Steatosis, defined as the accumulation of triacylglycerides in hepatocytes, is a frequent finding in most liver biopsies.¹ As much as 5% of the liver parenchyma may be composed of lipid. By convention, any quantity of lipid that occupies more than 5% of the liver mass is considered pathologic.

Macrovesicular steatosis is an accumulation of lipid droplets of various sizes in hepatocytes in which the cell nuclei are displaced peripherally. It is common to observe small and large droplets in the same cell and to find isolated droplets of lipid.

Microvesicular steatosis has a different appearance and clinical significance. The nuclei of affected hepatocytes are typically centrally located within abundant, foamy-appearing cytoplasm. Special stains, such as Oil Red O, may be necessary to confirm lipid accumulation, whereas in macrovesicular steatosis, the large and small lipid droplets are easily detectable by routine hematoxylin and eosin stain. Diseases associated with microvesicular steatosis are characterized by liver failure and hepatic encephalopathy rather than chronicity and potential fibrosis; the former features are the result of mitochondrial β-oxidation defects. The most common examples of purely microvesicular steatosis are acute fatty liver of pregnancy and Reye syndrome. This process has also been reported in the setting of drug toxicity. Drug-related entities are discussed in Chapter 48.

Alcohol-induced liver disease (ALD) refers to the spectrum of liver disorders directly related to excessive alcohol use. This chapter focuses primarily on the pathologic lesions of the ALD subset characterized by accumulation of fat in the liver parenchyma.

Nonalcoholic fatty liver disease (NAFLD) occurs without significant alcohol use. The clinicopathologic entity is characterized by a variety of hepatic parenchymal injury patterns with more than 5% macrovesicular steatosis.

The term nonalcoholic steatohepatitis (NASH) is in the same clinical setting as NAFLD, but NASH is associated with specific pathologic lesions in the liver. The lesions include various amounts of macrovesicular steatosis, hepatocellular injury (most commonly seen as ballooning), and inflammation in the hepatic lobules or portal tracts, or both. These lesions and others that may be seen in steatohepatitis are discussed in this chapter.

Alcohol-Induced Liver Disease

Clinical Features

The clinical features of ALD vary considerably. Table 49.1 highlights the primary clinical manifestations of the four main clinicopathologic subtypes of ALD: fatty liver, alcoholic hepatitis, alcoholic cirrhosis, and alcoholic foamy degeneration.

Table 49.1

 

Symptoms, Signs, and Laboratory Abnormalities of Alcohol-Induced Liver Disease

Symptoms and Signs	Fatty Liver	Steatohepatitis and Alcoholic Hepatitis	Cirrhosis	Foamy Degeneration
Asymptomatic	+++	+	Portal hypertension	−
Right upper quadrant discomfort	+++	Pain, not mild	+ to ++	+
Hepatomegaly	++	++	+ to ++	+
Elevated AST, normal ALT	++	+	+ (normal if abstinent)	++
AST/ALT ratio	AST > ALT	AST/ALT > 1-3 common	AST/ALT > 1	AST > ALT
Marked AST elevation	<5 times normal	As much as 5 times normal	−	++
Marked alkaline phosphatase elevation	−	+/−	−	++
Bilirubin elevation	−	>10 mg/dL	− (unless advanced)	++
Jaundice	−	Cardinal sign	As above	++
Elevated white blood cell count	−	>10,000/mm3	− (may be decreased)	−
Fever	−	+++	+ (with infection)	−
Ascites	−	+++	+ (advanced)	−

ALT, Alanine aminotransferase; AST, aspartate aminotransferase; −, does not occur; +, occurs uncommonly; ++, common; +++, very common.

 

Patients with ALD and fatty liver may be asymptomatic or may have right upper quadrant discomfort at presentation (see Table 49.1). Patients with alcoholic hepatitis usually have abdominal pain and pancreatitis, whereas patients with cirrhosis and alcoholic foamy degeneration often experience only vague abdominal discomfort. In all forms of ALD, the liver is typically enlarged. Fever, an elevated white blood cell count, and jaundice occur in various degrees with the different forms of ALD and may be related to ALD (e.g., alcoholic hepatitis) or to a secondary infection (e.g., peritonitis caused by alcoholic cirrhosis).

At clinical presentation, patients may have elevated serum levels of aminotransferases (i.e., aspartate aminotransferase [AST] and alanine aminotransferase [ALT]), and a ratio of AST to ALT greater than 1 is useful in the evaluation of patients with ALD. AST levels are typically greater than ALT levels in all forms of ALD. Radiographic evaluation plays a role in screening for hepatocellular carcinoma in patients with cirrhosis and in the clinical care of critically ill patients with pancreatitis and suspected intestinal ileus.

Epidemiology and Prevalence

Worldwide, ALD has no social or ethnic barriers, but it is uncommon in very young and very elderly individuals. Both genders may be affected, although women are at higher risk. In the United States, 7.4% of the population meet the criteria for alcohol dependence or abuse. It is the leading cause of end-stage liver disease and liver-related mortality and follows only heart disease and cancer as the major health issue in the United States.¹¹ The reported rates of per capita liters of alcohol consumed per year by individuals older than 15 years of age varies throughout the world: 13.9 in the Russian Federation; 12.9 in Germany, France, and the United Kingdom; 9.3 to 8.5 in North America, Japan, and Australia; 5.0 in China, Philippines, and Vietnam; 1.3 in Iran and Saudi Arabia; and 0.6 in Afghanistan and Pakistan.¹¹ Alcohol use may combine synergistically with other forms of chronic liver disease to generate progressive disease, malignancy, and morbidity.¹²

Familial Associations

Family, twin, and ethnic studies have confirmed a genetic susceptibility to ALD.¹¹ Factors under investigation include gender, genes involved in the metabolism of alcohol (particularly in East Asians), cytochrome P450 2E1 (CYP2E1) and other enzymes involved in the pathways of oxidative stress, various cytokines (e.g., tumor necrosis factor-α [TNF-α], interleukin 10 [IL10]), genes involved in the oxidation of hepatic fat, mechanisms of matrix deposition and degradation in fibrosis, and immune reactions to endotoxin and toxic adducts.¹³

Risk Factors

Liver disease does not develop in all individuals who abuse alcohol. The risk for ALD, even among heavy drinkers, is less than 10% to 15%. Known risk factors include the amount of alcohol consumed during a 10- to 20-year period (>60 g/day for men; 20 to 30 g/day for women), gender (female > male), central obesity, patterns of consumption (nonmealtime > mealtime; daily > weekend only; various types of beverages rather than one type), associated medications (acetaminophen), amount of coffee intake, and genes that regulate expression of proinflammatory cytokines and immune response mechanisms.^13–18 A sequence variation in the patatin-like phospholipase 3 (PNPLA3) gene, which encodes adiponutrin, is associated with the development of cirrhosis in white alcoholics.¹⁹ The same gene variation was previously found to correlate with steatosis in different populations around the world and with features of progressive disease in NAFLD patients.²⁰

Pathogenesis

Alcohol-induced liver damage is a multifactorial process that involves alterations of the gut, gut microbiota, and hepatocellular metabolic pathways and activation of the innate and adaptive immune systems. Resultant outcomes are activation of proinflammatory and profibrogenic molecular mechanisms in specific liver cells.

The primary mechanism of alcohol-induced liver injury is hepatocellular oxidative stress.^21,22 Ethanol is metabolized to acetate in the liver mainly by two enzyme systems: the dehydrogenases (i.e., alcohol and aldehyde), which produce acetate and result in steatosis, and the microsomal ethanol-oxidizing system (MEOS). The direct hepatotoxic effects of alcohol are caused primarily by acetaldehyde. Oxidative stress and free radical production result in mitochondrial damage, depletion of the antioxidants such as reduced glutathione, toxicity by free radicals, and induction of lipid peroxidation. Acetaldehyde-formed toxic adducts bind to proteins and lead to additional hepatocellular injury, serve as neoantigens for initiation of the adaptive immune response, and promote collagen production by hepatic stellate cells. The net effect of acetaldehyde production in hepatocytes is the accumulation of intracellular proteins, lipids, water, and electrolytes with the loss of the hepatocyte structural keratins 8 (K8) and 18 (K18). Histologically, they manifest as cellular enlargement, ballooning, empty-appearing cells, steatosis, and loss of immunoreactivity for K8 and K18.^23,24

The key enzyme of the MEOS is the ethanol-inducible CYP2E1, which is located primarily in the endoplasmic reticulum of acinar zone 3 hepatocytes. Activation of this enzyme system results in the production of reactive oxygen species, such as hydrogen peroxide (H₂O₂) and superoxide anions (O₂⁻), which cause further lipid peroxidation of cell membranes. This enzyme system also metabolizes acetaminophen and produces the toxic metabolite N-acetyl-p-benzoquinone imine (NAPQI). Even small amounts of acetaminophen may augment depletion of the antioxidant capabilities of the liver in chronic alcoholism.

Other sources of ethanol-induced oxidative stress include endotoxin-activated Kupffer cells, functionally impaired mitochondria, and ferric iron accumulation. Oxidative stress promotes hepatocellular injury, apoptosis, and necrosis; proinflammatory cytokine production (e.g., TNF-α, IL1, IL6); and parenchymal inflammation with polymorphonuclear leukocytes (PMNs) and mononuclear cells and perisinusoidal fibrosis.^25,26

Excessive alcohol intake leads to increased gut permeability and increased portal venous exposure to gut-derived endotoxins. This process results in Kupffer cell activation through the CD14/toll-like receptor 4 (TLR4) complex and activation of the innate immune response.^22,26

Increased lactate production, another downstream effect of ethanol metabolism, manifests as hyperuricemia and impaired carbohydrate metabolism. Hypoglycemia may occur. Chronic alcohol use impairs protein production and secretion. Multiple aberrations of lipid metabolism result in increased triglyceride production and subsequent accumulation (i.e., steatosis).

The mechanisms of fibrosis are under active scientific investigation. Ultimately, there is an imbalance between collagen deposition and degradation by activated hepatic stellate cells and portal myofibroblasts. Unique to ALD is the additional involvement of hepatocytes and sinusoidal endothelial cells in fibrogenesis through the production of transforming growth factor-β (TGF-β) and fibronectin isoforms.²⁷ Hypoxia induced by ethanol metabolism in zone 3 hepatocytes upregulates vascular endothelial growth factor.^26,27

Pathology

Grossly, steatosis enlarges the liver and imparts a greasy appearance. Livers with advanced fibrosis or cirrhosis may be enlarged and firm and contain micronodules (≤3 mm in diameter) scattered throughout the parenchyma. In some cases, fibrotic livers may be small and firm. With time, the nodules may coalesce to form larger ones, at which point determination of the underlying cause may not be possible. Hepatocellular carcinomas typically stand out on background cirrhosis as raised, green-tinged or darker nodules that usually are larger than 8 mm in diameter.

The histologic pattern of tissue injury in patients with alcohol-induced fatty liver disease initially involves acinar zone 3 (roughly equivalent to perivenular) hepatocytes. In patients with cirrhosis, steatosis or steatohepatitis may or may not persist.

Steatosis

Steatosis, the earliest pathologic finding in ALD, is not consistently found in all forms of ALD.^21,28 Alcoholic steatosis is typically macrovesicular and is characterized by large intracellular droplets of lipid in hepatocytes (single or multiple) that peripherally displace the cell nucleus. A minor component of lobular chronic inflammation or mild portal inflammation may be identified, but fibrosis is usually absent.

Steatohepatitis

Steatohepatitis is defined by the presence of both steatosis and hepatocyte injury. Injured hepatocytes may appear ballooned, apoptotic, or lytic. Confluent and bridging necrosis are rarely found in ALD.

Hepatocellular ballooning is considered an essential feature of steatohepatitis by most hepatopathologists.⁷ Ballooned hepatocytes are enlarged cells with rarefied, reticulated cytoplasm (Fig. 49.1) that may or may not contain fat droplets and intracytoplasmic material referred to as Mallory-Denk bodies (MDBs) or Mallory hyaline. Ballooned hepatocytes are located predominantly in zone 3 and are commonly associated with some degree of perisinusoidal fibrosis (Fig. 49.2).²⁹

Alcoholic Hepatitis

Patients with alcoholic hepatitis might not have steatosis. Histologic features of alcoholic hepatitis include hepatocyte ballooning, apoptotic bodies, MDBs with satellitosis, canalicular cholestasis, dense perisinusoidal fibrosis, and a perivenular lesion often referred to as sclerosing hyaline necrosis (i.e., MDBs, satellitosis, and obliterative fibrosis of the outflow vein). Portal hypertension may occur with sclerosing hyaline necrosis. Bridging fibrosis and a ductular reaction may be identified. The absence of steatosis does not rule out alcohol-induced hepatitis.

Alcoholic Foamy Degeneration

Alcoholic foamy degeneration is an unusual and often serious type of ALD associated with marked elevations of aminotransferases, γ-glutamyltransferase (GGT), alkaline phosphatase, and bilirubin.³⁵ Histologically, it is characterized by diffuse, primarily microvesicular steatosis without inflammation or ballooning or by marked fibrosis with or without canalicular cholestasis (Fig. 49.7). MDBs are uncommon. Perivenular and perisinusoidal fibrosis may be evident. Clinically, the disorder mimics extrahepatic biliary obstruction, but the liver biopsy findings can be used to distinguish the two entities. Alcoholic foamy degeneration is reversible with abstinence from alcohol.

Fibrosis and Cirrhosis

The characteristic pattern of early fibrosis in patients with noncirrhotic alcoholic steatohepatitis or alcoholic hepatitis is pericellular or perisinusoidal. This pattern results from deposition of collagen in the space of Disse and is commonly referred to as chicken wire fibrosis (Fig. 49.8).

Megamitochondria (i.e., giant mitochondria) are identified by light microscopy as intracytoplasmic, round or cigar-shaped, eosinophilic structures in hepatocytes, and they are most readily seen in hepatocytes that contain microvesicular steatosis (Fig. 49.13). Megamitochondria may also occur in normal pregnancy, acute fatty liver of pregnancy, and Wilson disease.

Iron Deposition.

Mildly increased iron deposition is common in most forms of ALD. However, in a minority of cases, it may be moderate or severe.²³ Possible reasons for hemosiderosis in ALD include increased dysregulation of hepcidin production,⁴³ intestinal iron absorption, iron in some types of alcoholic beverages, hemolysis related to spur cell anemia, and upregulation of the transferrin receptor.¹⁸ With a modified Perls stain, iron deposition is usually nonzonal.^30,37 In ALD-related cirrhosis,³⁷ iron deposition may vary in quantity between individual regenerative nodules (Fig. 49.14).

Acute and Chronic Cholestasis.

Any of the following pathologic processes related to ALD may result in intrahepatic cholestasis, which is characterized by intracanalicular bile plugs: severe fatty liver, alcoholic pancreatitis due to gallstones or alcohol, alcoholic foamy degeneration, alcoholic hepatitis, and decompensated alcoholic cirrhosis. Canalicular bile thrombi are observed in 15% to 32% of livers with ALD (Fig. 49.15). Severe fatty liver with portal tract features of cholangitis and cholestasis is a specific clinicopathologic cholestatic syndrome in patients with ALD. Bile stasis in ALD also may be a manifestation of superimposed viral hepatitis or drug-induced jaundice.²¹ In patients with established ALD-associated cirrhosis, a copper stain may show mild positivity, which represents chronic retention of bile caused by the cirrhotic remodeling.

Vascular Lesions.

Sclerosing hyaline necrosis, which is a marker of severe alcoholic hepatitis, and obliterative vein lesions may be responsible for the development of noncirrhotic portal hypertension in patients with ALD. Sclerosing hyaline necrosis consists of a combination of acinar zone 3 lesions, including dense perivenular and perisinusoidal fibrosis (i.e., sclerosis), occlusion of terminal hepatic venules, MDBs, and liver cell necrosis and loss, which results in the formation of large perivenular scars. Lesions of the terminal hepatic venules and sublobular veins include perivenular fibrosis, phlebosclerosis, and, less frequently, lymphocytic phlebitis (Fig. 49.16).⁴⁰

Treatment Effects.

Alcoholic steatosis may resolve completely within 4 weeks of alcohol abstinence.³⁰ However, lesions associated with alcoholic hepatitis or steatohepatitis may persist for as long as 6 months after alcohol withdrawal, albeit with decreased severity.⁴⁴ With abstinence, there is usually an increase in the degree of portal lymphocytic infiltration.⁴⁴

Prognostic Lesions.

Perivenular fibrosis in steatosis in the absence of cirrhosis portends progression unless abstinence occurs. Acute cholestasis correlates with poor survival of hospitalized patients with ALD.²¹ These patients have a reduced 5-year survival rate (22%) compared with patients with ALD who have no or mild cholestasis (54%).²³ The number of apoptotic bodies correlates positively with severe histologic and clinical disease activity.^45,46 Alcoholic hepatitis, sclerosing hyaline necrosis, and severe steatosis represent poor-prognostic lesions in patients with ALD, particularly in those with cirrhosis.^21,47 In noncirrhotic patients, mixed steatosis and the finding of megamitochondria in the initial liver biopsy of alcoholic patients without steatohepatitis correlate with an increased risk of subsequent fibrosis or cirrhosis on follow-up.⁴⁸ In acute-on-chronic liver failure caused by ALD, marked ductular bilirubinostasis and many MDBs are significant predictors of in-hospital mortality.⁴⁹

Lesions in the Allograft Liver.

Any of the previously described lesions may manifest as recurrent or de novo ALD in a liver allograft. Care is taken to evaluate other features of allografts, particularly acute and chronic rejection and recurrence of hepatitis C (see Chapter 46).

Natural History and Treatment

The financial and social burdens associated with ALD are significant.^11,50 Alcoholic hepatitis is associated with a significant risk of hospitalization and loss of time from work, morbidity and short-term mortality, and progression to cirrhosis within 5 years for as many as 70% of individuals. Few affected individuals are able to reverse the disease process with complete abstinence.¹¹ A 12-year historical study from a national registry of Danish subjects comparing biopsy-proven ALD with the general population showed that the cirrhosis risk was higher for affected women than men and twice as high for patients with steatohepatitis than those with pure steatosis.⁵¹

Chronic alcohol abuse is responsible for as many as 45% of cases of hepatocellular carcinoma in Western countries,⁵² a rate that is higher than that for chronic hepatitis C.¹⁸ Coexistent HCV infection has an additive effect on the risk of carcinoma,¹¹ and other comorbid factors, such as chronic hepatitis B, hemochromatosis, obesity, and cigarette smoking, contribute to an increased risk of hepatocellular carcinoma and mortality in patients with ALD.^53,54 Intrahepatic cholangiocarcinoma also has been linked to ALD.⁵⁵

The mainstay of treatment for ALD is abstinence from alcohol. One of the pharmacologic methods of treatment of alcoholism, cyanamide, may produce isolated, nonzonal eosinophilic globules in hepatocytes. Treatment of hospitalized patients with alcoholic hepatitis includes steroids and the antioxidants pentoxifylline and S-adenosyl-l-methionine (SAMe). For abstinent patients with cirrhosis, liver transplantation is an accepted treatment option. More controversial is a practice of liver transplantation for patients with acute alcoholic hepatitis.^56,57

Nonalcoholic Fatty Liver Disease

Clinical Features

Individuals with NAFLD may be asymptomatic or may show mild, nonspecific symptoms such as fatigue and right upper quadrant discomfort or pain (Table 49.2). Fever is not a feature of uncomplicated NAFLD; when present, it suggests a different or concomitant disorder.

Table 49.2

 

Symptoms, Signs, and Laboratory Abnormalities of Nonalcoholic Fatty Liver Disease

Symptoms and Signs	Fatty Liver	Steatohepatitis	Cirrhosis
Asymptomatic	++	+	Portal hypertension
Right upper quadrant discomfort	++	++	+
Hepatomegaly	++	++	+
Elevated AST, normal ALT	−	−	−
AST/ALT ratio	AST/ALT < 1	AST/ALT < 1	AST/ALT ≥ 1
Marked AST elevation	−	−	−
Marked alkaline phosphatase elevation	−	−	−
Bilirubin elevation	−	−	−
Jaundice	−	−	−
Elevated white blood cell count	−	−	−
Fever	−	−	−
Ascites	−	−	+ (advanced)

ALT, Alanine aminotransferase; AST, aspartate aminotransferase; −, does not occur; +, occurs uncommonly; ++, common; +++, very common.

 

The most common clinical feature of NAFLD is central (truncal or abdominal) obesity, which is best assessed by measurement of waist circumference. For adult men, central obesity is a waist circumference greater than 102 cm, and for adult women, it is greater than 88 cm. Obesity is defined by body mass index (BMI), which is the weight in kilograms divided by the height in meters squared (kg/m²). Compared with waist circumference, it is less specific and does not assess the anatomic location of the metabolically unique subcutaneous and visceral fat depots or shape of the body habitus. For non-Asian adults, the current definitions used are overweight (BMI > 25 kg/m²) and obesity (BMI > 30 kg/m²). Because the Asian body habitus is deceptively thin and able to conceal visceral adiposity, lower BMI values for overweight and obesity have been recommended.⁵⁸

Clinical evidence of hepatomegaly may be found in patients with an enlarged, fatty liver. Ascites and other features of portal hypertension may be identified in patients with NAFLD-associated cirrhosis. Obstructive sleep apnea and acanthosis nigricans are recognized associations of NAFLD.

Laboratory values commonly assessed for patients with NAFLD include ALT, AST, and GGT. Patients also undergo tests for hyperglycemia (glycated hemoglobin [HbA_1c] is used to evaluate long-term glycemic control), tests for insulin resistance (based on calculations of fasting glucose and insulin levels by using quantitative insulin sensitivity check index [QUICKI] and homeostasis model assessment [HOMA-IR]), and tests of lipid parameters.²¹

The overall sensitivity of liver enzymes for detection of fatty liver disease is poor.^21,59,60 For instance, patients with normal ALT levels may have steatosis or advanced steatohepatitis⁶¹ or silent cirrhosis.^62,63 If elevated, the ALT value is greater than the AST value, and an ALT/AST ratio greater than 1 is often found in cases of precirrhotic NAFLD; the reverse situation occurs in ALD. Autoantibodies, particularly antinuclear antibody and rarely antimitochondrial antibody, have been reported in as many as 34% of individuals with NAFLD.⁶⁴ In some, only further detailed clinical testing or liver biopsy can distinguish autoimmune hepatitis from concurrent autoimmune hepatitis with NAFLD or NAFLD alone.⁶⁴ A possible association of autoantibodies with more severe disease has been suggested but not confirmed.^65,66 In combination with the proven lack of reproducible gross evaluation of the liver by surgeons, authorities recommend a liver biopsy for all high-risk and bariatric patients,^67–69 especially when other causes of steatosis have been excluded.⁵⁹

Abnormal iron study results are a common feature in patients with NAFLD. For instance, elevated serum levels of ferritin (but not transferrin saturation) may be found in patients with NAFLD and may cause clinical concern about an iron overload disorder.⁷⁰ The serum ferritin level in NAFLD patients has been reported as an independent predictor of the histologic severity of underlying liver disease.⁷¹ An elevated serum ferritin level may indicate hepatocyte necrosis and systemic inflammation, and it is a recognized feature of the metabolic syndrome (discussed later). However, iron overload itself is not a feature of type 2 diabetes mellitus. The associations of iron dysregulation, HFE (hemochromatosis) mutations, and fibrosis with the metabolic syndrome remain subjects of debate.⁷⁰ A meta-analysis did not support an association between HFE genetic variants and NAFLD.⁷²

The proinflammatory iron regulator hepcidin is expressed in the liver and adipose tissue of obese patients, and this finding correlates with low transferrin values in 68% and anemia in 24% of these individuals.⁷³ Demonstration of hepcidin mRNA and protein in adipose tissue is a new finding and is the subject of iron and adipokine studies.⁷³

Laboratory tests are indicative of the complexity of this condition. Markers of systemic inflammation, such as elevated levels of C-reactive protein, fibrinogen, and plasminogen activator inhibitor 1, are found in the setting of NASH and fibrosis.⁷⁴ A depressed level of adiponectin, an endocrine product of visceral adipose tissue, is recognized as a pathologic feature of NAFLD or NASH.⁷⁴ Some work has implicated additional factors as being proinflammatory (e.g., hemoglobin) in subsets of individuals with NAFLD but without the metabolic syndrome.⁷⁵ In contrast with ALD, leukocytosis, cholestasis, and jaundice are not typical features of uncomplicated, noncirrhotic NASH.

By ultrasonography, computed tomography (CT), or magnetic resonance imaging (MRI), the characteristic finding in NAFLD is increased hepatic fat. The ultrasound finding is referred to as a bright liver. Newer types of imaging studies use methods to semiquantitate the lipid content of the liver.⁷⁶ However, when steatosis involves less than 33% of the liver parenchyma, imaging tests may not detect it.⁷⁷ Imaging is also not an adequate replacement for tissue analysis for determination of the degree of disease activity or the stage of fibrosis in patients with precirrhotic NASH.⁷⁸

Epidemiology and Prevalence

The epidemiology and prevalence of NAFLD are understood in terms of its relationship with obesity and increased risks of stroke and heart disease, a group of associations referred to as metabolic syndrome. The common underlying feature of this syndrome is systemic insulin resistance. Studies have confirmed associations of NAFLD with hepatic insulin resistance, cardiovascular diseases,⁷⁹ and the metabolic syndrome in adults⁸⁰ and children.⁸¹ In one definition of metabolic syndrome, a patient must have at least two of the following disorders plus type 2 diabetes mellitus, insulin resistance, or glucose intolerance: obesity (BMI ≥ 30); arterial hypertension (≥140/90 mm Hg); dyslipidemia as low levels of high-density lipoprotein (<0.9 mmol/L) or hypertriglyceridemia (>1.7 mmol/L); and microalbuminuria (>20 µg/min).⁸² The sole finding of fatty liver may indicate progression of the metabolic syndrome⁸³ and progression of liver disease.⁸⁴

NAFLD is a worldwide disorder and has been documented in almost all cultures. This global phenomenon is closely related to acquisition of a Western diet and lifestyle, which includes less physical activity. Initially thought to be a disease predominantly of overweight women, the entire spectrum of NAFLD is now recognized as common among men of all ages, but severe NASH and advanced fibrosis remain more common among adult women.^21,59 Several studies have confirmed an underrepresentation of this disease process in African Americans of all age groups and an overrepresentation in Hispanics.⁵⁹

Once referred to as a disease of the affluent, NAFLD has become recognized as everyman’s disease.⁸⁵ Future projections of the rates of obesity, type 2 diabetes mellitus, and their complications in the developing world for the year 2030 are staggering. They range from 18.6% in sub-Saharan Africa to 79.4% in India. The projected percent positive change for type 2 diabetes mellitus incidence is 32% in Europe, 72% in the United States, 150% in India, 162% in sub-Saharan Africa, and 164% in the Middle East.⁸⁶

Having considered the worldwide and multicultural nature of NAFLD, some studies have evaluated certain ethnic proclivities for development of this disease. For instance, an image-based study of hepatic steatosis from a large, urban, multiethnic population in the United States⁷⁶ found the following relative prevalence rates: Hispanic > white > African American. Results from a subsequent clinic-based study that used liver tests and liver biopsy results differed only slightly,⁸⁷ showing the following ethnic prevalence rates: white > Hispanic> Asian. African Americans represented only 3% of patients in this cohort. Others have observed an apparent underrepresentation of African Americans in cohorts of patients with NAFLD.⁸⁸ A small biopsy series from an inner-city hospital in the United States that primarily serves African Americans confirmed the relative scarcity of the disease in this patient population. Less than 2% of 320 liver biopsies showed clinicopathologic evidence of NAFLD, even though the BMI of the cohort was between 26.9 and 32.7.⁸⁹

Unfortunately, most of the early prevalence studies were performed without a full understanding of the link between the metabolic syndrome and fatty liver disease. In later studies, surrogate markers such as elevated ALT levels and a bright liver on ultrasound were used to examine study populations. However, similar to pathologic studies, the results vary widely, which is partly related to the criteria used to define the positivity of test values. For instance, in studies that relied on elevated serum aminotransferase levels, recognized shortcomings included underrecognition of positive cases with normal test values, determination of criteria for normal values in an increasingly obese population,⁸⁴ determination of reliable ALT values from frozen sera, and ascertainment of the levels of alcohol intake.⁹⁰ Methods of assessment of body habitus (i.e., BMI, waist-to-hip ratio, midthigh measurements, and more sophisticated and expensive calculations) also vary.⁷⁸ Commonly cited studies are listed in Table 49.3.

Table 49.3

 

Prevalence of Nonalcoholic Fatty Liver Disease among U.S. Adults Based on NHANES III Data*

Study	Criteria	Prevalence of NAFLD
Clark et al, 2002274	Any elevation of AST, ALT, GGT	23% of all U.S. adults
		31% male
		16% female
		39% of obese men, 40% of obese women
Clark et al, 2003275	± patients with diabetes	AST > 37 U/L, ALT > 40 U/L (men)
		AST, ALT > 31 U/L (women)
		7.9% of all U.S. adults; 69% unexplained (5.5% of adults)
Ruhl and Everhart, 2003276	Elevated ALT only; excluded patients with diabetes	ALT > 43 U/L (men and women)
		2.8% elevated ALT, 65% explained by elevated body mass index

^* Limitations of NHANES data: autoimmune hepatitis, primary biliary cirrhosis, Wilson’s disease, and α₁-antitrypsin deficiency not excluded rigorously and absence of liver pathology for confirmation or exclusion.

ALT, Alanine aminotransferase; AST, aspartate aminotransferase; GGT, γ-glutamyltransferase; NAFLD, nonalcoholic fatty liver disease; NHANES III, Third National Health and Nutrition Examination Survey.

 

Validation of the findings of the Third National Health and Nutrition Examination Survey (NHANES III) has emerged from subsequent studies. One calculated hepatic lipid levels by proton magnetic spectroscopy in a multiethnic population in the southwestern United States and found that 31% of 2287 unselected individuals had a greater than 5.5% rate of hepatic steatosis. Of these, 79% had normal ALT values.⁷⁶ A contemporaneous study from a large clinic population in California showed that of 742 individuals with newly diagnosed chronic liver disease, 21.4% had NAFLD.⁸⁷ NAFLD was found to be the most common cause of incidental liver function test abnormalities (26.4%) in a U.K. prospective primary care cohort of 1118 adults by a combination of ultrasound and biomarker assays; ALD was a close second (25.3%). Of the NAFLD cases, 7.6% had evidence of previously undetected advanced fibrosis.⁹¹

A U.S. biopsy-based prevalence study of asymptomatic adults found that 46% of 156 adult, unselected patients who had steatosis on ultrasound had NAFLD diagnosed on biopsy. Of the biopsied NAFLD patients, 30% had diagnostic NASH, and 2.7% of those with NASH had advanced fibrosis (≥stage 2). This finding allowed speculation that more than 2 million of the U.S. adult population was at risk for advanced fibrosis because of NASH. These findings led the researchers to conclude that the prevalence of NAFLD was greater than that of hepatitis C and posed a serious concern for the future of health care.⁹²

A study in a predominantly nonaffluent, nonobese population in India represented by 1911 inhabitants of a rural community showed the prevalence of NAFLD diagnosed by ultrasonography and CT to be 8.7%. NASH was seen in 31% of 36 biopsied persons with potentially significant disease, 4 of whom had cirrhosis.^93,94

The estimated prevalence rate of NAFLD in the United States is greater than that of ALD (Table 49.4). It may affect as many as 5% of the adult population, with an even higher rate among patients with type 2 diabetes.⁵⁹ Table 49.5 summarizes several published series focused on histologic documentation of fatty liver in adult autopsies. Correlations between body weight and diabetes were discussed in many of these studies long before the metabolic syndrome was identified as a specific entity. More than one investigator suggested that steatosis was a normal consequence of aging (see Pathogenesis).

Table 49.4

 

Prevalence Estimates of Fatty Liver Disease Compared with Other Chronic Liver Diseases

Disorder	Prevalence
Nonalcoholic fatty liver disease	25-75% of obese, diabetic adults 16% of 15- to 19-yr-old adolescents 38% of obese adolescents
Hepatitis C	1.3-2.0%
Alcohol-induced liver disease	1%
Hepatitis B	0.3-0.4%
Hereditary hemochromatosis	1/200-400 of northern European descent
Autoimmune hepatitis, primary biliary cirrhosis, primary sclerosing cholangitis	9-17/100,000
α1-Antitrypsin deficiency	1/1500-7600
Wilson disease	1/30,000

Data from McCullough AJ. Pathophysiology of nonalcoholic steatohepatitis. J Clin Gastroenterol. 2006;40(suppl 1):S17-S29 and Yu AS, Keeffe EB. Elevated AST or ALT to non-alcoholic fatty liver disease: accurate predictor of disease prevalence? Am J Gastroenterol. 2003;98:955-956.

 

Table 49.5

 

Prevalence of Fatty Liver Disease in Autopsy Studies

Study	Population Studied (N)	Prevalence of Fatty Liver	Associated Features
Hilden et al, 1977277	Motor vehicle accident victims; adults (503)	24% overall 0.5% had features of alcoholic hepatitis	Age: 1% < 20 yr 18% 20-40 yr 39% > 60 yr Overweight: associated with steatosis, not age
Underwood Ground, 1982278	Aircrew, accidental deaths, all men (199)	15.6% overall 2.1% had features of alcoholic hepatitis	All healthy; none had overt alcoholism
Hornboll and Olsen, 1982279	678 Consecutive autopsies, all > 20 yr of age (396 with alcohol and other exclusions)	54% overall 43% had < 33% 11% had > 33%	Age: no association after 40 yr Overweight: strong association with degree of fatty change; diabetic people were more common in this group Diabetes not independently associated
Underwood Ground, 1984280	Aircraft, motor vehicle accidents, all men; all accidents associated with alcohol, other illnesses excluded (166)	21%	Questioned possible irregular use of alcohol as factor; discussed increased carbohydrate intake relationship to fatty liver disease
Wanless, 1990176	Hospitalized, 207 obese patients, 144 matched, nonobese controls; alcoholic patients excluded (351)	Steatosis: 36.3% overall, 29.2 obese, 7.1% lean Steatohepatitis: 18.5% obese, 2.7% lean	Overweight: prevalence and grade of steatosis and steatohepatitis correlated with grade of obesity Diabetes: increased the risk of steatohepatitis and fibrosis
Schwimmer et al, 2006231	Consecutive pediatric and adolescent autopsies in a single county, nonselected (742)	Steatosis: 13% overall 2-4 yr: 0.7% 15-19 yr: 17.3%	Age: fatty liver incidence increased with age Obesity: 38% had fatty liver Ethnicity: Hispanic (11.8%), Asian (10.2%), white (8.6%), African American (1.5%)

 

In summary, the true prevalence rate of NAFLD remains unknown because there are no reliable or specific noninvasive tests available for an accurate diagnosis or exclusion of this process. A review of noninvasive markers in NAFLD, NASH, and fibrosis indicates the efforts in the field.²¹ Although liver biopsy evaluation is considered the gold standard of diagnostic tests, it cannot be considered an appropriate screening tool. Results from biopsy studies reinforce the observations that not all obese individuals have hepatic steatosis^8,95,96 and that unexplained elevated ALT values cannot be assumed to indicate fatty liver disease. For example, Skelly and colleagues⁹⁷ showed that as many as one third of 354 biopsies performed for unexplained elevated ALT levels were histologically normal or had abnormalities because of liver diseases other than NAFLD or NASH. Recommendations for the focused role of liver biopsy for patients with suspected NAFLD or NASH were discussed in the practice guidelines from the American Association for the Study of Liver Diseases⁵⁹ and in a consensus conference of the European Association for the Study of the Liver.⁶⁹

Familial Associations and Risk Factors

The genetic predispositions found in NAFLD are exemplified by differences in the incidence of other risk factors, such as obesity, diabetes, hypertension, and hyperlipidemia, among various ethnic populations. Studies have documented fatty liver disease and cirrhosis among kindreds.⁹⁸ One study of 20 patients showed a trend toward familial clustering and maternal linkage for insulin resistance among patients with NAFLD.⁹⁹

Type 2 diabetes and obesity are risk factors for progressive disease in clinical¹⁰⁰ and histologic studies.^8,101,102 Various possible environmental and genetic factors have been proposed for NAFLD,^13,103 including the type of diet, exercise, smoking,¹⁰⁴ the role of small bowel bacterial overgrowth, endotoxin and related immune or cytokine responses, polymorphisms for genes that control fatty acid oxidation (e.g., microsomal transfer protein), and fibrosis (i.e., angiotensinogen and TGF-β).

Broad, nonexclusive categories of genes under investigation as potential risk factors for NAFLD¹⁰⁵ include those involved in the control of adiposity and insulin resistance (e.g., peroxisome proliferator–activated receptor-γ [PPAR-γ], central versus peripheral fat depots); determinants of hepatic steatosis (i.e., free fatty acid delivery and de novo lipogenesis, processing, and egress); hepatic fatty acid oxidation pathways (e.g., mitochondrial, peroxisomal, microsomal); hepatic oxidative stress mechanisms (e.g., production, defense); cytokine effects (e.g., TNF-α, IL6, adipokines); and determinants of progression of fibrosis. One genomic and proteomic study of 98 bariatric patients found downregulation of genes involved in defense against oxidative stress in NAFLD cases and upregulation of genes associated with fibrogenesis and apoptosis in steatohepatitis cases.¹⁰⁶

Genome-wide association and candidate gene studies have identified a variant in PNPLA3, which encodes adiponutrin, an insulin-responsive phospholipase involved in lipolysis and lipogenesis that correlates with accumulation of hepatic triglyceride and some histologic features of severity of NASH with identical ethnic distributions in prevalence studies.^107–112 This association is not limited to adults.^112,113 The effects cannot be shown to be direct, and it is speculated the allelic variant results in sensitization of the liver to metabolic and environmental stressors.²⁰

Pathogenesis

Certain similarities (i.e., steatosis, oxidative damage, and cytokine response) and dissimilarities between the pathogenesis of NAFLD and ALD are highlighted in Figure 49.17. The most commonly discussed driver of NAFLD is insulin resistance and the associated lipotoxicity (i.e., fat in nonfat depots) and complex pathways of systemic and local inflammation.¹¹⁴ The metabolic system of insulin involves signaling among skeletal muscle, adipose tissue, and liver.^115–118

Pathology

Grossly, noncirrhotic fatty livers are typically enlarged and yellow and have a greasy consistency. The cirrhotic liver may be small or enlarged. Fat may be observed in an irregular pattern in liver nodules as yellow areas compared with neighboring tan nodules. Cirrhotic livers in patients with NAFLD may not be as firm, may have the same stroma-to-parenchyma ratio, or may be as micronodular as in ALD.

Features of NAFLD in affected morbidly obese and superobese (BMI ≥ 40 kg/m²) individuals¹²⁹ may not be the same as in non–morbidly obese patients. Most objective scientific studies are based on the latter, and the following discussion is based on that population unless stated otherwise.

The histologic injury pattern of noncirrhotic fatty liver disease in adults, regardless of cause, initially involves zone 3 hepatocytes (Fig. 49.19). In cirrhosis, steatosis or steatohepatitis may or may not persist. Table 49.6 outlines the key gross and microscopic features of NAFLD compared with ALD.

Steatosis

By definition, steatosis is identified in 100% of NAFLD cases. The minimum criterion for NAFLD is a finding of at least 5% macrosteatosis. In adults, steatosis commonly begins in zone 3 hepatocytes, which contrasts with the zone 1 steatosis in viral hepatitis C (Fig. 49.20) or the panacinar pattern seen in pediatric NAFLD.

Steatohepatitis

The minimal criteria for steatohepatitis are steatosis (≥5%), hepatocellular ballooning, and lobular inflammation (Fig. 49.21). Similar to other forms of chronic liver disease, fibrosis is not required to establish a diagnosis of steatohepatitis. In the initial phases, steatosis is centered in zone 3; with progression, steatosis may involve the entire hepatic lobule. Azonal steatosis may be seen in conjunction with advanced fibrosis.¹³⁵ Nonzonal patches of microvesicular steatosis may occur in NAFLD (see Fig. 49.13),¹³⁵ and this correlates with disease severity,¹³⁶ as do the degree and zonality of steatosis.¹³⁷

Fibrosis and Cirrhosis

The characteristic pattern of fibrosis in adult patients with noncirrhotic steatohepatitis is zone 3 pericellular or perisinusoidal (i.e., chicken wire fibrosis), which results from deposition of collagen and extracellular matrix in the space of Disse (see Figs. 49.6 and 49.7). This type of fibrosis may be associated with steatohepatitis, and when found in isolation, it probably suggests healed prior episodes of steatohepatitis. Dense, extensive perisinusoidal fibrosis associated with portal hypertension, as occurs in patients with ALD, has not been described in patients with NAFLD. Diabetic hepatosclerosis, another change that correlates with severe extrahepatic organ damage in patients with type 1 diabetes, is not seen in NASH. It is characterized by patchy, dense perisinusoidal collagen and basement membrane deposition in the absence of steatosis (Fig. 49.24).¹⁵² With progression, fibrosis develops in the periportal region (see Fig. 49.9), followed in some cases by bridging fibrosis, which can be in a central-central, central-portal, or portal-portal pattern.

In NAFLD, MDBs are usually identified in ballooned hepatocytes, in which they often appear wispy, poorly formed, or delicate. Masson trichrome stain may be used to aid identification of MDBs (see Fig. 49.6). Immunohistochemical staining for K8 or K18 (see Fig. 49.22) ubiquitin may be positive in MDBs.³² Increased numbers of MDBs correlate with higher degrees of necroinflammatory activity in patients with NAFLD¹³⁰ and confirm the diagnosis of steatohepatitis.⁶⁰

Hepatocellular Glycogen.

Hepatocellular glycogen can accumulate in the cytoplasm in NAFLD. Glycogen imparts a dull, grayish refractile appearance to the cytoplasm; this may occur in conjunction with steatosis or megamitochondria. In contrast to diffuse involvement by glycogenic hepatopathy, a process that occurs in poorly controlled type 1 diabetes (Fig. 49.27),^170,171 the histologic change in NAFLD may affect only focal groups of hepatocytes.

Vascular Alterations.

A small intraacinar arteriolar branch in perivenular areas in zone 3 that was observed in NASH correlated with disease severity. When associated with dense perivenular inflammation, the lack of a visible bile duct helps to avoid misinterpretation of the lesion as an inflamed portal tract.¹⁷²

Treatment Effects.

Resolution of steatosis, ballooning, lobular inflammation, steatohepatitis, fibrosis, and a decrease in the NAFLD activity score (NAS) have been reported as potential positive effects of treatment (e.g., diet and exercise, medical, surgical) in various clinical trials (see Fig. 49.23).¹⁷³ Clinical trials have produced two additional observations—spontaneous resolution of the histologic features of NAFLD may occur, and even after clinically successful treatment, the degree of portal inflammation may increase.¹⁷⁴ Laboratory values¹⁴⁷ and histologic findings¹⁷⁵ may provide evidence of recurrence several weeks after cessation of successful medical therapy, indicating the underlying systemic nature of the process.

Prognostic Lesions.

Table 49.7 summarizes the results of clinicopathologic studies that have correlated the histologic features of NAFLD with fibrosis.^{155,176–180} Using immunohistochemistry for activated hepatic stellate cells, others have demonstrated a correlation with advanced fibrosis in NAFLD biopsies.^181–183 Practical application of this test awaits further validation. A meta-analysis of studies done with various methods identified lobular inflammation as a prognostic feature for fibrosis.¹⁷⁹ Portal chronic inflammation is associated with advanced disease histologically and clinically.^150,151 Other studies have highlighted microvesicular steatosis or the amount and location of steatosis.^136,137 One immunohistochemical study highlighted the loss of K8 and K18 and positivity for ubiquitin in hepatocytes, which are markers of ballooning, as correlative with fibrosis.²⁹

There is strong interest in the development of serum biomarkers and noninvasive imaging tests to diagnose NASH and establish fibrosis in NAFLD.¹⁸⁴ In patients with NAFLD, the metabolic syndrome was identified as a marker of histologic activity.¹⁸⁵ Common features to many studies are older age, diabetes mellitus, degree of insulin resistance, and elevated aminotransferase levels in morbidly obese and nonmorbidly obese populations.^{8,178,179,186,187} The ratio of PMNs to lymphocytes correlates with histologic activity and fibrosis in liver biopsies.¹⁸⁸

A proteomic approach has shown weight-dependent variations in diagnosed NAFLD and NASH.¹⁸⁹ A meta-analysis of single nucleotide polymorphism (SNP) studies has shown that the I148M variant of the PNPLA3 gene is associated with increased histologic severity and fibrosis in NAFLD.²⁰ This area of investigation will continue to grow because liver biopsy is not a practical screening approach.

Natural History and Treatment

Long-term studies of the natural history of NAFLD are challenging and few, but most authorities agree that steatosis alone is nonprogressive,¹⁹⁰ whereas fibrosis progresses in 31% to 42% of individuals during a period of 3.2 to 13.7 years to cirrhosis and its complications of liver failure and hepatocellular carcinoma.^191–193 In other patients, fibrosis remains stable or regresses.^194,195 Some patients have had regression of NASH to NAFLD (i.e., steatosis) without specific treatment¹⁹⁶ or after weight reduction.¹⁸⁰

The literature is mixed in terms of the effect of NAFLD on overall survival. Some studies have shown no difference compared with the general population for individuals with cirrhosis from other forms of chronic liver disease.^195,197 In contrast, studies from multiple countries have shown that patients with NAFLD have a worse overall survival rate.^191,194,198 Compared with patients with advanced chronic hepatitis C, patients with NAFLD with advanced fibrosis or cirrhosis have lower rates of liver-related complications and hepatocellular carcinoma¹⁹⁷ but similar overall mortality rates.¹⁹⁹ Obesity,¹⁰¹ increasing patient age,^101,178 and diabetes^{8,101,102,200} are risk factors for progression of steatosis to steatohepatitis or cirrhosis. Caffeine consumption is protective against the development of fibrosis.^201,202 Cardiovascular disease may be the leading cause of death of patients with advanced NAFLD.^195,203,204

Whether nonfibrotic, non-NASH NAFLD is necessarily a precursor that progresses or steatosis and NASH with fibrosis are separate entities is debated.¹¹⁹ Among patients with steatosis, only 2% to 20% have steatohepatitis or cirrhosis.⁹² Moreover, Cirrhosis (15% to 25%), liver failure, or hepatocellular carcinoma do not develop in all patients with steatohepatitis.^205,206 Many succumb to cardiovascular complications.²⁰³

Problems related to biopsy sampling errors and the various definitions of steatosis, steatohepatitis, and NAFLD have created difficulties in natural history studies. NASH-related cirrhosis or liver failure is the third most common indication for liver transplantation in the United States (9.7% in 2009) and is on a trajectory to becoming the number one indication.²⁰⁷ NASH cirrhosis accounts for 30% to 40% of liver-related deaths,²⁰⁸ whereas steatosis without steatohepatitis or cirrhosis is considered a more benign process.^192,194

The risk of hepatocellular carcinoma among patients with NASH is not always recognized in general practice,²⁰⁹ but it is a growing concern.^4,205,206 Although it is lower than that for HCV-related cirrhosis in comparative series,^197,210,211 screening only cirrhotics is recognized as inadequate, and several groups have reported hepatocellular carcinoma in noncirrhotic patients.^{4,206,212,213} Benign liver tumors such as adenoma and adenomatosis have been reported for patients with various degrees of NAFLD.^214,215

Trials of NAFLD treatment have not been a great success, with the possible exception of weight loss surgery.²¹⁶ Less drastic therapy and trials are aimed primarily at the presumed causes of disease and have included lifestyle intervention with weight loss and exercise, control of insulin sensitivity, and interventions broadly directed at inflammation, hepatocyte injury and fibrosis, and hyperlipidemia. Several reviews detail the trials and results.^59,217,218

Some common forms of drug intervention include the insulin-sensitizing PPAR-γ agonist pioglitazone with or without antioxidants; antidiabetic drugs such as metformin and thiazolidinediones; the antioxidants betaine, ursodeoxycholic acid, vitamins E and C; antifibrotic angiotensin-converting enzyme inhibitors and receptor blockers; and the TNF-α antagonist pentoxifylline.

The coexistence of NAFLD and ALD has been recognized as a significant risk factor for progression.²¹⁹ However, the overlapping contributory components cannot be histologically separated.

Alcohol use by individuals with NAFLD remains controversial. Modest alcohol intake protects against the effects of insulin resistance in morbidly obese individuals⁸ and is associated with a decreased prevalence of steatohepatitis in nonmorbidly obese patients with NAFLD.⁹ However, moderate alcohol consumption correlates with progression of fibrosis in NAFLD.²²⁰

Fatty Liver Disease in Patients with Other Liver Disorders

Clinical complications of acute viral hepatitis A, B, and C; chronic hepatitis C; and acetaminophen toxicity are seen in patients with alcoholism. These disorders require careful consideration by the pathologist when examining liver biopsy material with unusual findings.

The outcome and histologic features of ALD with concurrent acute viral hepatitis depend on the stage of ALD. Acetaminophen toxicity may not result in the characteristic features of zone 3 bridging necrosis in patients with ALD. The effects of chronic HCV infection are synergistic with those of fibrosis in patients with ALD.¹⁸ The histologic features of superimposed hepatitis caused by HCV are similar to those seen in nonalcoholic patients (see Chapter 46). Patients with ALD are also at risk for chronic pancreatitis and biliary obstruction, both of which may manifest histologically with cholangiolitis. The coexistence of ALD with hereditary hepatic iron overload and with α₁-antitrypsin deficiency results in findings typical of those disorders in addition to those of ALD.

The coexistence of NAFLD with other chronic liver diseases is a growing area of interest because of the high prevalence of NAFLD and its possible influence on the progression of other liver diseases; the high prevalence of chronic HCV infection; the high incidence of steatosis among patients with HCV infection; and the challenges of diagnosing NAFLD in the setting of other chronic liver disorders.^148,149,221 Several studies have shown a negative impact of steatosis and insulin resistance on the progression of disease in patients with HCV infection or hereditary hemochromatosis.²²² In one series, 2.6% of more than 3000 nonallograft liver biopsies identified steatohepatitis (i.e., steatosis and zone 3 perisinusoidal fibrosis) with a non-ALD or non-NAFLD type of chronic liver disease, such as hepatitis C, hemochromatosis, α₁-antitrypsin deficiency, or autoimmune hepatitis.¹⁴⁸

Chronic hepatitis C is the most common chronic liver disease diagnosed with concurrent NASH (5% to 9% of chronic hepatitis C cases).^148,223 Criteria for the diagnosis of NASH with chronic hepatitis C have not been established, but one recommendation relies on the unique characteristic of NASH zone 3 perisinusoidal fibrosis and hepatocyte ballooning with a zone 3 predilection in addition to steatosis and lobular inflammation.^129,148

Rarely, clinical and histologic features of autoimmune hepatitis or primary biliary cirrhosis may be seen in patients with NAFLD.¹⁴⁸ Diagnosis of these types of concurrent liver disorders requires more than elevated levels of serum autoantibodies (Fig. 49.28).⁶⁶

Fatty Liver Disease in Children

A variety of childhood genetic disorders are characterized by hyperinsulinemia, insulin resistance, and chronic liver injury. They include Bardet-Biedl, Alström, and Prader-Willi syndromes; lipodystrophy; and polycystic ovary syndrome.¹¹⁶ Histologic evaluation alone cannot define the cause of fatty liver disease in these cases. Other childhood disorders characterized by steatosis include HCV infection, rapid weight loss or starvation, and inborn errors of metabolism (e.g., urea cycle deficiency, fatty acid oxidation defects, organic acidemia, carnitine deficiency, cystic fibrosis, Wilson disease).

The growing epidemic of childhood obesity has led to increases in the prevalence of obesity-related conditions such as the metabolic syndrome.^224,225 NAFLD, which strongly correlates with components of the metabolic syndrome such as visceral adiposity and insulin resistance,^81,226 is considered the most common cause of chronic liver disease in children throughout the world.¹²⁴ The prevalence of NAFLD among children increases with age, and it is higher among adolescent children.²²⁷ Changes in sex hormones, insulin sensitivity, and adipose tissue distribution and function²²⁸ during development and increased control over unhealthy food choices and increased sedentary lifestyle with age are considered influential factors.²²⁹ Equally disturbing for long-term liver disease is the convergence of binge alcohol drinking and adolescent obesity, both of which are increasing.²³⁰

A retrospective histologic study of autopsy livers from 742 unexplained deaths in individuals 2 to 19 years of age documented fatty liver disease in 13% overall, which corresponded to 17.3% of children in the 15- to 19-year age range and 38% of obese children.²³¹ The prevalence of fatty liver disease in children varied according to ethnicity: Hispanic (11.8%), Asian (10.2%), white (8.6%), and African American (1.5%).²³¹ This difference could be explained by the genetic background because a common variant of the PNPLA3 gene that is associated with NAFLD in obese children is more common in Hispanics than African Americans.^113,232 NAFLD may result in cirrhosis in as many as 5% of affected children,²¹ and it is associated with shorter survival compared with age- and gender-matched control populations.²³³

The histologic features of pediatric NAFLD, especially in the younger children, are different from those of adults (Fig. 49.29).^234,235 This fact has been documented repeatedly in studies from the United States,²³⁴ Europe,²³⁶ and Japan.²³⁷ For instance, compared with adults, liver biopsies from children with NAFLD show a predominance of portal-based chronic inflammation and fibrosis; more severe, predominantly panacinar (or zone 1) steatosis; lack of ballooning and MDBs; and lack of zone 3 perisinusoidal fibrosis.²¹ This pattern of injury was identified in 51% of patients in a study by Schwimmer and colleagues,²³⁴ who coined the term type 2 (pediatric-type) NAFLD. The histologic pattern of disease characterized by zone 3 accentuation of steatosis was called type 1 (adult-type) NAFLD and was found in only 17% of cases.²³⁴ The remainder of cases showed overlapping histologic features (Table 49.8).

Differentiating Alcohol-Induced from Nonalcoholic Fatty Liver Disease

A clinical model was developed to help distinguish ALD from NAFLD based on a variety of parameters, such as mean corpuscular volume, AST/ALT ratio, BMI, and gender. An ALD/NAFLD index (ANI) greater than 0 favors ALD, whereas an ANI less than 0 favors NAFLD.²³⁹ The same group proposed the use of immunohistochemical markers. Decreased expression of the insulin receptor and increased expression of protein tyrosine phosphatase 1B (PTPN1), a protein that acts as a negative regulator of insulin resistance, was demonstrated in patients with NAFLD compared with those with ALD.²⁴⁰ Neither the clinical model nor the immunohistochemical marker combination have been validated.

Table 49.6 highlights some of the pathologic differences of ALD and NAFLD. Some data suggest that the degree of steatohepatitis is often more severe in ALD than in NAFLD. Numerous MDBs, increased lobular infiltration by PMNs with satellitosis, and acute cholestasis are features more suggestive of ALD than NAFLD.^2,241,242 Reliable clinical information is ultimately essential in distinguishing ALD from NAFLD. Figure 49.30 shows their shared and distinct histologic features.

Features That Occur More Frequently in Alcoholic Liver Disease

Sclerosing hyaline necrosis, obliterative and inflammatory lesions of the hepatic outflow veins, alcoholic foamy degeneration, and acute cholestasis have been described in noncirrhotic ALD but not in patients with noncirrhotic NAFLD.¹²⁹ In contrast to NAFLD, alcoholic hepatitis may occur in the absence of steatosis in ALD.²¹ Portal lipogranulomas are common in ALD.³⁰ In cirrhosis caused by ALD, regions of parenchymal extinction (septa) are often quite broad, in contrast to cirrhosis caused by NAFLD (see Fig. 49.10).

Features That Occur More Frequently In Nonalcoholic Fatty Liver Disease

In comparative histologic studies, features that occur significantly more often in NAFLD than in ALD include marked steatosis, periportal hepatocellular glycogenated nuclei,^36,243 and lipogranulomas.²⁴³ However, on an individual basis, the specificity of these histologic features is questionable. There may also be qualitative differences in the type of collagen deposition in NAFLD or ALD. For instance, type I collagen is more common in NAFLD-associated steatohepatitis, whereas type III collagen is more common in ALD.²⁴³

Grading and Staging of Fatty Liver Disease

A proposal for a novel histologic score with prognostic value for alcoholic hepatitis is based on the semiquantitative assessment of histologic parameters that are independently associated with short-term survival. They include lobular infiltration by PMNs, bilirubinostasis, megamitochondria, and stage of fibrosis. The score has been validated in two independent cohorts from Europe and the UnitedStates.²⁴⁴

Another study proposed a histologic grading system with predictive ability for alcoholic steatohepatitis based on semiquantification of hepatocyte ballooning and lobular inflammation, but it awaits validation.⁴⁷ Although no formal semiquantitative pathologic scoring system has been accepted, adaptation of a system used in NAFLD may be applied to ALD.^30,37

One popular semiquantitative grading and staging system for assessment of histologic lesions in NASH is based on evaluation of necroinflammatory activity and fibrosis, respectively¹³⁰ (Tables 49.10 and 49.11). This grading system is based on a semiquantitative evaluation of the degree of steatosis, hepatocyte ballooning, and lobular and portal inflammation. The staging system takes into account the specific pattern of fibrosis unique to NAFLD and concomitant vascular architectural alterations.

The NASH CRN, a multicenter consortium sponsored by the National Institute of Diabetes and Digestive and Kidney Diseases, proposed and validated a feature-based scoring system for use in clinical trials (Table 49.12).¹³¹ The necroinflammatory grade is referred to as the NAFLD activity score (NAS). The NAS combines the unweighted scores of three individual histologic features: steatosis (0-3), ballooning (0-2), and lobular inflammation (0-2). The total score (0-8) encompasses the entire spectrum of NAFLD, from steatosis to steatohepatitis. Staging subdivides early fibrosis as follows: 1a for delicate zone 3 perisinusoidal fibrosis, 1b for dense zone 3 perisinusoidal fibrosis, and 1c for portal-only fibrosis, specifically for pediatric NAFLD and some cases of bariatric NAFLD. A concurrent validation study confirmed that a diagnosis of steatohepatitis correlated with a NAS of greater than 5 in most cases.

NAS has been validated outside the CRN in a retrospective study of liver biopsies from adults with NAFLD.²⁴⁵ The use of the NAS for prediction of subsequent fibrosis has been questioned.²⁴⁶ The numbers generated by NAS were not meant to be a substitute for a histologic diagnosis made by a pathologist.¹³¹ Results from a NASH CRN study of 976 adult patients with NAFLD have shown that the diagnoses of definite NASH and semiquantitative NAS are distinct, with correlative but separate meanings, as are the classification systems for grading and staging chronic hepatitis.²⁴⁷ A recently developed scoring system (SAF score) allowing the use of an algorithm (FLIP algorithm)^247a for the classification of liver injury in morbid obesity has been validated for use in NAFLD of diverse etiology.^247b

Sampling error is an important issue in the histologic diagnosis and semiquantitative assessment of NAFLD.^248,249 For example, studies that evaluated biopsies from the right lobe²⁴⁸ or from different lobes⁹⁵ showed excellent concordance for the grade of steatosis, moderate concordance for assessment of the degree of fibrosis, and fair agreement for the degree of necroinflammatory features. In NASH, as in chronic viral hepatitis,²⁵⁰ the length of the tissue core affects the diagnostic yield, with definite NASH more commonly diagnosed and more extensive steatosis, lobular inflammation, and fibrosis observed in longer than shorter samples.²⁵¹ Needle diameter also is important.²⁴⁹ Features of NAFLD, especially the degree of necroinflammation and fibrosis, may not be distributed uniformly throughout the liver parenchyma. Pathologists should encourage clinical colleagues to obtain adequate cores of liver tissue and preferably multiple cores. Caution is warranted in interpreting surgically obtained biopsies because the inflammatory foci of PMNs (i.e., surgical hepatitis) render semiquantitative assessment of inflammation challenging.^96,146

Four histologic reproducibility studies that assessed interobserver and intraobserver variability in NAFLD in adults have shown good to excellent kappa scores for evaluation of the extent of steatosis, finding of perisinusoidal fibrosis, and degree of fibrosis.^{131,247b,252,253} In all four studies, the degree of lobular inflammation showed the lowest level of agreement, which is similar to the results of reproducibility studies of patients with chronic hepatitis.²⁵⁴ One study evaluated interobserver and intraobserver variability between a community general pathologist and an experienced hepatopathologist by using the NASH CRN scoring system. Although there was good intraobserver agreement, only fair to good agreement for steatosis, lobular inflammation, and a definite diagnosis of NASH and poor agreement for ballooning and fibrosis staging were found.²⁵⁵ As in chronic hepatitis C, the level of experience of pathologists may play a role in variability.^256,247b

Less than 2% of cases of steatohepatitis are estimated to be attributable to drug exposure.⁶ In some instances, presumed drug-related steatohepatitis may be a manifestation of an underlying preexisting disorder. Table 49.13 lists some of the manifestations of fatty liver in non-ALD, non-NAFLD settings. Wilson disease may result in some or all of the histopathologic manifestations of NAFLD or NASH in any age group.

Direct hepatotoxicity or mitochondrial injury resulting in steatohepatitis or phospholipidosis has been reported with the use of cardiac medications such as Coralgil, amiodarone, nifedipine, and diltiazem. Tamoxifen and related drugs for breast cancer, estrogens, and corticosteroids can aggravate obesity-related hyperinsulinemia or lipid abnormalities. Methotrexate-induced steatosis may be exacerbated by obesity. The drug’s role in causing steatohepatitis is no longer controversial because the pattern of fibrosis caused by methotrexate is typically periportal, not pericentral, and other common features of steatohepatitis, such as ballooning and MDBs, are not usually associated with this agent. Toxins other than alcohol that may result in steatosis or steatohepatitis include industrial solvents (e.g., paint thinners), petrochemicals, aflatoxins, arsenic compounds, amanitin, rapeseed cooking oil, and cocaine.²¹