June 2001

Liver Diseases

Denny Meyer, DVM, DACVIM, DACVP
Gilead Sciences (Boulder, Colorado)

David C. Twedt, DVM, DACVIM
Colorado State University



PROFILING ACUTE AND CHRONIC HEPATIS: DIAGNOSIS AND MANAGEMENT
Denny Meyer, DVM, DACVIM, DACVP

Acute and chronic hepatitis have a wide differential diagnosis including idiopathic (most), drug-related, biliary tract disease, acute heart failure or shock, and "reactive" changes associated with extrahepatic disorders. Hepatic pathology that resolves within days to weeks is referred to as acute while chronic is generally defined as the persistence of abnormal liver tests for more than 3 to 4 weeks on two or more occasions of testing. The type of liver tests that are abnormal and the magnitude of change are generally different between the two classifications as well. These criteria define the terms from a clinical/clinical pathologic perspective. The definition of these terms based on microscopic findings is generally defined by the type of hepatocellular change and inflammatory cell infiltrate and alteration of the lobular architecture. The morphologic diagnosis of the histologic findings (acute or chronic) may or may not equate to the classification defined clinically. Furthermore, the morphologic diagnosis does not equate with an etiologic diagnosis but the microscopic alterations may suggest a specific cause or, more often, a differential diagnosis. We will discuss clinical pathology profiles that define acute and chronic hepatitis, illustrate the pros and cons of liver biopsy, and briefly outline their management.

Release of Pre-formed Enzymes.

Alanine aminotransferase (ALT); Aspartate aminotransferase (AST) . Species with high alanine aminotransferase (ALT) activity in the cytoplasm of the hepatocyte include the dog, cat, and primate whereas the liver of the equine, bovine, birds, and marmoset do not. One can think of each hepatocyte like a little balloon filled with ALT. Altered permeability of the hepatocellular membrane caused by injury or a metabolic disturbance result in a release of this soluble enzyme. Subsequent to an acute, diffuse injury, the magnitude of ALT increase in the plasma crudely reflects the number of affected hepatocytes. In chronic inflammatory liver disease, the magnitude of ALT rise does not relate to the degree of pathology. A variety of tissues, notably skeletal muscle and liver, contain high aspartate aminotransferase activity (AST). Because myositis and myocarditis are uncommon diseases in the dog and cat, a rise in the plasma AST activity generally indicates hepatic pathology. Skeletal muscle injury is assessed by the measurement of the serum creatine kinase (CK, CPK) activity, a specific skeletal muscle enzyme.

Is there added value in partnering AST with ALT in the assessment of liver pathology or is it redundant? Clinical experience suggests that interpretation of the both aminotransferases provides an additional insight into the evaluation of certain types of hepatic pathology. Following an acute injury resulting in a moderate to marked increase in the serum ALT and AST activities, the serum AST activity will return to normal more rapidly (hours to days) than the serum ALT activity (days) due to their difference in plasma half-lives and cellular location. By determining these values every 2 to 5 days for the dog following an acute insult, a sequential "biochemical picture" indicative of resolution is obtained. Persistent mild to moderately high serum ALT and AST values (documented multiple times over weeks to months) suggest a potentially serious, "smoldering" inflammatory process, chronic hepatitis. The persistent, often fluctuating abnormal aminotransferase values is probably a consequence of increased release subsequent to cell injury and on-going hepatocellular reparation (regeneration).

Enzymes Increased Secondary to Cholestasis or Effect of Drugs

Alkaline phosphatase (ALP) and Gamma glutamyltransferase (GGT) . Alkaline phosphatase and gamma glutamyltransferase (gamma glutamyltranspeptidase, GGTP) show minimal activity in normal hepatic tissue. These enzymes have a cell membrane location-ALP associated with the canalicular membrane and GGT associated with epithelial cells comprising the bile ductular system. There is minimal ALP increase in the plasma following an acute, severe insult in contrast to ALT and AST. Any initial rise is probably a reflection of enzyme activity on cell membrane fragments released to the plasma as a consequence of the damage and the initiation of increased synthesis due local cholestasis associated with damaged liver. During hepatic reparation following an injury, the serum aminotransferase activity slowly decreases while the serum ALP activity often increases until the intrahepatic architecture has recovered. Consequently, the serum ALP activity is usually the last serum hepatic enzyme test to return to the reference range following resolution of an acute insult.

Pathology that primarily obstructs the flow of bile, whether intrahepatic or extrahepatic, is initially associated with a rise in serum ALP activity followed by a raised serum total bile acid concentration and finally hyperbilirubinemia if the cholestatic process is protracted and sufficiently severe. None of these indicators of cholestasis alone are diagnostically reliable for differentiating extrahepatic and intrahepatic cholestatic disorders. An increase in the serum ALP activity is often associated with the use of glucocorticoid medications and hypercortisolemia. The high frequency of a raised ALP value in association with corticosteroids is unique to the dog and there is considerable individual variation in the hepatic response and magnitude of ALP rise. This corticosteroid-associated ALP isoenzyme can be distinguished from the cholestaticinduced liver ALP isoenzyme by several procedures and has been proposed for the differential diagnosis of hyperadrenocorticism. Unfortunately, a raised value has been shown to be associated with hepatobiliary disease, diabetes mellitus, hypothyroidism, acute pancreatitis, and phenobarbital treatment.

Liver Function Tests

Ammonia. The urea cycle is the major pathway for the conversion of intestinal-derived ammonia to urea nitrogen. Hepatic insufficiency can result in a reduced serum urea nitrogen (BUN) concentration relative to the serum creatinine concentration and a raised plasma ammonia concentration. Ammonia, along with other protein-derived metabolic products that escape hepatic metabolism, can alter the function of the central nervous system resulting in a syndrome referred to as hepatic encephalopathy. The determination of the plasma ammonia concentration can be used to support the presence of hepatic encephalopathy secondary to congenital portal vascular anomalies and cirrhosis.

Bile acids. The primary bile acids are synthesized from cholesterol in hepatocytes and secreted into the biliary system. Within the intestinal lumen, the primary bile acids, CA and CDCA, are dehydroxylated by bacteria to deoxycholic acid (DCA) and lithocholic acid (LCA), respectively. DCA and LCA are referred to as secondary bile acids. Only 5 to 10% of bile acids are lost in the feces. After their reabsorption into the portal circulation, they are carried to the liver and efficiently extracted from the sinusoidal blood by hepatocytes located in zone 1 and re-excreted into the biliary system completing an enterohepatic circulation.

The FBA and PPBA concentrations are a reflection of the efficiency and integrity of the enterohepatic circulation. Their measurement aids in the 1) detection of congenital portosystemic shunts, 2) identification of chronic hepatitis/cirrhosis prior to the development of jaundice, and 3) monitoring of the progression or resolution of hepatic disease with therapy (when ursodeoxycholic acid is not used). In a patient with raised liver enzyme tests, values of > 30 micromol/L for the dog and > 20 micromol/L for the cat have a high probability of microscopic finding in a liver biopsy. This does not necessarily imply that a morphological diagnosis of a specific disease entity will be achieved; only that there will be descriptive histopathologic findings.

Management of Hepatitis

The management of hepatic disease is based on four objectives: 1) treat the specific cause, 2) attend to the metabolic consequences of reduced hepatic function, 3) facilitate hepatocellular regeneration, and 4) impede the progression of hepatic injury and pathologic changes. We will discuss the general considerations for the management of hepatitis and define the role of ursodeoxycholic acid.

Suggested Reading

Hess P R, Bunch S E: Diagnostic approach to hepatobiliary disease. In Bonagura J D, ed: Kirk's Current Veterinary Therapy XIII. Philadelphia: W B Saunders, 2000, pp 659-664.

Meyer D J, Twedt D C: Effect of extrahepatic disease on the liver. In Bonagura J D, ed: Kirk's Current Veterinary Therapy XIII. Philadelphia: W B Saunders, 2000, pp 668-671.

Laflamme D P: Nutritional management of liver disease. In Bonagura J D, ed: Kirk's Current Veterinary Therapy XIII. Philadelphia: W B Saunders, 2000, pp 693-697.

Meyer D J, Harvey J W: Veterinary Laboratory Medicine: Interpretation and Diagnosis. Philadelphia, W B Saunders Co, 1998, pp 157-186.


Table 1.
Approximate Plasma Half-Life of Hepatic Enzymes in the Dog and Cat

EnzymeDogCat
ALT61 hours3.5 hours
AST12 hours1.5 hours
GLDH18 hours --
ALP
  -hepatobiliary isoenzyme66 hours6 hours
  -corticosteroid isoenzyme74 hours --
  -intestinal isoenzyme6 minutes2 minutes


EFFECTIVE USE OF HEPATIC CYTOLOGY AND LIVER BIOPSY
Denny Meyer, D.V.M., DACVIM, DACVP

"The establishment of an accurate diagnosis is dependent on sampling an adequate amount of tissue and, most important, on a histologic interpretation by someone well versed in liver histopathology."1

Abnormal liver tests are often identified in both sick and clinically healthy patients. The questions that one often asks are: "what is the nature of the hepatic pathology?" "Does the patient have primary liver disease?" "Should additional tests be determined?" "Will a sonogram be helpful?" "Will cytology be of value?" and/or "Should a liver biopsy be taken and is it generally a better investment than cytology?" Primary progressive hepatitis that marches on hepatic failure is an uncommon cause of abnormal liver tests for the majority of animals seen in private practice. Age-related changes and "reactive hepatopathy" secondary to an extrahepatic disorder are much more common. The advent of ultrasonographic examination of the liver has added another dimension of potential "abnormal" findings that contribute to the complexities of working up patients with abnormal liver tests and has facilitated the acquisition of cytologic specimens and needle biopsies.

Although cytology is practical and economical, it has defined (limited) diagnostic utility and its cavalier use can result in mis- (and missed) diagnosis. This is because the cytologic impression of a hepatic aspirate specimen is often projected to equate with a tissue-based diagnosis, an erroneous assumption since tissue architecture can not be assessed and changes cannot be quantified. It has value for the initial evaluation of hepatomegaly when caused by lipidosis, malignant lymphoma, myeloproliferative neoplasia, mast cell neoplasia, hepatocellular and cholangiocellular carcinomas, glucocorticoid hepatopathy, and amyloidosis. For the other types of liver disease, histopathologic examination is required to reach a final diagnosis, provide a prognosis, and monitor therapy.

The establishment of an accurate diagnosis is dependent on two important components -- sampling an adequate amount of tissue and interpretation of the histology by someone well-versed in liver pathology. The liver biopsy provides only a small "window" for viewing histopathologic changes. Biopsy specimens of the liver are usually obtained with needles provide a core of tissue that represents approximately 1/50,000 of the whole organ. The type of needle used impacts on the volume (amount) of tissue available for sectioning. Early or subtle changes in liver architecture associated with cirrhosis or nodular regeneration are likely to be missed with a needle biopsy specimen. The length and management of the specimen are important factors to ensure an adequate number of lobules for evaluation. In humans, 1.5 to 2.0 cm length of liver obtained by needle biopsy is generally considered reliable for the morphologic diagnosis of chronic progressive hepatitis and ensure that the prognostic indicators of bridging necrosis and/or bridging fibrosis may be observed.

The wedge biopsy provides ample tissue and contains larger portal tracts than those deeper in the hepatic parenchyma. Because of its superficial location relative to blood supply, the margin of the liver is predisposed to fibrosis that may mimic changes observed with cirrhosis. A biopsy taken hours after the beginning of a surgical procedure can contain neutrophils that accumulate under the capsule and focally in liver cell plates. Along with isolated liver cell necrosis, the artifactual process would result in a morphologic diagnosis of mild focal suppurative hepatitis. The changes are thought to be secondary to anoxia.

The liver may exhibit a variety of nonspecific secondary changes in response to disease elsewhere in the body. (Table 1) Liver test abnormalities are usually mild and the serum bile acid concentration is usually within the reference range. The histologic features of reactive hepatitis (reactive hepatopathy, nonspecific chronic hepatitis) encompass a variable combination of portal and parenchymal changes that are usually of minor degree and distributed in a patchy, uneven manner. The findings include portal infiltration by mononuclear cells (primarily lymphocytes), fatty change, focal hepatocellular necrosis, and lobular inflammation. The latter may consist of enlarged or hyperplastic Kupffer cells, small foci of monocyte-derived macrophages, or lymphocyte aggregates. The distinction from resolving acute hepatitis or a mild form chronic hepatitis may be difficult and arbitrary without clinical information.

Nodular hyperplasia is a relatively benign process may cause an increase hepatic tests and histopathological changes that may be suggestive of chronic hepatitis or an extrahepatic disease such as hyperadrenocorticism. The dog shows a propensity for the development of nodular hyperplasia of the liver. The cause of the proliferative change is not known. Nodules can be present as early as 6 years of age and are present in varying degrees in most dogs older than 14 years. The expansile process compresses existing parenchyma resulting in hepatocellular atrophy and approximation of the reticular fibers. Grossly, their appearance mimics macronodular cirrhosis and neoplasia. Microscopically, hepatocytes can develop a variety of cytoplasmic changes including lipidosis, hydropic degeneration, and glycogen accumulation. These microscopic findings are often problematic in needle biopsy specimens since the identification of a nodule is very difficult due to size limitations. The morphologic diagnosis of vacuolar hepatopathy results in a differential diagnosis of a metabolic disorder and unnecessary endocrine testing. In more advanced cases, the altered architecture contains varying degrees of mixed inflammatory cells that can result in a morphologic diagnosis of chronic active hepatitis, suppurative hepatitis, or granulomatous hepatitis. Nodular hyperplasia is associated with lipocyte prominence and the formation of lipogranulomata. Prussian blue staining demonstrates an abundance of iron accumulation in these foci of pigmented macrophages. An increase in serum hepatic enzyme activities, notably alkaline phosphatase, are often associated with nodular hyperplasia. The etiopathogenesis may be a reflection of two physioanatomical processes. First, the distorted hepatic architecture impairs bile flow and precludes adequate blood supply to hepatocytes and resulting in cholestatic induction of enzyme synthesis (alkaline phosphatase) and compromised membrane integrity (leakage of aminotransferases). Alternatively, the increase in at least the aminotransferases may be directly related to the hepatocyte proliferative process since an increase in the serum aminotransferase activities have been shown to be related to an increased production by proliferating hepatocytes during the reparative process.

Tables 2 and 3 list general criteria for determining the severity of the hepatic pathology and help different primary progressive hepatitis from reactive hepatopathy and age-related changes.

Suggested Reading/References

1. Schiff E R, Schiff L. Needle biopsy of the liver. In: Schiff L & Schiff E R, eds. Diseases of the liver. 7th ed. Philadelphia: J. B. Lippincott Co, 1993;216.

2. Ishak K. Chronic hepatitis: Morphology and nomenclature. Mod Pathol 1994;7:690-713.

Suggested Reading

Meyer D J, Twedt D C: Effect of extrahepatic disease on the liver. In Bonagura J D, ed: Kirk's Current Veterinary Therapy XIII. Philadelphia: W. B. Saunders, 2000, pp 668-671.

Meyer D J: Hepatic pathology. In Guilford W G, Center S A, Strombeck D R, et al: Strombeck's Small Animal Gastroenterology. Philadelphia, W. B. Saunders, 1996, pp 633-653.

Prause L C, Twedt D C: Hepatic nodular hyperplasia. In Bonagura J D, ed: Kirk's Current Veterinary Therapy XIII. Philadelphia: W. B. Saunders, 2000, pp 675-676.


Table 1. Extrahepatic Causes of Morphologic or Histopathologic Changes in the Liver

Inflammatory intestinal diseaseProlonged protein-restricted diet
Extrahepatic infections (bacterial)Septicemia
Rickettsial infectionsShock
Acute PancreatitisRight-sided heart failure
Diabetes mellitusHyperadrenocorticism
Congenital portosystemic shunt


Table 2. Degree of Activity in Chronic Hepatitis2

Lesions and degree of injury
CategoryPortal area inflammationPiecemeal necrosisSpotty necrosisBridging &/or multi-acinar necrosis
Mildmild, patchyabsent/mildmildabsent
Moderatemoderatemoderatemoderateabsent
Markedmarkedmarkedmarkedabsent
Very markedmarkedmarkedmarkedpresent


Table 3. Degree of Fibrosis in Chronic Hepatitis2

Component lesions
CategoryFibrous expansion of portal areasBridging fibrosis*Bridging with nodules (cirrhosis)
Mildabsent or mildabsentabsent
Moderatemoderateabsent**absent
Markedmarkedmarkedabsent***
Very Markedmarkedmarkedpresent


* Refer to Nomenclature for the definition
** Occasional bridging may be present
*** Occasional nodule may be present (incomplete cirrhosis)


CHALLENGING LIVER CASES WITH AN EMPHASIS ON THE BIOPSY FINDINGS
Denny Meyer, D.V.M., DACVIM, DACVP

The decision to pursue additional testing, imaging and/or biopsy of the liver after finding abnormal liver tests often are difficult to justify and the cost-benefit value unclear. Once a biopsy is obtained, the morphologic diagnosis, based on the microscopic findings, may be difficult to relate to a clinical diagnosis and/or the management of the patient. We will discuss cases in which a second pathologist, following a review of the biopsy, reads additional information into microscopic alterations. The key features of the biopsy will be illustrated with photomicrographs and the histologic information related to the clinical pathology and the patient's management.

Doberman pinscher, 4 years of age, female. History: acute development of an abdominal distention. The dog was quiet on physical examination but appeared clinical healthy except for a distended abdomen due to an abdominal effusion. Notable diagnostic findings: ALT 395 IU/L (<118); AST 93 IU/L (<66); ALP 291 IU/L (< 131); TB 0.7 mg/dL (<0.4); albumin 2.5 g/dL (2.3-3.9); TP (ascites) 2.8 g/dL with a few mesothelial cells. A normal to slightly enlarged liver was observed with ultrasound and a needle biopsy was obtained. Pathologist #1: moderate portal inflammation (lymphocytes, neutrophils and macrophages that contained a few pigmented granules suggestive of copper); marked vascular dilatation, chronic passive congestion in zone 1; scattered aggregates of inflammatory cells in lobules with occasional single cell necrosis; morphologic diagnosis: lobular dissecting hepatitis, moderate. Comment: consistent with copper-associated Doberman hepatitis along with significant hemodynamic alterations. Initial management: removed the abdominal fluid, zinc gluconate 100 mg bid; Actigall 300 mg/day; vitamin E 400 IU/day; U/D diet. Two weeks later the notable findings included: ALT 141 IU/L (<118); AST 135 IU/L (<66); ALP 374 IU/L (<131); TB 0.2 mg/dL (<0.4); albumin 2.3 g/dL (2.3-3.9); markedly distended abdomen due to an effusion. Pathologist #2 consulted and questions asked: Is the ascites secondary to Doberman hepatitis? Was there a sampling error ... more severe pathology missed? Should corticosteroids be used? Comments in response: most likely thrombosis of the hepatic vein(s) - Budd-Chiari-like syndrome with secondary portal hypertension with a component of reactive hepatitis. Recommendations: initiate furosemide plus spironolactone followed by only spironolactone -- anticipate resolution within another 2 to 3 weeks; liver diet; discontinue zinc gluconate; do not use corticosteroids; re-biopsy in 6 months. One month follow-up: clinical healthy with possibly slight ascites present. Question by owners: the dog is valuable and they want to know if is okay to have her bred? Six months later: remains clinically healthy with the following liver test values: ALT 70 IU/L (<118); AST 27 IU/L (<66); ALP 33 IU/L (< 131); TB 0.1 mg/dL (<0.4); albumin 3.4 g/dL (2.3-3.9). A second ultrasound-guided needle biopsy was obtained. Pathologist #1: mild portal fibrosis with mild portal infiltrates of macrophages, lymphocytes, plasma cells, & eosinophils with slight spillage of inflammation into periportal parenchyma although the majority of hepatic parenchyma is entirely normal; morphologic diagnosis: mild chronic active hepatitis. Pathologist #2: areas of parenchymal collapse with slight reactive inflammation, possibly a sequela of the previous pathology. The change may portend the development of nodular regenerative hyperplasia. There are occasional small foci of lymphocytes (2 to 5 cells) in lobules; possible harbinger of progressive Doberman hepatitis; continue to monitor the ALT and biopsy in 6 to 12 months.

Malamute/mix, 9 years of age, male. History: initially examined for nonlocalizing generalized pain; laboratory abnormalities consisted of: ALT 73 IU/L (<60) and albumin 2.4 g/dL (2.6-4.3). Radiographs of lumbar spine were negative. Carprofen was prescribed for 2 weeks and the pain resolved. Four months later the clinically healthy dog presented for a pre-dental evaluation. Notable diagnostic findings on the preanesthetic evaluation were: ALT 159 IU/L; albumin 2.3 g/dL; BUN 5 mg/dL (7-27). At this time, the question was "should we ignore or explore the abnormal laboratory findings?" Additional evaluation revealed: FSBA 6 micromol/L (<5); PPBA 9 micromol/L (<15); ultrasonography: small liver, no focal abnormalities and a distended gallbladder with a large amount of sludge that has an organized appearance suggestive of inspissation. Needle biopsy of the liver was obtained. Pathologist #1: mixed periportal inflammation that extends beyond the limiting plate with accompanying piecemeal necrosis. Prominent course yellow-brown granules in the hepatocytes are compatible with copper; morphologic diagnosis: chronic active hepatitis with copper accumulation secondary to cholestasis. Pathologist #2: copper-stained section demonstrates copper within hepatocytes of all 3 lobular zones. The mixed inflammation (neutrophils, lymphocytes, macrophages) appears to be intimately associated with the copper-laden cells. There are no morphologic findings suggestive of cholestasis. Questions by primary care veterinarian: Is the copper the cause of the pathology? Should it be treated? Would ursodeoxycholic acid &/or vitamin E, &/or milk thistle &/or SAMe be useful? What about the use of corticosteroids? Why are the albumin & BUN low? Management: copper chelator for at least 6 months followed by life-long zinc gluconate, liver diet (optional), vitamin E. Recommend a biopsy following 6 months of copper chelation to evaluate the effect.

Bichon Frise, 11 years of age, male, castrate. History: for approximately 1 year there has been slowly progressive polydipsia, polyuria, and urinary incontinence. During the past 6 months the urine specific gravity has ranged between 1.005 -1.012 with partial response to phenylpropanolamine & vasopressin. There has been mild weight loss and occasional lethargy. The dog appears thin with reduced muscle mass on physical examination. Notable diagnostic findings: hematology - target cells; clinical chemistry: ALT 143 IU/L (<50); AST 65 IU/L (<37) albumin 2.2 g/dL (2.3-3.9); BUN 6 mg/dL (7-30); creatinine 0.9 mg/dL (0.5-1.4); USG 1.014; FSBA 372 micromol/L (<5), PPBA 507 micromol/L (<15). A very small heteroechoic liver was observed with ultrasound and no shunts were visualized. At laparoscopy the small liver had a prominent yellow reticular pattern. A wedge biopsy was taken for histology and culture; there was grow (aerobic & anaerobic). Pathologist #1: prominent lipogranulomas, some with an infiltrate of lymphocytes and plasma cells. Comment: the findings are nonspecific but may have something to do with the clinical signs, however, how exactly is unknown. Pathologist #2: notable findings include lipogranuloma, prominent stellate cells (Ito cell, lipocytes), micronodule, iron-positive hepatocytes, and abnormal arteriolar structures. These findings are consistent with nodular regenerative hyperplasia possibly secondary to a portal vascular anomaly. Management: lactulose, liver diet (changed from W/D diet plus table scraps), vitamin E, and continued the phenylpropanolamine. One-month follow-up: approximately 0.4 kg (1 pound) weight gain, more alert and increased activity. The phenylpropanolamine was discontinued one week earlier with no recurrence of the urinary incontinence. Two-month follow-up: clinically healthy.

Whippet, female, spayed, 7 years of age. History: increasing episodes of intermittent lethargy, acts "lost" (old age?); periodic vomiting, +/- ( water intake ~2 months; abdominal enlargement ~2 to 3 weeks. Physical examination: thin - ( muscle mass, ascites. Notable laboratory findings: hematocrit 29% (37-54), MCV 56 fL (62-74),

ALT 154 IU/L (<58), ALP 278 IU/L (<73), albumin 2.1 g/dL (2.5-3.6), BUN 6 mg/dL (8-25), uSG 1.010, ascites: TP <2.5, few cells, FSBA 116 mol/L (<5; <25), PPBA 383 mol/L (<15; <25). Radiographs: poor contrast-effusion, +/- small liver; sonography: effusion/microhepatica with uniform surface & echogenicity + 1 or 2 extrahepatic shunts.

Needle biopsy attempted, small fragments û not submitted. Exploratory celiotomy: fluid; small liver with smooth slightly irregular surface & subtle nodular pattern; multiple extrahepatic shunt vessels; wedge biopsy obtained. Primary pathologist: hepatocellular vacuolation, moderate; periportal fibrosis moderate; multifocal pigmented/lipid-filled macrophages (lipogranulomatous hepatitis. Comment: appears to be an early or quiescent stage of cirrhosis with possible secondary metabolic disease. Macrophage foci may indicate a prior injury. No evidence of progressive disease. Second opinion: The smaller portal tracts have no identifiable portal vein, random arterioles in lobules, prominent lymphatics around THV. Stellate (Ito) cells prominent, macrophage foci (stain + iron), micronodule. Comment: consistent with an anomaly of the portal vasculature; age + ascites + multiple shunts; consider idiopathic noncirrhotic portal hypertension. Management: spironolactone +/- furosemide; restricted protein/sodium diet; vitamin E; +/- lactulose. One month later: ascites resolved, clinically healthy.

Rottweiler, 6 year old, male, castrate. History: inappetence and weight loss of several weeks duration. The dog appeared depressed on physical examination with loss of muscle mass and had icteric sclera. Notable diagnostic findings: PCV 33%; ALT 143 IU/L (<118); AST 137 IU/L (<66); ALP 505 IU/L (< 131); TB 2.2 mg/dL (<0.4); albumin 2.5 g/dL (2.3-3.9). A normal to heteroechoic liver was observed with ultrasound and the gallbladder was distended and "sludge" was possibly present. Two needle biopsy specimens were obtained. Pathologist #1: moderate portal fibrosis accompanied by infiltrates of lymphocytes, neutrophils, plasma cells, & macrophages (grouped into small granulomas) that extend into the parenchyma in which there is individual hepatocellular necrosis. Morphologic diagnosis: hepatitis, chronic, active, moderately severe. Comment: somewhat unusual inflammation à consider drug history such as carprofen. Pathologist #2: The microscopic alterations of one sample consist of multifocal mixed inflammation with immature plump, spindle-shaped cells. The other sample contains large foci of immature spindle-shaped cells (moderate anisocytosis) and plump immature histiocytes compatible with malignant fibrous histiocytosis. The neutrophils and hepatocellular necrosis is a secondary response (ôreactive hepatitis).

Siamese-cross, 12 years of age, female, spayed. History: May 15: 0.75 pound (0.3kg) weight loss in 3 months (see notable laboratory findings). Treated with prednisone, 5 mg qod. June 2: appetite still reduced (see notable laboratory findings); sonography: no abnormal findings. Treated with antibiotics. June 14: appetite & activity near normal (see notable laboratory findings). July 10: Physical exam: no abnormal findings. (see notable laboratory findings). Liver biopsied 4 days later.

Notable Laboratory Findings
  May15 June 2 June 14 July 10 
ALT (N<76)87253231364
AST (N<55)2811773136
ALP (N<62)39393448
GGT (<7)0000


Primary pathologist: Multifocal mild to moderate lymphoplasmacytic & neutrophilic periportal hepatitis with hepatocellular vacuolar degeneration, scattered extramedullary hematopoiesis, Ito cell hyperplasia. Comment (abbreviated): Changes are somewhat nonspecific:
  1. Ito cell: consider hypervitaminosis A, also linked to hepatic fibrosis,
  2. portal hepatitis: ascending insult from GI tract
  3. vacuolated hepatocytes: not typical of lipidosis; possibly degenerative change resulting from toxic insult or drug reaction.

Second opinion:
  1. lymphocytic portal hepatitis (not cholangiohepatitis) probably ôage-relatedö; not causative of clinical signs
  2. cytoplasmic rarefaction may be a consequence of extrahepatic disease (e.g., pancreatitis)
  3. lipid-filled macrophage foci: any oral lipid-based supplement given? May be a consequence of prior injury (e.g., pancreatitis)
  4. Telangiectatic foci may be age-related or consider bartonellosis
  5. Ito cell (lipocyte) prominence = an enigma!

Conclusion: Liver pathology not a clinical concern. Consider prior pancreatitis as a perpetrator of injury. Suggestion: Document baseline FBA & monitor ALT/AST/FBA every 3 to 6 months.

Chihuahua, 5 years of age, female, spayed. History: for approximately 3 months there have been recurrent episodes of ataxia that occur once or twice a month and are approximately 6 hours in duration. There were no significant findings on the physical examination. Notable diagnostic findings in November: FSBA 88 micromol/L (<), PPBA 35 micromol/L (<15). The remainder of the hematology and clinical chemistry and CSF analysis were unremarkable. The abdominal ultrasound, nuclear scan (transcolonic portal scintigraphy), and MRI were unremarkable (no intrahepatic anomalies or extrahepatic portosystemic shunt). In December there was another episode of ataxia (ôfaintingö). The FSBA and PPBA were 11 micromol/L (<5) and 89 micromol/L (<15), respectively, and the clinical chemistry profile was unremarkable. In January, an ultrasound-guided needle biopsy of the liver was obtained; its culture was negative (aerobic & anaerobic). The morphologic diagnosis by Pathologist #1 was lipogranulomatous hepatitis (fatty macrophages with a mixture of lymphocytes and neutrophils), and mild vacuolar hepatopathy. Comment: there are no obvious vascular anomalies. Pathologist #2: endothelial-like structures (juvenile vessels) are randomly distributed throughout parenchyma; some of the central veins have thick walls and are surrounded by dilated small vascular spaces (arteriolization?); mild multifocal diffuse hepatocellular swelling; multifocal hyperplasia/hypertrophy of fat-storage cells. Comment: these findings, along with the inability to visualize an extrahepatic portosystemic shunt, support a diagnosis of microvascular dysplasia of the intrahepatic portal circulation with secondary degenerative changes of the parenchyma. Another possibility is that an extrahepatic portosystemic shunt was missed. Management: low protein diet with adequate caloric intact (monitor weight), +/- vitamin E and follow the clinical response.

German shorthair pointer, 8 years of age, male, castrate. History: lethargic for 2-3 days, then an acute onset of ascites. Physical examination: distended abdomen-ascites. Notable laboratory findings: ALT 51 IU/L (<80); AST 51 IU/L (<55); ALP 19 IU/L (<100); albumin 1.8 g/dL (>2.5); calcium 7.5 mg/dL (9-11); FBA 5 mmol/L (<5; <25); PPBA 215 mmol/L (<15; <25); ascites: total protein: <2.5 g/dL, cell count = 735/mL (large mononuclear). Liver biopsy obtained: primary pathologist: Morphologic Dx: Vacuolar hepatopathy. Comment: a) fine vacuolation of hepatocytes suggestive of metabolic disease, b) small number of portal lymphocytes/plasma cells, c) random foci of fat-filled Kupffer cells & lipid granulomas, d) attenuation of hepatic cords suggestive of insufficient hepatotrophic stimulation from the animal's gut. Second opinion: a) ascitic transudate indicates impaired lymph flow -- possibilities include portal hypertension secondary to chronic hepatitis/cirrhosis or a mesenteric lesion -- however neither noted, b) mural hypertrophy of central veins is suggestive increased flow - "arteriolization" -- suggestive of a congenital portal vascular anomaly. The pigment-filled macrophage foci and the reduced albumin are supportive of a portal vascular anomaly. Conclusion: most likely thrombosis of a congenital vascular anomaly resulting in impaired venous-lymphatic flow. Manage the ascites and should resolve in 3 to 4 weeks. Three week follow-up: doing well, ascites resolving.

West Highland white terrier, 6 years of age, female, spayed. Pre-dental history and physical examination: clinically healthy; notable laboratory finding: ALP 1100 U/L (< 110) with progressive rise to 1600 U/L over several months. ACTH stimulation test: normal; sonogram of liver and adrenals: unremarkable; hepatic cytology û normal. Liver biopsy: marked hepatocellular swelling (vacuolation) "not typical of hyperadrenocorticalism" & minimal lymphoplasmactyic/ neutrophilic inflammation. Copper stain negative.

Questions?

( Can a pathologist tell if hepatocellular swelling (vacuolation) is or is not caused by an endocrine disorder?)

( No. Consider ballooning (hydropic) degeneration caused by a toxin; vacuole-like &/or rarefaction due to ( cytosolic water (hypoxia, ischemia, pancreatitis, drug-induced, toxins: cycad, endotoxins), glycogen accumulation (excessive corticosteroids - exogenous or endogenous), lipidosis: vacuoles û micro- and macrovesicular (feline syndrome, diabetes mellitus).

( What is the origin of the ALP? Liver? Bone? Other?)

( Most likely induced-ALP of liver origin. Liver-ALP = no microscopic cholestasis. Bone-ALP = ALP value too high, no clinical finding of bone pain.)

( Are ALP isoenzymes of value?)

( High values associated with a variety of extrahepatic diseases or medications (pancreatitis, hypothyroidism, diabetes mellitus, congestive heart failure, phenobarbital or primidone,). Generally are of limited, if any, additional diagnostic value.)

( Are additional tests of value: bile acids or GGT, T4, fT4, adrenal cortex sex hormone profile û (hyperadrenocorticalism = excessive secretion of one or more steroid hormones of the adrenal cortex)

( My plan: vitamin E and milk thistle and see what happens after a month à is this total or partial quackery? ( Primum non nocere. Hippocrates.)

Suggested Reading

Center S A: Chronic hepatitis, cirrhosis, breed-specific hepatopathies, copper storage hepatopathy, suppurative hepatitis, granulomatous hepatitis, and idiopathic hepatic fibrosis. In Guilford W G, Center S A, Strombeck D R, et al: Strombeck's Small Animal Gastroenterology. Philadelphia, W. B. Saunders, 1996, p 705-765.

Center S A, Schermerhorn T, Lyman R, Phillips L: Hepatoportal microvascular dysplasia. In Bonagura J D, ed: Kirk's Current Veterinary Therapy XIII. Philadelphia: W. B. Saunders, 2000, pp 682-686.

Laflamme D P: Nutritional management of liver disease. In Bonagura J D, ed: Kirk's Current Veterinary Therapy XIII. Philadelphia: W. B. Saunders, 2000, pp 693-697.

Meyer D J, Harvey J W: Evaluation of hepatobiliary system and skeletal muscle and lipid disorders, in Veterinary Laboratory Medicine û Interpretation and Diagnosis, W. B. Saunders, 2nd edition, 1998, pp 157-186.

Meyer D J: Hepatic pathology. In Guilford WG, Center SA, Strombeck D R, et al: Strombeck's Small Animal Gastroenterology. Philadelphia, W. B. Saunders, 1996, p 633-653.


REACTIVE HEPATOPATHIES: THE MOST COMMON LIVER DISEASE
David C. Twedt, DVM, DACVIM

One of the most common group of laboratory abnormalities encountered in small animal practice are abnormal liver enzymes (ALT, AST, ALP and GGT). Some animals are asymptomatic and the abnormalities are detected after performing a routing health screen. For example, the dog with dental disease that has a pre-anesthesia profile with elevations in the ALT and ALP, what should the clinician do: ignore them, perform additional diagnostic tests or consider a liver biopsy? The second scenario is the sick patient having abnormal liver enzymes. Does this animal have liver disease, should a diagnostic liver evaluation be the focus of attention or should a liver biopsy be considered? The answers to these questions are difficult and not absolute.

By far most common cause of abnormal liver enzymes in the sick patient are liver changes that arise secondary to a primary non-hepatic disease. These changes are collectively referred to as reactive hepatopathies. The reactive nature of the liver in response to other disease conditions is an important concept to understand. In these situations the hepatopathy is not specifically treated but rather the primary non-hepatic disease is. The table lists some common conditions that are often associated with reactive hepatopathies. Since reactive hepatopathies are the most common cause of abnormal liver enzymes those having abnormalities should first be evaluated for a primary non-hepatic disease in order to help explain the abnormal liver tests before beginning a comprehensive in-depth evaluation of the liver.

A secondary reactive hepatopathy poses two diagnostic problems: (1) it mimics primary hepatic disease and (2) it diverts attention from the primary extra-hepatic disease process. If an extra-hepatic condition is identified and there is evidence of good hepatic function the primary non-hepatic disease should be first addressed. Treating the primary disease usually results in resolution of the serum hepatic tests and histomorphologic abnormalities. If no explanation for the abnormal liver tests is identified, if there is evidence of altered hepatic function (such as abnormal serum bile acids), persistent abnormal liver enzymes continuing over time or deterioration of the condition of the patient then the liver should then be investigated as the primary disease. This often requires a liver biopsy to support ones suspicion of primary liver disease, which then will require specific therapy.

There are a variety of reasons why extra-hepatic diseases secondarily involve the liver. These can be divided into anatomical and functional relationships. The liver has two blood supplies, the hepatic artery and the portal vein. The former provides nutrition and oxygen. The later, comprises approximately 80% of the total hepatic blood flow, delivers substances absorbed from the gastrointestinal tract and hormones from the pancreas. Consequently hepatic integrity and function can be altered secondary to cardiovascular insufficiency, anemia, portosystemic shunts, and exposure to ingested xenobiotics and intestinal bacteria or their products when the intestinal barrier is violated by disease.

Hepatocytes, the principal parenchymal cell, reside in acini composed of three diverse metabolic zones. Blood flows from the portal triad passing through zones 1,2 and 3 before draining via the hepatic vein. Consequently, hepatocytes in zone 3 are most susceptible to hypoxic conditions such as heart failure and shock. The metabolic diversity of the hepatic zones is necessary to accommodate the numerous homeostatic activities. Many of these functions are related to the intermediary role of hepatocyte metabolism between dietary sources of energy and extra-hepatic tissue demands for energy. Therefore metabolic diseases often involve the liver. Examples include hyperadrenocorticism, diabetes mellitus, hyperthyroidism, perhaps hypothyroidism and nutritional disorders.

Another cell type that plays a role in the extra-hepatic manifestations of disease is the Kupffer cell; a member of the monocyte-macrophage system. It is involved in the hepatic immune response and "filters" toxins, acute phase reactant proteins, cytokines and bacteria that enter the portal circulation. When this role is amplified in response to extra-hepatic disease, focal hepatitis can develop. It may be helpful to conceptualize the liver as a large lymph node in the center of the body, which reacts to a multitude of insults. There are in fact more RE cells concentrated in the liver than any other part of the body.

CLINICAL FINDINGS

The following information presented is based on our interpretation after reviewing 150 consecutive liver biopsies performed at Colorado State University. Indications for these liver biopsies were based on clinical findings, laboratory abnormalities or imaging changes. The histological findings were grouped into a number of general categories. Of the three largest categories those grouped as reactive hepatopathies make up the largest number of cases (25% N=38). The two other major groups included those having chronic hepatitis and hepatic neoplasia. The reactive hepatopathy group was characterized by nonspecific hepatocellular degeneration or necrotic changes without evidence of chronic progressive inflammation. Histological findings often include descriptors such as vacuolar degeneration, hydropic degeneration, swollen hepatocytes, lipidosis, intracellular or intra-hepatic cholestasis, and periportal or variable hepatic necrosis. It should be noted that a majority of these changes were devoid of typical chronic inflammatory cell infiltrates or fibrosis characteristic of chronic hepatitis.

Of these cases we were later able to identify associated disease and the probable cause for the hepatic enzyme and histological changes. Concurrent diseases included non-hepatic neoplasia, gastrointestinal, renal, autoimmune, dermatologic, dental, infectious and cardiac disease as a few examples. In only 2 cases was an underlying disease not identified to explain the findings. The ALT values on the average were 1 X normal and the ALP values were 3-4 X normal. It is interesting to note that in all cases in which serum bile acids were run all were in the normal reference range suggesting hepatic function remained intact in most dogs having a secondary reactive hepatopathy.

CASE EXAMPLES

The following are some cases examples of reactive hepatopathies that occur in the dog that demonstrate reactive hepatopathies.

Cholestasis of Sepsis.

Cholestasis or impaired bile flow can be classified as extra-hepatic or intra-hepatic. Differential considerations include lesions that obstruct the flow of bile in the extra-hepatic biliary system. Intra-hepatic cholestasis can develop for a number of reasons and one is in association with extra-hepatic bacterial infections and sepsis. Clinically the patients may be icteric. The hepatic enzyme tests (especially ALP) show only mild to moderate increases despite sometimes a remarkable increase in the serum bilirubin concentration and relatively unremarkable histological findings. There may be accumulation of bile pigment, including canalicular plugs, and a mild inflammatory cell component, often round cell and periportal. The intra-hepatic architecture remains intact. Pathologic terms such as periportal hepatitis, focal hepatitis or hepatitis, chronic, active with bile retention may be used to characterize the microscopic findings. The successful treatment of the extra-hepatic infectious process is associated with spontaneous resolution of the cholestasis. The pathophysiologic mechanisms causing this functional cholestasis are multiple and poorly understood. Toxins derived from bacteria (especially E. coli) may either directly or indirectly by formation of antibodies to bacterial cell wall components that cross-react with the canalicular membrane and "paralyze" the energy-dependent transport systems of the membrane. It has also been shown that selected acute-phase reactant proteins can interfere with hepatocellular uptake of bile acids and other hepatocellular functions. The common pathophysiologic factor is the impaired excretion of bile acids a primary driving force for bile flow.

Gastrointestinal Disease

Occasionally dogs and cats with chronic intestinal inflammatory disease will have concurrent hepatic changes. The nature of the hepatic changes leads one to speculate that they are secondary to the portal uptake of bacteria or their products. The nature of the inflammatory infiltrate (periportal) secondary to intestinal disease has led to the hypothesis that the changes are secondary to bacteremia from intestinal bacterial translocation through the diseased intestinal tract. A concept now in favor is that the endotoxin lipopolysaccharide (LPS), a component of the cell wall of gram negative bacteria, transit the intestinal wall in significant amounts in states of intestinal inflammation. The hepatic reticuloendothelial system (Kupffer cell) is critical for LPS detoxification and impairment of this function may lead to hepatic injury/inflammation. Though endotoxins may directly damage hepatocytes, evidence suggests that a release of a number of substances from the macrophage mediates diverse biological effects. The macrophage products include superoxides, toxic oxygen radicals, tumor necrosis factor and platelet activating factor.

Serum hepatic enzyme tests may reflect intra-hepatic cholestasis and hepatocellular injury. Rarely will tests of hepatic function be altered. Changes are most often develop in association with long standing intestinal disease. The most common hepatic microscopic findings include mild to moderate periportal inflammatory cell infiltrates consisting of lymphocytes, plasma cells macrophages and neutrophils; these findings prompt the morphologic terms of hepatitis, chronic, active. When the inflammation primarily surrounds the bile ducts, pathologic terms such as cholangitis may be used. Ductular proliferation and mild fibrosis may be present. Vacuolated hepatocytes with lipid may be found but its presence is probably related more to the general nutritional debility of the patient.

Acute Pancreatitis

Acute pancreatitis can secondarily involve the liver through two pathologic processes: (1) impairment of bile flow in the common bile duct and (2) direct intra-hepatic damage. There is a variable amount of peripancreatic inflammation associated with acute pancreatitis. When the inflammation is sufficiently severe, it encompasses the common bile duct and can cause clinical, biochemical and histological findings compatible with extra-hepatic cholestasis. In most cases biochemical and microscopic findings consistent with hepatocellular injury and intra-hepatic cholestasis can also develop in association with acute pancreatitis. There are mild to moderate increases in the hepatic enzyme tests with or without hyperbilirubinemia. Microscopically there may be focal necrosis, Kupffer cell hyperplasia, mild periportal inflammatory cell infiltrates, and hepatocellular accumulation of bile pigment and vacuolar changes in the hepatocytes. The pathophysiologic mechanism responsible for these changes is not known. Since the portal circulation receives blood from the pancreas, it is tempting to speculate that the release of proteases directly into the portal circulation from the inflamed pancreas plays a pivotal role. Alternately, toxic substances released by the injured pancreas may be "filtered" by the Kupffer cell and stimulate the release of cytokines which amplify intra-hepatic injury.

Metabolic Disease

A variety of metabolic diseases can cause abnormal liver tests and, in some cases, alter hepatic morphology. One of the more widely recognized hormonally associated diseases that frequently affect the liver is hyperadrenocorticism. The canine liver is uniquely "responsive" to glucocorticoids; highlighted by moderate to markedly increased serum alkaline phosphatase activity (without hyperbilirubinemia) and a foamy change in the hepatocyte cytoplasm caused by glycogen accumulation. The glycogen accumulation can be diffuse enough to cause hepatomegaly. Less frequently the serum alanine aminotransferase and aspartate aminotransferase activities are mildly increased; presumably secondary to drug-stimulated production. There is minimal if any identifiable loss of hepatic function in cases with hyperadrenocorticism. The author believes that endogenous stress can also cause a mild steroid hepatopathy. The sick and stressed patient for any reason may be expected to have moderate increases in biochemical liver enzymes (ALT, ALP). For example a clinical study evaluating post operative ovariohysterectomy dogs found these previously normal patients to have increases in ALT and ALP concentrations for a two week period following surgery presumably due to secondary changes associated with anesthesia and surgery.

Insulin deficiency alters glucose and lipid metabolism. Patients with diabetes mellitus may develop hepatic lipidosis with abnormal liver tests. Hyperthyroidism in cats can consistently cause abnormal liver tests (ALT, ALP) and sometimes including hyperbilirubinemia, which spontaneously resolves following successful management of the disease. There is minimal to no microscopic hepatic lesions observed. In dogs with hypothyroidism, the authors have occasionally associated mild to moderately abnormal hepatic enzyme tests and hepatic hydropic degeneration and Ito cell prominence.

In summary, abnormal hepatic tests and histological findings can be associated with extra-hepatic diseases. The recognition of this relationship will focus attention on the diagnosis and management of appropriate disease process and curtail the misdiagnosis of primary hepatic disease.

Examples of Extra-hepatic Disorders that Can Cause Abnormal Liver Tests

Inflammatory bowel disease
Extra-hepatic infections (bacterial)
Rickettsial infections
Acute Pancreatitis
Diabetes mellitus
Hyperadrenocorticism, Hypoadrenocorticism
Hyperthyroidism, Hypothyroidism
Immune-mediated hemolytic anemia
Septicemia
Shock
Right-sided heart failure
Protein-losing enteropathy
Severe protein restriction
Starvation
Splenitis


SELECTED REFERENCES
  1. Anwer M S, Meyer D J. Bile acids in the diagnosis, pathology and therapy of hepatobiliary disease. Vet Clin North Am Small Anim Pract. 1995; 25: 503-518.
  2. Center S A. Pathophysiology and laboratory diagnosis of hepatobiliary disorders. In: S J Ettinger and E C Feldman eds. Textbook of Veterinary Internal Medicine. Philadelphia: W B Saunders, 1995; 1261-1312.
  3. Center S A, Baldwin M S, Hollis N, et al: Bile acid concentrations in the diagnosis of hepatobiliary disease. JAVMA 1985; 187:(9) 935-940.
  4. Center S A, Slater M R, Manwarren T, Prymak B S: Diagnostic efficacy of serum alkaline phosphatase and gamma glutamyltransferase in dogs with histologically confirmed hepatobiliary disease: 270 cases (1980-1990) JAVMA 1992; 201:(8) 1258-1264.
  5. Center S A, Warner K L, Corbett J R: PIVKA clotting times in dogs with suspected coagulopathies. JVIM 1998; 12:214.
  6. Dial SM. Clinicopathologic evaluation of the liver. Vet Clin North Am Small Anim Pract. 1995;25:273-274.
  7. Fondacaro J V, Guilpin V O, Powers B E, Twedt D C: Diagnostic correlation of liver aspiration cytology with histopathology in dogs and cats with liver disease. JVIM 1999; 13: 254.
  8. Meyer D J. Hepatic test abnormalities. In Meyer D J, Coles E H, Rich L J. eds. Veterinary Laboratory Medicine: Interpertation and diagnosis. Philadelphia: W B Saunders, 1992 83-92.
  9. Meyer D J, Williams D A: Diagnosis of hepatic and endocrine pancreatic disorders. Seminars in Veterinary Medicine and Surgery (Small Animal) 1992; 7:(4) 275-284.
  10. Roth L, Meyer D J. Interpretation of liver biopsies. Vet Clin North Am Small Anim Pract. 1995; 25: 293-304.
  11. Willard M D, Twedt D C: Gastrointestinal, pancreatic and hepatic Disorders. In Willard M D, Tvedten H, Turnwald G H eds. Small Animal Clinical Diagnosis by Laboratory Methods. 2nd Edition . Philadelphia: WB Saunders, 1994, 179-218.

Key Words: Reactive hepatopathies, Sepsis, Pancreatitis, Gastrointestinal disease, Steroid hepatopathy.


CHRONIC HEPATITIS: RECENT ADVANCES
David C. Twedt DVM, DACVIM

Chronic Hepatitis is an etiologic diverse and morphologically variable condition characterized by a mixed inflammatory cell infiltrates. Plasma cells, lymphocytes and macrophages predominate with a lesser number of neutrophils. The etiology of this chronic inflammatory condition is generally never determined.

ETIOLOGY

Copper associated chronic hepatitis has been documented and is discussed below. Drug-induced hepatitis specifically primidone, Phenobarbital and diethylcarbamazine oxibendazole has been reported to produce chronic hepatitis in some dogs. Recently we have also observed dogs receiving chronic carprofen to have changes representing chronic hepatitis. Infectious agents such as leptospirosis and infectious canine hepatitis are also a cause of chronic hepatitis. Possibly other infectious agents may be responsible as well. Alpha1-antitrypsin (AAT- also referred to as alpha one protease inhibitor) deficiency in the serum leading to accumulation of AAT in the hepatocytes, is known to cause chronic hepatitis and cirrhosis in man. Recent investigation by researchers in Sweden using immunostaining for AAT in hepatocytes found many dogs with chronic hepatitis to be positive. The breed most often associated with AAT accumulation was the cocker spaniel.

Circulating autoantibodies are important diagnostic markers used to identify autoimmune liver disease in humans. It appears that autoantibodies (ANA, antimitochrondial antibodies [AMA], smooth muscle antibodies [SMA], liver membrane autoantibodies [LMA]) are of a minor importance in classifying canine chronic hepatitis and thought to occur in most cases secondary to the liver damage. Nonetheless, immune mediated mechanisms are thought to be associated with certain cases of chronic hepatitis and are supported by the fact that some dogs respond favorably to immunosuppressive therapy. Secondary immune hepatitis may result from non-specific damage to hepatocytes that result in release of liver antigens. Antibodies are then formed against these liver specific antigens with continued damage intact hepatocytes.

BREED PREDISPOSITION

There are a number of breeds that have an increased incidence of chronic hepatitis, which suggests a genetic basis. Chronic hepatitis in Doberman Pinschers is more common in young to middle-aged female dogs. The CH is characterized by cholestasis, hypoalbuminemia with increased concentrations of copper and iron in the liver tissue. CH has been reported that Cocker Spaniels as an inherited liver disease that is specific but not associated with copper accumulation. These dogs have marked hypoalbuminemia, ascites and hepatic encephalopathy and generally have a grave prognosis. Other breeds that appear to have increased incidence include Labrador Retrievers and Standard Poodles.

COPPER ASSOCIATED LIVER DISEASE

Abnormal hepatic copper accumulation may be the result of either a primary metabolic defect in copper metabolism or as a secondary event as the result of abnormal hepatic function causing decreased copper excretion. When we reviewed a number of dogs having chronic hepatitis and not associated with known breed associated copper accumulation we found many dogs had increases in both copper and iron hepatic concentrations. A number of these dogs were also deficient in hepatic zinc. The interrelationship of the heavy metals and liver disease needs further investigation.

The diagnosis of abnormal Cu accumulation requires a liver biopsy. Excess Cu within the liver can be demonstrated using histochemical staining for hepatic Cu using rhodanine or rubeanic acid stain. Definitive determination of excess hepatic Cu requires a quantitative analysis of tissue Cu measured on the biopsy sample. Normal canine hepatic Cu concentrations are less than 400 ¦g/g dry weight liver. Hepatic Cu concentrations in dogs with secondary Cu accumulation generally fall in the range of 800 to 1,500 ¦g/g dry weight while breed associated hepatotoxicities generally have much higher concentrations.

Hepatic copper toxicity was first identified in Bedlington Terriers. It was subsequently shown that affected Bedlington terriers have an inherited autosomal recessive defect, which results in reduced biliary excretion of copper due to hepatic metallothionein sequestration of the metal in hepatic lysosomes. Clinically there is a progressive hepatic Cu accumulation with age ranging from 1,000 to 12,000 ¦g/g per dry weight of liver. The extent of hepatic damage tends to parallel the increasing hepatic Cu concentrations.

During the last decade an increasing number of breeds other than the Bedlington terrier have been identified with concurrent chronic hepatitis and increases in the hepatic Cu content. Liver disease with concurrent Cu accumulation is reported in the Doberman Pinscher, West Highland White Terrier, Skye Terrier as well as many other pure breed and mixed-breed dogs. Recently we have identified a number of Dalmatians having a suspected inherited copper associated liver disease.

CLINICAL FINDINGS

The incidence of chronic hepatitis makes up approximately one fourth of the cases having liver biopsies at Colorado State University (based on a review of 150 consecutive liver biopsies). Chronic hepatitis is more common in female dogs and the average age at presentation ranges from 4 to 10 years. It is interesting to note that in both our series and in studies by others it is uncommon to observe chronic hepatitis/cirrhosis in dogs older than 10 years of age. The clinical signs parallel the extent of hepatic damage. Early in the disease there are usually no or only minimal clinical signs. Only after the disease progresses do the clinical signs specific for liver disease becomes evident. Frequent early signs are gastrointestinal associated with vomiting, diarrhea and poor appetite or anorexia. Ascites, jaundice and hepatic encephalopathy may then occur as the disease progresses. With development of these late signs the long-term prognosis is generally poor. The laboratory findings include consistently elevated ALT, ALP and GGT. The magnitude of rise need not be marked however. One report found 75% of the cases of chronic hepatitis to have abnormal bilirubin increases (mean value of 2.6 mg/dl). Hyperbilirubinemia is however considered to be a later laboratory finding. Serum proteins are variable and as chronic hepatitis lesions become more severe albumin levels decline. We found serum bile acids concentrations to be abnormal in most all cases having chronic hepatitis. Early in the disease the fasted bile acids may be normal but postprandial bile acid concentrations are abnormal. We believe that identifying abnormal bile acid concentrations in the dog having abnormal liver enzymes to be a good predictor of significant liver disease which includes chronic hepatitis. In our study all dogs evaluated with chronic hepatitis had abnormal bile acid concentrations. In a second study only 8/26 dogs with chronic hepatitis had normal fasting bile acids. However in theses cases postprandial samples were not determined. Determining postprandial bile acids has been shown to increase the sensitivity of this test.

DIAGNOSIS

A presumptive diagnosis is made based on the clinical features and persistent increases of ALP and ALT values. The diagnosis is supported by abnormal bile acid concentrations, radiographic or ultrasound findings. A definitive diagnosis however requires a hepatic biopsy showing characteristic morphological patterns. Fine needle aspirates with cytology are not helpful in making the diagnosis of chronic hepatitis because it is important to see the architecture of the liver and location and extent of the inflammation. Recent evidence also suggests that a needle biopsy may not provide enough portal triads to absolutely make the diagnosis of hepatitis. One must work with the pathologist when making the diagnosis of chronic hepatitis and to be certain that characteristic abnormalities found in chronic hepatitis are present. That there is still considerable confusion in nomenclature and description of histological changes and terminology varies with pathologists. I believe that all cases suspected of having chronic hepatitis should also have hepatic copper, zinc and iron content determined. Finding abnormalities in metal content will help direct a course of therapy.

THERAPY

The therapy for chronic hepatitis involves removing the primary etiology. Short of treating the primary etiology all other therapies suggested are unproven in the management of chronic hepatitis in dogs. We are still waiting for good clinical studies proving efficacy in treatments. Such studies are hindered even from the start owing to the multiple etiologies of hepatitis and the inconsistent histological descriptions. To date we have only limited case studies and clinical impressions of efficacy in the management of chronic hepatitis.

Diet

Dietary therapy should be considered in all cases and general guidelines can be given. There is a major misconception about diet and liver disease that states all patients should be placed on a protein restricted diet. The goal of dietary therapy is to adjust the quantities and types of nutrients to provide needed nutrient requirements but to avoid the production of excess nitrogen by-products associated with liver disease and to provide factors that support liver function and regeneration. I believe that protein restriction should only be instituted in the patient that has clinical evidence of protein intolerance (i.e. hepatic encephalopathy). If the patient has hypoalbuminemia without hepatic encephalopathy the diet should be high in digestible protein. The diet should be selected that is of a good biological value protein. As a general recommendation the dietary protein should represent 17 to 22% of digestible Kcal. Lower amounts are used when hepatic encephalopathy signs are present. Home diets or commercial diets with or without protein supplements can be used. High carbohydrate and moderate fat content is suggested in addition to a good source of multiple vitamins. Mineral supplementation containing high concentrations of both copper and iron should be avoided. Potential benefits of zinc for enhanced ureagenesis and immune function and vitamin E for antioxidant effects is suggested.

Recently there has been evidence that fiber may have several beneficial effects in patients having liver disease. First, dietary fiber effectively binds bile acids in the intestinal tract and promotes their removal. Secondly soluble fiber appears to have some benefit in managing hepatic encephalopathy. Diets of vegetable protein sources have been effective in the management of patients with HE and some of the beneficial effects attributed to increases in fiber intake. In summary, high soluble fiber diets would seem a logical nutritional addition to the management of patients with hepatic encephalopathy. Psyllium as a source of soluble fiber given at a dose of 1-3 tsp/day can also be used as a dietary supplement.

Diets low in copper are recommended for breeds known to accumulate copper or that have documented increased concentrations on biopsy. The restriction of dietary copper however probably does little to lower hepatic copper concentrations in already diseased dogs with large amounts of hepatic copper. It is difficult to limit dietary copper because most all commercial dog foods contain supplemental copper that meet, or more frequently exceed the minimal dietary requirements. These standards allow for at least 7.3 mg/kg diets on a dry matter basis (DMB), a level considered to be far in excess of what is required for even the normal dog. Most commercial foods contain concentrations in excess of 7 mg/kg DMB. Unfortunately, many manufacturers do not list the copper content in their diets making dietary selections difficult.

Antiinflammatory Agents

Reducing Inflammation, as a specific therapy for chronic hepatitis is unproven though the clinical impression suggests antiinflammatory therapy is beneficial. The use of corticosteroids in the treatment of chronic hepatitis is quite controversial and there are as yet no good controlled studies in animals to support their use in every case. Corticosteroids have however proven beneficial in cases of immune mediated chronic hepatitis in humans.

Clinical experience from treating dogs with chronic hepatitis suggests that many dogs will improve following therapy. In a study by Strombeck found that dogs with chronic hepatitis tended to have a prolonged survival when treated with corticosteroids. This retrospective study is one with a wide diversity of diseases and concurrent therapies. But nonetheless, it appears that corticosteroids offer benefit in at least some cases. A suggested dose of 1 to 2 mg/kg/day using either prednisone or prednisolone should be instituted. When clinical improvement is suspected or after several weeks the dose is then gradually tapered eventually to a dose of 0.5 mg/kg/day or every other day. Since glucocorticoids alone cause elevations in ALP and ALT values enzyme evaluations are not helpful. The only accurate way to evaluate a response to any therapy is to re-biopsy the patient in 6 months to 1 year. The optimal duration for therapy is unknown and it appears that it may be life long in most cases.

Azathioprine (Imuran) is an effective immunosuppressant drug that has shown to increase survival in man when treated for chronic hepatitis. However azathioprine alone is not as effective as either remission with steroids then azathioprine or steroids and azathioprine combination. This therapy may also be beneficial in dogs by increasing the immunosuppressive response and enabling a reduction of both steroid dose and their side effects. A dose of 0.5 mg/kg/day is the suggested starting dose. The level of glucocorticoids can frequently be reduced when using azathioprine. The dose of azathioprine is generally tapered to an alternate day therapy after several weeks. The hemogram including platelet counts should be followed initially to detect and prevent azathioprine induced bone marrow suppression. One should also note that azathioprine can be associated with an idiosyncratic drug hepatotoxicity in dogs.

Copper reduction

If the liver biopsy of a dog with chronic hepatitis indicates abnormal hepatic copper accumulation, copper chelators should be considered. Hepatic copper stains will support copper retention but actual determination of hepatic copper is essential. Hepatic copper levels of greater than 750 ¦cg/g dry weight liver should have therapy to reduce copper concentrations. Animals having greater than 2,000 ¦cg/g dry weight copper content should all have chelator therapy.

Zinc therapy decreases intestinal absorption of Cu by inducing synthesis of intestinal mucosal protein metallothionein. Copper binds more tenaciously to metallothionein than zinc. This results in impeded serosal transfer of copper and eventual loss into the feces when the intestinal cells are shed. Thus, a mucosal block of Cu absorption is achieved. This block involves exogenous dietary copper and endogenously secreted copper found in the intestinal tract. There is also preliminary evidence that zinc therapy may also decrease hepatic stores of Cu though the mechanism is unknown.

Suggested doses of elemental zinc range from 5 to 10 mg/kg given every 12 hours. Several months after induction the dose was decreased by half. Serum zinc concentrations were monitored with a goal of no more than a twofold increase of serum concentrations (< 300 ¦g/ml). In this study serum zinc levels remained in the suggested therapeutic range and there was no evidence of toxicity. These results suggest zinc therapy may be beneficial in the management of dogs with increased hepatic Cu concentrations.

Chelating agents cause excretion of copper from the body and treatment using chelators is shown to have beneficial effects in dogs with increased hepatic Cu concentrations. These specific chelators bind with Cu in blood or tissues and promote their removal through the kidneys.

D-penicillamine (Cuprimine«) is the copper chelator recommended for the therapy of Wilson's disease in humans. In Wilson's disease patients, PCA depletes most but not all the hepatic copper. After a period of therapy urinary copper excretion levels decline even though there is continued abnormal hepatic copper concentrations. There is speculation that there may be several pools of hepatic copper in Wilson's disease, some of which may not be effected by PCA. There is now evidence that PCA may detoxify the remaining copper in the liver, either by of formation of a chelate that is nontoxic or by inducing the synthesis of hepatic metallothionine that binds and detoxifies copper. Trientine (Syprine«) is also a copper chelator that acts by increasing urinary excretion of copper. Both PCA and trientine have been used in dogs with increased hepatic copper concentrations and have been shown to be effective. PCA and trientine are given at 10 - 15 mg/kg bid. Either drug should be given 1-2 hours before a meal.

Antibiotic Therapy

Antibiotic therapy is generally not required for most cases of chronic hepatitis except for the treatment of primary hepatic infections, septicemia, prophylaxis and for the management of hepatic encephalopathy. In some situations where the specific indication for antibiotic therapy is not readily identified, the author has found in a few selected cases metronidazole to be of benefit for unknown reasons. Metronidazole is bactericidal, antiprotozoal and also has been shown to have immunosuppressive activity, specifically against cell-mediated immune responses and to also have some antioxidant effects. Approximately 30 to 60% of the drug relies on hepatic excretion and an empirical dose reduction has been recommended for patients with hepatic disease. A dose of 7.5 mg/kg bid is suggested in patients with liver disease. Though unproven metronidazole may be useful as an adjunct therapy in some cases. Metronidazole toxicity, associated with a central vestibulitis, should be a constant concern in the patient with liver disease.

Other and Supportive Therapy

Generally additional supportive therapy for chronic hepatitis involves treatment of secondary complications. These complications occur as the disease becomes advanced. Hepatic encephalopathy, gastrointestinal ulceration, ascites and fluid and electrolyte abnormalities are common clinical occurrences that must be managed. Other therapies can be used in chronic hepatitis however few studies have proven efficacy in managing chronic hepatitis in the dog. These other therapies will be covered in the manuscript entitled "A review of traditional and not so traditional therapies for liver disease."

Prognosis

There is little information of the prognosis with and without therapy. The prognosis in dogs with advanced chronic hepatitis and cirrhosis is guarded. In a study by Strombeck found mean survivals ranging from 6 to 16 months with therapy. This study also identified that dogs with hypoalbuminemia, hypoglycemia and coagulopathies have very guarded prognostic factors and many died within 1 week of diagnosis. A second study of 79 dogs found that dogs with cirrhosis had a survival of less than one week and dogs with chronic hepatitis had a mean survival in the range of about 20 to 30 months. Most of these dogs were not advanced in their disease and had concurrent corticosteroid therapy.

A Selected Reference List Is Available Upon Request

Key Words: Chronic hepatitis, copper hepatoxicity, Chronic hepatitis therapy, Copper chelators, Chronic hepatitis dietary management.


A REVIEW OF TRADITIONAL AND NOT SO TRADITIONAL THERAPIES FOR LIVER DISEASE
David C. Twedt DVM, DACVIM

There are no controlled studies evaluating therapies for liver disease in the dog. Consequently treatment recommendations are unsupported and based on case reports, retrospective reviews and information gathered from human studies. This information must be interpreted with care. A majority of the information presented includes therapy used for the management of chronic hepatitis. Because chronic hepatitis in the dog is a collection of undifferentiated sub-groups of multiple etiologies improvements noted in one case may not directly extrapolate to all cases. Furthermore, human diseases have no canine counterpart from which one can readily extrapolate information. The following discussion of various treatments is based on human reviews and clinical reports in dogs.

IMMUNOSUPPRESSIVE THERAPY

Glucocorticoids (steroids) have a number of potential favorable benefits in the management of chronic hepatitis. In addition to the antiinflammatory effects (lymphocyte reduction, reduced macrophage receptors for immunoglobulins, stabilizing lysosomes, neutrophil chemotaxis, and inhibition of prostaglandin and leukotrine production) they are also choleretic and inhibit fibrosis. They improve appetite and general well-being but can promote water retention and gastric ulceration. Steroids can also have a negative effect on survival in advanced cases by initiating decompensate or precipitating death.

Steroids have been used for the management of hepatitis in humans, however reports of responsiveness are variable as are the types of hepatitis that occur in man. There is evidence that corticosteroids have a beneficial effect on symptoms, hepatic histology and survival. These conclusions are derived from a highly select group of patients with autoimmune chronic hepatitis. The efficacy of corticosteroids in patients having other subgroups of hepatitis is quite variable. In a large study, patients having chronic active hepatitis were randomized to receive prednisone, prednisone and azathioprine or placebo therapy. Prednisone alone or in combination with azathioprine was superior to placebo. However, in this study 20% of the patients failed to respond to prednisone alone or in combination, and a similar number (20% of placebo group) experienced spontaneous remission. In summary, this study found corticosteroids improved survival and had the most pronounced effect to reduce mortality during the first 5 years of therapy.

Steroids appear to be of benefit in chronic lymphocytic-plasmacytic or autoimmune hepatitis. A recommended dosage of prednisone is 2.2 mg/kg for several weeks then tapered to 1.1 mg/kg every other day. Azathioprine in combination with prednisone is effective in humans with chronic hepatitis, however when used alone it is not as effective as prednisone. Azathioprine should be considered in dogs that cannot tolerate, or fail to respond to corticosteroids. Other immunosuppressant agents such as methotrexate or cyclosporin A have been used in hepatitis management in humans, however there are no clinical reports of their use in dogs with chronic hepatitis. Cholangiohepatitis in the cat is a chronic inflammatory liver disease that is generally treated with corticosteroid therapy. We have recently treated several patients that failed to respond to steroids but with the addition of chlorambucil at a dose of 2 to 3 mg per cat given orally three times a week seemed to improve. Chlorambucil is a slow alkylating agent that appears to be well tolerated in the cat.

URSODEOXYCHOLIC ACID

There is evidence that ursodeoxycholic acid UDCA is hepatoprotective for various types of liver disease. In cholestatic disease, hydrophobic bile acids concentrations increase and are potentially hepatotoxic. UDCA is a hydrophilic bile acid that displaces the more hydrophobic bile acid pool in cholestatic liver disease. Isolated hepatocyte studies have demonstrated the detergent properties of bile acids on membranes with emphasis on mitochondrial membrane damage leading to mitochondrial electron leakage and generation of oxygen free radicals. There is also preliminary evidence that UDCA may in its own right actually have antioxidant properties as well. UDCA acid appears to protect hepatocytes from oxidative injury by increasing levels of GSH and metallothionein. Other functions include induction of bile flow where unconjugated UDCA is secreted into the bile that subsequently promotes biliary water secretion. UDCA may also inhibit ileal uptake of secondary bile acids through competitive inhibition. Immunomodulating effects of UDCA have been reported to decrease immunoglobulin and interleukin responses and decrease hepatocyte cell surface membrane antigen expression (HLA) on liver and biliary cells.

Human clinical trials have shown that UDCA reduces biochemical enzyme markers of both cholestasis and hepatocellular damage in patients with chronic liver disease and improves liver histology in patients with primary biliary cirrhosis and sclerosing cholangitis. Data from placebo-controlled studies now supports the above observations, but it is yet unclear if there is a change in long-term survival. One multicenter study found no histological improvement in patients with chronic hepatitis that were treated with UDCA for one year. Several studies show UDCA not to be beneficial in viral hepatitis but of benefit in acute hepatotoxicity and autoimmune hepatitis, although combination with corticosteroids appeared to have a better response than UDCA alone.

In a canine case report, Meyer evaluated UDCA in a dog with chronic hepatitis and showed improvement in biochemical markers and a reduction in hepatotoxic bile acids during a 6 month follow-up. Others have shown no toxicity in UDCA therapy in dogs and cats. In summary, UDCA appears to have a place in the management of chronic hepatitis in the dog because it improves cholestasis, reduces hepatocellular damage and is not toxic. Dosage range is 10-15 mg/kg/day. Whether UDCA should be used as a sole therapy for chronic hepatitis in dogs is undetermined.

COLCHICINE

Colchicine is used in humans for the treatment of hepatic fibrosis and primary biliary cirrhosis. The actions of colchicine are many and include antifibrotic properties, by inhibiting the microtubular mediated transcellular movement of procollagen into the extracellular matrix. Colchicine also increases collagenase activity (at least in vitro) thus increasing collagen degradation. Colchicine also suppresses mediators of fibrogenesis from macrophages, has hepatoprotective effects stabilizing hepatocyte membranes, alters leukocyte locomotion and may act by modulating the function of circulating cytokines such as IL-1, IL-2 and TNF. There is now evidence that inflammation may be the principle mechanism for regulating metabolism of connective tissue and that colchicine functions by inhibiting both inflammation and fibrosis.

There are a number of reports showing colchicines effectiveness in chronic liver disease in humans. A randomized, double blind, placebo controlled study of 100 patients with early or mild cirrhosis of various causes found better survival in the colchicine group (11 years) versus the placebo group (3.5 years). Three studies of primary biliary cirrhosis using the Mayo survival model showed improvement of liver enzymes and increases in albumin concentrations as well as improved survival rate.

There are only three clinical papers reporting the use of colchicine in dogs with liver disease. One dog with chronic hepatitis showed clinical remission for 7 months and a biopsy taken at two months had some histological improvement. A dog with hepatoportal fibrosis survived 30 months with improvement of biochemical liver function and histology reflecting no progression of the hepatic fibrosis. A third Cocker Spaniel with fibrotic hepatopathy was managed in part with colchicine and had apparent benefit. There appears to be limited toxicity of colchicine, with the most common side effects being gastrointestinal signs. The dose recommendation for dogs is 0.03 mg/kg/day. Colchicine may be indicated in cases of chronic hepatitis with considerable fibrosis, biliary cirrhosis or in primary fibrotic diseases.

ZINC

Zinc is an essential co-factor involved in a number of biological reactions. Zinc is used predominately for management of Wilson's disease but is also reported to have anti-fibrotic properties and direct hepatoprotective effects by inhibiting lipid peroxidation and stabilizing lysosomal membranes. Zinc (Zn) deficiency produces low resistance to lipoprotein membrane oxidation. Zn has been suggested to have an antioxidant role by two mechanisms; the protection of sulfhydryl groups preventing oxidation and the inhibition of the production of ROS by transition metals. The latter may result from the specific induction of hepatic metallothionein (MT) proteins that binds the transitional metal copper. Zinc deficiency in many humans with advanced chronic liver disease correlates with the occurrence of hepatic encephalopathy. This finding supports the benefit of zinc for enhanced ureagenesis.

Hepatic zinc concentrations are reported decreased in some humans with liver disease, suggesting an indication for supplemental zinc therapy. We have also documented hepatic Zn deficiency in a number of dogs having chronic hepatitis. Zinc is advocated as a treatment for reducing hepatic copper concentrations. Zinc acts by inducing intestinal cell metallothionein (MT) production. Once MT is induced it has a high affinity for copper and prevents serosal transfer of copper into the blood. The intestinal cells turn over rapidly and take the MT-copper complex into the stool where it is excreted, thus resulting a mucosal block of copper uptake. Zinc is recommended as a maintenance therapy for patients with Wilson's disease that have been previously treated with chelators. Since both copper chelators penicillamine (PCA) and trientine have an affinity for zinc, concomitant administration has the theoretical potential of reducing the efficacy of both the chelator and zinc. A subsequent study showed that concomitant administration offers no benefit over single maintenance therapy.

Suggested doses of elemental zinc range from 2 to 3 mg/kg for zinc deficiency and 10 mg/kg given every 12 hours to reduce copper uptake. Zinc is available as an acetate, sulfate, gluconate and methionine. Zinc should be administered on an empty stomach to assure adequate absorption and not be given simultaneously with copper chelators. A common problem encountered with zinc therapy includes nausea, vomiting and anorexia shortly following administration of the drug. It is recommended to measure serum zinc concentrations after several weeks of therapy to assure levels are not in the toxic range. Based on the above studies zinc therapy would appear to be beneficial in the management of dogs with increased hepatic Cu concentrations or zinc deficiency. It is the authors experience however, zinc therapy alone has not been adequate to maintain some affected Bedlington terriers having high copper concentrations but may be of benefit in treatment of other breeds having copper hepatotoxicities, or in young affected Bedlington terriers before their copper concentrations become markedly abnormal.

VITAMIN E

There is evidence that oxidative damage that participates in the pathogenesis of liver injury. It has been shown that free radicals are generated in chronic hepatitis, cirrhosis, Wilson's disease and alcoholic liver disease in humans. The author and others have found dogs with chronic hepatitis to have abnormal levels of markers of hepatic oxidative damage. Increased concentration of malondialdehyde (TBARS) and decreased levels of reduced glutathione (GSH) support the concept of ROS formation in chronic hepatitis.

There is supporting evidence that many antioxidants have been beneficial in managing a number of types of chronic hepatitis in humans. Although most studies are small there is evidence to suggest in some types chronic hepatitis antioxidant or antioxidant ôcocktailö combinations reduce markers of oxidative damage, improve liver enzyme concentrations and improve clinical outcome.

In vitro evidence has demonstrated that vitamin E protects against oxidative damage from iron, copper, and bile acids. We have identified that Bedlington terriers with copper associated chronic hepatitis have oxidant damage associated with the mitochondria and reduced hepatic vitamin E concentrations. Vitamin E (d-alpha tocopherol) is also shown to protect the liver from copper related oxidant damage in vitro. The accumulation of hydrophobic bile acids is implicated in the pathogenesis of cholestatic liver disease. Evidence shows that mitochondrial lipid membrane peroxidation occurs from hydrophobic bile acids. The result is swelling of mitochondria, inhibition of normal respiration and flow of electrons through the respiratory chain and increased mitochondrial generation of hydrogen peroxide. We have also shown that vitamin E reduces or ameliorates hepatic oxidant injury when hepatocytes are exposed to cholestatic concentrations of bile acids.

Based on the above information, vitamin E therapy (given as d-alpha tocopherol) seems reasonable, and is given at a dose of 50 to 400 IU daily. Since bile acids are required for fat-soluble vitamin E absorption and may be reduced in obstructive cholestatic liver disease a water-soluble formulation of vitamin E is suggested for these cases. Although vitamin C also has antioxidant properties it increases the oxidative damage of copper in the liver and should be avoided in dogs with copper hepatotoxicity.

SILYMARIN

Milk thistle has been used for centuries as a natural remedy for diseases of the liver and biliary tract.

Silymarin the active extract consists of flavonolignans that have been reported to work as antioxidants, scavenging free radicals and inhibiting lipid peroxidation. As interest in alternative therapy has emerged in both human and veterinary medicine, so has the use of milk thistle and other natural remedies as adjunctive or principal therapy for patients with liver disease. In a review of several controlled clinical trials of patients with either acute or chronic liver disease, evidence is mounting that shows the benefit of milk thistle. These studies however must be interpreted with care because of variable experimental designs and limited number of cases. In one study of undifferentiated hepatitis of 170 patients, there was no significant change in biochemical parameters, however there was improved survival time and in a second study there was a trend toward histological improvement.

One experimental study showed that dogs pretreated with milk thistle and intoxicated with amanita mushrooms were protected from much of the toxic effects of the mushroom, while all placebo treated dogs died. Obviously further studies of milk thistle, as well as other natural remedies are needed before broad generalizations can be made.

Due to the lack of standardization of milk thistle preparations it is difficult to extrapolate an appropriate dosage. Suggestions have ranged anywhere from 50-250 mg/kg bid.

S-ADENOSYLMETHIONINE

S-Adenosylmethionine (SAMe) [DenosylÖ SD4. Nutramax Laboratories] is a naturally occurring molecule found in all living organisms and is involved in a number of metabolic pathways that appear to be beneficial to the liver as well as other tissues. SAMe is involved in three major biochemical pathways. It is involved in cell replication and protein synthesis, has a modulating influence on inflammation and plays a role as a precursor of antioxidants in the hepatocyte. Research has demonstrated that the exogenous administration of SAMe to have potential beneficial effects. SAMe results in methylation of liver plasma membranes phospholipids resulting in enhanced membrane function (membrane fluidity) and through the sulfation of accumulated endogenous bile acids. SAMe is also involved in the restoration of glutathione (GSH) concentrations. GSH participates in many metabolic processes and plays a critical role in detoxification mechanisms of the cell. Depletion of hepatic GSH can indirectly cause toxic effects in these cells by increasing oxidative stress. Exogenous administered SAM has been shown to increase intracellular GSH.

Certain drugs can behave as hepatotoxins and cause in oxidative damage. A classic example of drug toxicity and free radical formation is acetaminophen (ACA). Normally ACA is conjugated to nontoxic water-soluble compounds. When normal conjugation capacity is exceeded the bioformation by the cytochrome P450 to a toxic adduct occurs. This toxic compounds is deactivated by reduced glutathione (GSH) however when GSH depletion results severe oxidative stress occurs. During this process GSH becomes oxidized glutathione (GSSG) catalyzed by the glutathione peroxidase enzyme system. Evidence of ACA damage can be measured by increased TBARs production that reflects membrane peroxidation and a decreased ratio of GSH/GSSG as the consequence of GSSG accumulation. SAMe is shown to replace GSH stores and prevent oxidative damage in the ACA model.

Studies investigating naturally occurring liver disease in animals are required to determine the benefit of SAMe administration in liver disease. A recommended dose range is 20 mg/kg/day.

L-CARNITINE

There is evidence that L-carnitine supplementation in cats may protect against hepatic lipid accumulation and may be an appropriate adjunct for patients with liver disease. Carnitine is required for transport of long chain fatty acids into the mitochondria for subsequent oxidation to form acetyl-CoA fragments that enter the citric acid cycle for energy production. A deficiency of carnitine may lead to the accumulation of toxic acetyl-CoA metabolites with impaired mitochondrial function. Impaired function of the citrate cycle, fatty acid oxidation, and urea cycle leads to increased ammonia production. It appears that carnitine deficiency could occur in chronic liver disease and that supplementation may help protect against encephalopathy, hypoglycemia, and subcellular damage. Studies have shown that abundant dietary carnitine may protect cats from hepatic lipid accumulation and may be of benefit in hepatic lipidosis.

SUMMARY

There are a number of potential therapies that may be of benefit in the management of patients having chronic hepatitis. Until better classification of disease and controlled therapy trials are undertaken definitive recommendations cannot be made. Consequently, it is important that once treatment protocols is formulated for an individual patient, careful monitoring and repeat liver biopsies become essential to monitor response and adjust therapies.

Selected references are available on request.

KEY WORDS: Chronic hepatitis, Zinc, Colchicine, Vitamin E, S-Adenosylmethionine, Silymarin, L carnitine


DIFFICULT LIVER CASES: SOME NEW IDEAS
David C. Twedt DVM, DACVIM

The following are unusual liver cases that have either a unique clinical, histological or therapeutic implications. Some of the information presented lacks adequate clinical descriptions and some of the information presented here is opinion of the author.

HEPATOCUTANEOUS SYNDROME

Hepatocutaneous syndrome, also known as superficial necrolytic dermatitis or metabolic dermatosis is an uncommon disease observed in middle aged to older dogs. The skin lesions have characteristic histological changes and when combined with the hepatic changes typify this syndrome. The liver has mistakenly been described by some as cirrhosis because of the nodular appearance of the liver. The hepatic changes are best described as an idiopathic hepatocellular collapse. The typical hepatic histology shows hepatocyte collapse with hepatic nodular regeneration containing vacuolated hepatocytes. To date the pathogenesis of the hepatic disease is still controversial. In humans other liver diseases have been noted to produce the similar cutaneous lesions however the hepatocellular collapse described in the canine hepatocutaneous syndrome has not been reported. It is not known if the liver dysfunction is the major mediator of the necrolytic skin lesions or whether another metabolic disease produced both the skin and hepatic lesions.

A common abnormality observed in some affected dogs includes hypoaminoacidemia. It is thought that the necrolytic skin lesions are directly related to the hypoaminoacidemia. The hypoaminoacidemia may also be responsible for the hepatic changes as well. This is supported in part by our observation that dogs feed a protein deficient diet for a prolonged period of time develop hepatic changes with some resemblance to hepatic changes described in the hepatocutaneous syndrome. These protein deficient dogs did not however have the typical skin lesions.

The theory of hypoaminoacidemia in this disease is further supported through work done in our laboratory where we identified that the administration of intravenous amino acid solutions transiently improved the skin and hepatic lesions in most dogs. The cause of the amino acid deficiency is as yet unknown. The affected dogs appear to have been feed adequate protein content diets. The reported prognosis for this disease is grave in the dog and invariably most either succumb to the disease either due to liver dysfunction or from the severity of the skin lesions or both.

Our current therapy includes administration of intravenous amino acid solution. We give approximately 500 milliliters of Aminosyn (10% solution, Abbott) over an 8-12 hour period of time. Repeated infusions are given every 7-10 days. If after four amino acid infusions and there is no improvement it is unlikely the patient will improve. When a positive response is observed we will repeat the amino acid infusions as needed. In addition, we generally treat the patient with a dietary supplement of egg yolk (as an amino acid source), omega 3 fatty acids, vitamin E and zinc.

IDIOPATHIC VACUOLAR HEPATOPATHY

The histological report of vacuolar hepatopathy is often very frustrating and the underlying etiology may be difficult to determine. A vacuolar hepatopathy are basically hepatocytes containing vacuoles in their cytosolic compartment that may contain, fat, glycogen, intracellular water (edema) or other metabolic wastes or intermediates. Vacuolar hepatopathies may also occur in conjunction with hydropic degeneration in which there is cytosolic swelling but is devoid of distinct vacuoles. A number of clinical conditions can cause the above changes. If a secondary non-hepatic disease can be identified that could explain the hepatic changes then it is referred to as a reactive hepatopathy. Such changes can occur secondary to chronic stress (possibly endogenous steroid induced), inflammatory bowel disease, non-hepatic neoplasia, nutritional imbalances and hypothyroidism are but a few examples. Consequently, when a histological diagnosis of vacuolar hepatopathy is made one should investigate for other causes that could explain the reactive changes in the liver.

In some dogs we were unable to identify the cause of the vacuolar changes. These idiopathic vacuolar hepatopathies are frustrating to diagnose. We observe this condition in older dogs, often that are asymptomatic, but have increases in serum alkaline phosphatase (ALP). The typical liver histology report usually reads "diffuse hepatic vacuolar change suggestive of a steroid hepatopathy-check for Cushing's disease." Evaluating the liver with special stains find that the hepatocytes contain excess glycogen. In all intense purposes this group of dogs look like typical steroid hepatopathies based on histology and abnormal serum ALP concentrations. However, further investigation finds no clinical or laboratory evidence of Cushing's disease. Often the only laboratory abnormality that initiates liver investigation is the increase in serum ALP. Most cases have no clinical signs. Conventional adrenal testing (i.e. ACTH stimulation or LDDS) is normal. If one measures the glucocorticoid isoenzyme of alkaline phosphatase (G-ALP) will find that ALP is predominately G-ALP. Ultrasound imaging of the liver shows increased echogenicity and frequently, but not always, the adrenal glands appear normal. The clinical course in these dogs is usually unremarkable however we have observed a very small number of dogs develop typical Cushing's disease at a later date.

We have recently investigated a number of dogs having an idiopathic vacuolar hepatopathy. They have increased serum ALP concentrations but do not have Cushing's disease as we know it to be today. We speculate that this syndrome to be a type of abnormal steroid hepatopathy. Hepatic glycogen accumulation is generally a steroid associated event. Our speculation into this syndrome is supported by the fact that affected dogs given empirical ketoconazole or lysodren therapy has decreases in ALP concentrations and hepatic glycogen accumulation. Because adrenal nodular hyperplasia is a common finding in older dogs we question if it is possible that some of these dogs may have functional hyperplastic nodules producing some type of adrenal steroid. In typical Cushing's evaluation we measure only the corticosteroid cortisol. It is possible that abnormal concentrations of some of the "other" adrenal steroids (possibly one of the sex hormones such as progesterone, estradiol, DHEAS-S or 17a-Hydroxy-progesterone) may be responsible for the hepatic changes. We have identified a number of dogs having idiopathic vacuolar hepatopathy to have abnormal concentrations in one or more adrenal steroids following ACTH stimulation.

The clinical course of this syndrome needs further investigation. It appears most dogs do not progress in their disease syndrome and live a normal life with out adverse consequences from their liver changes. We have observed one dog later develop typical Cushing's disease. A question that needs answering is should these dogs be treated with lysodren or ketoconazole. At this time I have not been recommending therapy for asymptomatic dogs.

HEPATIC NODULAR HYPERPLASIA

Hepatic nodular hyperplasia (HNH) is a common benign lesion observed in livers of older dogs. It is characterized by a discrete accumulation of hyperplastic hepatocytes arising as either macroscopic or microscopic hepatic nodules. HNH does not appear to cause clinical signs of illness. The diagnosis of HNH is usually made as an incidental finding at necropsy or during a diagnostic work up for other concurrent medical problems. These lesions may result in laboratory, ultrasonographic, gross and microscopic hepatic changes which could lead to diagnostic errors in evaluating patients with diseases not related to the liver. The etiology is unknown although nutritional-metabolic factors may play a role.

In the dog, the lesion is common and appears to be age-related. Nodules may be present by 6 or 8 years of age and, in one study, were present in all dogs older than 14 years. We found in 57 dogs the mean age of diagnosis to be 11 years. Affected dogs with HNH generally have an increased serum alkaline phosphatase (ALP) above the normal reference range. There are generally no changes in mean total bilirubin, alanine aminotransferase (ALT), cholesterol, total protein or albumin concentrations. The serum ALP activity in affected dogs we investigated dogs was 2.5 times higher than the upper limits of the reference range. Several dogs with HNH had ALP concentrations as high as 10 to 14 times normal. Bile acid concentrations were determined in a few cases and were normal. The most likely explanations for the raised serum ALP activity is increased production secondary to focal intrahepatic cholestasis within the hyperplastic nodules or from the direct mechanical compression by the nodules on surrounding hepatic parenchyma. Occasionally a rise in the serum ALT activity was noted and may be the consequence of hepatocellular drop out due to altered microcirculation. It has also recently been shown that hepatocellular proliferation can result in a rise in the plasma aminotransferase activity; it is possible that the nodular proliferation is directly responsible for the rise.

Ultrasonographic findings are inconsistent due to the varied hepatocellular morphology and size of the nodules. The echogenicity is reported hypoechoic, hyperechoic, or mixed in appearance. The nodules may also have echogenicity identical to normal parenchyma, and thus be overlooked. Because some cases of HNH can present as multifocal larger macroscopic lesions, they may be mistaken for primary or secondary hepatic neoplasia.

HNH can occur as single, multiple or diffuse either macroscopic or microscopic nodules. The histological description of nodular hyperplasia is an expansile process of hepatocytes that compresses existing parenchyma resulting in hepatocellular atrophy and approximation of the reticular fibers. Grossly, their appearance may mimics macronodular cirrhosis if extensive. Microscopically, hepatocytes can develop a variety of cytoplasmic changes including lipidosis, hydropic degeneration, and glycogen accumulation. This may be problematic in needle biopsy specimens since the identification of nodular regeneration is very difficult due to size limitations and the histomorphologic findings can be suggestive of a metabolic disorder.

Clinically old dogs either healthy or sick having increases in biochemical liver enzymes must have nodular hyperplasia included in the differential diagnosis. Because of the varied ultrasonographic appearance and the need for larger wedge biopsy samples to make the diagnosis this common condition in older dogs may be easily misdiagnosed.

MICROVASCULAR PORTAL DYSPLASIA WITH PORTAL HYPERTENSION

Portal hypertension results from a number of etiologies. Portal hypertension is classified as being posthepatic (i.e. cardiac disease), prehepatic (i.e. portal vein obstruction), or hepatic. Hepatic associated portal hypertension is generally associated with inflammatory or fibrotic liver disease. Hepatic cirrhosis is the most often cause of hepatic portal hypertension. The usual result of portal hypertension is ascites and formation of acquired portosystemic shunts. The acquired shunts from portal hypertension are generally multiple extending most often to the left renal area with anastomosis to the caudal vena cava. As the degree of portosystemic shunting progresses hepatic encephalopathy often develops.

In contrast to acquired portosystemic shunts it is generally regarded that dogs having congenital portosystemic vascular anomalies from a single intra or extrahepatic shunting vessel have signs associated with hepatic encephalopathy but do not have portal hypertension and rarely ascites. A separate syndrome referred to as a microvascular portal dysplasia or idiopathic noncirrhotic portal hypertenison has been described. This condition is observed in young dogs. The clinical presentation is similar to dogs having either congenital intra or extrahepatic shunts. Work up on these patients fails to identify a single shunting vessel but rather these cases have marked portal hypertension associated with multiple acquired portosystemic shunts. The dogs often present with ascites and signs of hepatic encephalopathy that are both the result of hepatic portal hypertension. Hepatic biopsy of the liver in these dogs fail to demonstrate significant inflammatory or fibrotic liver disease as one would expected in patients having hepatic portal hypertension. The histological findings in these dogs often resemble the hepatic pathology of dogs having a congenital portosystemic shunts or microvascular portal dysplasia. A similar pathologic pattern is also observed in surgically created portal to vena cava shunts.

We suspect that these dogs have a nonreversible congenital malformation of the small intrahepatic portal vasculature. This severe form of microvascular portal dysplasia has been referred to as portal venous hypoplasia or noncirrhotic portal hypertension. The hepatic histology in these dogs demonstrate portal tracts associated with multiple arterioles, small or absent portal veins with variable fibrosis, lymphatic distention and bile duct proliferation. The pathology however is void of inflammatory infiltrates. There are also increased amounts of hepatic iron deposited in the liver. In some cases there is increased fibrosis around the portal triads and in a few cases bile duct hyperplasia. Some authors believe that the fibrosis can become extensive in some cases. Possibly dogs having what is referred to as congenital hepatic fibrosis may actually be a progression of the microvascular dysplasia syndrome or may be a separate syndrome.

The clinical manifestation of this syndrome is similar to that of congenital portosystemic shunt patients. Most dogs are under 2.5 years of age and there is no breed prevalence however Doberman Pinschers, Cocker Spaniels and Rottweilers may be over represented. The liver enzymes may be increased with a hypoalbuminemia and high bile acids concentrations. Ultrasound is often helpful showing microhepatia, hepatofugal portal blood flow and multiple abnormal extrahepatic veins. Portal contrast studies demonstrate acquired portal shunts and pressure measurements document portal hypertension. The prognosis for this condition is guarded but some dogs are reported to have a prolonged survival. Authors have reported treating some dogs with antifibrotic agents. The success of such therapy is unproven.

SELECTED REFERENCES
  1. Meyer D J: Hepatic pathology. In Guilford, Center, Strombeck, et al., ed.: Strombeck's Small Animal Gastroenterology. Philadelphia: W B Saunders Co., 1996, p 649.
  2. Gross T L et al: Superficial necrolytic dermatitis in dogs. Vet Pathol. 1993; 30:75-81.
  3. Rosychuk R A W: Superficial necrolytic dermatitis (Hepatocutaneous Syndrome) Proc 18th ACVIM Forum, Seattle, WA 2000. 651-653.
  4. Prause L C and Twedt D C: Hepatic Nodular Hyperplasia. In: Bonagura J D ed. Kirk's Current Veterinary Therapy XII, WB Saunders, Philadelphia, 2000; 675-676
  5. Schmeitzel L P, Lothrop C D, Rosenkrantz W S: Congenital adrenal hperplasia-likesyndrome. . In: Bonagura J D ed. Kirk's Current Veterinary Therapy XII, W B Saunders, Philadelphia, 1995; 600-604.
  6. Bunch S E, Johnson S E, Cullen J M: Idiopathic noncirrhotic portal hypertension in dogs: 33 cases (1982-1998). JAVMA , 2001; 218(3): 392-399.
  7. Van den Ingh T S, Rothuizen J, Meyer H P: Portal hypertension associated with primary hypoplasia of the hepatic portal vein in the dogs. Vet Rec. 1995; 137:424-427.

Key Words: Hepatocutaneous syndrome, Superficial necrolytic dermatitis, Hepatic nodular hyperplasia, Idiopathic vacuolar hepatopathy, Microvascular portal dysplasia, Portal hypertension


© 2001 Denny Meyer and David C. Twedt - All rights reserved