There is growing concern about nonalcoholic fatty liver disease (NAFLD), a major cause of chronic liver disease. The most serious manifestation, nonalcoholic steatohepatitis (NASH), is an increasingly common cause of end stage liver disease (Wanless and Lentz 1990
; Angulo 2002
; Schreuder et al. 2008
; Dowman et al. 2010
). Although NASH is known to be associated with the metabolic syndrome (obesity, insulin resistance and hypertriglyceridemia [Wanless and Lentz 1990
; Marchesini et al. 1999
; Charlton 2004
; Vuppalanchi and Chalasani 2009
]), the natural history of NAFLD progressing to NASH is incompletely understood. Because of the increasing incidence of fatty liver disease and also the important role fat (or “steatosis”) plays in the evaluation of liver donors for transplantation (Selzner and Clavien 2001
; Minervini et al. 2009
), it is critically important to improve our ability to diagnose the entire spectrum of NAFLD and to understand its pathophysiology. One essential and needed advance is the development of an inexpensive and easy-to-use instrument that could be widely available for researchers to assess the degree of steatosis in the liver repeatedly, painlessly and noninvasively.
The gold standard for assessing the degree of hepatic steatosis is biopsy (Strassburg and Manns 2006
). Although the risk of postprocedure bleeding is low and the risk of mortality is estimated to be between 0.01% and 0.1%, biopsy is not always logistically possible (especially in an organ donation setting) and the small amount of tissue procured during biopsy may not reflect the global degree of fatty infiltration. Furthermore, liver biopsies are disliked by patients and are sometimes misinterpreted due to processing artifacts or pathologist error. Therefore, a reliable noninvasive means of fat determination would be quite beneficial.
Ultrasound B-scan imaging is an inexpensive and readily available screening tool for steatosis (as determined by increased diffuse echogenicity due to parenchymal fat inclusions) but the sensitivity ranges from 60%–94% and specificity of 66%–95% in determining hepatic steatosis (Foster et al. 1980
; Debongnie et al. 1981
). Transient elastography, a technique that measures the velocity of propagation of shear waves through tissue to determine stiffness, has been shown to correlate with histologic stages of liver fibrosis between 3 and 5 (Castera et al. 2008
; Palmeri et al. 2008
). Transient elastography can be generated by a handheld mechanical vibration source placed over the liver (Sandrin et al. 2003
) or by radiation force excitation of shear waves (Sarvazyan et al. 1998
; Wang et al. 2009
). The shear wave speed is directly proportional to the elastic modulus, which increases with advanced liver fibrosis (Carstensen et al. 2008
). However, these methods cannot measure steatosis when the output is a single “stiffness” estimate. In fact, steatosis confounds shear wave measurements of fibrosis by adding a second variable (Yoneda et al. 2010
) and this issue is clinically significant given that NASH patients have varying degrees of these two variables. Magnetic resonance imaging (MRI) techniques show promise for assessment of steatosis (Bydder et al. 2008
; Ehman 2009
; Salameh et al. 2009
) but are in the research stage and would likely be more expensive and time-consuming than ultrasound techniques.
Although other methods exist to estimate steatosis such as proton magnetic resonance spectroscopy (1
H MRS)(Longo et al. 1995
) and bioimpedence (Hessheimer et al. 2009
), the former is logistically cumbersome in a clinical setting and the latter requires probes to be placed into the liver, thereby severely limiting its clinical utility because of safety issues. More recently, Sandrin and collaborators (Sasso et al. 2010
) have added a proprietary ultrasonic attenuation measurement at 3.5 Mhz to assess liver steatosis and to separate the effects of fat from fibrosis with two measurements (ultrasonic attenuation plus elastography). We agree that two measurements are required to estimate two independent variables, fat and fibrosis. However, we propose to evaluate the dispersion of shear wave properties to achieve this goal.
Dispersion refers to the frequency dependence of the speed of shear waves and attenuation of shear waves in a lossy medium. Generally, the speed of shear waves and the attenuation of shear waves (and other types of acoustic waves) will increase with frequency and can be modeled by a complex wave number (Oestreicher 1951
; Blackstock 2000
; Carstensen et al. 2008
). Whatever the precise mechanisms of loss are in a given medium, the dispersion of attenuation and speed are linked by Kramers-Kronig relations that are based on causality constraints (O’Donnell et al. 1981
; Szabo and Wu 2000
). With the development of elastographic imaging approaches over the past two decades (Parker et al. 2011
), some techniques have been extended to examine dispersion in tissues and pathologies including breast cancer (Tanter et al. 2008
), liver fibrosis (Muller et al. 2009
), muscle (Hoyt et al. 2007
; Chen et al. 2009
), normal mammalian livers (Chen et al. 2009
) and gelatins (Orescanin and Insana 2010
). We believe ours is the first effort to utilize the dispersion properties of elastography to measure steatosis as distinct from fibrosis.
We hypothesize that increasing amounts of fat in the normal liver will increase the dispersion (that is, the frequency dependence or slope) of the speed and attenuation of shear waves, while slightly reducing the speed of sound. illustrates our hypothesis. This is simply the consequence of adding a viscous (and highly lossy) component to the liver, which otherwise would exhibit a strong elastic component with lower dispersion (Carstensen et al. 2008
). Furthermore, as dispersion increases, so will shear wave attenuation. In this study, we employ crawling waves (CrWs), which are an interference pattern of shear waves, in the liver. The CrWs can be imaged by a Doppler ultrasound scanner, with high signal-to-noise over a large region-of-interest (ROI) (Wu et al. 2006
; Hoyt et al. 2008
). The analysis of the crawling wave pattern results in an estimate of the shear wave velocity. When repeated over multiple frequencies from 80 to 300 Hz (or higher in smaller animal livers), the resulting data provide the dispersion estimates that are correlated to steatosis.
Fig. 1 A two parameter assessment of liver viscoelastic properties and their dependence on steatosis and fibrosis: (a) shear velocity (vertical axis) is measured over an accessible bandwidth and plotted as a function of frequency (horizontal axis). Regardless (more ...)