Feline thymidine kinase 1: molecular characterization and evaluation of its serum form as a diagnostic biomarker.

BACKGROUND: Thymidine kinase 1 (TK1) catalyzes the initial phosphorylation of thymidine in the salvage pathway synthesis of dTTP, an essential building block of DNA. TK1 is a cytosolic enzyme with its highest level during the S-phase of the cell cycle. In cancer cells TK1 is upregulated and excess TK1 is leaked into the blood. Therefore, serum TK1 has been used as biomarker for cancer diagnosis and prognosis in human medicine. Feline TK1 shows high sequence similarity to TK1 from other species. The aim of this study was to characterize feline TK1 and evaluate if serum TK1 can be used as a diagnostic biomarker. RESULTS: Feline TK1 was cloned, expressed and affinity purified. The purified feline TK1 phosphorylated not only pyrimidine deoxyribonucleosides but also pyrimidine ribonucleosides and to some extent purine deoxynucleosides. A number of anticancer and antiviral nucleoside analogs also served as substrates with fairly high efficiency. ATP and dATP were the preferred phosphate donor. Serum TK1 activity in felines with malignant diseases was significantly higher than that in healthy individuals. ROC analysis revealed an area under the curve (AUC) of 0.98 with a sensitivity of 0.83 and a specificity of 0.95 for felines with lymphoma. Serum TK1 activity in felines with IBD or inflammatory disease was within the same range as healthy ones. Furthermore, in felines with lymphoma serum TK1 activity returned to normal levels in response to treatment. CONCLUSION: Feline TK1 has high specific activity and a broader substrate specificity in comparison with TK1 from other species. Serum TK1 activity in felines with malignant diseases is significantly higher than that in normal felines and in felines with inflammatory diseases. These results suggest that serum TK1 may be a promising biomarker for the diagnosis and monitoring of malignant diseases and for the differential diagnosis of certain inflammatory disease.

[Diagnostic utility of creatine kinase elevation in the routine biochemical profile in cats]. / Diagnostischer Nutzen einer Kreatin­kinase Erhöhung im Routine­chemogramm bei Katzen.

INTRODUCTION: Creatine kinase (CK) is a muscle enzyme that is very sensitive to muscle damage. Therefore, serum CK is measured particularly to confirm suspected myopathy. Since 2013, this enzyme has been included in the routine chemistry profile in our hospital. Soon thereafter, the subjective impression developed that its elevation did not correlate to and was not explainable with the actual clinical problem. Therefore, the aim of this retrospective study was to investigate in which clinical cases the CK elevation was adequate and in which cases without clinical evidence of muscle damage the CK was so markedly elevated that it implied a clinically relevant muscle damage. For this purpose, we evaluated the CK values of 1641 cats presented in the years 2013/2014 at our university animal hospital. The CK was comprehensibly elevated in cats with trauma and various diseases with obvious and traceable muscle damage like thrombo-embolic damage or seizures. In addition, the CK was elevated in diseases where concomitant muscle damage is perceivable like in cats with hypertrophic cardiomyopathy. However, the CK also was commonly and sometimes dramatically elevated in cats of essentially any disease group without any comprehensible skeletal muscular lesion. These results confirm the hypothesis that the diagnostic value of this parameter is most questionable. A CK elevation does not allow any conclusion regarding its original diagnostic purpose, i.e. to confirm the presence of a clinically relevant myopathy.

INTRODUCTION: La créatine kinase (CK) est une enzyme musculaire très sensible lors de dommages musculaires. Par conséquent, la CK sérique est mesurée en particulier pour confirmer une myopathie suspectée. Depuis 2013, cette enzyme fait partie du chimiogramme de routine de notre hôpital. Après peu de temps, l'impression subjective s'est développée que son élévation n'était pas corrélée ni explicable avec le problème clinique réel. Par conséquent, le but de cette étude rétrospective était d'étudier dans quels cas cliniques l'élévation de la CK était adéquate et dans quels cas sans signe clinique de lésion musculaire, la CK était si nettement élevée qu'elle impliquait une lésion musculaire cliniquement pertinente. À cette fin, nous avons évalué les valeurs CK de 1641 chats présentés dans les années 2013/2014 à notre hôpital universitaire pour animaux. La CK était sensiblement élevée chez les chats souffrant de traumatismes et de diverses affections avec des lésions musculaires évidentes et traçables comme des dommages thrombo-emboliques ou des convulsions. De plus, la CK était élevée dans les maladies où des lésions musculaires concomitantes étaient décelables comme chez les chats atteints de cardiomyopathie hypertrophique. Cependant la CK était également couramment et parfois considérablement élevée chez les chats de pratiquement n'importe quel groupe de pathologies sans aucune lésion musculaire squelettique explicable. Ces résultats confirment l'hypothèse que la valeur diagnostique de ce paramètre est très discutable. Une élévation de la CK ne permet aucune conclusion concernant son objectif diagnostique d'origine, c'est-à-dire de confirmer la présence d'une myopathie cliniquement significative.

Retrospective study of the diagnostic utility of Spec fPLin the assessment of 274 sick cats.

BACKGROUND: Serum feline pancreatic lipase immunoreactivity (fPL) commonly is used in the assessment of sick cats suspected to have pancreatitis but its diagnostic utility is debated. OBJECTIVES: To evaluate the diagnostic utility of the Spec fPL test and selected serum biochemistry tests in the diagnosis of pancreatitis in cats. ANIMALS: Two hundred seventy-four client-owned cats presented to a university teaching hospital in the United Kingdom, from April 2013 to May 2017, in which Spec fPL was measured. METHODS: Cats were classified into 1 of 4 groups based on clinical signs (all cats), ultrasonographic findings (all cats) and histopathological or cytological assessment of the pancreas where available (9 cats) regardless of Spec fPL concentration. The groups were (a) definite pancreatitis (n = 9), (b) probable pancreatitis (n = 49), (c) possible pancreatitis (n = 139), and (d) unlikely pancreatitis (n = 77). Spec fPL and selected serum biochemistry test results were compared among groups. RESULTS: Serum fPL concentrations >5.3 µg/L were classified as positive and concentrations <3.5 µg/L were classified as negative. There was a significantly (P = .03) lower proportion of false-positive results (cats unlikely to have pancreatitis, n = 77, with a positive fPL, n = 8, 10%) than false-negative results (cats with definite or probable pancreatitis, n = 58, with a negative fPL result, n = 14, 24%). None of the selected biochemical tests were helpful diagnostically. CONCLUSION AND CLINICAL IMPORTANCE: A positive Spec fPL result indicates that pancreatitis is a probable diagnosis, but the test cannot be used to rule the diagnosis out.

Antioxidant defence and oxidative stress markers in cats with asymptomatic and symptomatic hypertrophic cardiomyopathy: a pilot study.

BACKGROUND: Hypertrophic cardiomyopathy is the most common cardiovascular cause of death in cats. Although the majority of cats remain asymptomatic, some may develop signs of chronic heart failure due to diastolic failure, arterial thromboembolism (ATE) or sudden cardiac death. Therefore, it is crucial to identify individuals that are in high risk of developing cardiac complications before the onset of life-threatening signs. Oxidative stress is the imbalance between the production and neutralisation of reactive oxygen species. Uncontrolled reactive oxygen species overproduction leads to protein and lipid peroxidation and damages the DNA strands, injuring the cells and leading to their death. The aim of the study was to evaluate the oxidative state in cats with hypertrophic cardiomyopathy and healthy controls. RESULTS: In total, 30 cats divided into three groups were assessed: animals with clinically evident hypertrophic cardiomyopathy (HCM; n = 8), subclinical hypertrophic cardiomyopathy (SUB-HCM; n = 11) and healthy controls (n = 11). The activity of superoxide dismutase was statistically significantly lower in animals with symptomatic and asymptomatic hypertrophic cardiomyopathy (HCM 0.99 ± 0.35 U/mL; SUB-HCM 1.39 ± 0.4 U/mL) compared to healthy cats (2.07 ± 0.76 U/mL, p < 0.01). The activity of catalase was significantly lower in the SUB-HCM group (19.4 ± 4.2 nmol/min/mL) compared to the HCM (23.6 ± 5.9 nmol/min/mL) and the control (30 ± 7.5 nmol/min/mL, p < 0.01) group. The activity of glutathione peroxidase was 4196 ± 353 nmol/min/mL in the HCM group, 4331 ± 451 nmol/min/mL in the SUB-HCM group and 4037 ± 341 nmol/min/mL in the control group and did not differ significantly between groups. The total antioxidant capacity of plasma was 602 ± 65.5 copper reducing equivalents (CRE) in the HCM group, 605.9 ± 39.9 CRE in the SUB-HCM group and 629 ± 77.5 CRE in the healthy cats and did not differ significantly between the groups. CONCLUSIONS: Activities of superoxide dismutase and catalase differed in cats with hypertrophic cardiomyopathy, however the activity of the latter was only significantly lower in asymptomatic stage of the disease. The potentially beneficial effect of antioxidative substances on the disease progression in the asymptomatic and symptomatic stage of this disease should also be examined.

Histopathology and Feline Pancreatic Lipase Immunoreactivity in Inflammatory, Hyperplastic and Neoplastic Pancreatic Diseases in Cats.

The most common pancreatic diseases in cats are pancreatitis and exocrine pancreatic insufficiency (EPI). Non-invasive methods, such as serological quantification of feline pancreatic lipase immunoreactivity (fPLI), are often used in the diagnosis of pancreatitis. Previous studies have compared fPLI concentrations with histopathology, considered to be the gold standard for diagnosis of feline pancreatitis. However, fPLI concentrations in cats suffering from pancreatic tumours were rarely described. The aim of the present study was to determine the sensitivity and specificity of an in-house enzyme-linked immunosorbent assay (ELISA) for the quantification of fPLI in serum samples based on histopathological findings in cats diagnosed with various pancreatic diseases. Pancreatic biopsy samples from 80 cats were included. Five groups were defined on the basis of pancreatic histopathology: group 1, normal pancreas; group 2, nodular hyperplasia; group 3, mild pancreatitis; group 4, marked (moderate/severe) pancreatitis; and group 5, pancreatic neoplasia. Serum samples from all cats were tested by fPLI ELISA (<3.6 µg/l normal, 3.6-5.3 µg/l questionable, >5.3 µg/l pancreatitis). In group 1 (n = 19), serum fPLI values were within the reference interval in 74% of cases and in group 2 (n = 9) in 78%. Cats with mild pancreatitis (n = 23), marked pancreatitis (n = 11) and pancreatic neoplasms (n = 18) had significantly increased fPLI concentrations compared with group 1 (P = 0.004/0.001/≤0.0001). Cats with nodular hyperplasia had significantly lower fPLI values than cats with marked pancreatitis (P = 0.048) or tumours (P = 0.002). Serum fPLI concentrations in group 3 were <3.6 µg/l (n = 6), 3.6-5.3 µg/l (n = 4) and >5.3 µg/l (n = 13). Calculated test sensitivity for mild pancreatitis was fPLI >3.5 µg/l: 73.9% and fPLI >5.3 µg/l: 56.5%. In group 4 (n = 11), seven of nine cats (77.8%) with marked purulent pancreatitis had elevated fPLI. In group 4, a sensitivity of 81.8% was detected for fPLI >3.5 µg/l and 63.6% for fPLI >5.3 µg/l. Two cats with marked non-purulent pancreatitis had elevated fPLI, while two cats with marked purulent pancreatitis had normal fPLI values (<3.6 µg/l). In group 5, one cat with pancreatic adenoma and one with pancreatic acinar carcinoma had normal fPLI concentrations. The other cats with pancreatic adenoma (solid, n = 1; cystic, n = 4) or carcinoma (solid, n = 9; cystic, n = 2) had elevated or high fPLI values (4.1 to >40 µg/l, median 21.2 µg/l), probably caused by additional inflammation. The results of the present study confirm the importance of detailed histopathological characterization for the interpretation of clinical signs and fPLI values in feline pancreatitis. Primary pancreatic neoplasms may also lead to elevated fPLI concentrations as there is concurrent pancreatitis in most cases. However, severe pancreatic diseases, such as chronic non-purulent pancreatitis or tumours without inflammation, may result in normal fPLI values.

Alpha-enolase staining patterns in the renal tissues of cats with and without chronic kidney disease.

Renal α-enolase has variable expression in inflammatory and neoplastic diseases. Therefore, in order to define the distribution of α-enolase in renal tissues of cats, an immunohistochemistry assay was validated and described here. Tissues from 29 cats with IRIS Stage 2-4 CKD, 8 control cats < 2 years of age, and 4 control cats> 10 years of age were assessed. Interstitial nephritis was the predominant histopathological finding in the CKD group. The control cats < 2 years of age had moderate α-enolase immunoreactivity in tubular epithelium but staining was absent to mild in glomeruli. In contrast, α-enolase was moderate to high in tubular epithelium and glomeruli in control cats > 10 years of age. In cats with CKD, α-enolase was decreased in tubules that were degenerative or atrophic, similar to normal tubules in control groups, and moderate to high in glomeruli. When compared between the study groups, the results suggest that alpha-enolase decreases in damaged tubules and increases in the glomeruli of older cats prior to the development of detectable CKD. Further studies will be required to determine whether these findings relate to the pathogenesis or could be used in the diagnosis of feline CKD.

Significance of sphingosine kinase 1 expression in feline mammary tumors.

BACKGROUND: Sphingosine kinase 1 (SPHK1) is an enzyme that converts pro-apoptotic ceramide and sphingosine into anti-apoptotic sphingosine-1-phosphate. There is growing evidence that SPHK1 activation promotes oncogenic transformation, tumor growth, chemotherapy resistance, and metastatic spread. High SPHK1 expression has been associated with a poor prognosis in several human cancers. RESULTS: In the present study, the expression level of SPHK1 was examined in feline mammary tumor (FMT) specimens, and the IHC expression level of SPHK1 was associated with the histological grade of FMTs. IHC analysis of 88 FMT cases revealed that the expression level of SPHK1 was upregulated in 53 tumor tissues (60.2%) compared to adjacent mammary tissues. SPHK1 expression in FMTs was significantly associated with histological grade, presence of lymphovascular invasion, and estrogen receptor negativity. Treatment of primary FMT cells with SPHK1 inhibitors reduced cell viability, indicating that SPHK1 acts to promote FMT cell survival. These results indicate that SPHK1 may play an important role in FMTs and may be a therapeutic target in cats with FMT. CONCLUSIONS: SPHK1 over-expression in breast cancer tissues is associated with a poor prognosis in humans. SPHK1 over-expression in more aggressive FMTs provides support for a potential role of SPHK1 inhibitors for the treatment of FMTs. Targeting SPHK1 has potent cytotoxic effects in primary FMT cells. These findings suggest that further examination of the role SPHK1 plays in FMTs will pave the way for the investigation of SPHK1 inhibitors in future clinical applications.

Diagnosis of feline pancreatitis with SNAP fPL and Spec fPL.

OBJECTIVES: Pancreatitis is a frequent disease in cats for which the ante-mortem diagnosis remains challenging. Feline pancreatic lipase immunoreactivity (fPLI) has been reported to have a high sensitivity for the diagnosis of pancreatitis. The aim of this study was to compare the rapid in-house test SNAP fPL with the standard test Spec fPL and to evaluate the use of SNAP fPL to diagnose pancreatitis in an emergency setting. METHODS: fPLI of 111 cats with a clinical suspicion of pancreatitis was measured with both SNAP fPL and Spec fPL. Furthermore, clinical signs, haematological and biochemical changes, and abdominal ultrasound findings were recorded. RESULTS: Seventy-eight of 111 cats (70.3%) were tested below the cut-off level for pancreatitis with SNAP, as well as Spec fPL, whereas 21/111 (18.9%) were tested with values above the cut-off level with both tests. In 12/111 (10.8%) cats the results were discordant. The comparison of both tests revealed an agreement of 78/80 (97.5%) when Spec fPL was ⩽3.5 µg/l (negative) and 18/20 (90%) when Spec fPL was ⩾5.4 µg/l (positive). The most common clinical signs in cats with suspected pancreatitis (n = 21) were lethargy (95.2%), reduced appetite and vomiting (90.5% each), dehydration (81.0%), diarrhoea (57.1%), abdominal pain and weight loss (47.6% each). Hyperglycaemia and hyperbilirubinaemia (85.7% each), increased aspartate transaminase (76.2%) and alanine transaminase (47.6%), leucocytosis (61.9%), lymphopenia (57.1%), decreased sodium and chloride (57.1% each), and increased urea (52.4%) were the most common abnormalities in blood work. CONCLUSIONS AND RELEVANCE: Clinical signs, as well as routine blood-work changes, were non-specific and thus proved to be insufficient to diagnose pancreatitis. The combination of SNAP fPL and subsequent Spec fPL, if indicated, provided the opportunity to rule out or to diagnose pancreatitis with a higher certainty than previously known test methods. This study proved SNAP fPL to be a valuable tool to exclude or include pancreatitis in an emergency setting.

Identification and quantification of domestic feline cytochrome P450 transcriptome across multiple tissues.

Understanding of cytochrome P450 (CYP) isoform distribution and function in the domestic feline is limited. Only a few studies have defined individual CYP isoforms across metabolically relevant tissues, hampering the ability to predict drug metabolism and potential drug-drug interactions. Using RNA sequencing (RNA-seq), transcriptomes from the 99 Lives Cat Genome Sequencing Initiative databank combined with experimentally acquired whole transcriptome sequencing of healthy, adult male (n = 2) and female (n = 2) domestic felines, expression of 42 CYP isoforms were identified in 20 different tissues. Thirty-seven of these isoforms had not been previously reported in cats. Depending on the tissue, three to twenty-nine CYP isoform transcripts were expressed. The feline genome annotations did not differentiate CYP2E1 and 2E2 genes, demonstrating poor annotation for this gene using the reference genome. As the majority of the sequences are based on automated pipelines, complete cDNA sequences for translation into CYP protein sequences could not be determined. This study is the first to identify and characterize 37 additional CYP isoforms in feline tissues, increasing the number of identified CYP from the previously reported seven isoforms to 42 across 20 tissues.

A GNPTAB nonsense variant is associated with feline mucolipidosis II (I-cell disease).

BACKGROUND: Mucolipidosis II (ML II; I-cell disease) is caused by a deficiency of N-acetylglucosamine-1-phosphotransferase (GNPTAB; EC 2.7.8.17), which leads to a failure to internalize acid hydrolases into lysosomes for proper catabolism of various substances. This is an autosomal recessive lysosomal storage disease and causes severe progressive neuropathy and oculoskeletal dysfunction in humans (OMIM 252500). A naturally occurring disease model has been reported in juvenile domestic cats (OMIA 001248-9685) with clinical signs similar to human patients. We investigated the molecular genetic basis of ML II in a colony of affected cats by sequencing the coding and regulatory regions of GNPTAB from affected and clinically healthy related and unrelated domestic cats and compared the sequences to the published feline genome sequence (NCBI-RefSeq accession no. XM_003989173.4, Gene ID: 101100231). RESULTS: All affected cats were homozygous for a single base substitution (c.2644C > T) in exon 13 of GNPTAB. This variant results in a premature stop codon (p.Gln882*) which predicts severe truncation and complete dysfunction of the GNPTAB enzyme. About 140 GNPTAB variants have been described in human ML II patients, with 41.3% nonsense/missense mutations, nine occurring in the same gene region as in this feline model. Restriction fragment length polymorphism and allelic discrimination real-time polymerase chain reaction assays accurately differentiated between clear, asymptomatic carriers and homozygous affected cats. CONCLUSION: Molecular genetic characterization advances this large animal model of ML II for use to further define the pathophysiology of the disease and evaluate novel therapeutic approaches for this fatal lysosomal storage disease in humans.

Cyclooxygenase-2 immunoexpression in intestinal epithelium and lamina propria of cats with inflammatory bowel disease and low grade alimentary lymphoma.

BACKGROUND: Cyclooxygenase 2 (COX-2) is an inducible isoform by cellular activation, proinflammatory cytokines and growth factors. The aims of the current study were to evaluate COX-2 immunoexpression in epithelial and lamina propria (LP) of cats with inflammatory bowel disease (IBD) and low grade alimentary lymphoma (LGAL), as well as to correlate them with clinical signs and histopathological scoring. Cats diagnosed with IBD and LGAL (2007-2013) were included in the current study. Feline chronic enteropathy activity index (FCEAI) was calculated for all cases. Control group was composed by 3 healthy indoor cats and 5 sick cats died or were euthanized (non-gastrointestinal illness). Diagnosis and classification of IBD and LGAL was established according to the WSAVA gastrointestinal standardization group template and the National Cancer Institute formulation, respectively. Furthermore, a modified WSAVA template was applied for LGAL evaluation. Immunolabelling for COX-2 (polyclonal rabbit anti-murine antibody) was performed on biopsy samples. Epithelial and LP (inflammatory or neoplastic cells) COX-2 immunolabelling was calculated according to the grade and intensity. The most representative segment scored by the WSAVA and the modified WSAVA were used for statistical analysis. RESULTS: Significant difference was found regarding COX-2 intensity overexpression in the epithelial cells of IBD and LGAL groups when compared to control cats, but not between the groups of sick cats, whereas no differences were found regarding the grade of immunoreactivity between groups. No difference was found for COX-2 immunoexpression at the LP between all groups. However, 3 cats from LGAL group showed COX-2 expression in neoplastic cells at the LP. There were no correlations between epithelial or LP COX-2 expression and FCEAI and histological alterations. CONCLUSIONS: Increased COX-2 intensity at the epithelial cells observed in cats with IBD and LGAL may be secondary to the inflammatory response or a protective function in the intestinal reparation. COX-2 expression at the LP was presented in 33% of LGAL. This result provides a reason for further investigation concerning the role of COX-2 expression in feline alimentary lymphoma.

Fatty acid synthase as a potential therapeutic target in feline oral squamous cell carcinoma.

Oral squamous cell carcinoma (OSCC) is an aggressive and treatment-resistant malignancy in both feline and human patients. Recent work has demonstrated aberrant expression of fatty acid synthase (FASN) and an increased capacity for lipogenesis in human OSCC and other cancers. In human OSCC, inhibition of FASN decreased cell viability and growth in vitro, and diminished tumour growth and metastasis in murine preclinical models. This study aimed to characterize FASN as a therapeutic target in feline OSCC. Immunohistochemistry revealed high FASN expression in primary feline OSCC tumours, and FASN expression was detected in OSCC cell lines (3 feline and 3 human) by immunoblotting and quantitative real-time-polymerase chain reaction (qRT-PCR). Orlistat, a FASN inhibitor, substantially reduced cell viability in both feline and human OSCC lines, although feline cell lines consistently displayed higher sensitivity to the drug. FASN mRNA expression among cell lines mirrored sensitivity to orlistat, with feline cell lines expressing higher levels of FASN. Consistent with this observation, diminished sensitivity to orlistat treatment and decreased FASN mRNA expression were observed in feline OSCC cells following incubation under hypoxic conditions. Treatment with orlistat did not potentiate sensitivity to carboplatin in the cell lines investigated; instead, combinations of the 2 drugs resulted in additive to antagonistic effects. Our results suggest that FASN inhibition is a viable therapeutic target for feline OSCC. Furthermore, cats may serve as a spontaneous large animal model for human oral cancer, although differences in the regulation of lipogenesis between these 2 species require further investigation.

Estimates of biologic variation in specific feline pancreatic lipase concentrations in cats without clinical or ultrasonographic evidence of pancreatitis.

BACKGROUND: Serum concentrations of feline-specific pancreatic lipase are commonly used for diagnosis and monitoring of pancreatitis in cats, but little is known regarding biologic variation of this analyte. OBJECTIVES: The purpose of the study was to determine biologic variation, index of individuality, and reference change values (RCV) for specific feline pancreatic lipase concentrations (Spec fPL) in apparently healthy cats. METHODS: Four blood samples were collected prospectively from 12 apparently healthy cats at 2-week intervals. The Spec fPL was measured in all serum samples by a reference laboratory. RESULTS: Intra-individual variation for Spec fPL was 33.5% (95% CI 27.1-43.8%); inter-individual variation was 99.9% (69.2-169.9%) and analytic variation was 7.3% (6.1-9.2%). Reciprocal index of individuality for Spec fPL was 2.9 (2.5-3.8), the 2-sided RCV was 95.1% (77.0-123.9%). CONCLUSIONS: Due to high individuality, use of a population-based RI for Spec fPL may fail to detect clinically significant elevations in individual cats. The use of subject-based RIs for Spec fPL may have greater sensitivity for the detection of a change in pancreatic physiology in cats; however, this would require serial assessment of apparently healthy cats.

Feline platynosomiasis: analysis of the association of infection levels with pathological and biochemical findings.

Platynosomiasis is a common feline hepatic disease caused by Platynosomum fastosum (Trematoda - Dicrocoelidae), which is also known as 'lizard poisoning'. Most reports of feline platynosomiasis show that this disease is sporadic and manifests with uncommon lesions; its pathogenicity is still not well understood. This study aimed to describe liver injuries and enzymatic changes associated with natural P. fastosum infection in 47 stray cats in an endemic area. Overall, 38.3% (18/47) of cats were parasitized, and 2,358 flukes (P. fastosum) were collected (131 - mean intensity of parasitism; 50.2 - mean abundance). The alanine transaminase (ALT) measure was significantly higher in parasitized animals, while alkaline phosphatase (ALP) showed no statistical difference between parasitized and non-parasitized animals. In infected animals, gross pathological lesions and microscopic liver injuries ranged from mild to severe, and were similar to those in previous descriptions of feline platynosomiasis. Nonetheless, the intensity of parasitism was not related to the severity of macroscopic or microscopic hepatic injuries. However, feline platynosomiasis should be considered in the differential diagnosis of feline liver disorders, as well as, in any program of helminth control, even if no clinical abnormalities are present.

Feline platynosomiasis: analysis of the association of infection levels with pathological and biochemical findings / Platinossomiase felina: análise da associação dos níveis de infecção com achados patológicos e bioquímicos

Abstract Platynosomiasis is a common feline hepatic disease caused by Platynosomum fastosum (Trematoda - Dicrocoelidae), which is also known as ‘lizard poisoning’. Most reports of feline platynosomiasis show that this disease is sporadic and manifests with uncommon lesions; its pathogenicity is still not well understood. This study aimed to describe liver injuries and enzymatic changes associated with natural P. fastosum infection in 47 stray cats in an endemic area. Overall, 38.3% (18/47) of cats were parasitized, and 2,358 flukes (P. fastosum) were collected (131 – mean intensity of parasitism; 50.2 – mean abundance). The alanine transaminase (ALT) measure was significantly higher in parasitized animals, while alkaline phosphatase (ALP) showed no statistical difference between parasitized and non-parasitized animals. In infected animals, gross pathological lesions and microscopic liver injuries ranged from mild to severe, and were similar to those in previous descriptions of feline platynosomiasis. Nonetheless, the intensity of parasitism was not related to the severity of macroscopic or microscopic hepatic injuries. However, feline platynosomiasis should be considered in the differential diagnosis of feline liver disorders, as well as, in any program of helminth control, even if no clinical abnormalities are present.

Resumo Platinossomiase é uma doença hepática felina comum causada por Platynosomum fastosum (Trematoda - Dicrocoelidae), também é conhecida como “envenenamento por lagartixa”. A maioria dos relatos de platinossomiase felina mostra que esta doença é esporádica e se manifesta com lesões incomuns; sua patogenicidade ainda não é bem compreendida. Este estudo objetivou descrever as lesões no fígado e alterações enzimáticas associadas à infecção natural por P. fastosum em 47 gatos errantes em uma área endêmica. No total, 38,3% (18/47) dos gatos estavam parasitados, e 2.358 trematódeos (P. fastosum) foram coletados (131 – intensidade média de parasitismo; 50,2 – abundância média). A quantidade de alanina transaminase (ALT) foi significativamente maior nos animais parasitados, enquanto a fosfatase alcalina (ALP) não apresentou diferença estatística entre os animais parasitados e não parasitados. Nos animais infectados, lesões patológicas macroscópicas e microscópicas hepáticas variaram de leve a grave, e foram semelhantes a descrições anteriores de platinossomiase felina. No entanto, a intensidade do parasitismo não foi relacionada à gravidade das lesões hepáticas macroscópicas ou microscópicas. Contudo, a platinossomiase felina deve ser considerada no diagnóstico diferencial de distúrbios hepáticos em felinos, assim como, em qualquer programa de controle de helmintos, mesmo que nenhuma anormalidade clínica esteja presente.

Kit receptor tyrosine kinase dysregulations in feline splenic mast cell tumours.

This study investigated Kit receptor dysregulations (cytoplasmic immunohistochemical expression and/or c-KIT mutations) in cats affected with splenic mast cell tumours. Twenty-two cats were included. Median survival time was 780 days (range: 1-1219). An exclusive splenic involvement was significantly (P = 0.042) associated with longer survival (807 versus 120 days). Eighteen tumours (85.7%) showed Kit cytoplasmic expression (Kit pattern 2, 3). Mutation analysis was successful in 20 cases. Fourteen missense mutations were detected in 13 out of 20 tumours (65%). Eleven (78.6%) were located in exon 8, and three (21.6%) in exon 9. No mutations were detected in exons 11 and 17. Seven mutations corresponded to the same internal tandem duplication in exon 8 (c.1245_1256dup). Although the association between Kit cytoplasmic expression and mutations was significant, immunohistochemistry cannot be considered a surrogate marker for mutation analysis. No correlation was observed between c-Kit mutations and tumour differentiation, mitotic activity or survival.

AKT capture by feline leukemia virus.

Oncogene-containing retroviruses are generated by recombination events between viral and cellular sequences, a phenomenon called "oncogene capture". The captured cellular genes, referred to as "v-onc" genes, then acquire new oncogenic properties. We report a novel feline leukemia virus (FeLV), designated "FeLV-AKT", that has captured feline c-AKT1 in feline lymphoma. FeLV-AKT contains a gag-AKT fusion gene that encodes the myristoylated Gag matrix protein and the kinase domain of feline c-AKT1, but not its pleckstrin homology domain. Therefore, it differs structurally from the v-Akt gene of murine retrovirus AKT8. AKT may be involved in the mechanisms underlying malignant diseases in cats.

Comparison of Serum Spec fPL(™) and 1,2-o-Dilauryl-Rac-Glycero-3-Glutaric Acid-(6'-Methylresorufin) Ester Assay in 60 Cats Using Standardized Assessment of Pancreatic Histology.

BACKGROUND: Feline pancreas-specific lipase (Spec fPL) is considered a useful test for the antemortem diagnosis of pancreatitis in cats. A recent study found good agreement between the results of the Spec fPL and catalytic 1,2-o-dilauryl-rac-glycero-3-glutaric acid-(6'-methylresorufin) ester (DGGR) lipase assay. Prospective studies evaluating their sensitivity and specificity are lacking. OBJECTIVES: To compare the results of the Spec fPL and the DGGR assays with a standardized histologic assessment of the pancreas. ANIMALS: Sixty client-owned cats presented for necropsy. PROSPECTIVE STUDY: Spec fPL concentrations and serum DGGR lipase activity were measured from the same blood sample. The pancreas was removed within 3 hours after euthanasia; serial transverse sections were made every 0.5 cm throughout the entire pancreas and reviewed using a histologic grading scheme. Sensitivity and specificity for the Spec fPL and DGGR assay results were determined. RESULTS: The sensitivity and specificity for the Spec fPL assay (cutoff value ≥5.4 µg/L) was 42.1 [95% confidence interval (95% CI), 29.4-55.9%] and 100% (95% CI, 31.0-100.0%). The sensitivity and specificity for the DGGR assay (cutoff value >26 U/L) was 36.8 (95% CI, 24.7-50.7%) and 100% (95% CI, 31.0-100.0%). When lymphocytic inflammation up to 10% of a section was considered normal, the sensitivity and specificity for Spec fPL assay (cutoff value ≥5.4 µg/L) was 61.1 (95% CI, 36.1-81.7%) and 69.0% (95% CI, 52.8-81.9%) and the sensitivity and specificity for the DGGR assay (cutoff value >26 U/L) was 66.7 (95% CI, 41.2-85.6%) and 78.6% (95% CI, 62.8-89.2%). CONCLUSIONS AND CLINICAL IMPORTANCE: Both lipase assays performed similarly well, but their agreement with histologic pancreatic inflammation was limited.

Is Serum Total LDH Evaluation Able to Differentiate between Alimentary Lymphoma and Inflammatory Bowel Disease in a Real World Clinical Setting?

CONTEXT: An increase in enzyme lactate dehydrogenase (LDH) in serum is a negative prognostic factor for survival in cats affected by lymphoma. Measuring LDH at the time of diagnosis has been studied for differentiating neoplastic disease from non-neoplastic disease in dogs. Inflammatory bowel disease (IBD) and alimentary lymphoma are common diseases in cats. OBJECTIVE: The aim of this study was to determine whether elevation of total LDH occurred in cats with alimentary lymphoma and non-neoplastic gastrointestinal disease, such as IBD, and to evaluate whether this enzyme is useful in supporting the differential diagnosis of these specific diseases. MATERIALS AND METHODS: A prospective non-randomized controlled study was carried-out in a real world setting of three Italian private veterinary clinics. Seventy-one client-owned cats with a history of chronic gastrointestinal symptoms were enrolled; 33 cats were histologically diagnosed as having alimentary lymphoma and 38 cats as having IBD. Serum samples of total LDH analysis were measured. RESULTS: Gender (P = 0.016) and age (P = 0.046) were found to be significant factors influencing the differentiation of serum total LDH between cats with alimentary lymphoma and those with IBD. Despite low diagnostic accuracy in the overall population (63%), a cut-off value of serum total LDH ranging from 0.85- to 1.04-times the upper reference limit showed good capability (accuracy >80%) of differentiating these two conditions in neutered males and cats younger than 8 years of age (AUC: 0.805, 0.833; sensitivities: 76.9%, 83.3%; specificities: 80.0%, 76.5%; PPV: 76.9%, 55.6%; NPV: 80.0%, 92.9%; respectively). CONCLUSIONS: Although our study showed that gender and age are significant factors in differentiating serum total LDH between cats with alimentary lymphoma and those with IBD, this test had poor diagnostic accuracy in differentiating between these two conditions in the overall population.

Serum HER2 levels are increased in cats with mammary carcinomas and predict tissue HER2 status.

HER2 is overexpressed in about 30% of feline mammary carcinomas (FMC) and in 15-30% of breast cancers. Women with HER2-positive breast tumors are associated with shorter survival. This study aimed to optimize the detection and quantification of serum HER2 (sHER2) in cats and to evaluate its potential in diagnosing cats with mammary carcinomas (MC) overexpressing HER2. A prospective study was conducted in 60 queens showing MC and 20 healthy animals. Pre-operative serum samples were collected for sHER2 quantification using two immunoassays: ELISA and Dot blot assay. sHER2 levels were compared with tissue HER2 status assessed by immunohistochemistry. Queens with FMC showed significantly higher mean levels of sHER2 by both ELISA and Dot blot assay. A significant difference in the sHER2 levels was also found between cats with HER2-positive MC and those with low-expressing HER2 MC. A significant correlation between sHER2 levels and tumor HER2 status was also found, particularly when ELISA was used (r = 0.58, p < 0.0001). The value of 10 ng/ml was proposed as the optimal cutoff for both immunoassays by ROC analysis. Like in humans, sHER2 levels are increased in cats with MC HER2-positive, strongly suggesting that evaluation of sHER2 levels can be very useful in feline oncology. The results show that ELISA and Dot blot assay can replace the immunohistochemistry technique, due to their efficacy and lower costs for diagnostic purposes and for monitoring the response to anti-HER2 therapies in cats.