Practical Updates in Liver Pathology: Grading, Staging, and Nomenclature
Section 2 -
Moderate Steatosis and Steatohepatitis
Neil Theise, M.D.
Romil Saxena, M.D.
33 year old woman with morbid obesity. Liver biopsy during bariatric surgery.
Diagnosis: Moderate steatosis and steatohepatitis, compatible with
obesity-associated fatty liver disease.
22 year old woman with morbid obesity. Liver biopsy during bariatric surgery.
Diagnosis: Moderate steatosis with steatohepatitis and extensive
steatofibrosis consistent with obesity-associated fatty liver disease.
28 year old man with morbid obesity. Liver biopsy during bariatric surgery
Diagnosis: No pathologic change
The term NASH (non-alcoholic steatohepatitis) was coined in 1980 by Ludwig to describe biopsy findings
resembling those seen in chronic alcohol-related liver disease in patients who did not drink. While the
hepatic lesions associated with obesity and diabetes, and their similarity to alcoholic liver disease had
been recognized for a long time, this landmark publication highlighted these findings and named a
"hitherto unnamed disease". Recent improvement in our understanding of the pathophysiology of
non-alcoholic fatty liver disease (NAFLD) has brought into question the appropriateness of the term NASH,
as it defines a disease by what it is not. On the other hand, it has become increasingly clear that in
most individuals, NAFLD is a metabolic syndrome in which obesity, diabetes, hyperinsulinemia and insulin
resistance (metabolic syndrome or syndrome X) play an important role.
The wide range of histological findings of non-alcoholic fatty liver disease ranging from steatosis to
steatosis with inflammation and fibrosis, with variable progression from one stage to another, have
further complicated the use of uniform terminology. Currently, two terms dominate medical literature,
NAFLD (non-alcoholic fatty liver disease) and NASH. The former is an all-inclusive term encompassing the
entire range of histological findings, while NASH is restricted to those cases that show inflammation and
fibrosis. The surgical pathologist faces a third scenario of steatosis with fibrosis and no
inflammation; "steatofibrosis" is often used for this situation.
The advantage conferred on the surgical pathologist to visualize the changes of NAFLD, allows better
terminology in the diagnostic report. We therefore use the following terms in our pathology reports:
Steatosis - with an assessement of the extent of steatosis
Steatohepatitis - reflects additional changes of cellular injury namely, Mallory hyaline, inflammatory
Steatofibrosis - reflects the distinctive pericellular fibrosis seen in NAFLD
With an increasing trend towards obesity in both children and adults, and a projected incidence of 40%
in adults by 2025, 10% of whom will have insulin resistance and diabetes, the focus on NASH is
The difficulty in estimating the true incidence or prevalence of NAFLD
arises from the following facts:
1) The disease is silent in the majority of patients and may only be
discovered during routine medical examination (slightly elevated ALT) or investigation for other medical
conditions (abnormal liver on imaging).
2) There are no diagnostic laboratory tests (no gold standard). LFTs
are neither sensitive nor specific for NAFLD
3) Imaging studies similarly lack sensitivity and specificity for
diagnosis of fatty liver disease.
4) There is uncertainty about the minimal histological criteria
necessary for diagnosis of NAFLD. This reflects uncertainty regarding the natural history and
progression of findings on the lower end of the spectrum, like minimal steatosis.
5) The diagnosis requires the exclusion of alcohol as an etiologic
agent, and there is disagreement on what constitutes medically acceptable alcohol use. Furthermore,
there is wide variability in individual metabolism of alcohol.
6) Studies on prevalence and incidence use variable criteria for
diagnosis of NAFLD, as well as variable levels of alcohol use.
With these limitations in mind, prevalence of NAFLD is either based on imaging studies, biochemical
studies or autopsy studies on selected or unselected populations; the former have focused on patients
most at risk, e.g. patients with diabetes or obesity. Two large population-based studies used data from
15,000 subjects in the third US National Health and Nutrition Examination Survey (NHANES III, 1988-1994).
One group (Ruhl and Everhart) looked at non-diabetic patients with ALT >43 U/L, no clinically
documented liver disease and alcohol consumption of not more than 2 drinks/ day. ALT was raised in 2.8%
of adult non-diabetic subjects; of these 65% were overweight (BMI > 25 kg/m2) or obese (BMI
>30kg/ m2). Other features associated with increased ALT in this analysis included younger
age (39 yrs vs 45 yrs), male gender, Mexican American ethnicity, impaired fasting glucose, increased
waist to hip ratio, increased fasting serum leptin, triglycerides and insulin levels. Forty subjects
with increased ALT fit criteria for the metabolic syndrome which is defined by truncal obesity,
hyperglycemia, low levels of HDL, hypertriglyceridemia and hypertension.
A second study (Clark et al) used the same database but applied different criteria to identify
patients with NAFLD: AST > 37 IU/L and ALT >40U/L for men, AST and ALT > 31 IU/L for women.
They excluded patients with transferrin saturation > 50%, positive viral markers and alcohol
consumption of > 1 drink per day for women and 2 drinks per day for men. The study included diabetic
patients. NAFLD, defined by unexplained elevation in transaminases, was present in 5.5% of the
The prevalence of NAFLD in studies based on histological examination varies, partly because of the
varied populations studied and the varying criteria used for diagnosis.
Hilde found steatosis in 24% of 503 livers from car accident victims at postmortem, of these 6 of 120
cases had features of alcoholic hepatitis. Wanless, in an autopsy series of 351 non-alcoholic patients,
found steatosis in 70% obese and 35% lean patients; steatohepatitis in 18.5% obese and 2.7% lean
patients, and fibrosis in 13.8% vs 6.6% obese and lean patients respectively. Charlton found NASH in
2.6% of 1207 hepatectomies with end-stage liver disease at transplantation. In a biopsy study for
unexplained enzyme elevations, steatosis was found in 34% and steatohepatitis in 32% of 354 biopsy
specimens. Steatosis was found in 14.5% of healthy donors for live related liver transplantation without
enzyme elevations (Marcos, 2000).
Conditions Associated with NAFLD
(a non-comprehensive list of the commoner associations)
- Diabetes mellitus
- Metabolic syndrome X
- Rapid weight loss (jejuno-ileal, gastric bypass)
- Total parenteral nutrition
- Drugs (Amiodarone)
Insulin resistance and hyperinsulinemia are central to the etiopathogenesis of NAFLD.
In the liver, insulin promotes glycogen synthesis, and inhibits gluconeogenesis as well as
mitochondrial β-oxidation of fatty acids. In peripheral tissues, insulin promotes lipogenesis and
inhibits lipolysis. Insulin resistance is associated with a failure to suppress lipolysis, thus
increasing the delivery and uptake of fatty acids in the liver. The net effect is to increase metabolic
turnover of fatty acids either by mitochondrial β-oxidation, packaging and secretion into VLDL or storage
in the liver as triglycerides.
Free fatty acids in blood further worsen insulin resistance. Both free fatty acids and TNF-α are
increased in obesity and both down regulate IRS-1 (insulin receptor substrate-1). IRS-1, which is
activated by tyrosine phosphorylation, promotes translocation of the glucose transporter protein GLUT4 to
the cell membrane where it facilitates uptake of glucose from blood. Both free fatty acids and TNF-a
hinder phosphorylation, and therefore the activity of IRS-1. TNF-alpha is released from adipose tissue
in the absence of inflammation or infection.
The development of steatohepatitis requires factors additional to those that cause fatty liver - a
"second hit". This second hit is mediated by oxidant stress and production of free radicals that cause
lipid peroxidation within hepatocytes. The P450 cytochrome system, TNF, hepatic iron stores, and
lipoxygenase, are all potential sources of free radicals. Fatty acids themselves induce the Cyp2E1
enzyme. In addition, the increased mitochondrial β-oxidation of fatty acids is a source of free radical
Fatty acids also exert a direct deleterious effect on hepatocytes, which explains the existence of
myriad mechanisms to guard against excess accumulation of free fatty acids by binding, transforming,
catabolizing and exporting free fatty acids. These mechanisms include FFA binding proteins, triglyceride
synthesis, secretion as VLDL, mitochondrial β-oxidation and enzymatic removal of lipid peroxidation
The Role of the Liver Biopsy in NAFLD
In the absence of non-invasive markers with acceptable predictive and discriminative value for the
diagnosis of NAFLD, liver biopsy offers the only means of making a definitive diagnosis. The clinical
diagnosis of NAFLD is suspected in individuals with asymptomatic elevations of liver enzymes, increased
echogenicity on ultrasonography, unexplained hepatomegaly, or the presence of diabetes, obesity or
dyslipidemia. Exclusion of other causes of chronic liver disease, like viral, autoimmune, metabolic and
genetic, is a necessary step in the clinical diagnosis of NAFLD. Elevations of liver enzymes are minimal
and non-specific, and may be entirely absent in some cases. Imaging studies are similarly
non-discriminatory, and cannot accurately detect hepatitis and fibrosis. Fibrosis, including septal
fibrosis and cirrhosis, is present in a significant number of NAFLD patients in the initial biopsy.
Therefore, besides being a diagnostic tool, the liver biopsy provides valuable prognostic information as
Although the finding of steatosis was initially considered to be important for diagnosis of NASH, the
greater awareness of this condition and the finding of NASH as a cause of end-stage cryptogenic
cirrhosis, ahs led to a re-evaluation of the necessity of requiring steatosis for a diagnosis of NASH.
It is now considered that steatosis is not an absolute necessity as long as there are other features of
cellular injury like ballooning injury or MH. Steatosis when present is macrovesicular or large-droplet
fat; the presence of exclusively or predominantly microvesicular fat should prompt exclusion of other
pathologic processes. At least 5-10% of fat with a predominant zone 3 distribution is indicative of
Ballooning degeneration is a useful, and in the opinion of most pathologists, a necessary feature for
diagnosis of NASH. Ballooning change consists of enlarged hepatocytes that are usually intermixed with
areas of steatosis and perisinusoidal fibrosis. They are a manifestation of cellular injury and
Lobular inflammation is an integral part of the pathology of NASH. Since the term arose by
extrapolation from alcohol-related liver disease, it was necessarily assumed that neutrophils should form
an integral part of NASH. However, with increasing experience, it has become evident that neutrophils
are not the only inflammatory component present in patients with NASH; the inflammatory component very
often consists of lymphocytes, with or without neutrophils. In fact, portal inflammation with
lymphocytes is the predominant component of NASH in children. Experts in the field are not unanimous
about the requirement or the frequency of PMNs or lymphocytes in NASH. If portal inflammation is in
excess of lobular inflammation, it is imperative to rule out concurrent liver disease, eg hepatitis C.
The presence of perisinusoidal and perivenular fibrosis is an important diagnostic feature of NAFLD.
This type of fibrosis is present predominantly in zone 3. However, it has been increasingly observed that
NAFLD is also associated with portal fibrosis which may show thin incomplete radiating septa. Portal
fibrosis predominates over perisinusoidal fibrosis in pediatric NAFLD.
NAFLD in Children
The incidence of NASH in children is increasing in concert with a sedentary lifestyle and exposure to
diets rich in fats and refined sugars. As in adults, pediatric NAFLD is associated with obesity,
hyperinsulinemia, insulin resistance and the metabolic syndrome. The exclusion of alcohol as an
etiologic factor is a lesser problem in this group of patients, especially if adolescents are not
included. Pediatric NAFLD shows male preponderance and strong association with Hispanic ethnicity.
Histological differences with adults include more severe steatosis; little or no ballooning, Mallory
hyaline, or neutrophils. In addition, pediatric NAFLD shows chronic portal inflammation in preference to
lobular inflammation. Fibrosis occurs predominantly in a portal and periportal distribution, and central
perisinusoidal distribution is seen less often than in adults. There is also a lesser incidence of
centrilobular distribution of NAFLD changes like steatosis, ballooning and Mallory hyaline.
Grading and Staging the Histopathologic Lesions of Nonalcoholic Steatohepatitis
Brunt EM Seminars in Liver Disease 2001; 21: 3-16
|Grade 1, Mild|
| ||Steatosis: predominantly macrovesicular, involves <33% up to 66% of the lobules|
| ||Ballooning: occasionally observed; zone 3 hepatocytes|
| ||Lobular inflammation: scattered and mild acute (polymorphs) inflammation and occasional chronic inflammation (mononuclear cells)|
| ||Portal inflammation: none or mild|
|Grade 2, Moderate|
| ||Steatosis: any degree and usually mixed macrovesicular and microvesicular|
| ||Ballooning: obvious and present in zone 3|
| ||Lobular inflammation: polymorphs may be noted associated with ballooned hepatocytes, pericellular fibrosis; mild chronic inflammation may be seen|
| ||Portal inflammation: mild to moderate|
|Grade 3, Severe|
| ||Steatosis: typically >66% (panacinar); commonly mixed steatosis|
| ||Ballooning: predominantly zone 3; marked|
| ||Lobular inflammation: scattered acute and chronic inflammation; polymorphs may appear concentrated in zone 3 areas of ballooning and perisinusoidal fibrosis|
| ||Portal inflammation: mild to moderate|
|Staging Fibrosis in NASH|
| ||Stage 1: Zone 3 perivenular perisinusoidal/pericellular fibrosis, focal or extensive|
| ||Stage 2: As above with focal or extensive periportal fibrosis|
| ||Stage 3: Bridging fibrosis, focal or extensive|
| ||Stage 4: Cirrhosis|
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