Magnetic resonance spectroscopy (MRS) is a powerful tool that allows direct quantification of metabolites in tissue or areas of interest. MRS has been applied in both research and clinical studies to assess liver fat noninvasively in vivo. MRS has also demonstrated excellent performance in liver fat assessment with high sensitivity and specificity compared to biopsy and other imaging modalities. Because of these qualities, MRS has been generally accepted as the reference standard for the noninvasive measurement of liver steatosis. MRS is an evolving technique with high potential as a diagnostic tool in the clinical setting.
Hepatic steatosis or the accumulation of liver fat is the pathological hallmark of NAFLD and has many other clinical implications.
The criterion for the diagnosis of NAFLD is excess fat accumulation in the liver that affects more than 5% of hepatocytes . The degree of liver steatosis is stratified by the percentage of hepatocytes affected by steatosis, as follows: S0 (<5%), S1 (5–33%), S2 (>33–66%), and S3 (>66%) .
Currently, the gold standard for identifying more aggressive NASH is a liver biopsy in staging fibrosis and liver steatosis . In addition to providing information for staging liver steatosis, a liver biopsy also assists with identification of the manifestation of other liver diseases that might coexist or have similar characteristics to fatty liver, such as chronic hepatitis C infection . However, the percutaneous liver biopsy is limited by its invasive nature and is not suitable for clinical applications that require real-time monitoring of liver fat levels throughout therapeutic intervention.
Notably, an early stage of NAFLD can easily be reversed with lifestyle modification . While the identification of this early stage may help prevent disease progression, a sensitive and noninvasive method for liver steatosis will prove to be more useful in later stages of NAFLD. Considering the prevalence and severe consequences of advanced NAFLD, sensitive and real-time monitoring tools would help with the evaluation of the therapeutic response that might lead to small changes in liver fat in the early stage of intervention. At present, serum biomarkers and imaging techniques have been proposed as two main approaches for noninvasive liver steatosis assessment , and MRS is one of those techniques.
MRS utilizes the MR principle to identify and quantify the metabolite from the tissue of interest. The signal in MRS is obtained in the same way as MR imaging, that is, a radiofrequency (RF) at specific resonance is applied to nuclei (e.g., 1H, 13C, 31P, etc.) in a static magnetic field to generate a signal . The pulse sequence and MR signal acquisition are shown in Figure 1. This signal comes from a specific area of interest or the voxel that is then Fourier transformed from the MR signal to the MR spectrum. Unique chemical properties and environments lead to the unique proton resonance frequency and peak shape of each metabolite. This slight shift of the resonance position along the x-axis of the spectrum is termed the chemical shift, which is measured in ppm . The calculation of ppm is obtained from the distance in Hertz (Hz) relative to a reference peak such as water, divided by the operating frequency of the MR system . The proton resonance frequency is proportional to the MR field strength at 63.9 MHz at 1.5 Tesla, 127.8 MHz at 3 Tesla, and 298.2 MHz at 7 Tesla. Therefore, the chemical shift in ppm can be compared across studies irrespective of MR field strength. The MR field strength is also proportional to the improved signal-to-noise ratio (SNR). Thus, the increased field strength of MR machines improves spectral resolution and the separation of metabolite peaks .
Most of the visible peaks in the MRS liver spectrum obtained from the clinical MR scanner (1.5-3 Tesla) are fat and water. While water shows a single peak at approximately 4.7 ppm, fat shows multiple peaks due to its complex chemical components (Table 1) . Six resonances of fat are usually detected with the main lipid peak at approximately 0.9 to 2.75 ppm. There are also unresolved fat resonances at 4.2 and 5.3 ppm from glycerol and olefinic acid, respectively (Figure 2) . These two peaks overlap with the water peak signal at 4.7 ppm. While the correct identification of the liver fat peak is possible in MR systems with a high field, it is less feasible in a lower field with lower spectral resolution and broader linewidth. The misidentification of lipid peaks leads to quantification errors in liver fat; therefore, these unresolvable peaks are not qualified for diagnostic purposes .
|Peak||Chemical Shift (ppm)||Type||Hydrogen Atom Position (Bold)|
MRS liver spectra are often obtained using a single-voxel technique . The advantage of the single-voxel technique is that it provides a high SNR from a large volume of liver sampled. While multivoxel spectroscopy allows larger coverage of the liver than other techniques, the distance from the coil to the organ, longer acquisition time, and reduced shim quality limit its application .
A coil with a multichannel coil array receiver is recommended over a body coil for MRS acquisition to maximize the SNR . The quality of the MRS spectrum is sensitive to inhomogeneous magnetic fields. Good magnetic field homogeneity is required for good spectral resolution or small line width to distinguish peaks from each other. The use of shimming of the magnetic field is therefore necessary to minimize field inhomogeneity across the voxel. While most commercially available MR machines have automated shimming prior to MRS acquisition, manual shimming can also be performed to improve field homogeneity.
The most common pulse sequences for MRS spectral acquisition are stimulated-echo acquisition mode (STEAM) and point-resolved spectroscopy (PRESS). STEAM is a stimulated echo-based technique that utilizes three 90° angles to create well-defined voxels and reduce contaminating signals outside the voxel . PRESS is a spin echo-based technique that uses a 90° pulse followed by 180°–180° (Figure 1).
MRS has been used in clinical trials to investigate liver steatosis grading. Previous studies have applied MRS to investigate liver steatosis grading in a large group of subjects . This illustrates the feasibility of MRS for hepatic steatosis grading in the general population.
Moreover, MRS can estimate the subspecies of liver fat from MRS from the lipid subspecies index using an equation based on the oil spectra model . It has been suggested that the degree of saturation of liver lipids may be associated with liver fat accumulation on hepatocellular damage and disease progression . Each peak of lipid spectra reflects the different chemical positions within the triglyceride molecule, including unsaturated, saturated, monounsaturated, and polyunsaturated fatty acids . The fatty acid composition quantification obtained from MRS has also shown good agreement with other MRI-based methods .
Several studies have demonstrated that MRS can evaluate the efficiency of therapeutic intervention . MRS was previously used in a clinical trial of drugs for NAFLD and was able to evaluate the reduction in liver fat content in a dose-dependent manner . In a double-blind study of NAFLD patients, MRS-assessed liver fat showed no alteration from symbiotic treatment while reducing fecal dysbiosis . Additionally, MRS can also be used to assess the effect of dietary and lifestyle changes on liver fat content. A previous MRS study demonstrated that short-term exercise improved liver lipid saturation, insulin sensitivity, and oxidative stress in individuals with known NAFLD . Additionally, MRS is regarded as an accurate noninvasive tool for liver fat quantification in NASH, the more severe form of NALD .
MRS has also been used to study liver-related diseases that have fat accumulation within hepatocytes, such as human immunodeficiency virus (HIV), hepatitis C virus (HCV), excessive alcohol consumption, and hepatotoxic effects from chemotherapeutic agents or antiretroviral therapy .
Due to the high accuracy and sensitivity of MRS, it has also been used in liver fat assessment for liver transplants. Hepatic steatosis not only influences the outcome of translation but also increases the complication risk of both participants and donors .