Research ArticleTranslocator protein (18 kDa), a potential molecular imaging biomarker for non-invasively distinguishing non-alcoholic fatty liver disease
Introduction
Non-alcoholic fatty liver disease (NAFLD) is becoming a public health concern worldwide. It represents a wide spectrum of conditions, ranging from simple steatosis, which generally follows a benign clinical course, to non-alcoholic steatohepatitis (NASH), which progresses to fibrosis in 30–40% of patients and to cirrhosis in 10–15% of patients [1], [2]. Due to the limitations of liver biopsy and currently available non-invasive imaging techniques, new non-invasive biomarkers, that accurately distinguish NASH from NAFLD and allow staging NAFLD, have become an urgent requirement in hepatology [3], [4]. Researchers have tried to identify serum biomarkers on the basis of the current knowledge of the pathophysiological mechanisms of NAFLD, including markers of reactive oxygen species (ROS), inflammation, apoptosis, and fibrosis. However, the results of these studies are controversial [4], suggesting that histological evaluation of NASH may be difficult on the basis of serum biomarker measurements only. Unlike blood tests and histological testing, imaging biomarkers applied to positron emission tomography (PET) technology enable direct, quantitative, and multispatial visualization of physiological and cellular processes at multiple time points and at the whole organism level.
Translocator protein (18 kDa) (TSPO) (also known as peripheral-type benzodiazepine receptor, PBR), a nucleus-encoded mitochondrial target transmembrane protein, has been indicated as an active participant in the modulation of mitochondrial function [5], [6]. PET with radiolabeled TSPO probes has allowed non-invasive and reliable investigation of TSPO in neuropathological damages of experimental animals and humans [7], [8], [9], [10], [11], [12]. Mitochondrial dysfunction plays a key role in the physiopathology of NASH irrespective of the initial cause. Ultrastructural abnormalities and impaired mitochondrial function occur in the liver of patients with NASH, as well as in animal models of NASH [13], [14]. Numerous studies have considered the close relationship between TSPO and mitochondrial dysfunction in conditions such as cardiovascular disease, ischemia, and reperfusion injury [15], [16]. However, to date, no study has examined TSPO expression in the liver during NAFLD progression. Thus, TSPO needs to be explored as a novel imaging marker for non-invasive diagnosis and staging of NAFLD.
N-Benzyl-N-methyl-2-[7,8-dihydro-7-(2-[18F]fluoroethyl)-8-oxo-2-phenyl-9H-purin-9-yl]acetamide([18F]FEDAC) is a specific PET probe for TSPO imaging, developed by our research group [7], [9]. Here, TSPO expression and NAFLD progression were evaluated and quantified using [18F]FEDAC-PET, computerized tomography (CT), histology, and gene analysis in methionine and choline-deficient (MCD) diet-fed mice, one of the most commonly used NASH animal models with severe oxidative stress and hepatocellular injury [17], [18]. We demonstrated the feasibility of TSPO as an imaging biomarker for the diagnosis and staging of NAFLD.
Section snippets
Animals and dietary treatments
All animal experiment protocols were approved by the Animal Ethics Committee of the National Institute of Radiological Sciences and were carried out according to the recommendations of the Committee for the Care and Use of Laboratory Animals, National Institute of Radiological Sciences. Male C57BL/6 mice, aged 6–8 weeks, were purchased from Shizuoka Laboratory Animal Center (Shizuoka, Japan) and maintained in temperature- and light-controlled chambers. The mice were studied at 2, 4, and 8 weeks
Physiological characteristics and liver pathology
Mice fed the MCD diet had lower weights than those fed the normal diet (p <0.05, Table 1). They also had a slightly lower liver weight, but the difference was not statistically significant. When the liver weight was expressed as a percent of the body weight, no difference was found between MCD-fed mice and controls throughout the course of dietary feeding (p >0.05, Table 1). Further, serum ALT and AST levels were significantly greater in MCD-fed mice compared to controls (p <0.05, Table 1).
Discussion
To our knowledge, this is the first study to directly investigate TSPO involved in the modulation of mitochondrial function in MCD diet-fed NAFLD models using PET with a radiolabeled TSPO ligand. TSPO expression increased with NAFLD progression and showed heterogeneous distribution in the liver. The PET signals of [18F]FEDAC-TSPO corresponded to the pathological features observed by histology and CT. Strikingly, the uptake ratio of radioactivity exhibited statistically significant differences
Financial support
This study was supported by research grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan (Grant-in-Aid 24790543).
Conflict of interest
The authors who have taken part in this study declared that they do not have anything to disclose regarding funding or conflict of interest with respect to this manuscript.
Acknowledgements
The authors are grateful to Dr. M. Higuchi for the NP155 antibody and Dr. A. Tsuji for his advice on CT scanning. We thank the staff of the National Institute of Radiological Sciences for their support with cyclotron operation, radioisotope production, radiosynthesis, and animal experiments.
References (28)
- et al.
Non-invasive assessment and quantification of liver steatosis by ultrasound, computed tomography and magnetic resonance
J Hepatol
(2009) - et al.
Translocator protein (18kDa): new nomenclature for the peripheral-type benzodiazepine receptor based on its structure and molecular function
Trends Pharmacol Sci
(2006) - et al.
Peripheral benzodiazepine receptors and mitochondrial function
Neurochem Int
(2002) - et al.
Mitochondrial dysfunction in NASH: causes, consequences and possible means to prevent it
Mitochondrion
(2006) - et al.
Mitochondrial abnormalities in non-alcoholic steatohepatitis
J Hepatol
(1999) - et al.
The peripheral-type benzodiazepine receptor and the cardiovascular system. Implications for drug development
Pharmacol Ther
(2006) - et al.
Translocator protein 18 kDa (TSPO): molecular sensor of brain injury and repair
Pharmacol Ther
(2008) - et al.
Expression of the peripheral-type benzodiazepine receptor and apoptosis induction in hepatic stellate cells
Gastroenterology
(2001) - et al.
Enhanced free cholesterol, SREBP-2 and StAR expression in human NASH
J Hepatol
(2009) - et al.
Hyperferritinemia, iron overload, and multiple metabolic alterations identify patients at risk for nonalcoholic steatohepatitis
Am J Gastroenterol
(2001)
Liver iron excess in patients with hepatocellular carcinoma developed on non-alcoholic steato-hepatitis
J Hepatol
Protein–protein interactions mediate mitochondrial cholesterol transport and steroid biosynthesis
J Biol Chem
The natural history of nonalcoholic fatty liver disease with advanced fibrosis or cirrhosis: an international collaborative study
Hepatology
Decreased survival of subjects with elevated liver function tests during a 28-year follow-up
Hepatology
Cited by (52)
The involvement of the mitochondrial membrane in drug delivery
2024, Acta BiomaterialiaTranslocator protein imaging with <sup>18</sup>F-FEDAC-positron emission tomography in rabbit atherosclerosis and its presence in human coronary vulnerable plaques
2021, AtherosclerosisCitation Excerpt :In a rat model of pulmonary inflammation, 18F-FEDAC showed high uptake at the inflammation site and low uptake in the heart and blood [17]. In a mouse model of nonalcoholic steatohepatitis, increased liver uptake of 18F-FEDAC was correlated with an increased amount of macrophages and closely linked to disease progression [18]. 18F-FEDAC can also be used for in vivo PET imaging of macrophage-induced joint inflammation in mice [19].
Radiosynthesis of <sup>18</sup>F-fluoroethylated tracers via a simplified one-pot <sup>18</sup>F-fluoroethylation method using [<sup>18</sup>F]fluoroethyl tosylate
2021, Applied Radiation and IsotopesCitation Excerpt :In this study, we have attempted to simplify the radiosynthesis of two [18F]fluoroethylated tracers using each desmethyl labeling precursor and [18F]fluoroethyl tosylate, based on the above-mentioned hydrous 18F-fluoroethylation. N-Benzyl-N-methyl-2-[7,8-dihydro-7-(2-[18F]fluoroethyl)-8-oxo-2-phenyl-9H-purin-9-yl]acetamide ([18F]FEDAC) exhibits high binding affinity and selectivity for translocator protein (18 kDa, TSPO), and has been used for the noninvasive visualization of neuroinflammation (Yanamoto et al., 2009; Yui et al., 2010), lung inflammation (Hatori et al., 2012), liver inflammation (Hatori et al., 2014, 2015; Xie et al., 2012), and rheumatoid arthritis (Chung et al., 2018). We have previously reported the synthesis of [18F]FEDAC in two steps using its N-desmethyl labeling precursor and [18F]fluoroethyl bromide (Yanamoto et al., 2009).