Case 5 -

Acute Hepatitis associated with SLE and Macrophage Activation Syndrome Jay H. Lefkowitch Columbia University New York, N.Y.

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Liver pathology shows acute hepatitis? THINK VIRUS & DRUGS (the most common causes) Don't forget LESS COMMON causes: unusual microbes; alternative/herbal/recreational agents; autoimmune hepatitis; "bystander hepatitis" induced by infiltrating immune cells activated in extra-hepatic disorders ERYTHROPHAGOCYTOSIS IS ABNORMAL: systemic viremias, unusual microbial agents and hemophagocytic syndromes should be considered IMMUNOHISTOCHEMISTRY adds useful information regarding the etiology and pathogenesis of acute hepatitis Case History: A 19 year old woman with known SLE was transferred from an outside hospital to our medical center for management of her marked transaminitis (AST = 1318 IU/L and ALT = 2375 IU/L), episodic thrombocytopenia, fever and persistently high ferritin. Her SLE had been diagnosed 4 years earlier, with positive smooth muscle and ribonucleoprotein antibodies and complications including lupus cerebritis and avascular necrosis of both hips. Six months before transfer she presented at another hospital with AST/ALT 9000 and respiratory, kidney and liver failure with uncontrolled bleeding. She was diagnosed with macrophage activation syndrome (MAS) (bone marrow biopsy failed to demonstrate erythrophagocytosis or other diagnostic features, but liver biopsy was reported positive) and responded with aggressive medical management, including immunosuppression (steroids, cyclosporine A, then rituximab—anti-CD20 antibody, and kineret—anakinra, an interleukin-1 receptor antagonist). At transfer, in addition to the marked aminotransferase elevations she had an INR of 1.29, total bilirubin of 2.9 mg/dl (direct of 1.7) and albumin of 3.7 g/dl. Her 3-week hospitalization was characterized by rising aminotransferases to >3000 IU/L, hypotensive shock with positive blood cultures for enterococcus, serum ammonia of 785, free air in the abdomen and gastrointestinal hemorrhage requiring laparotomy and segmental colectomy (intraluminal blood was found, but no perforation was identified). During her final week of life there was diffuse cerebral edema and impending transtentorial herniation, INR of 2.83 with elevated fibrinogen and D-dimers and continued marked transaminitis. She remained in liver failure, critically ill and her family authorized withdrawal of support. Case 5 - Figure 1 The initial liver biopsy performed at an outside hospital shows a diffuse hepatitis with mild portal tract inflammation and abundant lobular necroinflammation. Case 5 - Figure 2 Mild mononuclear inflammatory cell infiltrates, predominantly lymphocytes, are present in portal tracts. Note the intact native bile duct (BD). Case 5 - Figure 3 The lobular hepatitis is characterized by heterogeneous collections of intrasinusoidal lymphocytes, Kupffer cells and neutrophils. Note the prominent in-plate apoptosis of several hepatocytes to the left of center. Inset: Intrasinusoidal Kupffer cell erythrophagocytosis of small fragmented red blood cells (arrows) is present. Case 5 - Figure 4 The portal tract inflammatory infiltrates actively extend into periportal sinusoids producing hepatocyte apoptosis. Case 5 - Figure 5 CD68 immunostain shows numerous prominent portal and sinusoidal macrophages. Case 5 - Figure 6 CD68 immunostain shows enlarged intrasinusoidal Kupffer cells accompanied by lymphocytes and prominent neutrophils. Case 5 - Figure 7 Postmortem liver. Marked hemorrhagic necrosis involving nearly entire lobules is present, with preservation of small rims of periportal hepatocytes (PT = portal tract). This distribution is consistent with pre-terminal severe hypotensive and/or septic shock. Case 5 - Figure 8 A portal tract and partially preserved periportal liver-cell plates contrast with the extensive hemorrhagic necrosis in the inner portions of the lobule. Kupffer cells (some of which contain finely dispersed lipid vacuoles derived from cell breakdown) appear prominent within the sinusoids. Case 5 - Figure 9 Sinusoidal Kupffer cells contain conspicuous red blood cells and RBC fragments (erythrophagocytosis). Case 5 - Figure 10 Upper panel: CD68 immunostain of postmortem liver at low and medium power shows the continued prominence of intrasinusoidal Kupffer cells, notably in the periportal regions. (This localization is unusual, since hypotensive/septic shock with ischemic necrosis of any duration typically leads to activation of centrilobular rather than periportal Kupffer cells.) Lower panel: CK7 immunostain shows a brisk periportal activation of progenitor/stem cells with the appearance of many intermediate hepatobiliary cells (reflecting the degree of severity of centrilobular-to-midzonal necrosis present in this case. Case 5 - Figure 11 CD8 immunostain shows scattered intra-portal and periportal sinusoidal T cytotoxic lymphocytes. Inset: A CD8-positive cytotoxic T cell has infiltrated the bile duct, but there is no significant duct damage or bile duct loss elsewhere. Case 5 - Figure 12 C4d immunostain shows no obvious staining along vessel walls, but there is extensive positivity within intrasinusoidal Kupffer cells. In the context of the clinical history of SLE, the local immunologic response to antigen-antibody complexes formed may have entailed Kupffer cell phagocytosis of the complexes, with later complement fixation and local generation of C4d. Case 5 Discussion Introduction: Kupffer Cells and the Liver Kupffer cells constitute 15% of all of the cells within the liver and, impressively, these phagocytic and cytokine-secretory cells represent 80-90% of all reticuloendothelial macrophages [1]. Kupffer cells have 3 major functions: Clearance (antigen sequestration; immune complex clearance) Cytokine release (e.g., IL-1, IL-6, TNF-α, IFN-γ) Antigen-presentation (much less efficient than macrophages elsewhere) Kupffer cells reside within the vascular lumens of hepatic sinusoids where they rest on the endothelial cells, often anchoring their ruffled surface pseudopodia into the endothelial sieve plates. Kupffer cells originate from circulating monocytes from bone marrow, primitive macrophages in the yolk sac and from division of resident Kupffer cells within the liver. A major and critical function is surveillance for and destruction of deleterious macromolecules and infective organisms delivered to the liver in portal blood draining from the intestines. The abnormal vascular flow patterns in cirrhosis—including vascular shunting from portal vessels directly into central veins—by which bacteria gain direct access to systemic blood without Kupffer cell clearance, may result in bacteremia and sepsis in cirrhotic patients. Kupffer cells are distributed throughout the liver, with the majority (43%) localized in periportal regions [2]. They are also capable of migrating along sinusoidal walls at approximately 4-5 microns/minute [3]. Liver diseases in which Kupffer cells assume prominence include acute hepatitis and metabolic storage disorders [4]. In the former, the histologic features include disarray of liver-cell plates, hepatocyte ballooning and apoptosis, intrasinusoidal lymphocytic infiltrates. Ceroid-laden Kupffer cells are often visible within sinusoids following ingestion of effete hepatocytes; these cells stain positively with DPAS stain. In the centrilobular regions of steatohepatitis, activated Kupffer cells are also evident, particularly with immunostaining for CD68 [5]. In chronic hepatitis, periodic fluctuations in lobular necroinflammatory activity are often evident within sinusoids as clusters of lymphocytes and Kupffer cells containing phagocytic debris (often highlighted with DPAS stain). Lysosomes of Kupffer cells are natural sites for storage of a variety of abnormal products in inherited diseases such as Niemann-Pick disease [6]. Macrophage Activation Syndrome (MAS) Histiocytic disorders have been recently categorized by the WHO in 3 classes [7, 8]: Class 1 (dendritic cell-related disorders); Class 2 (macrophage-related disorders) and Class 3 (malignant disorders). MAS is a hemophagocytic syndrome [9] which falls in Class 2 (which also encompasses hemophagocytic lymphohistiocytosis or HLH [10], a very similar condition to MAS). Hemophagocytic syndromes may be further classified as primary (familial or sporadic, associated with family history and autosomal recessive transmission and genetic mutations such as perforin gene mutations) or secondary (triggered by viral infections, malignancy or rheumatologic disorders—most commonly juvenile rheumatoid arthritis but also rheumatoid arthritis, SLE, Sjogren's syndrome, dermatomyositis, Kawasaki disease, mixed connective tissue disease and systemic sclerosis). MAS therefore represents a secondary hemophagocytic syndrome. For patients with SLE, one hypothesis is that immune complexes deposit near/on bone marrow hematopoietic cells followed by complement activation, thereby precipitating excessive and dysregulated histiocytic activity, hemophagocytosis and engagement of CD8-positive T-cells [8]. Alternatively, an autoantibody in SLE may bind to histiocytes and trigger the MAS pathway. For both hypotheses, a network of cytokinhes (IL-1β, IL-2, IFN-γ, IL-6, M-CSF and TNF-α) is also engaged [8]. The mortality in MAS ranges from 8-22%. The diagnostic criteria for MAS [11] include persistent high fever, mucosal bleeding, neurologic abnormalities, pulmonary failure, lymphadenopathy and hepatosplenomegaly, cardiac and renal involvement, increased serum ferritin and lactate dehydrogenase, pancytopenia and DIC/elevated PT-PTT and INR. The syndrome complex of MAS must clinically be distinguished from an acute exacerbation of the underlying connective tissue disorder (when present as the trigger of MAS). The Liver in MAS Several studies have addressed involvement of the liver in MAS [12, 13, 14, 15]. In a condition characterized by dysregulated T-cell and Kupffer cell activity, not unsurprisingly the most diagnostic pathology includes portal and sinusoidal infiltrates of Kupffer cells (with erythrophagocytosis in the latter site) accompanied by CD8+ cytotoxic T-lymphocytes in both sites [12]. In the study of Billiau et al. [12], a key diagnostic criterion, erythrophagocytosis, was not identified in bone marrow specimens from 3 of their 5 reported cases, but was present in the liver. Portal infiltrates of lymphocytes were interpreted as "chronic persistent hepatitis" in one study from 1998 [14]. Focus on the lobular hepatitis (similar to the present case), including frequent hepatocellular apoptotic bodies, and the potential for bile duct damage and destruction and (in one case, resultant biliary cirrhosis) was presented by Bihl et al [13]. Immunostaining may be essential for establishing the diagnosis (including identification of erythrophagocytosis which may be inapparent on H&E stain) [12]. As pointed out by Billiau et al., the density of portal macrophage infiltrates can vary considerably, with the most diagnostic cases including numerous macrophages. The tendency of portal tract lymphocytes to extend into periportal regions as interface hepatitis was cited by Bihl et al. [13]. The liver in SLE The most common finding in the liver in patients with SLE is steatosis [16, 17]. Primary biliary cirrhosis occurs significantly more often than either chronic hepatitis or cirrhosis [18]. While autoimmune hepatitis and SLE are characterized by autoantibodies, the specificities differ between the two conditions. Other hepatic disorders identified in individuals with SLE include nodular regenerative hyperplasia, arteritis and granulomas [16, 18]. Liver Pathology of Case 5 The patient of this case, a young woman with systemic lupus erythematosus, had previously suffered several complications of this connective tissue disease and was first suspected of and diagnosed with MAS six months before her terminal admission to our center. The liver biopsy provided in this case had provided the diagnostic evidence of erythrophagocytosis which was lacking in a bone marrow biopsy. This biopsy demonstrated activated macrophages within portal tracts and Kupffer cells within sinusoids, well shown on CD68 immunostain. The degree of macrophage infiltration in the biopsy was relatively mild, perhaps readily overlooked had not the clinical suspicion of MAS been present. The lobular hepatitis was striking, however, particularly the degree of apoptosis. This is consistent with the heightened cytokine secretory activity of the infiltrating CD8+ T-cells. Such excessive destruction of hepatocytes is likely to have been a major contributing factor to the major elevations in serum aminotransferases in the patient's previous (and terminal) admission. One of the major clinical questions at the time of the patient's demise concerned the state of activity of the MAS. The postmortem liver specimen demonstrated multifactorial liver disease, with the massive hemorrhagic necrosis linked to the patient's extensive blood loss prior to death. However, evidence of aggregation of abundant aggregated Kupffer cells in periportal sinusoids (as demonstrated with CD68 staining) remained present even in the postmortem liver, indicative of ongoing MAS. The predilection for periportal sinusoidal Kupffer cells is consistent with the greater distribution of Kupffer cells in these sites [2]. Whether or not the activation of Kupffer cells at the time of death was due to immune complex clearance related to the underlying SLE is uncertain, but evidence supporting this concept was present in the positive immunostain results for C4d (presumably generated locally in proximity to these complexes). The bile duct damage which may occur in MAS [13] was not present in either the liver biopsy or postmortem specimens in this case. References Kolios G, Valatas V, Kouroumalis E. Role of Kupffer cells in the pathogenesis of liver disease. World Journal of Gastroenterology 2006; 14: 7413-7420. Bouwens L, Baekeland M, DeZanger R et al. Quantitation, tissue distribution and proliferation kinetics of Kupffer cells in normal rat liver. Hepatology 1986; 6: 718-722. MacPhee PJ, Schmidt EE, Groom AC. Evidence for Kupffer cell migration along liver sinusoids, from high-resolution in vivo microscopy. American Journal of Physiology 1992; 263: G17-G23. Ishak,KG and Sharp,HL: Ch. 4. 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