Research Article[18F]-Alfatide PET imaging of integrin αvβ3 for the non-invasive quantification of liver fibrosis
Graphical abstract
Introduction
Liver fibrosis is a wound-healing process characterized by the accumulation of extracellular matrix (ECM) proteins in response to chronic injury.1 The accumulation of ECM proteins (mainly collagen type I and III) distorts the normal hepatic architecture and forms fibrous scars. As collagen deposition increases, nodules of regenerating hepatocytes result in cirrhosis with its high morbidity and mortality.2 Current methods for evaluating liver fibrosis often provide an incomplete picture of disease. Procedures and biomarkers for determining fibrosis include ultrasonography, elastography, age, platelet count, albumin, and serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) values.3,4 Even when these techniques are incorporated into multifactorial scoring systems, they still suffer from low sensitivity and specificity, and often do not correlate with the degree of fibrosis.5,6 Liver biopsy is the gold standard for assessing fibrosis, but unfortunately this technique samples a limited hepatic volume and can only be performed at a limited frequency due to concerns regarding complication rates.7
There is reason to believe that hepatic integrin αvβ3 may be a useful surrogate diagnostic biomarker for the extent of fibrosis. Hepatic stellate cells (HSCs) are important collagen-producing cells and αvβ3 expression on activated HSCs is associated with HSC proliferation during fibrogenesis.8,9 In addition, imaging hepatic integrin αvβ3 may provide information about therapeutic strategies designed to reduce collagen-producing activated HSCs (aHSCs), either by disrupting αvβ3-ECM interactions or through other approaches.10
The PET tracer used here to image hepatic integrin αvβ3, [18F]-Alfatide, has several advantages. Developed by Chen and coworkers at the NIH,11,12 [18F]-Alfatide is a dimer of RGD peptides, utilizing multivalent affinity enhancement to increase its interactions with its integrin target.[13], [14], [15] Second, [18F]-Alfatide features a radiosynthesis whereby the fluoride ion (18F-) tenaciously binds the aluminum of an Al3+: NOTA complex rather than undergoing formation of covalent 18F-C bond.12,16,17 [18F]-Alfatide's relatively simple radiosynthesis is an advantage when multisite clinical trials are considered. Finally, [18F]-Alfatide has been used in several clinical studies.[18], [19], [20]
Thus, a series of factors, the shortcomings of current liver fibrosis diagnostic methods, the importance of αvβ3 expression on aHSCs for collagen synthesis, the simplicity of [18F]-Alfatide radiochemistry and the prior clinical uses of [18F]-Alfatide, all suggest an [18F]-Alfatide/PET combination might be useful for imaging hepatic integrin αvβ3. To further this possibility, we employed [18F]-Alfatide/PET with the carbon tetrachloride (CCl4) and bile duct ligation (BDL) mouse models of hepatic disease. With both models, the 18F-Alfatide/PET technique imaged hepatic integrin αvβ3, and integrin αvβ3 expression correlated with disease severity and the degree of fibrosis. These results suggest [18F]-Alfatide/PET may offer a non-invasive imaging modality for quantifying hepatic αvβ3, which serves as a surrogate PET biomarker for liver fibrosis in clinical settings.
Section snippets
Animal models of liver fibrosis
We studied liver fibrosis in 2 experimental models. In the first model, 10-week-old CD1 male mice were treated with CCl4 (Sigma, St. Louis, MO) at a dose of 0.2 ml/kg in a 1:5 CCl4: olive oil mixture, administered by means of intraperitoneal (i.p.) injection twice a week for 12 weeks. Control animals were treated with vehicle (olive oil). In the second, BDL-induced biliary fibrosis model, mice were transected at the common bile duct after midline laparotomy. Sham-treated mice underwent midline
Designation of mild and severe levels of liver fibrosis in animal models
Following 6 weeks of CCl4 administration, a mild infiltration of inflammatory cells along with aggregated lymphocytes was observed in the portal areas (indicated by arrows, Fig. 1A,b) by H&E staining. With Sirius red, fibrotic lesions stained weakly in the portal area (mild fibrosis, Fig. 1A,e), and inflammatory cells and formation of regenerative nodules of liver parenchyma separated by fibrotic septa were observed (severe fibrosis, Fig. 1A,c,f). For the BDL model, at 10-day post operation,
Discussion
To our knowledge, this is the first study that systematically validates [18F]-Alfatide imaging in liver fibrosis by performing a spatially precise correlative analysis of PET/CT imaging, histopathology, integrin αvβ3 expression and autoradiography examination in both preclinical rodent models and clinical fibrotic liver specimens. We employed 2 commonly used fibrosis models: CCl4 intoxication and surgical BDL. CCl4 toxicity results from the generation of trichloromethyl (CCl3) radicals by
Financial support
The work presented in this study was supported by NIH grants R01AI136715, R01DK108370.
Authors' contributions
T.S. designed the study, performed experiments and drafted manuscript, Z.C., V.B., H.F., X.D., J.R., Q.Y., L.L., S.R., W.L., and L.J. performed experiments, X.W., V.K. and A.S. performed shear wave elastography experiments, S.R. provided some samples. X.C., R.C. and S.L. conceived project, designed the study and wrote the manuscript.
Conflict of interest
The authors declare no conflicts of interest that pertain to this work.
Please refer to the accompanying ICMJE disclosure forms for further details.
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