse sequence, (2) the frequency ranges it is desired to perturb, (3) the desired perturbation, and (4) the frequency range it is desired not to perturb. We can then synthesize a hard pulse sequence which will yield that desired perturbation with the minimal possible error. The minimum error is global, in the sense that no pulse sequence can do better at achieving the specifications, and is not just a local minimum, around the pulse sequences close to the derived pulse.

The synthesis of soft pulses with a specified frequency response.

We propose a new approach to synthesizing shaped pulses, by first synthesizing a hard pulse sequence and then converting it to a soft pulse. In previous papers, we developed a new approach to synthesizing hard pulse sequences by exactly inverting the Bloch equation, which yields the optimal frequency response. These results can now be applied to shaped pulses. Specifically, one can specify (1) the total duration of the pulse, (2) the frequency range it is desired to perturb, (3) the perturbation desired and, (4) the frequency range it is desired not to perturb. One can then synthesize a shaped pulse which has the desired effect with the least possible error. This enables us to obtain very sharp inversion and pi/2 pulses.

Turnover of succinyl-CoA:3-oxoacid CoA-transferase in glioma and neuroblastoma cells. Specific influence of acetoacetate in neuroblastoma cells.

The specific activity of succinyl-CoA:3-oxo-acid CoA-transferase (3-oxoacid CoA-transferase, EC 2.8.3.5) increases significantly during growth in culture in both mouse neuroblastoma N2a and rat glioma C6 cells. To investigate the mechanism(s) responsible for this, antibody specific for rat brain 3-oxoacid CoA-transferase was raised in rabbits. Immunotitrations of 3-oxoacid CoA-transferase from neuroblastoma and glioma cells on days 3 and 7 of growth after subculture showed that the ratio of 3-oxoacid CoA-transferase activity to immunoprecipitable enzyme protein remained constant, indicating that differences in specific activity of the enzyme at these times in both cell types reflect differences in concentration of enzyme protein. In glioma cells, the relative rate of 3-oxoacid CoA-transferase synthesis was about 0.04-0.05% throughout 9 days in culture. In contrast, the relative rate of synthesis of 3-oxo-acid CoA-transferase in neuroblastoma cells was about 0.07-0.08% on days 3, 5 and 7 after subculture, but fell to 0.052% on day 9. The degradation rates of total cellular protein (t1/2 = 28 h) and 3-oxoacid CoA-transferase (t1/2 = 46-50 h) were similar in both cell lines. The rise in specific activity of the enzyme in both cell lines from days 3 to 7 without a significant increase in the relative rate of synthesis reflects a slow approach to steady-state conditions for the enzyme secondary to its slow degradation. Differences in 3-oxoacid CoA-transferase specific activity between the two cell lines are apparently due to a difference of about 60% in relative rates of enzyme synthesis. The presence of 0.5 mM-acetoacetate in the medium significantly increased the specific activity of 3-oxoacid CoA-transferase in neuroblastoma cells during the early exponential growth phase. This treatment increased the relative rate of synthesis of 3-oxoacid CoA-transferase by 23% (P less than 0.025) in these cells on day 3, suggesting that substrate-mediated induction of enzyme synthesis is a mechanism of regulation of 3-oxoacid CoA-transferase.

Detection of residual quadrupolar interaction in human skeletal muscle and brain in vivo via multiple quantum filtered sodium NMR spectra.

Using sodium multiple quantum filtered methods, we have, for the first time, demonstrated the presence of residual quadrupolar interaction in human skeletal muscle and brain in vivo. Surface coils were used in both skeletal muscle and brain studies on healthy human volunteers. Theoretical analysis shows that even with arbitrary flip angles the double quantum filter retains filtering capability; it passes even rank two quantum coherence more efficiently than odd rank two quantum coherence. Multiple quantum filtered spectra were obtained from the gastrocnemius muscle in the leg and from the parietal lobe region of the brain. In double quantum filtered spectra, we observed different proportions of narrow and broad components with preparation time, while the line shape of triple quantum filtered spectra remained similar to the one observed in isotropic environment. These results suggest that, in human skeletal muscle and brain, sodium ions are bound to some ordered structures.

Fatty tissue on opposed-phase MR images: paradoxical suppression of signal intensity by paramagnetic contrast agents.

PURPOSE: To evaluate paradoxically decreased signal intensity on gadolinium-enhanced opposed-phase magnetic resonance (MR) images of fatty tissues. MATERIALS AND METHODS: Unenhanced and gadolinium-enhanced axial, opposed-phase, gradient-echo images were analyzed visually and with region-of-interest measurements. Tissues measured included adipose tissue (n = 10), angiomyolipomas (n = 8), and vertebral hemangiomas (n = 7). Additionally, a phantom of mayonnaise, soybean oil, agarose, and water (63% lipid signal) with variable concentrations of gadolinium chelate was imaged with similar technique. RESULTS: After administration of gadolinium chelate, signal intensity reduction averaged 18% for adipose tissue, 34% (72-48 units) for predominately fatty angiomyolipomas, and 39% (85-52 units) for vertebral hemangiomas. Imaging of the phantom showed a maximum of 79% reduction in signal intensity with gadolinium chelate (227-47 units). DISCUSSION: Gadolinium-enhanced opposed-phase images depict a significant loss in signal intensity in tissues with MR signal predominately from lipid. Gadolinium chelate increases the signal of water within fatty tissues, which increases the amount of lipid signal suppression due to destructive interference between water and lipid proton magnetizations.

Metastatic lesions of the brain: imaging with magnetization transfer.

PURPOSE: To study solitary metastatic lesions of the brain with routine spin-echo (SE) and magnetization transfer (MT) magnetic resonance (MR) imaging. MATERIALS AND METHODS: In 15 patients with such lesions, the MT ratio (MTR) was calculated in the center and at the periphery of the metastatic lesion, and distally in adjacent white matter, from the periphery of the lesion radially to the most distant cortex. It also was calculated for mirror-image locations in the opposite hemisphere to provide control values. RESULTS: MTRs were decreased in and immediately around the site of the metastatic focus. MTRs also were lower than control values far distal to the metastatic focus, even when no abnormality was seen on SE MR images obtained before and those obtained after administration of gadopentetate dimeglumine. CONCLUSION: MT enables demonstration of white matter abnormalities in patients with metastatic lesions not seen on SE MR images. These changes can be found in white matter far distal to the lesion and surrounding areas of edema.

Quantitative magnetic resonance imaging of human brain perfusion at 1.5 T using steady-state inversion of arterial water.

We report our experience using a noninvasive magnetic resonance technique for quantitative imaging of human brain perfusion at 1.5 T. This technique uses magnetically inverted arterial water as a freely diffusible blood flow tracer. A perfusion image is calculated from magnetic resonance images acquired with and without arterial blood inversion and from an image of the apparent spin-lattice relaxation time. Single-slice perfusion maps were obtained from nine volunteers with approximately 1 x 2 x 5-mm resolution in an acquisition time of 15 min. Analysis yielded average perfusion rates of 93 +/- 16 ml.100 g-1.min-1 for gray matter, 38 +/- 10 ml.100 g-1.min-1 for white matter, and 52 +/- 8 ml.100 g-1.min-1 for whole brain. Significant changes in perfusion were observed during hyperventilation and breath holding. This technique may be used for quantitative measurement of perfusion in human brain without the risks and expense of methods which use exogenous tracers.

Real-time multiple linear regression for fMRI supported by time-aware acquisition and processing.

Real-time parametric statistical analysis of functional MRI (fMRI) data would potentially enlarge the scope of experimentation and facilitate its application to clinical populations. A system is described that addresses the need for rapid analysis of fMRI data and lays the foundation for dealing with problems that impede the application of fMRI to clinical populations. The system, I/OWA (Input/Output time-aWare Architecture), combines a general architecture for sampling and time-stamping relevant information channels in fMRI (image acquisition, stimulation, subject responses, cardiac and respiratory monitors, etc.) and an efficient approach to manipulating these data, featuring incremental subsecond multiple linear regression. The advantages of the system are the simplification of event timing and efficient and unified data formatting. Substantial parametric analysis can be performed and displayed in real-time. Immediate (replay) and delayed off-line analysis can also be performed with the same interface. The capabilities of the system are demonstrated in normal subjects using a polar visual angle phase mapping paradigm. The system provides a time-accounting infrastructure that readily supports standard and innovative approaches to fMRI. Magn Reson Med 45:289-298, 2001.