Breed-related expression patterns of Ki67, γH2AX, and p21 during ageing in the canine liver

Abstract: Cellular senescence is a molecular hallmark of ageing that is associated with multiple pathologies, and DNA damage marker γH2AX, together with cell cycle inhibitor p21, have been used as senescence markers in multiple species including dogs. Idiopathic canine chronic hepatitis has recognised breed-related differences in predisposition and prognosis, but reasons behind this are poorly understood. This retrospective study using archived post mortem tissue aimed to provide insight into liver ageing in 51 microscopically normal canine livers across seven breed categories, including those with and without increased risk of chronic hepatitis. Immunohistochemistry was conducted for γH2AX, p21, and cell proliferation marker Ki67, and the mean number of positive hepatocytes per high power field was determined. All three markers were strongly correlated to each other, but no age-dependent expression was seen in the combined study population. Overall expression levels were low in most dogs, with median values representing less than 1.5% of hepatocytes, but this increased to 20–30% in individual dogs at the upper end of the range. Individual breed differences were noted in two breeds that have increased risk of chronic hepatitis, with English Springer Spaniels having lower expression of Ki67 than other dogs, and Labradors having higher expression of Ki67 and γH2AX than other dogs. These results warrant further investigation in these breeds and highlight a need to validate reliable markers of cellular senescence in dogs

Clinical features and long-term follow-up of 70 cases of canine idiopathic eosinophilic lung disease.

BACKGROUND: Canine idiopathic eosinophilic lung disease (ELD) is sparsely documented in the literature. METHODS: Clinical presentation and outcome of dogs diagnosed with ELD (eosinophilic bronchitis or eosinophilic bronchopneumonia) were reviewed. Subgroups were made based on chronicity of clinical signs and findings of thoracic imaging: NCI (no changes in thoracic imaging), BRON (bronchial/peribronchial pattern), INT (bronchointerstitial/interstitial/alveolar). RESULTS: Seventy cases were included. There were more young to adult, crossbreed and female dogs. Compared with the other two groups NCI dogs showed lower bronchoalveolar lavage fluid eosinophilic pleocytosis and absence of circulating eosinophilia, bronchiectasis or death due to respiratory disease. All dogs responded clinically to corticosteroids. Median treatment duration was four months. Remission (no clinical signs after treatment discontinuation for >one month) and long-term remission (>six months) was achieved in 60 per cent, and 51 per cent of patients, respectively. Relapse occurred in 26 per cent of cases after remission but was rare (3 per cent) after long-term remission. The one-year, two-year and four-year survival to death due to respiratory disease was 98 per cent, 97 per cent and 91 per cent, respectively. CONCLUSION: Prognosis and initial clinical response for ELD was generally good although achievement of long-term remission was only seen in 51 per cent of dogs. Different outcomes based on chronicity of signs, corticosteroid dose, thoracic imaging abnormalities and other clinical variables were not appreciated

Hepatic leptospiral infections in dogs without obvious renal involvement.

BACKGROUND: Reports of chronic hepatitis in dogs caused by Leptospira spp. are confined to small case series. Fluorescence in situ hybridization (FISH) allows the identification of spirochetes in liver samples. Consequently, this technique may help elucidate the role of Leptospira spp. in cases of chronic hepatitis. OBJECTIVES: To describe cases of hepatic leptospirosis in dogs diagnosed by FISH and subsequent polymerase chain reaction (PCR) speciation, with the absence of clinically relevant renal involvement. ANIMALS: Ten client-owned dogs. METHODS: Retrospective case series from the University of Cambridge presented between 2013 and 2016 or cases consulted by telephone advice during this time period. Cases were selected based on histopathologically confirmed granulomatous hepatitis and leptospiral organisms identified by FISH and PCR speciation (Leptospira interrogans/kirschneri). RESULTS: All cases had increased liver enzyme activities, and FISH in combination with PCR speciation-confirmed infection with L. interrogans/kirschneri. Four dogs underwent repeat liver biopsy, FISH and PCR speciation 4-15 months after initial presentation and doxycycline treatment with 1 dog undergoing repeat sampling at necropsy. Three dogs that underwent repeat biopsy remained positive for L. interrogans/kirschneri infection. Six dogs were alive at the time of manuscript preparation and 4 dogs were euthanized as a result of progressive liver disease. CONCLUSIONS AND CLINICAL IMPORTANCE: The presence of hepatic leptospiral organisms may be associated with chronic granulomatous hepatitis without clinical evidence of renal involvement. Further studies are necessary to elucidate the etiological role of these organisms in the disease

Artifactually increased serum bicarbonate in a cat with rhabdomyolysis

A 3-year-old male neutered domestic shorthair cat presented with lethargy, hyporexia and pyrexia of unknown origin. Biochemical analysis using a Beckman Coulter AU480 demonstrated marked increases in creatine kinase and aspartate aminotransferase, indicative of severe muscle injury, with concurrent presumptive myoglobinuria on urinalysis. A marked, non-physiological increase in measured bicarbonate and resultant negative anion gap were documented; however, calculated bicarbonate obtained via a point-of-care blood gas analyzer was within normal limits. Laboratory error due to interference by lactate dehydrogenase was suspected and supported by the results of subsequent biochemical testing. Artefactual increases in bicarbonate have been documented in cases of rhabdomyolysis in horses, cows, and a bird. However, to the best of the authors’ knowledge, this is the first report to demonstrate this spurious change in a cat

Breed-related expression patterns of Ki67, γH2AX, and p21 during ageing in the canine liver

Abstract: Cellular senescence is a molecular hallmark of ageing that is associated with multiple pathologies, and DNA damage marker γH2AX, together with cell cycle inhibitor p21, have been used as senescence markers in multiple species including dogs. Idiopathic canine chronic hepatitis has recognised breed-related differences in predisposition and prognosis, but reasons behind this are poorly understood. This retrospective study using archived post mortem tissue aimed to provide insight into liver ageing in 51 microscopically normal canine livers across seven breed categories, including those with and without increased risk of chronic hepatitis. Immunohistochemistry was conducted for γH2AX, p21, and cell proliferation marker Ki67, and the mean number of positive hepatocytes per high power field was determined. All three markers were strongly correlated to each other, but no age-dependent expression was seen in the combined study population. Overall expression levels were low in most dogs, with median values representing less than 1.5% of hepatocytes, but this increased to 20–30% in individual dogs at the upper end of the range. Individual breed differences were noted in two breeds that have increased risk of chronic hepatitis, with English Springer Spaniels having lower expression of Ki67 than other dogs, and Labradors having higher expression of Ki67 and γH2AX than other dogs. These results warrant further investigation in these breeds and highlight a need to validate reliable markers of cellular senescence in dogs

Artifactually increased serum bicarbonate in a cat with rhabdomyolysis

Publication status: PublishedA 3‐year‐old male neutered domestic shorthair cat presented with lethargy, hyporexia, and pyrexia of unknown origin. Biochemical analysis using a Beckman Coulter AU480 demonstrated marked increases in creatine kinase and aspartate aminotransferase, indicative of severe muscle injury, with concurrent presumptive myoglobinuria on urinalysis. A marked, non‐physiologic increase in measured bicarbonate and resultant negative anion gap was documented; however, calculated bicarbonate obtained via a point‐of‐care blood gas analyzer was within normal limits. Laboratory error due to interference by lactate dehydrogenase was suspected and supported by the results of subsequent biochemical testing. Artifactual increases in bicarbonate have been documented in cases of rhabdomyolysis in horses, cows, and a bird. However, to the best of the authors' knowledge, this is the first report to demonstrate this spurious change in a cat

Hepatocyte expression and prognostic importance of senescence marker p21 in liver histopathology samples from dogs with chronic hepatitis.

BACKGROUND: Chronic hepatitis (CH) occurs commonly in dogs but is associated with a variable and largely unpredictable prognosis. p21, a cell-cycle inhibitor and marker of cellular senescence, is upregulated in human liver disease and is a better prognostic marker than histological or clinical scoring systems. OBJECTIVE: To quantify hepatocyte p21 immunopositivity in histopathology samples from dogs with CH and determine its association with outcome. ANIMALS: Twenty-six client-owned dogs with histologically confirmed CH, and 15 dogs with normal liver histology. METHODS: Medical records and liver histopathology samples were retrospectively reviewed to identify cases of CH. Immunohistochemistry for p21 was performed on all samples and hepatocyte immunopositivity was visually quantified. Relationships between p21 and dog age and dog survival time were statistically evaluated. RESULTS: Hepatocyte p21 immunopositivity in dogs with CH was high (median percentage of positive hepatocytes: 90%, range: 20%-98%) and exceeded 70% in 23/26 cases with no association with age. In control dogs, p21 immunopositivity was low (≤15% positive hepatocytes in 12/15 cases) and was positively correlated with age (rs  = 0.63; P = .011). Dogs with p21 immunopositivity exceeding 91.8% (upper tercile) had significantly shorter survival compared to dogs with less than 88.9% immunopositivity (lowest tercile; 218 versus 874 days, P = .006). Increasing hepatocyte p21 immunopositivity was significantly negatively associated with survival time (HR 4.12; 95% CI 1.34-12.63; P = .013). CONCLUSIONS AND CLINICAL IMPORTANCE: Marked p21 immunopositivity in dogs with CH might be indicative of widespread hepatocellular senescence. A significant association with survival time also suggests a potential value for p21 quantification in determining prognosis.Petplan Charitable Trust (Grant no 217-263)