T1rho Values Research Articles

In vivo measurement of T1rho dispersion in the human brain at 1.5 tesla.

To measure T1rho relaxation times and T1rho dispersion in the human brain in vivo. Magnetic resonance imaging (MRI) was performed on a 1.5-T GE Signa clinical scanner using the standard GE head coil. A fast spin-echo (FSE)-based T1rho-weighted MR pulse sequence was employed to obtain images from five healthy male volunteers. Optimal imaging parameters were determined while considering both the objective of the study and the guarantee that radio-frequency (RF) power deposition during MR did not exceed Food and Drug Administration (FDA)-mandated safety levels. T1rho-weighted MR images showed excellent contrast between different brain tissues. These images were less blurred than corresponding T2-weighted images obtained with similar contrast, especially in regions between brain parenchyma and cerebrospinal fluid (CSF). Average T1rho values for white matter (WM), gray matter (GM), and CSF were 85 +/- 3, 99 +/- 1, and 637 +/- 78 msec, respectively, at a spin-locking field of 500 Hz. T1rho is 30% higher in the parenchyma and 78% higher in CSF compared to the corresponding T2 values. T1rho dispersion was observed between spin-locking frequencies 0 and 500 Hz. T1rho-weighted MRI provides images of the brain with superb contrast and detail. T1rho values measured in the different brain tissues will serve as useful baseline values for analysis of T1rho changes associated with pathology.

T2 and T1rho MRI in articular cartilage systems.

T2 and T1rho have potential to nondestructively detect cartilage degeneration. However, reports in the literature regarding their diagnostic interpretation are conflicting. In this study, T2 and T1rho were measured at 8.5 T in several systems: 1) Molecular suspensions of collagen and GAG (pure concentration effects): T2 and T1rho demonstrated an exponential decrease with increasing [collagen] and [GAG], with [collagen] dominating. T2 varied from 90 to 35 ms and T1rho from 125 to 55 ms in the range of 15-20% [collagen], indicating that hydration may be a more important contributor to these parameters than previously appreciated. 2) Macromolecules in an unoriented matrix (young bovine cartilage): In collagen matrices (trypsinized cartilage) T2 and T1rho values were consistent with the expected [collagen], suggesting that the matrix per se does not dominate relaxation effects. Collagen/GAG matrices (native cartilage) had 13% lower T2 and 17% lower T1rho than collagen matrices, consistent with their higher macromolecular concentration. Complex matrix degradation (interleukin-1 treatment) showed lower T2 and unchanged T1rho relative to native tissue, consistent with competing effects of concentration and molecular-level changes. In addition, the heterogeneous GAG profile in these samples was not reflected in T2 or T1rho. 3) Macromolecules in an oriented matrix (mature human tissue): An oriented collagen matrix (GAG-depleted human cartilage) showed T2 and T(1rho) variation with depth consistent with 16-21% [collagen] and/or fibril orientation (magic angle effects) seen on polarized light microscopy, suggesting that both hydration and structure comprise important factors. In other human cartilage regions, T2 and T1rho abnormalities were observed unrelated to GAG or collagen orientation differences, demonstrating that hydration and/or molecular-level changes are important. Overall, these studies illustrate that T2 and T1rho are sensitive to biologically meaningful changes in cartilage. However, contrary to some previous reports, they are not specific to any one inherent tissue parameter.

In vivo proton MR three-dimensional T1rho mapping of human articular cartilage: initial experience.

The purpose of this study was to demonstrate the feasibility of computing three-dimensional relaxation maps of spin-lattice relaxation time in the rotating frame (T1rho) from in vivo magnetic resonance (MR) images of the human patellofemoral joint. T1rho was measured by applying a three-dimensional gradient-echo pulse sequence in six healthy subjects and one symptomatic subject by using a 1.5-T MR imager and a 15-cm-diameter transmit-receive quadrature birdcage radiofrequency coil. Average T1rho measured in healthy patellar cartilage was 49.7 msec +/- 3.2 (mean +/- SD). Two-dimensional T1rho-weighted images were obtained with a fast spin-echo pulse sequence for comparison. There was good correlation between two-dimensional and three-dimensional T1rho values for the six healthy subjects (R2 = 0.88, slope = 1.16).

Early gene therapy-induced apoptotic response in BT4C gliomas by magnetic resonance relaxation contrast T1 in the rotating frame.

The design and evaluation of therapeutic gene transfection protocols and vectors are under extensive development. Magnetic resonance imaging (MRI) techniques can aid considerably in the development of experimental treatment approaches, as well as in determining treatment response by observing gross tissue morphology. However, through a unique set of contrast parameters, namely T1, T2, and diffusion, more information about tissue status can be obtained while delineating and classifying tumor characteristics in more detail. We show here that T1 relaxation in the rotating frame, T1rho, provides unique in vivo MRI contrast. Ganciclovir treatment of HSV-tk+BT4C gliomas, which effectively eradicates these tumors, resulted in significantly prolonged T1rho relaxation times in MRI already after 3 days of treatment, whereas conventional contrast parameters were elevated after 6-8 days of therapy. Interestingly, the prolonged T1rho values were observed while an increase in tumor volume was still taking place. The regions of elevated T1rho relaxation coincided with high apoptotic activity as determined by histology, suggesting that T1rho MRI contrast could be used as a novel early indicator of cytotoxic cell damage in gliomas.

Proteoglycan-induced changes in T1rho-relaxation of articular cartilage at 4T.

Proteoglycan (PG) depletion-induced changes in T1rho (spin-lattice relaxation in rotating frame) relaxation and dispersion in articular cartilage were studied at 4T. Using a spin-lock cluster pre-encoded fast spin echo sequence, T1rho maps of healthy bovine specimens and specimens that were subjected to PG depletion were computed at varying spin-lock frequencies. Sequential PG depletion was induced by trypsinization of cartilage for varying amounts of time. Results demonstrated that over 50% depletion of PG from bovine articular cartilage resulted in average T1rho increases from 110-170 ms. Regression analysis of the data showed a strong correlation (R2 = 0.987) between changes in PG and T1rho. T1rho values were highest at the superficial zone and decreased gradually in the middle zone and again showed an increasing trend in the region near the subchondral bone. The potentials of this method in detecting early degenerative changes of cartilage are discussed. Also, T(1rho)-dispersion changes as a function of PG depletion are described.

Assessing potential bias in the determination of rotational correlation times of proteins by NMR relaxation.

The various factors that influence the reliable and efficient determination of the correlation time describing molecular reorientation of proteins by NMR relaxation methods are examined. Nuclear Overhauser effects, spin-lattice, and spin-spin relaxation parameters of 15N NMR relaxation in ubiquitin have been determined at 17.6, 14.1, 11.7 and 9.4 Tesla. This unusually broad set of relaxation parameters has allowed the examination of the influence of chemical shift anisotropy, the functional form of the model-free spectral density, and the reliability of determined spin-spin relaxation parameters on the characterization of global tumbling of the protein. Treating the 15N chemical shift anisotropy (CSA) as an adjustable parameter, a consensus value of -170 +/- 15 ppm for the breadth of the chemical shift tensor and a global isotropic correlation time of 4.1 ns are found when using the model-free spectral density to fit T1 and NOE data from all fields. The inclusion of T2 relaxation parameters in the determination of the global correlation time results in its increase to 4.6 ns. This apparent inconsistency may explain a large portion of the discrepancy often found between NMR- and fluorescence-derived tau m values for proteins. The near identity of observed T2 and T1 rho values suggests that contributions from slow motions are not the origin of the apparent inconsistency with obtained T1 and NOE data. Various considerations suggest that the origin of this apparent discrepancy may reside in a contribution to the spectral density at zero frequency that is not represented by the simple model-free formalism in addition to the usual experimental difficulties associated with the measurement of these relaxation parameters. Finally, an axially symmetric diffusion tensor for ubiquitin is obtained using exclusively T1 and NOE data. A recommendation is reached on the types and combinations of relaxation data that can be used to reliably determine tau m values. It is also noted that the reliable determination of tau m values from 15N T1 and NOE relaxation parameters will become increasingly difficult as tau m increases.

13C relaxation and dynamics of the purine bases in the iron responsive element RNA hairpin.

The iron responsive element (IRE) RNA hairpin contains a conserved six-nucleotide loop. The NMR structure of this loop showed that the positions of four of its bases are not tightly constrained, while the remaining two are hydrogen-bonded [Laing, L. G., and Hall, K. B. (1996) Biochemistry 35, 13586]. To investigate the flexibility of the RNA in the loop and in the stem, 13C NMR relaxation methods have been used to describe the dynamics of the purine bases. IRE hairpins containing [13C]guanosine and [13C]adenosine are used in NMR experiments to measure T1, T1rho, and NOE values of the bases as a function of temperature (20-37 degreesC). Data are analyzed using the Lipari-Szabo model-free formalism [Lipari, G., and Szabo, A. (1982) J. Am. Chem. Soc. 104, 4546] to determine order parameters and time scales of the motion. Results indicate that the purine bases in the stem have order parameters that are independent of temperature, although they show evidence of both fast (6-40 ps) motions and slower motions at 37 degreesC. The three purines in the loop exhibit increasingly complex motions with long (nanoseconds) correlation times as the temperature increases, suggesting that the loop structure has become disordered.

T1rho-relaxation in articular cartilage: effects of enzymatic degradation.

Spin-lattice relaxation in the rotating frame (T1rho) dispersion spectroscopy and imaging were used to study normal and enzymatically degraded bovine articular cartilage. Normal specimens demonstrate significant T1rho "dispersion" (approximately 60 to approximately 130 ms) in the 100 Hz to 9 kHz frequency range. Proteoglycan-degraded specimens have 33% greater T1rho values than collagen-degraded or normal samples. T1rho-weighted images reveal structure not found in conventional T1- or T2-weighted images. Our results suggest that T1rho measurements are selectively sensitive to proteoglycan content. The potential of this method in distinguishing the early degenerative changes in cartilage associated with osteoarthritis is discussed.

Rotational dynamics of calcium-free calmodulin studied by 15N-NMR relaxation measurements.

The backbone motions of calcium-free Xenopus calmodulin have been characterized by measurements of the 15N longitudinal relaxation times (T1) at 51 and 61 MHz, and by conducting transverse relaxation (T2), spin-locked transverse relaxation (T1 rho), and 15N-[1H] heteronuclear NOE measurements at 61 MHz 15N frequency. Although backbone amide hydrogen exchange experiments indicate that the N-terminal domain is more stable than calmodulin's C-terminal half, slowly exchanging backbone amide protons are found in all eight alpha-helices and in three of the four short beta-strands. This confirms that the calcium-free form consists of stable secondary structure and does not adopt a 'molten globule' type of structure. However, the C-terminal domain of calmodulin is subject to conformational exchange on a time scale of about 350 microseconds, which affects many of the C-terminal domain residues. This results in significant shortening of the 15N T2 values relative to T1 rho, whereas the T1 rho and T2 values are of similar magnitude in the N-terminal half of the protein. A model in which the motion of the protein is assumed to be isotropic suggests a rotational correlation time for the protein of about 8 ns but quantitatively does not agree with the magnetic field dependence of the T1 values and does not explain the different T2 values found for different alpha-helices in the N-terminal domain. These latter parameters are compatible with a flexible dumb-bell model in which each of calmodulin's two domains freely diffuse in a cone with a semi-angle of about 30 degrees and a time constant of about 3 ns, whereas the overall rotation of the protein occurs on a much slower time scale of about 12 ns. The difference in the transverse relaxation rates observed between the amides in helices C and D suggests that the change in interhelical angle upon calcium binding is less than predicted by Herzberg et al. Strynadka and James [Strynadka, N. C. J. & James, M. N. G. (1988) Proteins Struct. Funct. Genet. 3, 1-17].

A T1 rho-filtered two-dimensional transferred NOE spectrum for studying antibody interactions with peptide antigens

Transferred nuclear Overhauser effect (TRNOE) spectroscopy can be used to study intra- and intermolecular interactions of bound ligands complexed with large proteins. However, the 2D NOE (NOESY) spectra of large proteins are very poorly resolved and it is very difficult to discriminate the TRNOE cross peaks, especially those due to intermolecular interactions, from the numerous cross peaks due to intramolecular interactions in the protein. In previous studies we measured two-dimensional difference spectra that show exclusively TRNOE and exchange cross-peaks (Anglister, J., 1990. Quart. Rev. Biophys. 23:175-203). Here we show that a filtering method based on the difference between the T1rho values of the ligand and the protein protons can be used to directly obtain a two-dimensional transferred NOE spectrum in which the background cross-peaks due to intramolecular interactions in the protein are very effectively removed. The usefulness of this technique to study protein ligand interactions is demonstrated for two different antibodies complexed with a peptide of cholera toxin (CTP3). It is shown that the T1 rho-filtering alleviates t problems encountered in our previous measurements of TRNOE by the difference method. These problems were due to imperfections in the subtraction of two spectra measured for two different samples.

Magic angle spinning carbon-13 NMR of tobacco mosaic virus. An application of the high-resolution solid-state NMR spectroscopy to very large biological systems

Magic angle spinning 13C NMR was used to study tobacco mosaic virus (TMV) in solution. Well-resolved 13C NMR spectra were obtained, in which several carbon resonances of amino acids of the TMV coat protein subunits that are not observable by conventional high-resolution NMR spectroscopy can be designed. RNA resonance were absent, however, in the magic angle spinning 13C NMR spectra. Since three different binding sites are available for each nucleotide of the RNA, this is probably due to a line broadening caused by distributions of isotropic chemical shift values. In 13C-enriched TM 13C-13C dipolar interactions also gave rise to line broadening. By suitable pulse techniques that discriminate carbon resonances on the basis of their T1 and T1 rho values, it was possible to select particular groups of carbon nuclei with characteristic motional properties. Magic angle spinning 13C NMR spectra obtained with these pulse techniques are extremely well resolved.