Identification of new resonances in downfield 1 H MRS of human calf muscle in vivo: Potentially metabolite precursors for skeletal muscle NAD.

PURPOSE: The purpose of this study was to identify and characterize newly discovered resonances appearing in the downfield proton MR spectrum (DF 1 H MRS) of the human calf muscle in vivo at 7T. METHODS: Downfield 1 H MRS was performed on the calf muscle of five healthy volunteers at 7T. A spectrally selective 90° E-BURP RF pulse with an excitation center frequency at 10.3 ppm and an excitation bandwidth of 2 ppm was used for DF 1 H MRS acquisition. RESULTS: In all participants, we observed new resonances at 9.7, 10.1, 10.3, and 10.9 ppm in the DF 1 H MRS. Phantom experiments at 37°C strongly suggest the new resonance at 9.7 ppm could be from H2-proton of the nicotinamide rings in nicotinamide riboside (NR) and nicotinamide mononucleotide (NMN) while the resonance at 10.1 ppm could be attributed to the indole -NH proton of L-tryptophan. We observed that the resonances at 10.1 and 10.9 ppm are significantly suppressed when the water resonance is saturated, indicating that these peaks have either 1 H chemical exchange or cross-relaxation with water. Conversely, the resonances at 9.7 and 10.3 ppm exhibit moderate signal reduction in the presence of water saturation. CONCLUSION: We have identified new proton resonances in vivo in human calf muscle occurring at chemical shifts of 9.7, 10.1, 10.3, and 10.9 ppm. These preliminary results are promising for investigating the role of NR/NMN and L-tryptophan metabolism in understanding the de novo and salvage pathways of NAD+ synthesis in skeletal muscle.

Quantification of cross-relaxation in downfield 1 H MRS at 7 T in human calf muscle.

PURPOSE: The purpose of this study was to characterize the 1 H downfield MR spectrum from 8.0 to 10.0 ppm of human skeletal muscle at 7 T and determine the T1 and cross-relaxation rates of observed resonances. METHODS: We performed downfield MRS in the calf muscle of 7 healthy volunteers. Single-voxel downfield MRS was collected using alternately selective or broadband inversion-recovery sequences and spectrally selective 90° E-BURP RF pulse excitation centered at 9.0 ppm with bandwidth = 600 Hz (2.0 ppm). MRS was collected using TIs of 50-2500 ms. We modeled recovery of the longitudinal magnetization of three observable resonances using two models: (1) a three-parameter model accounting for the apparent T1 recovery and (2) a Solomon model explicitly including cross-relaxation effects. RESULTS: Three resonances were observed in human calf muscle at 7 T at 8.0, 8.2, and 8.5 ppm. We found broadband (broad) and selective (sel) inversion recovery T1 = mean ± SD (ms): T1-broad,8.0ppm = 2108.2 ± 664.5, T1-sel,8.0ppm = 753.6 ± 141.0 (p = 0.003); T1-broad,8.2ppm = 2033.5 ± 338.4, T1-sel,8.2ppm = 135.3 ± 35.3 (p < 0.0001); and T1-broad,8.5ppm = 1395.4 ± 75.4, T1-sel,8.5ppm = 107.1 ± 40.0 (p < 0.0001). Using the Solomon model, we found T1 = mean ± SD (ms): T1-8.0ppm = 1595.6 ± 491.1, T1-8.2ppm = 1737.2 ± 963.7, and T1-8.5ppm = 849.8 ± 282.0 (p = 0.04). Post hoc tests corrected for multiple comparisons showed no significant difference in T1 between peaks. The cross-relaxation rate σAB = mean ± SD (Hz) of each peak was σAB,8.0ppm = 0.76 ± 0.20, σAB,8.2ppm = 5.31 ± 2.27, and σAB,8.5ppm = 7.90 ± 2.74 (p < 0.0001); post hoc t-tests revealed the cross-relaxation rate of the 8.0 ppm peak was significantly slower than the peaks at 8.2 ppm (p = 0.0018) and 8.5 ppm (p = 0.0005). CONCLUSION: We found significant differences in effective T1 and cross-relaxation rates of 1 H resonances between 8.0 and 8.5 ppm in the healthy human calf muscle at 7 T.

Identification of l-Tryptophan by down-field 1 H MRS: A precursor for brain NAD+ and serotonin syntheses.

PURPOSE: To explore the presence of new resonances beyond 9.4 ppm from the human brain, down-field proton MRS was performed in vivo in the human brain on 6 healthy volunteers at 7 T. METHODS: To maximize the SNR, a large voxel was placed within the brain to cover the maximal area in such a way that sinus cavities were avoided. A spectrally selective 90° E-BURP pulse with an excitation bandwidth of 2 ppm was used to probe the spectral chemical shift range between 9.1 and 10.5 ppm. The E-BURP pulse was integrated with PRESS spatial localization to obtain non-water-suppressed proton MR spectra from the desired spectral region. RESULTS: In the down-field proton MRS obtained from all of the volunteers scanned, we identified a new peak consistently resonating at 10.1 ppm. Protons associated with this resonance are in cross-relaxation with the bulk water, as demonstrated by the water saturation and deuterium exchange experiments. CONCLUSION: Based on the chemical shift, this new peak was identified as the indole (-NH) proton of l-tryptophan (l-TRP) and was further confirmed from phantom experiments on l-TRP. These promising preliminary results potentially pave the way to investigate the role of cerebral metabolism of l-TRP in healthy and disease conditions.

Fluorescence excitation analysis by two-photon confocal laser scanning microscopy: a new method to identify fluorescent nanoparticles on histological tissue sections.

In the present study, we make use of the ability of two-photon confocal laser scanning microscopes (CLSMs) equipped with tunable lasers to produce spectral excitation image sequences. Furthermore, unmixing, which is usually performed on emission image sequences, is performed on these excitation image sequences. We use factor analysis of medical image sequences (FAMIS), which produces factor images, to unmix spectral image sequences of stained structures in tissue sections to provide images of characterized stained cellular structures. This new approach is applied to histological tissue sections of mouse aorta containing labeled iron nanoparticles stained with Texas Red and counterstained with SYTO13, to obtain visual information about the accumulation of these nanoparticles in the arterial wall. The possible presence of Texas Red is determined using a two-photon CLSM associated with FAMIS via the excitation spectra. Texas Red and SYTO13 are thus differentiated, and corresponding factor images specify their possible presence and cellular localization. In conclusion, the designed protocol shows that sequences of images obtained by excitation in a two-photon CLSM enables characterization of Texas Red-stained nanoparticles and other markers. This methodology offers an alternative and complementary solution to the conventional use of emission spectra unmixing to localize fluorescent nanoparticles in tissue samples.