Antiviral activity of clevudine [L-FMAU, (1-(2-fluoro-5-methyl-beta, L-arabinofuranosyl) uracil)] against woodchuck hepatitis virus replication and gene expression in chronically infected woodchucks (Marmota monax).

L: -FMAU [1-(2-fluoro-5-methyl-beta,L-arabinofuranosyl) uracil] has been shown to be an effective inhibitor of hepatitis B virus (HBV) and duck hepatitis B virus replication in cell culture and duck hepatitis B virus replication in acutely infected Peking ducks. The woodchuck hepatitis virus (WHV) and its natural host, the Eastern woodchuck (Marmota monax), have been established as a predictive model for the evaluation of antiviral therapies against chronic HBV infection. In this report, the antiviral activity of l-FMAU against WHV replication in chronically infected woodchucks is described. Four weeks of once-daily oral administration of L-FMAU significantly reduced viremia, antigenemia, intrahepatic WHV replication, and intrahepatic expression of woodchuck hepatitis virus core antigen (WHcAg) in a dose-dependent manner. At the highest dose administered (10 mg/kg/d), significant reductions of intrahepatic WHV RNA and covalently closed circular (ccc)WHV-DNA levels also were observed. The reduction in viremia was remarkably rapid at the higher doses of L-FMAU, with greater than 1,000-fold reductions in WHV-DNA serum levels observed after as little as 2 to 3 days of therapy. Following the withdrawal of therapy, a dose-related delay in viremia rebound was observed. At the highest doses used, viremia remained significantly suppressed in at least one half of the treated animals for 10 to 12 weeks' posttreatment. No evidence of drug-related toxicity was observed in the treated animals. L-FMAU is an exceptionally potent antihepadnaviral agent in vitro and in vivo, and is a suitable candidate for antiviral therapy of chronic HBV infection.

Temporal pathogenesis of experimental neonatal woodchuck hepatitis virus infection: increased initial viral load and decreased severity of acute hepatitis during the development of chronic viral infection.

Acute hepatitis B virus (HBV) infections either resolve or progress to chronicity. Identification of early deviations in host-virus responses associated with these outcomes can further differentiate cause-effect mechanisms that initiate and maintain chronicity. Neonatal woodchucks were infected experimentally with the woodchuck hepatitis virus (WHV) at 3 days of age. At 8 or 14 weeks of age (i.e. , the early- or mid-acute stage of infection), whole blood and large surgical biopsies of the liver were obtained from infected animals and uninfected controls. These were stored for later correlating histopathologic responses and viral load with the subsequently determined outcome of infection. As of 1 year postinfection, half of the surgically treated infected woodchucks had developed self-limited infections, while the other half developed chronic infections. The self-limited outcome was characterized by decreased viral load in acute-phase liver and plasma and a generally robust acute hepatic inflammatory response. Comparisons at the same early time points revealed that the chronic outcome was characterized by increasing initial viral load in liver and plasma, and a detectable, but diminished, acute hepatic inflammation. These cotemporal comparisons indicate that there is an early host-response deviation during the acute phase of a developing chronic infection. Continued analysis of the tissues banked from this study will facilitate further temporal characterization of acute-phase mechanisms that determine resolution versus chronicity in WHV infection. Understanding such mechanisms may be useful in the rational design of therapy for established chronic HBV infection.

Effects of age and viral determinants on chronicity as an outcome of experimental woodchuck hepatitis virus infection.

Acute hepadnavirus infections either resolve or progress to chronicity. Factors that influence chronicity as an outcome of hepatitis B virus (HBV) infection in humans can be studied experimentally in the woodchuck model. Accordingly, several woodchuck hepatitis virus (WHV) inocula were characterized. Representative inocula had high titers of infectious virus (approximately 10(7.7)-10(9.5) woodchuck 50% infectious doses per milliliter [WID(50%)/mL] by subcutaneous inoculation), with 1 WID(50%) ranging between 21 and 357 physical virion particles. WHV7P1 (standard high dose, 5 x 10(6) WID(50%)) produced a 72% chronicity rate (i.e., percent chronic of total infected) in neonatal woodchucks (1-3 days old). Comparable doses of WHV8P1 resulted in a lower chronicity rate in neonates (34% chronic) indicating that it represented a strain different from WHV7P1. Neonatal woodchucks were more susceptible to chronic infection by high doses of WHV7P1 (range, 65%-75% chronic) compared with 8-week-old weanlings (33% chronic) and adult woodchucks (0% chronic; i.e., all resolved). High doses of cloned wild-type viruses also induced high rates of chronicity in neonates (70%-80% chronic). Chronicity rates in neonates were decreased for low doses of WHV7P1 (500 WID(50%), 9% chronic) and for high doses of a precore WHeAg-minus mutant WHV8 clone (17% chronic). Thus, both age and viral determinants can influence chronicity as an outcome of experimental WHV infection. Standardized inocula will enable the study of mechanisms that initiate and maintain chronic hepadnavirus infection and also provide a means for developing WHV carriers for therapeutic studies.

Antiviral activity and toxicity of fialuridine in the woodchuck model of hepatitis B virus infection.

Woodchucks were used to study the antiviral activity and toxicity of fialuridine (FIAU; 1,-2'deoxy-2'fluoro-1-beta-D-arabinofuranosyl-5-iodo-uracil). In an initial experiment, groups of six chronic woodchuck hepatitis virus (WHV) carrier woodchucks received daily doses of FIAU by intraperitoneal injection for 4 weeks. At 0.3 mg/kg/d, the antiviral effect was equivocal, but at 1.5 mg/kg/d, FIAU had significant antiviral activity. No evidence of drug toxicity was observed during the 4-week period of treatment or during posttreatment follow-up. In a second experiment, groups of nine WHV carriers or uninfected woodchucks were given 1.5 mg/kg/d of FIAU orally for 12 weeks, and the results compared with placebo-treated controls. After 4 weeks, the serum WHV-DNA concentration in the FIAU-treated carrier group was two to three logs lower than that in the placebo-treated group. After 12 weeks of FIAU treatment, serum WHV DNA was not detectable by conventional dot-blot analysis, hepatic WHV-DNA replicative intermediates (RI) had decreased 100-fold, and hepatic expression of WHV core antigen was remarkably decreased. No evidence of toxicity was observed after 4 weeks, but, after 6 to 7 weeks, food intake decreased and, after 8 weeks, the mean body weights of woodchucks treated with FIAU were significantly lower than controls. Anorexia, weight loss, muscle wasting, and lethargy became progressively severe, and all FIAU-treated woodchucks died or were euthanized 78 to 111 days after treatment began. Hepatic insufficiency (hyperbilirubinemia, decreased serum fibrinogen, elevated prothrombin time), lactic acidosis, and hepatic steatosis were characteristic findings in the final stages of FIAU toxicity in woodchucks. The syndrome of delayed toxicity in woodchucks was similar to that observed previously in humans treated with FIAU, suggesting that the woodchuck should be valuable in future investigations of the molecular mechanisms of FIAU toxicity in vivo and for preclinical toxicological evaluation of other nucleoside analogs before use in patients.

Pharmacokinetics of 1-(2-fluoro-5-methyl-beta-L-arabinofuranosyl)uracil in woodchucks.

1-(2-Fluoro-5-methyl-beta-L-arabinofuranosyl)uracil (L-FMAU) is a nucleoside analog with potent in vitro activity against hepatitis B virus (HBV) and Epstein-Barr virus. The purpose of this study was to characterize the disposition of L-FMAU following oral and intravenous administration in the woodchuck animal model. The numerous similarities between woodchuck hepatitis virus and HBV infection justify the use of the woodchuck as an animal model for preclinical studies of anti-HBV agents in vivo. Woodchucks were given 25 mg of L-FMAU per kg of body weight intravenously and orally. Concentrations of L-FMAU in urine and plasma were determined by high-performance liquid chromatography. Following intravenous administration of 25 mg of L-FMAU per kg to woodchucks, total clearance was moderate, averaging 0.23 +/- 0.07 liter/h/kg. Renal clearance and nonrenal clearance averaged 0.13 +/- 0.08 and 0.10 +/- 0.06 liter/h/kg, respectively. The steady-state volume of distribution averaged 0.99 +/- 0.17 liter/kg, indicative of intracellular distribution of the nucleoside. The terminal-phase half-life of L-FMAU following intravenous administration averaged 6.2 +/- 2.0 h, and mean residence time averaged 4.5 +/- 0.8 h. Absorption of L-FMAU after oral administration was incomplete, and bioavailability was approximately 20%. Concentrations of L-FMAU in plasma remained above the in vitro 50% effective concentration of 0.026 microg/ml for HBV (C. K. Chu, T. Ma, K. Shanmuganathan, C. Wang, Y. Xiang, S. B. Pai, G.-Q. Yao, J.-P. Sommadossi, and Y.-C. Cheng, Antimicrob. Agents Chemother. 39:979-981, 1995) for 24 h after both intravenous and oral administration of 25 mg of L-FMAU per kg.

Pharmacokinetics of (-)-2'-3'-dideoxy-3'-thiacytidine in woodchucks.

The woodchuck (Marmota monax) has proven to be a suitable animal model for studying hepatitis B virus (HBV) infection owing to similarities in the course of infection between woodchuck hepatitis virus (WHV) in woodchucks and HBV in humans. (-)-beta-L-2',3'-Dideoxy-3'-thiacytidine (3TC; lamivudine) is a nucleoside analog which has demonstrated antiviral activity against HBV as well as human immunodeficiency virus (HIV). The purpose of the present investigation was to characterize the pharmacokinetics of 3TC following intravenous and oral administration of 20 mg of 3TC per kg of body weight to woodchucks. Following intravenous administration, the concentrations of 3TC in plasma declined, with a terminal half-life of 2.84 +/- 0.85 h (mean +/- standard deviation). The systemic clearance and steady-state volume of distribution of 3TC were 0.22 +/- 0.078 liters/h/kg and 0.75 +/- 0.13 liters/kg, respectively. The renal clearance of the nucleoside analog was 0.063 +/- 0.016 liters/h/kg. The oral bioavailability of 3TC ranged from 18 to 54%. Allometric relationships between pharmacokinetic parameters and body weight developed by Hussey et al. (E.K. Hussey, K.H. Donn, M.J. Daniel, S.T. Hall, A.J. Harker, and G.L. Evans, J. Clin. Pharmacol. 34:975-977, 1994) were augmented by including data from woodchucks, monkeys (S.M. Blaney, M.J. Daniel, A.J. Harker, K. Godwin, and F.M. Balis, Antimicrob. Agents Chemother. 39:2779-2782, 1995), and additional data from rats (P. Rajagopalan, L. Moore, C.K. Chu, R.F. Schinazi, and F.D. Boudinot, submitted for publication). Interspecies scaling of the pharmacokinetic parameters of 3TC demonstrated a good correlation between clearance (0.74 . W0.76 [where W is body weight]; r = 0.93; P < 0.025), apparent volume of distribution (1.62 . W0.81; r = 0.98; P < 0.005), and steady-state volume of distribution (1.09 . W0.94; r = 0.99; P < 0.05) and species body weight. The allometric relationships for clearance and volume of distribution at steady state predicted the observed pharmacokinetic parameters in humans quite well; however, the apparent volume of distribution was underestimated in humans. Thus, the pharmacokinetic data obtained with the woodchuck HBV animal model should be useful for designing clinical trials.

Heterogeneity of the woodchuck hepatitis virus genome in a chronically infected woodchuck.

The nucleotide sequence of an isolate of woodchuck hepatitis virus (WHV) from the serum of a woodchuck trapped in New York state (WHVNY) was compared with the sequences of previously published isolates. The nucleotide sequence of WHVNY was closest to that of an isolate originating from New Jersey: the two genomes shared a 15 nucleotide in-frame deletion in the region where the presurface and polymerase genes overlap (nucleotides 3260-3274) and differed by 54 point mutations (1.6% of genome). Amino acid differences ranged from 0.4% in the surface gene to 5.7% in the X gene. Three isolates from woodchucks that originated in Pennsylvania and Maryland did not contain the deletion and differed from WHVNY by 102 to 106 point mutations (3.0% to 3.2% of nucleotides). Amino acid changes ranged from 0.5% in the core gene to 5.7% in the X-gene. Thus, WHVNY differed little from previous isolates. Next, the genomes from 102 independent clones of WHVNY were compared to ascertain the extent of sequence variation among WHV genomes in a chronically infected animal. A total of 98 clones had genomes of unit length while 2 clones had genomes shorter than unit length and 2 clones had genomes longer than unit length. The clones not of unit length possessed deletions or inverted duplications of sequence. The rate of mutation in the viral genes was 2.65 mutations per 10,000 nucleotides in the precore domain, 1.27 per 10,000 in the X-gene, 0.98 per 10,000 in the presurface gene, and 3.77 per 10,000 at the 5' end of the core gene.(ABSTRACT TRUNCATED AT 250 WORDS)

Experimental hepatocellular carcinoma: MR receptor imaging.

Relaxation time measurements and magnetic resonance (MR) imaging were performed in three different animal models of hepatocellular carcinoma (HCC). After intravenous administration of asialoglycoprotein-directed arabinogalactan-stabilized ultrasmall superparamagnetic iron oxide (10 mumol Fe/kg receptor agent), T2 of normal liver decreased from 41.6 msec +/- 1.0 to 19.4 msec +/- 1.7 (P less than .05) in rats. T2 of HCC implanted in normal liver or liver with chronic hepatitis was essentially unchanged. These results were similar to those obtained by administration of a reticuloendothelial cell-directed conventional iron oxide; however, the required dose of receptor agent was lower. MR imaging in a woodchuck model of virally induced HCC confirmed the distribution of the hepatocyte-directed agent to regions of functioning and differentiated hepatocytes but not to malignant tumor tissue. The results suggest that MR receptor imaging may play a role in the differentiation between primary liver tumor and functional liver tissue such as that in normal liver hepatitis or regenerating nodules.

Evidence against a requisite role for defective virus in the establishment of persistent hepadnavirus infections.

The factors involved in the establishment of persistent hepadnavirus infection are poorly understood. Recent findings demonstrate that the sequence of the genome of hepatitis B virus (HBV) is variable in infected individuals and that, in some cases, virus mutants predominate. Our objectives in the present study were to analyze the variability of woodchuck hepatitis virus (WHV) genomes in an infected animal and to determine whether sequence heterogeneity played a critical role in the ability of WHV to induce chronic infection. We cloned and determined the complete nucleotide sequence of three supercoiled genomes from an animal that became infected after inoculation with a standardized WHV serum pool (i.e., the WHV7 virus pool). We found that there were four nucleotide substitutions among the three genome sequences as well as a 73-nucleotide deletion in one of the recombinants. DNA transfection experiments revealed that only one of the three recombinants was capable of independent replication. These data suggest that a significant proportion of replicative templates in woodchucks that are infected with WHV are defective virus genomes. Next, we compared the outcome of acute infection after inoculation with a serum pool containing a uniform population of replication competent virus (i.e., the WHV7R pool) with a serum pool composed of WHV genomes of variable sequence. The WHV7R serum pool originated from a woodchuck that became a chronic carrier after in vivo transfection of the liver with the infectious WHV7 recombinant. Neonatal woodchucks were inoculated with 5 x 10(6) WHV genome equivalents of either the WHV7 pool or the WHV7R pool. All animals in the study became acutely infected with WHV. Of the animals infected with the WHV7 serum pool, 65% became chronic carriers, while 80% of the animals infected with the WHV7R serum pool developed chronic infection. Thus, infection of woodchucks with a serum pool containing defective virus resulted in a rate of chronic WHV infection that was similar to, or even lower than, a rate from a pool containing only wild-type virus. This suggests that the presence of defective virus in the inoculum is not a prerequisite for the establishment of persistent hepadnavirus infections.

Effects of induction versus prevention of hibernation on reproduction in captive male and female woodchucks (Marmota monax).

Reproductive performance and gonadal function were studied in two groups of woodchucks containing 16 females and 8-9 males each. The control group was maintained indoors under conditions that do not induce hibernation, including fluctuating temperature of 6-12 degrees C, ad libitum feed, and exposure to natural and incandescent light. The hibernaculum-housed group was, in addition, provided conditions that do induce hibernation, including temperature reduced to 5.6 +/- 1.7 degrees C, no food, and no light from 19 November to 21 February. Hibernaculum housing caused 67% of males and 94% of females to hibernate with reduced body temperature for significant periods of times. Compared to control conditions, hibernaculum housing caused (p less than 0.05) a delay in testis recrudescence, a reduction in mean peak testis size (1.8 vs. 3.0 cc) and its mean date of occurrence (14 March vs. 15 February), a reduction in mean testosterone concentrations in February, and a reduced incidence of fertile matings (22 vs. 88%). In females, hibernation resulted (p less than 0.05) in a greater loss in body weight and a reduced pregnancy rate (31 vs. 82%), which was related to post-hibernation body weights that, in turn, were related to prehibernation body weights. Among hibernaculum-housed females, the pregnancy rate was lower in smaller females (0%; 2.0 +/- 0.1 kg) than in the larger ones (63%; 2.9 +/- 0.1 kg). The results demonstrate that the experimental imposition of hibernation-inducing conditions for 3 mo prior to the expected breeding season can alter subsequent reproductive performance in laboratory-maintained woodchucks, whereas a reproductive rate of 88% can be obtained in animals prevented from hibernating during their first year in captivity.

Serum bile acid determination for assessment of hepatic injury in the woodchuck.

A direct spectrophotometric assay for determination of the serum bile acid concentration in the woodchuck (Marmota monax) has been validated. The assay relies on the conversion of 3-hydroxy bile acids to 3-oxo bile acids by 3 alpha-hydroxysteroid dehydrogenase with concomitant reduction of NAD+ to NADH. Reduction of NAD+ is coupled via a diaphorase catalyst to the formation of a diformazan dye from nitrotetrazolium blue and the diformazan product is measured spectrophotometrically at 540 nm. Interfering endogenous dehydrogenase activity present in woodchuck sera was inactivated with sodium pyruvate. Mean recovery of seven exogenous bile acids added to woodchuck sera was 102.0 +/- 2.2%. Intra-assay precision was determined with ten replicate samples giving a mean +/- standard error of the mean of 1.94 +/- 0.12 micron/L with a coefficient of variation of 3.9%. The mean serum bile acid concentration determined in 33 clinically healthy animals was 5.52 +/- 0.81 micron/L. The serum bile acid concentration increased following surgical ligation of the bile duct from 3.78 +/- 0.58 micron/L to a maximum value of 148.0 +/- 30.7 micron/L and remained increased for the 42 day study period. In woodchucks treated with carbon tetrachloride, the serum bile acid concentration peaked at 16 hours following treatment at 72.7 +/- 29.3 micron/L, and returned to pretreatment concentration within 6 days. The serum bile acid concentration therefore appears to be a sensitive biochemical test of cholestasis and hepatocellular forms of hepatic injury and of potential value in the clinical assessment of hepatic disease associated with woodchuck hepatitis virus infection.

Reproductive patterns in cyclic and pregnant thyroidectomized mares.

Three Quarter-horse mares were thyroidectomized at about 1.5 years of age. Three similar intact mares served as controls. The study continued through two breeding seasons. The thyroidectomized mares were lethargic, rear limbs were oedematous and hair coats were coarse. They displayed a tranquil oestrous behaviour when exposed to a stallion and were only mildly antagonistic when not in oestrus. Length of oestrous cycles varied but most often they were 19-24 days long. Duration of oestrus (mean +/- s.e.m.) for the control and thyroidectomized mares was 12.9 +/- 2.9 and 11.7 +/- 2.2 days respectively (P greater than 0.05). The peak of LH during oestrus was as high as 60 ng/ml blood serum with no difference between the two groups. Peak progestagen on Day 7 after ovulation for controls was 9.0 +/- 1.6 ng/ml and was not different (P greater than 0.05) from the peak of 6.3 +/- 1.7 ng/ml for thyroidectomized mares on Day 8. Pregnancy was achieved in both groups of mares, including the use of semen from a thyroidectomized stallion. Thyroxine was detectable in one pregnant thyroidectomized mare during the last two-thirds of pregnancy only.

Bile acid concentrations in the diagnosis of hepatobiliary disease in the cat.

The clinical usefulness of measuring serum bile acid concentrations as a diagnostic test for hepatobiliary disease was examined in 80 cats that were suspected of having hepatic disease. Serum values of total bilirubin, alkaline phosphatase (ALP), alanine transaminase (ALT), and aspartate transaminase (AST) also were measured. Fasting serum bile acid values were determined by use of solid-phase radioimmunoassay for total conjugated bile acids or by a direct enzymatic spectrophotometric method. A definitive diagnosis was established by histologic examination of the liver, and on the basis of these findings, cats were assigned to groups (1 to 8, respectively) including: extrahepatic bile duct obstruction, hepatic lipidosis, cirrhosis, intrahepatic cholestasis (cholangiohepatitis, cholangitis), neoplasia, hepatic necrosis, portosystemic vascular anomalies, and miscellaneous. Cats in group 8 had no morphologic evidence of hepatobiliary disease or had hepatic lesions that were mild. Test efficacy of fasting serum bile acids, total bilirubin, ALP, ALT, and AST were expressed by use of 4 indices: sensitivity, specificity, positive predictive value, and negative predictive value. The diagnostic efficacy of fasting serum bile acids was examined alone and in combinations with the other tests. There was wide overlapping of values of fasting serum bile acids, total bilirubin, ALP, ALT, and AST among cats in groups 1 to 7. The specificity of fasting serum bile acids for the diagnosis of hepatic disease exceeded 90% at values greater than or equal to 5 mumol/L and reached 100% at greater than or equal to 15 mumol/L.(ABSTRACT TRUNCATED AT 250 WORDS)

Light microscopic and electron microscopic changes in the livers of cats with extrahepatic bile duct obstruction.

Light microscopic and transmission electron microscopic changes were studied in the livers of 6 cats at 25 to 54 days after their extrahepatic bile ducts were experimentally obstructed. Histologic findings included various degrees of bile duct dilatation, ductular proliferation, and peribiliary fibrosis. Concentric layers of dense, birefringent connective tissue surrounded midsized bile ducts. The degrees of bile duct proliferation, hepatocellular degeneration, and bile retention were similar in each cat, but the amount of periductular connective tissue increased with chronicity of bile duct obstruction. Ultrastructural changes included marked swelling of endoplasmic reticulum and mitochondrial inner compartments, marked dilatation and distention of the canaliculi by bile casts, and numerical reduction of canalicular microvilli. Microvilli were often swollen and blunt. Mitochondria were swollen and long, and the cristae lacked normal distribution and density. Cytoplasmic accumulations of granular, electron-dense material and concentrically laminated arrays of material indicative of bile substances were also observed.

Diagnostic value of serum gamma-glutamyl transferase and alkaline phosphatase activities in hepatobiliary disease in the cat.

The diagnostic value of serum gamma-glutamyl transferase (GGT) activity and serum alkaline phosphatase (ALP) activity in the detection of liver disease in the cat (n = 69) was compared. On the basis of histologic examination of the liver, cats were assigned to 8 groups: group 1--complete extrahepatic bile duct obstruction (n = 5), group 2--cholangiohepatitis-cholangitis syndrome (n = 11), group 3--hepatic lipidosis (n = 15), group 4--neoplasia, including lymphosarcoma and myeloproliferative disease (n = 9), group 5--hepatic necrosis (n = 7), group 6--cirrhosis (n = 3), group 7--portosystemic vascular anomaly (n = 4), and group 8--miscellaneous (n = 15). Cats assigned to group 8 lacked substantial histologic abnormalities of the liver. The mean value +/- SD of GGT in 20 clinically normal cats was 0.44 +/- 0.26 IU/L. The highest GGT activity in clinical patients developed in groups 1, 2, and 6. The highest ALP activity developed in groups 1 to 4. Significant correlations between GGT and ALP activities were detected only in groups 2 (P less than 0.001) and 5 (P less than 0.10). Among 54 cats with hepatic disease, only 11% had both the GGT and ALP activities within the normal ranges. Comparatively, 52% had ALP activities within the normal range, and 17% had GGT activities within the normal range.(ABSTRACT TRUNCATED AT 250 WORDS)

Bile acid concentrations in the diagnosis of hepatobiliary disease in the dog.

The clinical usefulness of measuring serum bile acid concentrations as a diagnostic test for hepatobiliary disease, was examined in 150 dogs that were suspected of having hepatic disease. Serum values of total bilirubin (TB), alkaline phosphatase (ALP), alanine transaminase (ALT), and albumin were also measured. Fasting serum bile acid (FSBA) values were determined, using a solid-phase radioimmunoassay for total conjugated bile acids or a direct enzymatic spectrophotometric method. A definitive diagnosis was established by histologic examination of the liver. On the basis of histologic findings, dogs were assigned to groups (1 to 8, respectively) including: extrahepatic bile duct obstruction, cirrhosis, portal systemic vascular anastomosis (PSVA), hepatic necrosis, intrahepatic cholestasis, steroid hepatopathy, neoplasia, and secondary disease. Dogs in group 8 had no morphologic evidence of hepatobiliary disease or had mild hepatic lesions. Test efficacies of FSBA, TB, ALP, ALT, and albumin were expressed using 4 indices: sensitivity, specificity, and positive-predictive and negative-predictive values. The diagnostic efficacy of FSBA was examined alone and in combinations with the other tests. There was wide overlapping of FSBA values among dogs in groups 1 to 7, and there was wide overlapping of ALT and ALP values among dogs in all groups. The specificity of FSBA for the diagnosis of liver disease exceeded 90% at values greater than or equal to 30 mumol/L and reached 100% at greater than or equal to 50 mumol/L. Individual liver tests with the best sensitivity for each group were:FSBA and ALP for extrahepatic bile duct obstruction; FSBA for cirrhosis and PSVA; ALT for hepatic necrosis; and ALP for intrahepatic cholestasis, steroid hepatopathy, and neoplasia. Combinations of tests with the best sensitivity for each group were: FSBA + ALP for extrahepatic bile duct obstruction; FSBA + ALT for cirrhosis and PSVA; FSBA + ALT and TB + ALT for hepatic necrosis; and FSBA + ALP for intrahepatic cholestasis, steroid hepatopathy, and neoplasia. Individual tests had the best sensitivity.(ABSTRACT TRUNCATED AT 250 WORDS)

Laboratory assessment of hepatic injury in the woodchuck (Marmota monax).

Normal reference values for total serum protein, albumin, cholesterol, alanine aminotransferase (ALT), aspartate aminotransferase (AST), sorbitol dehydrogenase (SDH), gamma glutamyl transferase (GGT), alkaline phosphatase (AP), and total bilirubin were established in 48 clinically healthy woodchucks. To validate the use of these biochemical tests in the woodchuck for assessment of liver injury, carbon tetrachloride was administered to produce hepatocellular necrosis and the common bile duct was surgically occluded to produce cholestasis. Biochemical tests were performed prior to experimental treatment and thereafter in surviving woodchucks for a period of 6 weeks. There were marked increases in the serum activities of AST, ALT, and SDH following carbon tetrachloride administration and all 3 enzymes appeared to be useful markers of acute hepatocellular injury. The predominate biochemical abnormalities in woodchucks with bile duct obstruction were hyperbilirubinemia, hypercholesterolemia and increased serum AP and GGT activities. The increase of GGT occurred earlier following bile duct obstruction and the magnitude of increase was greater than that of AP, suggesting that GGT would be the preferred serum enzyme test in the woodchuck for assessment of cholestatic liver injury.

Evaluation of serum bile acid concentrations for the diagnosis of portosystemic venous anomalies in the dog and cat.

The serum concentration of bile acids was measured in dogs and cats with portosystemic venous anomalies (PSVA). In 14 dogs, the mean serum bile acid concentration after 12 hours of fasting was 61.7 +/- 68.7 mumol/L (normal, 2.3 +/- 0.4 mumol/L (SEM) and when measured 2 hours after a meal in 15 dogs was 229.9 +/- 87.7 mumol/L (normal, 8.3 +/- 2.2 mumol/L). The fasting serum bile acid concentration was within the normal range in 5 of 14 dogs. The postprandial concentration was determined in 3 of the 5 and in each case increased more than tenfold above the fasting value. The mean fasting serum bile acid concentration in 4 cats was 24.4 +/- 10.1 mumol/L (normal, 1.7 +/- 0.3 mumol/L) and in 2 of the cats increased to a mean of 120.6 mumol/L (normal, 8.3 +/- 0.8 mumol/L) 2 hours after feeding. The bile acid values in patients with PSVA were correlated with values for blood ammonia content, sulfobromophthalein (BSP) retention, and results of conventional tests of hepatic function. Bile acid concentrations were more sensitive than abnormalities in serum enzyme activities or BSP retention and equal in sensitivity to the ammonia tolerance test in detecting hepatobiliary insufficiency. Bile acid measurements were accomplished with less inconvenience to the patient and clinician, than tests of BSP excretion or ammonia tolerance. Used in combination with conventional laboratory tests for hepatic disease, pre- and postprandial serum bile acid concentrations appear to be a sensitive and specific indicator of hepatobiliary dysfunction of value in the diagnosis of PSVA in the dog and cat.

Circannual changes in serum testosterone concentrations of adult and yearling woodchucks (Marmota monax).

Testicular volumes and serum testosterone concentrations were determined biweekly in 5 adult and 4 yearling woodchucks maintained indoors from December through August. Food and water were provided ad libitum except for 2 mo beginning at the winter solstice when feeding and lighting (12L:12D) supplemental to available natural light were discontinued. Temperatures fluctuated with outdoor temperatures greater than 4 degrees C. No significant hibernation occurred. Testes in adults were small in December (0.3-1.8 cm3), largest in February and March (3.5-5.6 cm3), and smallest in late June (0.1-0.5 cm3). Testosterone was basal in December (less than 0.6 ng/ml), maximal (3.4-6.6 ng/ml) between early January and late March, and minimal from April through August (less than 0.8 ng/ml). In yearlings, maximum testes volumes (1.6 cm3) and serum testosterone (0.9 +/- 0.2 ng/ml) were less, and occurred later, than in adults. Testosterone levels and testis volumes measured in newly captured woodchucks in March and April and again 2-3 mo later were generally similar to those of their laboratory counterparts. Thus, in woodchucks: annual cycles of testosterone production and of testes recrudescence and regression parallel each other with maxima during the short, late-winter breeding season; those cycles are not altered significantly by the absence of hibernation or the present conditions of captivity; and yearling males apparently are not an important part of the breeding population.