T h e Im p a c t o f O b e s i t y o n L i v e r Histology Zachary D. Goodman,
MD, PhD
KEYWORDS Nonalcoholic fatty liver disease Steatosis Nonalcoholic steatohepatitis Cryptogenic cirrhosis Hepatocellular adenoma Hepatocellular carcinoma KEY POINTS Obesity and metabolic syndrome produce changes in the liver’s normal role in lipid and energy metabolism that cause a sequence of histopathologic changes. Steatosis is caused by an increase in hepatocellular fat vacuoles that parallels increased body mass index. Death of a small number of steatotic hepatocytes can produce liver enzyme elevation and focal nonspecific inflammation. Steatohepatitis occurs when there is cytoskeletal damage in genetically susceptible individuals resulting in loss of normal keratin filaments, ballooning degeneration of affected liver cells, and formation of Mallory-Denk bodies. In patients with steatohepatitis, activation of hepatic stellate cells produces intralobular fibrosis in the perisinusoidal spaces, whereas periportal ductular reaction causes activation of portal myofibroblasts and periportal fibrosis. With continuing fibrogenesis, there is progression to bridging fibrosis and cirrhosis. Hepatocellular carcinoma may develop in the cirrhotic liver, but both hepatocellular adenoma and hepatocellular carcinoma may occur in fatty liver disease before cirrhosis develops.
NORMAL LIVER AND NONSPECIFIC OR PHYSIOLOGIC STEATOSIS
The liver plays a central role in lipid metabolism, and consequently lipids (primarily triglycerides) may accumulate in the liver (primarily in hepatocytes) whenever there is an imbalance between the delivery of fat to the liver from the diet or from adipose tissue stores and the export of fat as a component of very-low-density lipoproteins. Small lipid droplets, identifiable only by electron microscopy or fat stains, are normally present in the cytoplasm of hepatocytes,1 but under conditions of metabolic imbalance, stress, or cellular injury in many pathologic processes, lipid droplets become large enough to visualize by light microscopy as clear vacuoles in hepatocytes cytoplasm.
Disclosure Statement: The author has nothing to disclose. Center for Liver Diseases, Inova Fairfax Hospital, 3300 Gallows Road, Falls Church, VA 22042, USA E-mail address: [email protected] Clin Liver Dis 18 (2014) 33–40 http://dx.doi.org/10.1016/j.cld.2013.09.010 1089-3261/14/$ – see front matter Ó 2014 Elsevier Inc. All rights reserved.
liver.theclinics.com
34
Goodman
Fat stains performed on frozen sections of livers from autopsies of hospitalized adults2 and children dying of trauma3 have shown that some degree of microvesicular steatosis is nearly always present, even though it is often not appreciated with routine paraffin-embedded sections. Similar population-based histologic studies of liver biopsies have not been performed in normal living subjects, because liver biopsies are seldom performed without a clinical indication, but proton nuclear magnetic resonance spectroscopy detects some hepatic triglyceride in all normal individuals, with a minimum of 1.9% of tissue by weight.4 OBESITY-RELATED STEATOSIS AND NONALCOHOLIC FATTY LIVER DISEASE
The presence of hepatocyte fat vacuoles that can be detected in routine hematoxylineosin–stained sections can be considered steatosis. Any alteration of the lipid transport and lipoprotein secretion may cause sufficient enlargement of normal lipid droplets to qualify, and minor degrees of unexplained steatosis are common. Traditionally, steatosis has been graded histologically by the proportion of affected parenchyma as mild (2/3) (Fig. 1).5 This grading has the disadvantage of including many cases with only a few affected hepatocytes in the same category as those that have 32% fat; so, in recent years, a lower limit of 5% fat, as estimated by a pathologist, has often been used as a defining feature of NAFLD.6 This practice is supported by the nuclear magnetic resonance studies showing that 95% of individuals with normal body mass index (BMI) and no risk factors for fatty liver disease had less than 5.5% hepatic triglyceride by weight.4 Even experienced pathologists, however, routinely overestimate the amount of fat when compared with quantitative measurements by digital image analysis and computer-assisted morphometry,7 with estimates typically doubling the amount of fat that is actually present, so liver biopsies estimated to have 5% fat may actually have much less.
Fig. 1. Degrees of steatosis. None (upper left)—hepatocytes have no visible cytoplasmic fat vacuoles. Mild (upper right)—fat vacuoles occupy greater than 5% but less than 33% of hepatocyte cytoplasm. Moderate (lower right)—fat vacuoles occupy greater than 33% of hepatocyte cytoplasm. Marked (lower left)—fat vacuoles occupy greater than 66% of hepatocyte cytoplasm.
The Impact of Obesity on Liver Histology
Although many genetic, environmental, and other factors seem to play a role in the pathogenesis of hepatocellular fat accumulation, and there is wide variation among individuals, the degree of histologic hepatic steatosis seems to correlate approximately with BMI. As previously noted, population-based data are impossible to obtain, and studies of obese patients are typically limited to those with an indication for liver biopsy, usually elevated liver-associated enzymes, or to morbidly obese patients undergoing bariatric surgery. There is evidence, however, of a correlation of steatosis with BMI in patients with chronic hepatitis C biopsied before therapeutic trials. In one study of more than 1400 patients, those with absolutely no steatosis on liver biopsy had a mean BMI of 26; those with microscopic steatosis up to 5% had a mean BMI of 28; those with 5% to 33% steatosis had average mean BMI of 30; and those with greater that 33% fat, as estimated by a pathologist, had a mean BMI of 31.8 Similarly, another study of 400 hepatitis C patients found a mean BMI of 27; fewer than 5% had mean BMI of 29, and those with 5% or greater had mean BMI of 32.9 In contrast, in patients with morbid obesity, the degree of steatosis does not correlate significantly with the BMI,10 perhaps because both weight and steatosis are skewed toward the high end of the scale in this population. Liver biopsies performed during bariatric surgery routinely detect some degree of steatosis in approximate 90% of morbidly obese patients. Metabolic syndrome, in particular insulin resistance, is much more important than obesity alone in the development of hepatic steatosis and steatohepatitis.11,12 Many other genetically determined metabolic diseases are characterized by hepatocellular fat accumulation,13 but these are generally rare compared with common metabolic syndrome–associated nonalcoholic fatty liver disease (NAFLD). Hyperinsulinemia promotes both triglyceride synthesis with hepatocellular fat accumulation and lipoprotein secretion leading to hyperlipidemia. The resulting steatosis is the first hit of the two-hit theory of the pathogenesis of steatohepatitis.14 Oxidative stress or other poorly understood factors may cause minor degrees of hepatocellular injury in the fatty liver with focal hepatocyte dropout and minor degrees of inflammation characterized by focal clusters of inflammatory cells, mainly lymphocytes and hypertrophied Kupffer cells. Depending on the degree of inflammation, these changes may be regarded as simple steatosis or steatosis with nonspecific inflammation. NONALCOHOLIC STEATOHEPATITIS
In 1979 and 1980 there were several reports of series of patients who did not consume alcohol but who had liver disease that histologically mimicked alcoholic hepatitis.15–17 Most of these patients were obese, diabetic, or both. There had been earlier descriptions of the same phenomenon, but these received little attention, and the prevailing view up to that time had been that if the liver biopsy showed alcoholic hepatitis, then the patient must have been secretly drinking. Ludwig and colleagues16 were especially instrumental in establishing this as a disease entity, and their name, nonalcoholic steatohepatitis (NASH), became the preferred term for what is now considered the severe form of NAFLD. Although obesity plays a role in NASH, metabolic syndrome and insulin resistance are considered more important in its pathogenesis in combination with oxidative stress and genetic predisposition.14 In addition to hepatocellular fat accumulation, livers with steatohepatitis contain variable numbers of liver cells with cytoskeletal damage that can be recognized by the presence of hepatocellular ballooning and Mallory-Denk bodies (Fig. 2). Epithelial cell cytoskeletons are composed of keratin proteins, which in hepatocytes are predominantly K8 and K18, that can be demonstrated by specific immunostaining.
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Fig. 2. NASH. Ballooned hepatocytes that contain Mallory-Denk bodies (arrows).
Oxidative stress–related cytoskeletal damage causes the normal keratin filaments to aggregate and form Mallory-Denk bodies associated with stress-related proteins, such as ubiquitin and p62. The cell that has lost its skeleton swells like an inflated balloon, losing the normal cytoplasmic staining for K8/18, which is now confined to the Mallory-Denk body.18 The injury in steatohepatitis is most severe in zone 3 of the hepatic acinus (ie, centrilobular areas, where ballooned cells and Mallory-Denk bodies are most readily found). Because the ballooned hepatocytes are enlarged, up to 100 mm in diameter, the Mallory-Denk body may be out of the plane of a 4-mm–thick microscopic section, which is the most likely reason why every ballooned cell does not seem to contain one of these inclusions. Immunostaining for ubiquitin or p62 can be helpful in identification of Malory-Denk bodies and can establish a diagnosis of NASH in equivocal cases (Fig. 3). Apoptotic bodies may also be present, and there is typically an inflammatory response with a predominance of lymphocytes and Kupffer cells and occasionally some neutrophils in more severe cases. Most patients who undergo liver biopsy have some degree of fibrosis. Centrilobular pericellular/perisinusoidal fibrosis produced by activated stellate cells in association with inflammation and
Fig. 3. NASH. Hepatocellular ballooning in NASH is characterized by swollen liver cells with thin strands of cytoplasm (left). The Masson trichrome stain shows deposition of delicate strands of blue-staining collagen in the perisinusoidal spaces surrounding ballooned liver cells (center), and an immunostain for ubiquitin demonstrates small Mallory-Denk bodies (MDB) (arrows) in some of the ballooned hepatocytes (right).
The Impact of Obesity on Liver Histology
Fig. 4. Bridging fibrosis in NASH (Masson trichrome stain). Collagenous fibrous tissue, stained blue, extends from a centrilobular area at lower right, to a portal area at upper left. Trapped hepatocytes are incorporated into the scar.
ballooning is the earliest stage (see Fig. 3) and is characteristic of the steatohepatitis pattern of injury. Portal fibrosis associated with ductular reaction follows the centrilobular fibrosis,19 and both processes seem to play a role in the development of bridging fibrosis (Fig. 4) and cirrhosis (Fig. 5), whereas chronic portal inflammation also increases and may play a role in advancing fibrosis.20 Population-based data on the relative incidence of these lesions are not available, and some degree of selection bias is inevitable in published series based on liver biopsies. In a large series of adults with NAFLD accumulated by the Nonalcoholic Steatohepatitis Clinical Research Network, of 693 patients, 26% had no fibrosis, 49% had mild or moderate fibrosis, 17% had bridging fibrosis, and 8% had cirrhosis.21 CRYPTOGENIC CIRRHOSIS
Some obese or diabetic patients are found to have otherwise unexplained cirrhosis on liver biopsy or at autopsy. In one autopsy series, 14% of obese patients had cirrhosis,22 and cirrhosis is 2 to 3 times more frequent in diabetics than in the general population. Although the cause is often uncertain, it is likely that most of these are the result of clinically silent NASH. Furthermore, more than 70% of patients with so-called cryptogenic cirrhosis are obese and/or diabetic, suggesting previous NASH that had
Fig. 5. Cirrhosis in NASH (Masson trichrome stain). Nodules of hepatic parenchyma are surrounded by bands of blue-staining fibrous scars with loss of normal architecture.
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Fig. 6. Liver biopsy showing hepatocellular carcinoma (right) in a liver with NASH (left).
become histologically inapparent after the development of cirrhosis.23,24 A recent investigation found that loss of fat correlated well with elevation of serum adiponectin levels in patients with cirrhosis, suggesting that this may be the mechanism leading to cryptogenic cirrhosis as a form of burned-out NASH.25 PRIMARY HEPATIC NEOPLASMS—HEPATOCELLULAR ADENOMA AND HEPATOCELLULAR CARCINOMA
Hepatocellular adenoma is an uncommon benign neoplasm that occurs most often in women with long-term exposure to contraceptive steroids. Three subtypes have been described, and all have been noted to be increasing in frequency in association with overweight and obesity.26,27 Some of these have evolved into hepatocellular carcinoma.28 Hepatocellular carcinoma (Fig. 6) has an increased incidence in patients with obesity and diabetes.29,30 A majority of these are in individuals with NASH-related cirrhosis or cryptogenic cirrhosis presumably due to NASH.31 A recent review, however, documented 116 reported cases of hepatocellular carcinoma in precirrhotic NAFLD, including some patients with no fibrosis at all.32 Such cases can be expected to become more common in future years with the growing epidemic of obesity. REFERENCES
1. Phillips MJ, Poucell S, Patterson J, et al. The Liver: an atlas and text of ultrastructural pathology. New York: Raven Press; 1987. 2. Fraser JL, Antonioli DA, Chopra S, et al. Prevalence and nonspecificity of microvesicular fatty change in the liver. Mod Pathol 1995;8:65–70. 3. Bonnell HJ, Beckwith JB. Fatty liver in sudden childhood death. Am J Dis Child 1986;140:30–3.
The Impact of Obesity on Liver Histology
4. Browning JD, Szczepaniak LS, Dobbins R, et al. Prevalence of hepatic steatosis in an urban population in the United States: impact of ethnicity. Hepatology 2004; 40:1387–95. 5. Burt AD, Mutton A, Day CP. Diagnosis and interpretation of steatosis and steatohepatitis. Semin Diagn Pathol 1998;15:246–58. 6. Kleiner DE, Brunt EM, Van Natta M, et al. Design and validation of a histological scoring system for nonalcoholic fatty liver disease. Hepatology 2005;41:1313–21. 7. Hall AR, Dhillon AP, Green AC, et al. Hepatic steatosis estimated microscopically versus digital image analysis. Liver Int 2013;33:926–35. 8. Poynard T, Ratziu V, McHutchison J, et al. Effect of treatment with peginterferon or interferon alfa-2b and ribavirin on steatosis in patients infected with hepatitis C. Hepatology 2003;38:75–85. 9. Conjeevaram HS, Kleiner DE, Everhart JE, et al. Race, insulin resistance and hepatic steatosis in chronic hepatitis C. Hepatology 2007;45:80–7. 10. Dixon JB, Bhathal PS, O’Brien PE. Nonalcoholic fatty liver disease: predictors of nonalcoholic steatohepatitis and liver fibrosis in the severely obese. Gastroenterology 2001;121:91–100. 11. Marceau P, Biron S, Hould FS, et al. Liver pathology and the metabolic syndrome X in severe obesity. J Clin Endocrinol Metab 1999;84:1513–7. 12. Bugianesi E, McCullough AJ, Marchesini G. Insulin resistance: a metabolic pathway to chronic liver disease. Hepatology 2005;42:987–1000. 13. Hooper AJ, Adams LA, Burnett JR. Genetic determinants of hepatic steatosis in man. J Lipid Res 2011;52:593–617. 14. Day CP. Pathogenesis of steatohepatitis. Best Pract Res Clin Gastroenterol 2002; 16:663–78. 15. Adler M, Schaffner F. Fatty liver hepatitis and cirrhosis in obese patients. Am J Med 1979;67:811–6. 16. Ludwig J, Viggiano TR, McGill DB, et al. Nonalcoholic steatohepatitis. Mayo Clinic experiences with a hitherto unnamed disease. Mayo Clin Proc 1980;55: 434–8. 17. Falchuk KR, Fiske SC, Haggitt RC, et al. Pericentral hepatic fibrosis and intracellular hyalin in diabetes mellitus. Gastroenterology 1980;78:535–41. 18. Lackner C, Gogg-Kamerer M, Zatloukal K, et al. Ballooned hepatocytes in steatohepatitis: the value of keratin immunohistochemistry for diagnosis. J Hepatol 2008;48:821–8. 19. Skoien R, Richardson MM, Jonsson JR, et al. Heterogeneity of fibrosis patterns in non-alcoholic fatty liver disease supports the presence of multiple fibrogenic pathways. Liver Int 2013;33:624–32. 20. Brunt EM, Kleiner DE, Wilson LA, et al. Portal chronic inflammation in nonalcoholic fatty liver disease (NAFLD): a histologic marker of advanced NAFLDClinicopathologic correlations from the nonalcoholic steatohepatitis clinical research network. Hepatology 2009;49:809–20. 21. Neuschwander-Tetri BA, Clark JM, Bass NM, et al. Clinical, laboratory and histological associations in adults with nonalcoholic fatty liver disease. Hepatology 2010;52:913–24. 22. Wanless IR, Lentz JS. Fatty liver hepatitis (steatohepatitis) and obesity: an autopsy study with analysis of risk factors. Hepatology 1990;12:1106–10. 23. Caldwell SH, Oelsner DH, Iezzoni JC, et al. Cryptogenic cirrhosis: clinical characterization and risk factors for underlying disease. Hepatology 1999;29:664–9. 24. Caldwell SH, Crespo DM. The spectrum expanded: cryptogenic cirrhosis and the natural history of nonalcoholic fatty liver disease. J Hepatol 2004;40:578–84.
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25. van der Poorten D, Samer CF, Ramezani-Moghadam M, et al. Hepatic fat loss in advanced nonalcoholic steatohepatitis: are alterations in serum adiponectin the cause? Hepatology 2013;57:2180–8. 26. Paradis V, Champault A, Ronot M, et al. Telangiectatic adenoma: an entity associated with increased body mass index and inflammation. Hepatology 2007;46: 140–6. 27. Biolulac-Sage P, Taouji S, Possenti L, et al. Hepatocellular adenoma subtypes: the impact of overweight and obesity. Liver Int 2012;32:1217–21. 28. Paradis V, Zalinski S, Chelbi E, et al. Hepatocellular carcinomas in patients with metabolic syndrome often develop without significant liver fibrosis: a pathological analysis. Hepatology 2009;49:851–9. 29. Larsson SC, Wolk A. Overweight, obesity and risk of liver cancer: a meta-analysis of cohort studies. Br J Cancer 2007;97:1005–8. 30. El-Serag HB, Hampel H, Javadi F. The association between diabetes and hepatocellular carcinoma: a systematic review of epidemiologic evidence. Clin Gastroenterol Hepatol 2006;4:369–80. 31. White DL, Kanwal F, El-Serag HB. Association between nonalcoholic fatty liver disease and risk for hepatocellular cancer, based on systematic review. Clin Gastroenterol Hepatol 2012;10:1342–59. 32. Baffy G, Brunt EM, Caldwell SH. Hepatocellular carcinoma in non-alcoholic fatty liver disease: an emerging menace. J Hepatol 2012;56:1384–91.
MD, PhD
KEYWORDS Nonalcoholic fatty liver disease Steatosis Nonalcoholic steatohepatitis Cryptogenic cirrhosis Hepatocellular adenoma Hepatocellular carcinoma KEY POINTS Obesity and metabolic syndrome produce changes in the liver’s normal role in lipid and energy metabolism that cause a sequence of histopathologic changes. Steatosis is caused by an increase in hepatocellular fat vacuoles that parallels increased body mass index. Death of a small number of steatotic hepatocytes can produce liver enzyme elevation and focal nonspecific inflammation. Steatohepatitis occurs when there is cytoskeletal damage in genetically susceptible individuals resulting in loss of normal keratin filaments, ballooning degeneration of affected liver cells, and formation of Mallory-Denk bodies. In patients with steatohepatitis, activation of hepatic stellate cells produces intralobular fibrosis in the perisinusoidal spaces, whereas periportal ductular reaction causes activation of portal myofibroblasts and periportal fibrosis. With continuing fibrogenesis, there is progression to bridging fibrosis and cirrhosis. Hepatocellular carcinoma may develop in the cirrhotic liver, but both hepatocellular adenoma and hepatocellular carcinoma may occur in fatty liver disease before cirrhosis develops.
NORMAL LIVER AND NONSPECIFIC OR PHYSIOLOGIC STEATOSIS
The liver plays a central role in lipid metabolism, and consequently lipids (primarily triglycerides) may accumulate in the liver (primarily in hepatocytes) whenever there is an imbalance between the delivery of fat to the liver from the diet or from adipose tissue stores and the export of fat as a component of very-low-density lipoproteins. Small lipid droplets, identifiable only by electron microscopy or fat stains, are normally present in the cytoplasm of hepatocytes,1 but under conditions of metabolic imbalance, stress, or cellular injury in many pathologic processes, lipid droplets become large enough to visualize by light microscopy as clear vacuoles in hepatocytes cytoplasm.
Disclosure Statement: The author has nothing to disclose. Center for Liver Diseases, Inova Fairfax Hospital, 3300 Gallows Road, Falls Church, VA 22042, USA E-mail address: [email protected] Clin Liver Dis 18 (2014) 33–40 http://dx.doi.org/10.1016/j.cld.2013.09.010 1089-3261/14/$ – see front matter Ó 2014 Elsevier Inc. All rights reserved.
liver.theclinics.com
34
Goodman
Fat stains performed on frozen sections of livers from autopsies of hospitalized adults2 and children dying of trauma3 have shown that some degree of microvesicular steatosis is nearly always present, even though it is often not appreciated with routine paraffin-embedded sections. Similar population-based histologic studies of liver biopsies have not been performed in normal living subjects, because liver biopsies are seldom performed without a clinical indication, but proton nuclear magnetic resonance spectroscopy detects some hepatic triglyceride in all normal individuals, with a minimum of 1.9% of tissue by weight.4 OBESITY-RELATED STEATOSIS AND NONALCOHOLIC FATTY LIVER DISEASE
The presence of hepatocyte fat vacuoles that can be detected in routine hematoxylineosin–stained sections can be considered steatosis. Any alteration of the lipid transport and lipoprotein secretion may cause sufficient enlargement of normal lipid droplets to qualify, and minor degrees of unexplained steatosis are common. Traditionally, steatosis has been graded histologically by the proportion of affected parenchyma as mild (2/3) (Fig. 1).5 This grading has the disadvantage of including many cases with only a few affected hepatocytes in the same category as those that have 32% fat; so, in recent years, a lower limit of 5% fat, as estimated by a pathologist, has often been used as a defining feature of NAFLD.6 This practice is supported by the nuclear magnetic resonance studies showing that 95% of individuals with normal body mass index (BMI) and no risk factors for fatty liver disease had less than 5.5% hepatic triglyceride by weight.4 Even experienced pathologists, however, routinely overestimate the amount of fat when compared with quantitative measurements by digital image analysis and computer-assisted morphometry,7 with estimates typically doubling the amount of fat that is actually present, so liver biopsies estimated to have 5% fat may actually have much less.
Fig. 1. Degrees of steatosis. None (upper left)—hepatocytes have no visible cytoplasmic fat vacuoles. Mild (upper right)—fat vacuoles occupy greater than 5% but less than 33% of hepatocyte cytoplasm. Moderate (lower right)—fat vacuoles occupy greater than 33% of hepatocyte cytoplasm. Marked (lower left)—fat vacuoles occupy greater than 66% of hepatocyte cytoplasm.
The Impact of Obesity on Liver Histology
Although many genetic, environmental, and other factors seem to play a role in the pathogenesis of hepatocellular fat accumulation, and there is wide variation among individuals, the degree of histologic hepatic steatosis seems to correlate approximately with BMI. As previously noted, population-based data are impossible to obtain, and studies of obese patients are typically limited to those with an indication for liver biopsy, usually elevated liver-associated enzymes, or to morbidly obese patients undergoing bariatric surgery. There is evidence, however, of a correlation of steatosis with BMI in patients with chronic hepatitis C biopsied before therapeutic trials. In one study of more than 1400 patients, those with absolutely no steatosis on liver biopsy had a mean BMI of 26; those with microscopic steatosis up to 5% had a mean BMI of 28; those with 5% to 33% steatosis had average mean BMI of 30; and those with greater that 33% fat, as estimated by a pathologist, had a mean BMI of 31.8 Similarly, another study of 400 hepatitis C patients found a mean BMI of 27; fewer than 5% had mean BMI of 29, and those with 5% or greater had mean BMI of 32.9 In contrast, in patients with morbid obesity, the degree of steatosis does not correlate significantly with the BMI,10 perhaps because both weight and steatosis are skewed toward the high end of the scale in this population. Liver biopsies performed during bariatric surgery routinely detect some degree of steatosis in approximate 90% of morbidly obese patients. Metabolic syndrome, in particular insulin resistance, is much more important than obesity alone in the development of hepatic steatosis and steatohepatitis.11,12 Many other genetically determined metabolic diseases are characterized by hepatocellular fat accumulation,13 but these are generally rare compared with common metabolic syndrome–associated nonalcoholic fatty liver disease (NAFLD). Hyperinsulinemia promotes both triglyceride synthesis with hepatocellular fat accumulation and lipoprotein secretion leading to hyperlipidemia. The resulting steatosis is the first hit of the two-hit theory of the pathogenesis of steatohepatitis.14 Oxidative stress or other poorly understood factors may cause minor degrees of hepatocellular injury in the fatty liver with focal hepatocyte dropout and minor degrees of inflammation characterized by focal clusters of inflammatory cells, mainly lymphocytes and hypertrophied Kupffer cells. Depending on the degree of inflammation, these changes may be regarded as simple steatosis or steatosis with nonspecific inflammation. NONALCOHOLIC STEATOHEPATITIS
In 1979 and 1980 there were several reports of series of patients who did not consume alcohol but who had liver disease that histologically mimicked alcoholic hepatitis.15–17 Most of these patients were obese, diabetic, or both. There had been earlier descriptions of the same phenomenon, but these received little attention, and the prevailing view up to that time had been that if the liver biopsy showed alcoholic hepatitis, then the patient must have been secretly drinking. Ludwig and colleagues16 were especially instrumental in establishing this as a disease entity, and their name, nonalcoholic steatohepatitis (NASH), became the preferred term for what is now considered the severe form of NAFLD. Although obesity plays a role in NASH, metabolic syndrome and insulin resistance are considered more important in its pathogenesis in combination with oxidative stress and genetic predisposition.14 In addition to hepatocellular fat accumulation, livers with steatohepatitis contain variable numbers of liver cells with cytoskeletal damage that can be recognized by the presence of hepatocellular ballooning and Mallory-Denk bodies (Fig. 2). Epithelial cell cytoskeletons are composed of keratin proteins, which in hepatocytes are predominantly K8 and K18, that can be demonstrated by specific immunostaining.
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Fig. 2. NASH. Ballooned hepatocytes that contain Mallory-Denk bodies (arrows).
Oxidative stress–related cytoskeletal damage causes the normal keratin filaments to aggregate and form Mallory-Denk bodies associated with stress-related proteins, such as ubiquitin and p62. The cell that has lost its skeleton swells like an inflated balloon, losing the normal cytoplasmic staining for K8/18, which is now confined to the Mallory-Denk body.18 The injury in steatohepatitis is most severe in zone 3 of the hepatic acinus (ie, centrilobular areas, where ballooned cells and Mallory-Denk bodies are most readily found). Because the ballooned hepatocytes are enlarged, up to 100 mm in diameter, the Mallory-Denk body may be out of the plane of a 4-mm–thick microscopic section, which is the most likely reason why every ballooned cell does not seem to contain one of these inclusions. Immunostaining for ubiquitin or p62 can be helpful in identification of Malory-Denk bodies and can establish a diagnosis of NASH in equivocal cases (Fig. 3). Apoptotic bodies may also be present, and there is typically an inflammatory response with a predominance of lymphocytes and Kupffer cells and occasionally some neutrophils in more severe cases. Most patients who undergo liver biopsy have some degree of fibrosis. Centrilobular pericellular/perisinusoidal fibrosis produced by activated stellate cells in association with inflammation and
Fig. 3. NASH. Hepatocellular ballooning in NASH is characterized by swollen liver cells with thin strands of cytoplasm (left). The Masson trichrome stain shows deposition of delicate strands of blue-staining collagen in the perisinusoidal spaces surrounding ballooned liver cells (center), and an immunostain for ubiquitin demonstrates small Mallory-Denk bodies (MDB) (arrows) in some of the ballooned hepatocytes (right).
The Impact of Obesity on Liver Histology
Fig. 4. Bridging fibrosis in NASH (Masson trichrome stain). Collagenous fibrous tissue, stained blue, extends from a centrilobular area at lower right, to a portal area at upper left. Trapped hepatocytes are incorporated into the scar.
ballooning is the earliest stage (see Fig. 3) and is characteristic of the steatohepatitis pattern of injury. Portal fibrosis associated with ductular reaction follows the centrilobular fibrosis,19 and both processes seem to play a role in the development of bridging fibrosis (Fig. 4) and cirrhosis (Fig. 5), whereas chronic portal inflammation also increases and may play a role in advancing fibrosis.20 Population-based data on the relative incidence of these lesions are not available, and some degree of selection bias is inevitable in published series based on liver biopsies. In a large series of adults with NAFLD accumulated by the Nonalcoholic Steatohepatitis Clinical Research Network, of 693 patients, 26% had no fibrosis, 49% had mild or moderate fibrosis, 17% had bridging fibrosis, and 8% had cirrhosis.21 CRYPTOGENIC CIRRHOSIS
Some obese or diabetic patients are found to have otherwise unexplained cirrhosis on liver biopsy or at autopsy. In one autopsy series, 14% of obese patients had cirrhosis,22 and cirrhosis is 2 to 3 times more frequent in diabetics than in the general population. Although the cause is often uncertain, it is likely that most of these are the result of clinically silent NASH. Furthermore, more than 70% of patients with so-called cryptogenic cirrhosis are obese and/or diabetic, suggesting previous NASH that had
Fig. 5. Cirrhosis in NASH (Masson trichrome stain). Nodules of hepatic parenchyma are surrounded by bands of blue-staining fibrous scars with loss of normal architecture.
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Fig. 6. Liver biopsy showing hepatocellular carcinoma (right) in a liver with NASH (left).
become histologically inapparent after the development of cirrhosis.23,24 A recent investigation found that loss of fat correlated well with elevation of serum adiponectin levels in patients with cirrhosis, suggesting that this may be the mechanism leading to cryptogenic cirrhosis as a form of burned-out NASH.25 PRIMARY HEPATIC NEOPLASMS—HEPATOCELLULAR ADENOMA AND HEPATOCELLULAR CARCINOMA
Hepatocellular adenoma is an uncommon benign neoplasm that occurs most often in women with long-term exposure to contraceptive steroids. Three subtypes have been described, and all have been noted to be increasing in frequency in association with overweight and obesity.26,27 Some of these have evolved into hepatocellular carcinoma.28 Hepatocellular carcinoma (Fig. 6) has an increased incidence in patients with obesity and diabetes.29,30 A majority of these are in individuals with NASH-related cirrhosis or cryptogenic cirrhosis presumably due to NASH.31 A recent review, however, documented 116 reported cases of hepatocellular carcinoma in precirrhotic NAFLD, including some patients with no fibrosis at all.32 Such cases can be expected to become more common in future years with the growing epidemic of obesity. REFERENCES
1. Phillips MJ, Poucell S, Patterson J, et al. The Liver: an atlas and text of ultrastructural pathology. New York: Raven Press; 1987. 2. Fraser JL, Antonioli DA, Chopra S, et al. Prevalence and nonspecificity of microvesicular fatty change in the liver. Mod Pathol 1995;8:65–70. 3. Bonnell HJ, Beckwith JB. Fatty liver in sudden childhood death. Am J Dis Child 1986;140:30–3.
The Impact of Obesity on Liver Histology
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