CHAPTER 7 Steatosis, Steatohepatitis and Related Conditions

Steatosis

Steatosis (fatty change, fatty liver) is the accumulation of abnormal amounts of lipid in hepatocytes. Most steatosis is of the macrovesicular type, in which a single large fat vacuole or several smaller ones occupy the greater part of the cell, pushing the nucleus to the periphery (Fig. 7.1). The less common and often more serious type is microvesicular steatosis (Fig. 7.2). The fat in this type is finely divided and the nucleus remains central. The two types of steatosis are sometimes found together, though one type usually predominates.

Figure 7.1 Macrovesicular steatosis.

There are large fat vacuoles in perivenular hepatocytes, displacing the nuclei to the edges of the cells. (Needle biopsy, H&E.)

Figure 7.2 Microvesicular steatosis.

Swollen hepatocytes near an efferent venule (right) contain numerous small vacuoles. The nuclei have maintained their central position. Some large fat vacuoles are also present. (Needle biopsy, H&E.)

Macrovesicular steatosis

Macrovesicular steatosis is common. It is frequently apparent by non-invasive imaging and may be accompanied by moderate abnormalities of serum aminotransferases, alkaline phosphatase and γ-glutamyl transpeptidase. Liver tests may be normal.¹ There are many causes of macrovesicular steatosis, of which the most common are listed in Table 7.1. It is usually not possible to determine the cause of uncomplicated large-droplet steatosis from histological examination alone.

Table 7.1 Common causes of macrovesicular steatosis

Obesity and diabetes mellitus

Protein-calorie malnutrition

Total parenteral nutrition

Drugs and toxins (e.g. alcohol, corticosteroids)

Metabolic disorders (e.g. Wilson’s disease)

Infections (e.g. hepatitis C)

The lipid in macrovesicular steatosis accumulates in hepatocytes because of increased triglyceride synthesis or decreased excretion.² Increased synthesis results from availability of excess free fatty acids and fatty acid precursors and from reduced fatty acid oxidation. Reduced excretion is a result of diminished apoprotein production, seen for example in protein malnutrition and alcohol abuse. Macrovesicular steatosis provides the background on which the important lesions of alcoholic and non-alcoholic steatohepatitis develop.³

Most steatosis is perivenular. Alcohol use, adult obesity, diabetes and corticosteroid therapy typically show this location. Increasing amounts of steatosis extend to progressively involve mid-zonal and periportal regions (acinar zones 2 and 1, respectively). In contrast, periportal steatosis is more common in children with non-alcoholic fatty liver disease (discussed later), in patients on parenteral nutrition, in kwashiorkor and protein-calorie malnutrition, and it is sometimes seen in AIDS (Fig. 7.3). In focal fatty change⁴ more or less rounded foci of steatosis are seen in an otherwise normal liver and may be mistaken for neoplasms on imaging.

Figure 7.3 Periportal steatosis.

Liver biopsy from a patient with AIDS and portal tract infiltration by large-cell lymphoma. Periportal hepatocytes contain large fat vacuoles. (Needle biopsy, H&E.)

The histological grade of steatosis should be reported based on the percentage of hepatocytes which contain lipid vacuoles. One commonly used scoring system includes grades of minimal (<5%), mild (5–30%), moderate (30–60%) and marked (>60%). Provision of a numerical assessment to the nearest percentile is also recommended (e.g. ‘marked macrovesicular steatosis is present involving approximately 90% of the parenchyma’). Periodic acid–Schiff and trichrome stains can be helpful in the assessment, providing contrast of the large lipid vacuoles against the more darkly stained background cytoplasm of hepatocytes. Digitised computer image analysis ⁵ is an alternative method of grading but from a practical standpoint is better suited to research settings.⁶

Occasionally lipid-laden hepatocytes rupture and the fat is then taken up by macrophages. The resulting lesion is a lipogranuloma (Fig. 7.4). Lipogranulomas are situated within the lobules, often near terminal venules. Serial sectioning may be needed to identify the fat in the centre of the lesion. Lipogranulomas may undergo fibrosis, but this does not appear to contribute to progressive liver disease and must be distinguished from the more important pericellular fibrosis characteristic of steatohepatitis (see below). Globules within portal tracts are usually the result of uptake of ingested or injected mineral oils by macrophages, rather than uptake of lipids⁷ (Fig. 7.5). Lipopeliosis – the formation of large intrasinusoidal fat cysts following release of lipid from hepatocytes after transplantation – is described in Chapter 16.

Figure 7.4 Lipogranuloma.

A lipogranuloma has formed near the terminal venule (V) in this moderately steatotic liver. Inset: The lipogranuloma contains large lipid vacuoles and aggregated Kupffer cells with scattered lymphocytes and a few eosinophils. (Needle biopsy, H&E.)

Figure 7.5 Mineral oil globules.

A row of vacuoles within macrophages is seen to the right of a portal venule. (Needle biopsy, H&E.)

The differential diagnosis of macrovesicular steatosis includes microvesicular steatosis. The presence of several fat vacuoles in one hepatocyte has to be distinguished from true microvesicular steatosis (below) in which vacuoles are generally less than 1 µm in diameter and may even be invisible in paraffin sections by light microscopy. The distinction is clinically important. The location of the nucleus helps to differentiate the two conditions. A second differential diagnosis is from stellate cell hyperplasia (Fig. 7.6) in which the vacuoles are not in hepatocytes but in perisinusoidally located stellate cells.⁸ Their nuclei are compressed into a crescentic shape by the vitamin A-rich globules. Stellate cell hyperplasia may be unexplained, but should lead to investigation of possible overuse of vitamin A or other retinoids.

Figure 7.6 Stellate cell hyperplasia.

Stellate cells with single or multiple lipid vacuoles lie in the space of Disse between hepatocytes (arrows). Stellate-cell nuclei are small, intensely basophilic and indented by the cytoplasmic vacuoles. Hepatocyte nuclei are larger, less dense and rounded. (Needle biopsy, H&E.)

Microvesicular steatosis

In this serious and sometimes fatal condition, finely divided fat accumulates in hepatocyte cytoplasm as a result of mitochondrial damage leading to impaired β-oxidation.⁹ Causes include acute fatty liver of pregnancy (Ch. 15), hepatotoxic drugs such as valproate and nucleoside analogues (Ch. 8), mitochondrial DNA depletion and deletion syndromes,¹⁰ foamy degeneration in the alcoholic (see below) and total parenteral nutrition (Table 7.2). Another cause, Reye’s syndrome, has declined sharply in incidence in recent years. In neonates and children, mitochondrial hepatopathies may need consideration.¹⁰ Viral infections occasionally give rise to similar changes.¹¹

Table 7.2 Main causes of microvesicular steatosis

Acute fatty liver of pregnancy

Alcoholic foamy degeneration

Drugs (e.g. nucleoside analogues, valproate)

Toxins (e.g. in Jamaican vomiting disease)

Total parenteral nutrition

Inborn errors of metabolism (e.g. urea cycle disorders)

Reye’s syndrome

Infections

Histologically, the cytoplasmic lipid is seen to be very finely divided and is not always obvious in paraffin sections. It can be stained with oil red O in frozen sections. The affected hepatocytes are often swollen. Their nuclei remain central (see Fig. 7.2).

The differential diagnosis is from macrovesicular steatosis and from conditions in which hepatocytes are swollen for other reasons, such as hepatitis. As discussed in Chapter 13, phospholipids and sphingolipids accumulate in various metabolic disorders. Cholesterol esters accumulate in hepatocytes in Wolman’s disease and cholesterol ester storage disease, and glycogen accumulates in glycogen storage disease.

Alcoholic and non-alcoholic fatty liver disease

The terms alcoholic fatty liver disease (AFLD) and non-alcoholic fatty liver disease (NAFLD) are used to describe the complete range of changes from uncomplicated macrovesicular steatosis to steatohepatitis and cirrhosis seen, respectively, in alcohol abuse and in obesity, diabetes, hyperlipidaemia and the metabolic syndrome. Insulin resistance, central (truncal) obesity, type 2 diabetes, hyperlipidaemia and systemic hypertension constitute the metabolic syndrome. NAFLD is considered the hepatic expression of the metabolic syndrome.¹² The recent upsurge in obesity and diabetes in industrialised countries and in other populations has brought NAFLD to increased attention in clinical and basic science. In the United States, NAFLD is currently the leading cause of abnormal serum aminotransferases and chronic liver disease.¹³^,¹⁴ A similar impact is likely in other Western countries and in other populations where the risk factors for NAFLD are prevalent. Emphasis on the histological evaluation of macrovesicular steatosis and related changes in liver biopsy, explant and postmortem specimens has consequently grown.

Systematic histological approach to fatty liver disease

Histological evaluation in AFLD and NAFLD should take into account not only the presence of large-droplet steatosis, but also evidence of hepatocellular damage, inflammation, fibrosis and siderosis which may also be present. The diagnosis of steatohepatitis should be rendered based on specific histological criteria (described in detail below). In AFLD and NAFLD there may be relatively inconspicuous apoptotic bodies.¹⁵^–¹⁷ Focal lobular inflammation (usually clusters of lymphocytes and activated Kupffer cells) may be seen but does not constitute steatohepatitis. Uncomplicated steatosis in the majority of cases is not associated with significant portal tract inflammation. However, focal minimal or mild portal lymphocytic infiltrates are sometimes present in either condition.¹⁸^,¹⁹ These must be distinguished from the portal inflammation seen in chronic hepatitis which is denser, regularly involves most or all portal tracts, includes plasma cells and eosinophils in addition to lymphocytes and may also be accompanied by interface hepatitis and lymphoid aggregates or follicles. Any histological doubt on this issue should be resolved by discussion with the clinician and investigations to exclude causes of chronic hepatitis when necessary. Some adult and paediatric patients with NAFLD show positive serum anti-nuclear and/or anti-smooth muscle antibodies,²⁰^–²² raising a clinical suspicion of autoimmune hepatitis (AIH). However, these autoantibodies are usually low-titer and are considered non-specific. The characteristic portal lymphoplasmacytic inflammation, interface hepatitis and regenerative rosettes of AIH are typically absent in such instances. Rarely, anti-mitochondrial antibody may be positive.²³^,²⁴

A connective tissue stain is important to evaluate the extent of any fibrosis and its distribution in the steatotic liver. When there is fibrosis in AFLD and NAFLD, it is usually present as a feature of steatohepatitis and is seen in centrilobular regions (acinar zone 3), as described below. Alternatively (and less frequently), there may be portal and periportal fibrosis. This distribution is more common in paediatric NAFLD ²⁵ and in morbidly obese individuals.²⁶

An iron stain should also be reviewed for siderosis. Mild iron overload in Kupffer cells and/or hepatocytes may be seen in alcoholic patients because of altered intestinal iron absorption and in NAFLD ²⁷^,²⁸ due to dysmetabolic hepatic iron overload (DHIO)²⁹ (Fig. 7.7). Significant hepatocellular siderosis should always prompt consideration of possible primary (genetic) iron overload.

Figure 7.7 Dysmetabolic hepatic iron overload.

This case of non-alcoholic fatty liver disease with marked steatosis is associated with mild siderosis of periportal hepatocytes (long arrows) and sinusoidal Kupffer cells (short arrow). (Needle biopsy, Prussian blue iron stain.)

Several systems have been used to assess steatosis,³⁰ inflammation and hepatocellular damage in NAFLD.³¹ A useful approach was provided by Matteoni and colleagues,³² who characterised steatotic changes in NAFLD according to four types: NAFLD type 1 (simple steatosis); NAFLD type 2 (steatosis with inflammation); NAFLD type 3 (steatosis with hepatocellular ballooning degeneration); and NAFLD type 4 (non-alcoholic steatohepatitis or NASH) (Fig. 7.8). Increased morbidity and mortality was associated with types 3 and 4. Swollen and ballooned hepatocytes in livers with macrovesicular steatosis are an indication for further careful examination to exclude frank steatohepatitis. The pattern of portal/periportal fibrosis with chronic inflammation (paediatric NAFLD, morbid obesity) might be considered to be NAFLD type 5 (Fig. 7.9).

Figure 7.8 Non-alcoholic fatty liver disease (NAFLD) types 1–4.

A: NAFLD type 1 (simple steatosis). B: NAFLD type 2 (steatosis with focal inflammation). C: NAFLD type 3 (steatosis with ballooned hepatocytes). Swollen hepatocytes with rarefied cytoplasm are seen at right. D: NAFLD type 4 (non-alcoholic steatohepatitis or NASH). The terminal venule (V) is surrounded by fibrosis and inflammation. Perivenular hepatocytes are swollen and show cytoplasmic Mallory bodies (arrows). (Needle biopsies, H&E.)

Figure 7.9 Steatosis with mild periportal fibrosis and chronic inflammation.

This biopsy from an obese child shows large-droplet steatosis with mild periportal fibrosis and chronic inflammation (at right). This pattern in children with non-alcoholic fatty liver disease is referred to as NASH type 2. Note the absence of liver-cell ballooning, Mallory bodies, inflammation or fibrosis near the terminal venule at lower left. (Needle biopsy, H&E.)

Diabetes mellitus

In patients with diabetes mellitus, glycogen vacuolation of hepatocyte nuclei is common ³³ (Fig. 7.10). These ‘glycogen nuclei’ are also seen in Wilson’s disease (Ch. 14), in non-alcoholic steatohepatitis and in biopsies from children and adolescents less than 14–15 years of age. Hepatomegaly in diabetics is not always attributable to steatosis: rarely, diabetics who are poorly controlled may develop Mauriac’s syndrome, with abnormal serum liver tests and massive accumulation of glycogen in hepatocytes (glycogenic hepatopathy³⁴) giving rise to a picture closely resembling inherited glycogen storage disease (see Fig. 13.12). Some diabetics with diabetic hepatosclerosis³⁵ (Fig. 7.11) show an increase in perisinusoidal type IV collagen³⁶ without a zonal predilection.