Congenital portosystemic shunts are naturally occurring vascular anomalies that allow portal blood to bypass the liver and to enter the systemic circulation directly.1 Extrahepatic CPSSs arise from the main trunk of the portal vein or one of its major tributaries and most frequently empty into the caudal vena cava or azygos vein.2 Intrahepatic CPSSs arise from 1 of the 3 main branches of the portal vein and enter the caudal vena cava directly or via one of the hepatic veins.3 Congenital portosystemic shunts in dogs are often associated with clinical signs of hepatic encephalopathy and clinicopathologic findings indicative of decreased liver function, including hyperammonemia, increased serum bile acids concentrations, decreased SUN, hypoalbuminemia, prolonged PTTs, and ammonium biurate crystalluria.1,4–8
Definitive treatment for dogs with a CPSS is surgical attenuation of the shunting vessel to redirect portal blood into the liver.9 However, hypoplasia of the intrahepatic portal veins may restrict portal blood flow into the liver, leading to portal hypertension and the development of multiple acquired portosystemic shunts or death.10–14 We have previously used operative mesenteric portovenography as an indicator of intrahepatic portal blood flow to support this idea.15 In that study,15 we found that dogs with a greater degree of opacification of their intrahepatic portal vessels were more likely to tolerate complete surgical attenuation of a CPSS. Moreover, degree of opacification of intrahepatic portal vessels was positively associated with improvements in clinical and serum biochemical indicators of liver function. Histologic examination of hepatic tissues from dogs with a CPSS may provide further information to predict outcome in dogs undergoing surgical attenuation of a CPSS and to identify the mechanisms underlying improvements in liver function following surgical attenuation.
Hepatic histologic features of dogs with a CPSS were reviewed in 2003 by the World Small Animal Veterinary Association Liver Standardization Group and comprise loss of portal vein profiles, increased numbers of arteriolar profiles, hepatocellular atrophy with lipogranulomas, and sometimes periportal sinusoidal dilatation.16 In addition, bile duct proliferation, Kupffer cell hyperplasia, hemosiderosis, and hepatic fibrosis have been reported.5,11,17,18 Baade et al17 considered the histologic examination findings of liver biopsy specimens before and after surgical attenuation of a CPSS. They found that bile duct proliferation, portal fibrosis, and hepatocellular damage appeared to increase with age and that surgical attenuation of a CPSS resulted in a decrease in hepatocyte damage in 44% of dogs. However, they did not investigate the association between these histologic factors and clinical outcomes. Parker et al18 found no relationship between hepatic histopathologic lesions and survival time following surgical attenuation of a CPSS. However, they did not look at other clinical measures of outcome following surgical attenuation.
The purpose of the study reported here was to investigate the relationship between hepatic histopathologic lesions and clinical, serum biochemical, and portovenographic indicators of liver function in a group of dogs undergoing surgical attenuation of a CPSS according to a standardized protocol. We hypothesized that hepatic histologic examination findings prior to surgical attenuation of a CPSS are related to the ability to tolerate complete surgical attenuation and to improvements in liver function following surgical attenuation. In addition, we hypothesized that hepatic histologic features would provide information regarding the mechanisms underlying improvements in liver function following surgical attenuation of a CPSS.
Materials and Methods
Study dogs—Liver biopsy specimens and medical records of dogs undergoing surgical attenuation of a single CPSS at the Royal Veterinary College of the University of London between August 2000 and July 2004 were reviewed. Surgical treatment of dogs with a CPSS followed a previously published protocol,15 which included obtaining a wedge liver biopsy specimen during the procedure for surgical attenuation of a CPSS. Initial surgical attenuation of a CPSS was performed by use of 2–0 ligatures of silk or polypropylene suture material via a midline celiotomy under general anesthesia for intrahepatic and extrahepatic shunts. For intrahepatic shunts, blunt dissection through the liver parenchyma was performed as necessary to access the shunt. Complete surgical attenuation of a CPSS was always attempted, but partial surgical attenuation was performed if temporary complete CPSS occlusion was associated with portal pressures > 18 mm Hg or that doubled from baseline or persistent evidence of portal hypertension on visual inspection of the gastrointestinal tract, including cyanosis, hyperperistalsis, and excessive pulsation of the mesenteric blood vessels.19 In addition, when these findings were variable or considered borderline and temporary complete CPSS occlusion was associated with a decrease in central venous pressure > 1 mm Hg or a decrease in arterial pressure > 5 mm Hg, a partial rather than complete surgical attenuation of a CPSS was performed.19 For dogs undergoing partial portosystemic shunt attenuation at initial surgery, follow-up surgery was recommended 2 to 3 months later to attempt further surgical CPSS attenuation. To this end, an extra ligature with 2–0 polypropylene suture material was tied loosely around the CPSS at initial surgery and then tightened around the CPSS at follow-up surgery.
Information obtained from the hospital records for each study dog regarding initial surgical and follow-up visits included clinical history; results of clinical examination, serum biochemical analysis, and intraoperative mesenteric portovenography; PT and PTT; and the outcome of surgical attenuation of a CPSS. Portovenography was performed before and after temporary complete CPSS occlusion at initial and follow-up surgeries. In addition, for dogs that received complete surgical attenuation of a CPSS and subsequently underwent a neutering procedure at the Royal Veterinary College, portovenography was offered to confirm complete CPSS occlusion. Portovenograms were graded according to the degree of opacification of the intrahepatic portal vessels: grade 1 = an absence of visible intrahepatic portal vasculature, grade 2 = faint opacification of vestigial portal vessels, grade 3 = faint opacification of a few second- or third-generation portal vessels, and grade 4 = good opacification of third- and fourth-generation portal vessels, as in clinically normal (unaffected) dogs.15
Liver necropsy specimens—Liver necropsy specimens from dogs without liver disease were included for histologic examination to act as negative control tissues and to validate the accuracy of the hepatic histologic examination findings of study dogs. Liver necropsy specimens were found by searching the pathological reports archive between 2004 and 2008 for canine liver necropsy specimens that had been reported to be histologically normal by a veterinary pathologist and that had been obtained from dogs with no clinical evidence of liver disease. Only liver necropsy specimens from dogs < 2 years of age were included in the study to approximate the age at surgery of most study dogs with a CPSS. Liver tissue specimens from only 9 dogs undergoing necropsy met all of these criteria.
Histologic examination—Intraoperative liver biopsy specimens from dogs with a CPSS and liver necropsy specimens from dogs without liver disease were fixed in formalin and embedded in paraffin wax. For this study, 3-μm sections of paraffin-embedded liver specimens were cut and stained with H&E, periodic acid–Schiff, and Perls' Prussian blue. Sections from each liver biopsy or necropsy specimen were assigned an arbitrary number so that investigators would be blinded to clinical data on subsequent histologic examination. Histologic examination of liver biopsy and necropsy specimens was performed by a veterinary pathologist (JVH) and specialist human liver pathologist (AW). Findings on each liver biopsy or necropsy specimen were then discussed to achieve a consensus histologic description. Histologic descriptions were subsequently matched to the clinical records for each study dog for statistical analysis.
Histologic descriptions were qualitative and included assessment of the portal veins, hepatic arteries, bile ducts, iron deposits, and steatosis. Portal veins were identified in H&E-stained sections by their thin wall and large size relative to other portal tract structures, flattened endothelial lining, and the frequent presence of RBCs within their lumen. The presence or absence of an identifiable portal vein within the portal tracts was reported. Hepatic arteries within portal tracts were identified in H&E-stained sections by their thick wall containing a circular layer of smooth muscle, endothelial lining, round outline, and variable presence of RBCs within their lumen. Hepatic arteriolar proliferation was identified as clusters of small to medium arterioles within the portal tracts. Some of these clusters appeared to be associated with a small bile duct. Hepatic arteriolar proliferation was graded as mild if 3 to 5 small to medium arterioles were found within a single portal tract, affecting less than half of portal tracts examined; mild to moderate if 3 to 5 small to medium arterioles were found within a single portal tract, affecting over half of the portal tracts examined; and moderate to severe if ≥ 3 small to medium arterioles were found within a single portal tract, affecting almost all portal tracts examined and with ≥ 6 small to medium arterioles noted in at least 1 portal tract. Bile ducts within portal tracts were identified in H&E-stained sections. Primary bile ducts were identified by their paired relationship with the hepatic artery, cuboidal epithelium, well-formed lumen, and central location within the portal tract. Ductular reaction was defined as the presence of irregular bile ducts, most often without a discernible lumen, at the limiting plate (the interface between the portal tract and surrounding hepatocytes). Immunohistochemical analysis for bile duct epithelium (mouse monoclonal cytokeratin 19 antibody [1:100 dilution]a) was used to confirm ductular reaction. Intracellular iron accumulation was reported as present or absent following examination of Perls' Prussian blue–stained sections. Steatosis was assessed by review of H&E-stained sections. Steatosis was characterized by clear cytoplasmic vacuoles within hepatocytes. Steatosis was graded as mild, moderate, or severe when approximately < 33%, 33% to 66%, or > 66% of hepatocytes were affected, respectively. Intermediate grades were assigned when the degree of steatosis varied throughout the section (eg, mild to moderate meant that some areas had mild steatosis, whereas others had moderate steatosis).
Statistical analysis—Descriptive statistics were obtained for all of the histologic examination findings for liver biopsy specimens obtained from dogs with a CPSS during initial surgery (preattenuation), liver biopsy specimens obtained from dogs with a CPSS during follow-up surgery (postattenuation), and liver necropsy specimens from dogs without liver disease. The effect of surgical attenuation of a CPSS on histopathologic lesions was assessed with paired comparisons between pre- and postattenuation liver biopsy specimens by use of the McNemar test for 2-category variables and the Wilcoxon signed rank test for other ordinal nonparametric variables. To investigate the relationship between histopathologic lesions prior to surgical attenuation of a CPSS and clinical variables, all histologic abnormalities were decreased to 2 categories (present or absent). This allowed associations between histologic variables and clinical data to be tested by use of the Fisher exact test for 2-category clinical variables, χ2 test for trends for ordinal clinical variables, and Mann-Whitney U test for continuous clinical variables. All statistical analyses were performed by use of statistical software.b Values of P ≤ 0.05 were considered significant.
Results
Liver biopsy specimens—Preattenuation liver biopsy specimens were available from 38 dogs with a CPSS. Postattenuation liver biopsy specimens were obtained from 13 of these dogs a median of 3 months (range, 2 to 13 months) after initial surgical attenuation of a CPSS. Archived liver necropsy specimens from 9 dogs without liver disease were obtained. These 9 dogs had undergone necropsy following euthanasia for reasons other than liver disease, including meningitis, meningoencephalitis, myelomeningitis, nephritis, familial nephropathy, ocular lymphoma, soft tissue sarcoma of the zygoma, and lung lobe torsion. Median age of these dogs at the time of euthanasia and liver necropsy specimen collection was 15 months (range, 3 to 23 months).
Dogs with a CPSS—The 38 dogs with a CPSS included 29 dogs with an extrahepatic CPSS and 9 with an intrahepatic CPSS. Median age of the dogs with a CPSS at the time of initial surgical attenuation was 12.5 months (range, 2 to 91 months). Sixteen were female, and 22 were male. Thirty-four of the 38 (89%) dogs had clinical signs consistent with hepatic encephalopathy before surgical attenuation of a CPSS, including depression, abnormal behaviors, head pressing, circling, ataxia, aimless wandering, insomnia, muscle tremors, seizures, blindness, deafness, and ptyalism. Findings on serum biochemical analysis and PT and PTT values of dogs with a CPSS prior to surgical attenuation were consistent with decreased liver function (Table 1).
Mean ± SD and median baseline serum biochemical and coagulation variables for 38 dogs at the time of initial evaluation for surgical attenuation of a CPSS.
| Variable | No. of dogs | |||
|---|---|---|---|---|
| Albumin concentration (g/L) | 31 | 25.1 ± 3.8 | 26 | 25–39 |
| Urea concentration (mmol/L) | 31 | 2.4 ± 0.7 | 2.5 | 3.0–10.0 |
| Creatinine concentration (mmol/L) | 31 | 53.7 ± 14.1 | 55 | 50–140 |
| Cholesterol concentration (mmol/L) | 31 | 3.3 ± 1.9 | 2.8 | 2.5–8.0 |
| Total bilirubin concentration (mmol/L) | 31 | 5.8 ± 3.5 | 5.6 | 2.5–12.0 |
| Alanine transferase activity (U/L) | 31 | 221.1 ± 268.5 | 133 | 22–220 |
| Alkaline phosphatase activity (U/L) | 31 | 432.9 ± 378.8 | 282 | 30–250 |
| Baseline bile acids (mmol/L) | 26 | 146.8 ± 101.9 | 110 | 0–10 |
| 2-hour postprandial bile acids (mmol/L) | 27 | 305.9 ± 165.4 | 273 | < 30 |
| Ammonia concentration (mmol/L) | 24 | 139.6 ± 80.5 | 121 | 0–70 |
| PT (s) | 22 | 11 ± 5 | 11 | 9 ± 2* |
| PTT (s) | 22 | 19 ± 8 | 19 | 17 ± 4* |
Prothrombin time and PTT for control dogs were measured concurrently with test samples.
At initial surgery, portovenograms performed before and after temporary complete CPSS occlusion had median grades of 2 and 4, respectively. Of the 9 dogs with an intrahepatic CPSS, 7 received partial and 2 received complete surgical attenuation of a CPSS, and of the 29 dogs with an extrahepatic CPSS, 15 (52%) received partial and 14 (48%) received complete surgical attenuation of a CPSS. Four dogs (2 with partial attenuation of an intrahepatic CPSS and 2 with partial attenuation of an extrahepatic CPSS) died within the first 2 weeks after surgery. Three of these dogs had signs of hypovolemic shock associated with melena, hematemesis, and anemia, despite intraoperative portal pressures < 18 mm Hg and an absence of other intraoperative signs suggestive of excessive portal pressures. One dog developed uncontrolled status epilepticus and was euthanized.
Thirty-two of the 38 study dogs were returned to the Royal Veterinary College for a follow-up examination a median of 3 months (range, 1 to 13 months) after initial surgical attenuation of a CPSS. At follow-up examination, clinical improvement was evident in most dogs (P < 0.001); only 6 of 32 (19%) dogs continued to have signs of hepatic encephalopathy. These 6 dogs with continued hepatic encephalopathy included 2 dogs with an intrahepatic CPSS (one underwent complete attenuation and the other partial attenuation) and 4 with an extrahepatic CPSS (one underwent complete attenuation and the other 3 partial attenuation). Follow-up serum biochemical analysis results were available for 17 dogs, and these were indicative of significant improvements in liver function (Table 2).
Mean ± SD (median) serum biochemical variables in 17 dogs for which data were available before and after surgical attenuation of a CPSS.
| Variable | Before surgical attenuation | 3 months after surgical attenuation | P value |
|---|---|---|---|
| Albumin concentration (g/L) | 25 ± 4 (26) | 30 ± 4(29) | 0.003 |
| Urea concentration (mmol/L) | 2.5 ± 0.7(2.7) | 4.1 ± 2.1(4.0) | 0.004 |
| Creatinine concentration (mmol/L) | 50 ± 15(48) | 76 ± 19(68) | 0.001 |
| Cholesterol concentration (mmol/L) | 2.9 ± 1.3(2.6) | 6.0 ± 2.6(6.3) | 0.001 |
| Total bilirubin concentration (mmol/L) | 5.2 ± 3.7(4.5) | 4.4 ± 4.1(3.0) | 0.414 |
| Alanine transferase activity (U/L) | 236 ± 259(150) | 66 ± 49(51) | 0.018 |
| Alkaline phosphatase activity (U/L) | 471 ± 365(435) | 184 ± 162(113) | 0.002 |
| Baseline serum bile acids (mmol/L) | 158 6113 (148) | 42 ± 47(26) | 0.003 |
| 2-hour postprandial serum bile acids (mmol/L) | 328 ± 188(273) | 98 ± 81(78) | 0.001 |
P values are given for paired comparisons of mean values.
Follow-up portovenography was performed in 16 dogs. Two of these dogs had received complete surgical attenuation of a CPSS; complete CPSS occlusion was confirmed by use of portovenography. The remaining 14 dogs received partial surgical attenuation of a CPSS initially, but portovenography failed to show continued blood flow through the CPSS in 6 of these dogs, suggestive of spontaneous CPSS occlusion. Follow-up portovenographic grades were significantly (P = 0.001) higher, compared with those before surgery; median portovenographic grade was 4 before and after temporary complete CPSS occlusion.
Thirteen dogs receiving partial surgical attenuation of a CPSS at initial surgery underwent further surgical attenuation of a CPSS at follow-up surgery, and 11 tolerated complete surgical attenuation of a CPSS without complications. One dog receiving partial surgical attenuation of a CPSS at initial surgery had a lack of blood flow through the CPSS on a follow-up portovenogram; for this dog, the surgeon elected not to perform further CPSS attenuation. The owners of the other surviving 4 dogs receiving partial surgical attenuation of a CPSS at initial surgery declined further surgical attenuation.
Histologic examination—Of the 38 preattenuation liver biopsy specimens, 2 contained < 5 portal tracts on histologic examination and were therefore judged to be inadequate for evaluation of portal tracts. Portal tracts of the remaining 36 preattenuation liver biopsy specimens were characterized by multiple small hepatic arterioles in 25 (69%), inability to identify a portal vein in 13 (36%), and ductular reaction in 5 (14%; Figure 1). Hepatic arteriolar proliferation was graded as mild in 12 preattenuation liver biopsy specimens, mild to moderate in 4, and moderate to severe in 9. Eleven of the 13 liver biopsy specimens without identifiable portal veins had hepatic arteriolar proliferation, compared with 14 of 23 liver biopsy specimens with identifiable portal veins. Ductular reaction was always observed concurrently with arteriolar proliferation. The parenchyma of the 38 preattenuation liver biopsy specimens was characterized by steatosis in 16 (42%) and iron accumulation in Kupffer cells in 32 (84%). Steatosis was graded as mild in 11 preattenuation liver biopsy specimens, mild to moderate in 4, and moderate to severe in 1.
Photomicrographs of sections of liver biopsy specimens obtained from a dog with a CPSS at the time of initial surgery for CPSS attenuation. A—Notice that the portal area has arteriolar proliferation (arrows), ductular reaction (arrowheads), and absence of a prominent portal vein (H&E stain; bar = 100 μm). B—Notice a cluster of small arterioles (arrows), defined as arteriolar proliferation, within a portal tract (H&E stain; bar = 50 μm). C—Results of immunohistochemical staining for cytokeratin 19, featuring bile duct epithelium and thus ductular reaction within the portal areas (bar = 50 μm). D—Results of the Perls' Prussian blue stain, featuring iron accumulation in Kupffer cells within the perivenous zone of the hepatic parenchyma (bar = 100 μm).
Citation: Journal of the American Veterinary Medical Association 239, 5; 10.2460/javma.239.5.638
For the 13 postattenuation liver biopsy specimens, portal veins were not identified in 3, hepatic arteriolar proliferation was present in 7, and ductular reaction was present in 1. Hepatic arteriolar proliferation was graded as mild in 5 postattenuation liver biopsy specimens, mild to moderate in 1, and moderate to severe in 1. Steatosis was present and was graded as mild in 4 of 13 postattenuation liver biopsy specimens. Iron accumulation in Kupffer cells was present in 9 of 13 postattenuation liver biopsy specimens.
For the 9 liver necropsy specimens from dogs without liver disease, the only abnormality reported was the absence of identifiable portal veins in 1. Hepatic arteriolar proliferation, ductular reaction, steatosis, and iron accumulation were not present.
Both pre- and postattenuation liver biopsy specimens were available for 13 dogs: 10 dogs receiving partial attenuation of an extrahepatic CPSS at initial surgery, 1 dog receiving partial attenuation of an intrahepatic CPSS at initial surgery, 1 dog receiving complete attenuation of an intrahepatic CPSS, and 1 dog receiving complete attenuation of an extrahepatic CPSS. Paired comparisons of the results of histologic examination of these liver biopsy specimens were performed and revealed a significant (P = 0.048) decrease in the grade of steatosis following surgical attenuation of a CPSS. There were no significant differences with respect to portal vascular and bile duct findings for the paired comparisons.
Associations between histologic examination findings and clinical findings—Ability to tolerate complete CPSS attenuation at the initial surgery was significantly associated with the type of preattenuation hepatic histopathologic lesion. Eleven of 13 dogs in which a portal vein could not be identified did not tolerate complete surgical attenuation of a CPSS, whereas only 10 of 23 (43%) dogs in which a portal vein could be identified did not tolerate complete surgical attenuation of a CPSS (P = 0.018). In addition, all 5 dogs with a ductular reaction were unable to tolerate complete surgical attenuation of a CPSS. Despite differences in the percentages of dogs with an intrahepatic CPSS and an extrahepatic CPSS that tolerated complete surgical attenuation of a CPSS, there were no significant associations between histopathologic lesions and shunt type.
Portovenographic grade after temporary complete CPSS occlusion at initial surgery was also significantly (P = 0.050) associated with type of preattenuation hepatic histopathologic lesion. Portovenograms were scored as grade 4 for 15 of 23 (65%) dogs with preattenuation liver biopsy specimens with identifiable portal veins; by comparison, portovenograms were scored as grade 4 for only 4 of 13 (31%) dogs with preattenuation liver biopsy specimens without identifiable portal veins.
There was a prolonged PT with an increase in hepatic arteriolar proliferation, but this finding was not significant (P = 0.070); median PT for dogs with no, mild, mild to moderate, and moderate to severe hepatic arteriolar proliferation was 9.5, 10.1, 10.7, and 11.1 seconds, respectively. Four dogs with preattenuation liver biopsy specimens that had a ductular reaction had a median PT and PTT of 11.1 seconds (range, 10.5 to 31.0 seconds) and 23.9 seconds (range, 19.4 to 50.7 seconds), respectively. Median PT and PTT for 18 dogs with preattenuation liver biopsy specimens that did not have a ductular reaction were 10.1 seconds (range, 8.5 to 13.0 seconds) and 17.7 seconds (range, 8.3 to 25.7 seconds), respectively. The difference in PTT between dogs with a ductular reaction and those without was significant (P = 0.010), but the difference in PT between the same groups of dogs was not significant (P = 0.053). Serum alkaline phosphatase activities were significantly (P = 0.031) higher in dogs with a ductular reaction; median serum alkaline phosphatase activities of dogs with a ductular reaction was 711 U/L (range, 265 to 999 U/L), compared with 237 U/L (range, 42 to 1,508 U/L) for dogs without a ductular reaction.
Discussion
Findings in this study confirm and extend previous descriptions of the histologic features of the liver of dogs with a single CPSS and document associations between histologic features and clinical, radiographic, serum biochemical, and hematologic variables. Previously reported characteristic histologic features of dogs with a CPSS include lack of portal veins, increased numbers of hepatic arteries, and hepatocellular atrophy with or without lipogranulomas.16 In addition, periportal sinusoidal dilatation, bile duct proliferation, Kupffer cell hyperplasia, hemosiderosis, and hepatic fibrosis have been variably reported for dogs with a CPSS.5,11,17,18,20 In the present study, hepatic arteriolar proliferation was found to be the most frequently reported feature of the portal tract of dogs with a CPSS, and the degree of hepatic arteriolar proliferation was positively associated with prolongation in PT. Dogs for which portal veins could not be identified histologically in liver biopsy specimens were more likely to have poor opacification of intrahepatic portal vessels on portovenography and were less likely to tolerate complete surgical attenuation of a CPSS, compared with dogs in which portal veins could be identified. Histologic identification of a ductular reaction in liver biopsy specimens of dogs with a CPSS always occurred in combination with hepatic arteriolar proliferation and was associated with higher serum alkaline phosphatase activities and longer clotting times, compared with those dogs without a ductular reaction. Moreover, no dog with a ductular reaction tolerated complete attenuation of a CPSS at initial surgery. Attenuation of a CPSS resulted in significant clinical and serum biochemical changes indicative of improvements in liver function that were associated with a more subtle decrease in hepatocyte steatosis a median of 3 months after surgery.
The association between portosystemic shunting and hepatic arteriolar proliferation is believed to be due to a chronic hepatic arterial buffer response, which attempts to maintain total liver blood flow. The mechanism for this proliferation in dogs remains unknown. It has been proposed that the acute increase in hepatic artery blood flow that develops secondary to an acute decrease in portal vein blood flow is due to accumulation of the vasodilator adenosine within the space of Mall, which surrounds the hepatic arteries and portal veins.21 Hepatic arteriolar proliferation in dogs with a CPSS may be due to chronic hypoxia resulting from continued failure of hepatic artery blood flow to meet the oxygen demands of the liver or a chronic insufficiency of gastrointestinal tract–derived growth factors, hormones, and nutrients. However, a previous study22 found no significant difference in the partial pressure of oxygen in the portal vein of dogs with a CPSS, compared with clinically normal dogs.
Ductular reaction was observed in a subset of dogs with a CPSS and was associated with serum biochemical abnormalities, suggestive of more severe liver dysfunction or bile duct injury than in dogs without a ductular reaction. Ductular reaction occurs in response to various forms of liver injury and has been reported for liver diseases that feature cholestasis or necrosis followed by hepatic regeneration.23 Ductular reaction has been reported for dogs with acute and chronic hepatitis, primary vascular disorders, biliary atresia, extra-hepatic biliary obstruction, inflammatory cholangitis, and cholangiocellular carcinoma.24,25 The stimulus for ductular reaction in dogs with a CPSS cannot be determined from the present study. However, as ductular reaction was always associated with hepatic artery proliferation, it may have been stimulated by a critical level of hepatic ischemia, which was present in only those dogs in which portal blood flow into the liver was most decreased. If this is correct, hepatic arteriolar proliferation in these dogs was unable to fully compensate for a loss of portal blood flow with respect to liver oxygenation. Further studies are required to investigate oxygen delivery to the liver in the presence of portosystemic shunting.
In the present study, hepatic steatosis was a frequent finding in dogs with a CPSS and decreased following surgical CPSS attenuation. Steatosis is a reversible change in response to mild liver injury16 and has been reported previously for dogs with a CPSS.17,18 Steatosis in dogs with a CPSS may result from hepatic hypoxia or a defect in fatty acid metabolism due to failure of transport of gastrointestinal-derived hormones, such as insulin, to the liver. A decrease in the grade of steatosis following successful surgical CPSS attenuation and subsequent redirection of portal blood into the liver would therefore be expected. This finding agrees with results of a previous study,17 which found a decrease in hepatocyte damage in 11 of 25 dogs following surgical attenuation of a CPSS.
Hepatic hemosiderosis was another common finding in dogs with a CPSS of this report. Hepatic hemosiderosis represents an excessive accumulation of hemosiderin (iron [III] oxide-hydroxide) in hepatocytes and Kupffer cells.26 In the presence of portosystemic shunting, liver hypoxia may promote hepatic iron sequestration attributable to the critical role iron plays in oxygen binding and transport.27 This agrees with results of previous studies7,28,29 that investigated iron status in dogs with a CPSS and found hepatic hemosiderosis in association with normal to low serum iron concentrations and normal to low total iron binding capacities. Regardless of the pathogenesis for increased hepatic iron accumulation, there is no effective means for the body to increase iron excretion.26 It is therefore unsurprising that attenuation of CPSS was not associated with a decrease in hepatic hemosiderosis.
This study demonstrates a natural variation in histopathologic lesions among dogs with a CPSS; the abnormal findings of hepatic arteriolar proliferation, apparent lack of identifiable portal veins, ductular reaction, iron accumulation, and steatosis were not present in all liver biopsy specimens or uniform in degree. This variation may partly reflect differences in the shunt fraction (ie, the proportion of total portal blood flow that passes through the shunt). Variable preoperative shunt fraction in dogs with a CPSS has been observed fluoroscopically15 and when rectal scintigraphy is used,30–32 but preoperative shunt fraction consistently exceeds 90% when portal scintigraphy is used.33,34 Any true differences in shunt fraction would presumably depend on the number and caliber of the intrahepatic portal vessels as well as the caliber of the shunting vessel itself. However, we found that surgical attenuation of a CPSS increased portal blood flow into the liver and significantly improved liver function without a significant change in the appearance of the hepatic vasculature on histologic examination. This suggests that variations in hepatic portal blood flow may occur because of physiologic rather than structural differences in intrahepatic hemodynamics. Further investigation of dogs with a CPSS undergoing surgical attenuation may provide more information regarding the long-term effects of changes in portal blood flow on the hepatic arterial buffer response. Moreover, an improved understanding of the mechanisms regulating intrahepatic portal blood flow may allow us to manipulate intrahepatic portal blood flow in dogs with a CPSS to achieve complete surgical attenuation without the complication of portal hypertension.
The results of this study are based on qualitative histologic descriptions compiled by a veterinary pathologist and human liver pathologist. It is unlikely that these results were biased by the fact that the study was assessing dogs with a CPSS because the histologic review of tissues was conducted in a blinded fashion with regard to the identity of the dogs and included 9 liver necropsy specimens from dogs without liver disease, of which only 1 was reported to be abnormal with respect to only 1 variable. Histologic examination findings for the liver necropsy specimens were not compared with those of the preattenuation liver biopsy specimens by use of hypothesis testing statistics because the inclusion of the liver necropsy specimens in this study was for the purpose of validating the accuracy of the histologic examination findings in liver biopsy specimens from dogs with CPSS and because of the small number of archived liver necropsy specimens that met the inclusion criteria for normal tissue.
Unfortunately, the data collected for this study were not sufficient to allow for analysis of the relationship between types of hepatic histopathologic lesions and survival time. Parker et al18 reported that hepatic histologic examination findings of 64 dogs undergoing primarily partial attenuation of a CPSS were not associated with survival time, which somewhat contradicts our findings that histopathologic lesions are related to the ability to tolerate complete surgical attenuation of a CPSS and thus to a better clinical outcome.10,35 However, it is possible that complete surgical attenuation of a CPSS results in clinical improvements in the short term without long-term improvements in survival time. Long-term survival data for dogs treated by use of the protocol reported in the present study, which aims to achieve complete surgical attenuation of a CPSS, are presently being collected. An additional limitation of this study is the absence of obtaining liver biopsy specimens beyond the time of the second surgery. A larger number of CPSS liver biopsy specimens may also have increased the significance of our findings.
For the dogs with a CPSS included in this study, portovenography and liver biopsy were performed to document the changes to the intrahepatic portal vasculature and hepatic histologic examination findings, which occur following surgical attenuation of a CPSS and which may account for the clinical and serum biochemical changes indicative of improvements in liver function. In addition, at initial surgery, portovenography was performed to confirm the diagnosis of a CPSS, CPSS anatomy and morphology, and correct placement of the ligature around the CPSS, and histologic examination was performed to rule out other causes for the clinical signs in dogs with a CPSS, which may have been especially valuable for those dogs that did not respond well to surgical attenuation of a CPSS. Detailed analysis of the portovenograms has been reported previously; portovenography confirmed immediate and long-term increases in intrahepatic portal blood flow following surgical attenuation of a CPSS, which occurred alongside clinical and serum biochemical changes indicative of improvements in liver function.15 The results of the present study do not reveal substantial concurrent improvements in hepatic histopathologic lesions when only routine histologic staining techniques are applied. Ongoing studies are investigating further techniques for examining liver biopsy specimens from dogs with a CPSS.
Findings of the present study indicate that dogs with a CPSS and histopathologic lesions that include ductular reaction and absence of identifiable portal veins are less likely to tolerate complete CPSS attenuation at the initial surgery than are dogs without these histologic features. Attenuation of a CPSS is associated with significant clinical and serum biochemical changes indicative of improvements in liver function and portovenographic evidence of increased intrahepatic portal blood flow over a median of 3 months. However, our observations of 13 dogs (11 receiving partial surgical attenuation of a CPSS and 2 receiving complete surgical attenuation) demonstrated that a decrease in steatosis was the only significant histologic change over the same time frame. Further research is required to increase our understanding of the improvements in liver function and intrahepatic portal blood flow after surgical attenuation of a CPSS to improve outcomes in those dogs for which portal hypertension limits surgical attenuation and thus complete resolution of clinical signs, including hepatic encephalopathy.
ABBREVIATIONS
| CPSS | Congenital portosystemic shunt |
| PT | Prothrombin time |
| PTT | Partial thromboplastin time |
Novocastra Laboratories Ltd, Newcastle upon Tyne, Northumberland, England.
PASW Statistics 18, SPSS Inc, Chicago, Ill.
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