Generation and characterization of cytochrome P450 3A74 CRISPR/Cas9 knockout bovine foetal hepatocyte cell line (BFH12).

In human, the cytochrome P450 3A (CYP3A) subfamily of drug-metabolizing enzymes (DMEs) is responsible for a significant number of phase I reactions, with the CYP3A4 isoform superintending the hepatic and intestinal metabolism of diverse endobiotic and xenobiotic compounds. The CYP3A4-dependent bioactivation of chemicals may result in hepatotoxicity and trigger carcinogenesis. In cattle, four CYP3A genes (CYP3A74, CYP3A76, CYP3A28 and CYP3A24) have been identified. Despite cattle being daily exposed to xenobiotics (e.g., mycotoxins, food additives, drugs and pesticides), the existing knowledge about the contribution of CYP3A in bovine hepatic metabolism is still incomplete. Nowadays, CRISPR/Cas9 mediated knockout (KO) is a valuable method to generate in vivo and in vitro models for studying the metabolism of xenobiotics. In the present study, we successfully performed CRISPR/Cas9-mediated KO of bovine CYP3A74, human CYP3A4-like, in a bovine foetal hepatocyte cell line (BFH12). After clonal expansion and selection, CYP3A74 ablation was confirmed at the DNA, mRNA, and protein level. The subsequent characterization of the CYP3A74 KO clone highlighted significant transcriptomic changes (RNA-sequencing) associated with the regulation of cell cycle and proliferation, immune and inflammatory response, as well as metabolic processes. Overall, this study successfully developed a new CYP3A74 KO in vitro model by using CRISPR/Cas9 technology, which represents a novel resource for xenobiotic metabolism studies in cattle. Furthermore, the transcriptomic analysis suggests a key role of CYP3A74 in bovine hepatocyte cell cycle regulation and metabolic homeostasis.

Antitumor Effect of Berberine Analogs in a Canine Mammary Tumor Cell Line and in Zebrafish Reporters via Wnt/ß-Catenin and Hippo Pathways.

The heterogeneous nature of human breast cancer (HBC) can still lead to therapy inefficacy and high lethality, and new therapeutics as well as new spontaneous animal models are needed to benefit translational HBC research. Dogs are primarily investigated since they spontaneously develop tumors that share many features with human cancers. In recent years, different natural phytochemicals including berberine, a plant alkaloid, have been reported to have antiproliferative activity in vitro in human cancers and rodent animal models. In this study, we report the antiproliferative activity and mechanism of action of berberine, its active metabolite berberrubine, and eight analogs, on a canine mammary carcinoma cell line and in transgenic zebrafish models. We demonstrate both in vitro and in vivo the significant effects of specific analogs on cell viability via the induction of apoptosis, also identifying their role in inhibiting the Wnt/ß-catenin pathway and activating the Hippo signals with a downstream reduction in CTGF expression. In particular, the berberine analogs NAX035 and NAX057 show the highest therapeutic efficacy, deserving further analyses to elucidate their mechanism of action more in detail, and in vivo studies on spontaneous neoplastic diseases are needed, aiming at improving veterinary treatments of cancer as well as translational cancer research.

Contrast-enhanced ultrasound features of adrenal lesions in dogs.

BACKGROUND: The contrast-enhanced ultrasound (CEUS) features of adrenal lesions are poorly reported in veterinary literature. METHODS: Qualitative and quantitative B-mode ultrasound and CEUS features of 186 benign (adenoma) and malignant (adenocarcinoma and pheochromocytoma) adrenal lesions were evaluated. RESULTS: Adenocarcinomas (n = 72) and pheochromocytomas (n = 32) had mixed echogenicity with B-mode, and a non-homogeneous aspect with a diffused or peripheral enhancement pattern, hypoperfused areas, intralesional microcirculation and non-homogeneous wash-out with CEUS. Adenomas (n = 82) had mixed echogenicity, isoechogenicity or hypoechogenicity with B-mode, and a homogeneous or non-homogeneous aspect with a diffused enhancement pattern, hypoperfused areas, intralesional microcirculation and homogeneous wash-out with CEUS. With CEUS, a non-homogeneous aspect and the presence of hypoperfused areas and intralesional microcirculation can be used to distinguish between malignant (adenocarcinoma and pheochromocytoma) and benign (adenoma) adrenal lesions. LIMITATIONS: Lesions were characterised only by means of cytology. CONCLUSIONS: CEUS examination is a valuable tool for distinction between benign and malignant adrenal lesions and can potentially differentiate pheochromocytomas from adenocarcinomas and adenomas. However, cytology and histology are necessary to obtain the final diagnosis.

Contrast-enhanced ultrasound features of focal pancreatic lesions in cats.

A large overlap in the ultrasound (US) features of focal pancreatic lesions (FPLs) in cats is reported. Furthermore, only a small number of studies describing the contrast-enhanced ultrasound (CEUS) features of FPLs in cats have been conducted today. The aim of this study is to describe the B-mode US and CEUS features of FPLs in cats. Ninety-eight cats cytologically diagnosed with FPL were included. The lesions were classified as adenocarcinoma (n = 40), lymphoma (n = 11), nodular hyperplasia (n = 17), other benign lesion (OBL) (n = 20), cyst (n = 4) or other malignant lesion (OML) (n = 6). Several qualitative and quantitative B-mode and CEUS features were described in each case. OMLs and cysts were not included in the statistical analysis. A decision tree to classify the lesions based on their B-mode and CEUS features was developed. The overall accuracy of the cross-validation of the decision tree was 0.74 (95% CI: 0.63-0.83). The developed decision tree had a very high sensitivity and specificity for nodular hyperplasia (1 and 0.94, respectively) as well as good sensitivity and specificity for both adenocarcinomas (0.85 and 0.77, respectively) and OBLs also (0.70 and 0.93, respectively). The algorithm was unable to detect any specific feature for classifying lymphomas, and almost all the lymphomas were classified as adenocarcinomas. The combination between CEUS and B-mode US is very accurate in the classification of some FPLs, especially nodular hyperplasia and adenocarcinomas. Cytopathology and or histopathology is still a fundamental step FPL diagnostic workflow.

Contrast-enhanced ultrasound features of focal pancreatic lesions in dogs.

BACKGROUND: Contrast-enhanced ultrasound (CEUS) features of pancreatic lesions are poorly reported in veterinary literature. METHODS: Qualitative and quantitative features of pancreatic benign (nodular hyperplasia [NH], cyst and abscess) and malignant (adenocarcinoma and insulinoma) lesions during B-mode and CEUS examinations are described in 75 dogs. RESULTS: Adenocarcinomas (n = 23) had mixed echogenicity at B-mode, and they were hypoenhancing or non-enhancing at CEUS, with a non-homogeneous and cystic enhancement pattern. Insulinomas (n = 23) appeared as hypoechoic lesions at B-mode, and as hyperenhancing, homogeneous and solid lesions at CEUS. NH (n = 17) had an constant appearance, being hypoechoic at ultrasound (US) and isoenhancing at CEUS. Cysts (n = 7) were all anechoic, with acoustic enhancement clearly detectable at US, but were non-enhancing at CEUS. Lastly, abscesses (n = 5) had mixed echogenicity, and they showed both hyperenhancement and non-enhancement at CEUS. Hypoenhancement and non-homogeneous appearance had a moderate diagnostic accuracy in the detection of adenocarcinomas. In particular, hyperenhancement was evident only in malignant lesions (adenocarcinomas and insulinomas). CONCLUSION: CEUS, in combination with B-mode US features, is a valuable tool for distinction of benign and malignant abnormalities of the pancreas and can potentially differentiate insulinomas from adenocarcinomas.

Deepening the Whole Transcriptomics of Bovine Liver Cells Exposed to AFB1: A Spotlight on Toll-like Receptor 2.

Aflatoxin B1 (AFB1) is a food contaminant metabolized mostly in the liver and leading to hepatic damage. Livestock species are differently susceptible to AFB1, but the underlying mechanisms of toxicity have not yet been fully investigated, especially in ruminants. Thus, the aim of the present study was to better characterize the molecular mechanism by which AFB1 exerts hepatotoxicity in cattle. The bovine fetal hepatocyte cell line (BFH12) was exposed for 48 h to three different AFB1 concentrations (0.9 µM, 1.8 µM and 3.6 µM). Whole-transcriptomic changes were measured by RNA-seq analysis, showing significant differences in the expression of genes mainly involved in inflammatory response, oxidative stress, drug metabolism, apoptosis and cancer. As a confirmatory step, post-translational investigations on genes of interest were implemented. Cell death associated with necrosis rather than apoptosis events was noted. As far as the toxicity mechanism is concerned, a molecular pathway linking inflammatory response and oxidative stress was postulated. Toll-Like Receptor 2 (TLR2) activation, consequent to AFB1 exposure, triggers an intracellular signaling cascade involving a kinase (p38ß MAPK), which in turn allows the nuclear translocation of the activator protein-1 (AP-1) and NF-κB, finally leading to the release of pro-inflammatory cytokines. Furthermore, a p38ß MAPK negative role in cytoprotective genes regulation was postulated. Overall, our investigations improved the actual knowledge on the molecular effects of this worldwide relevant natural toxin in cattle.

A Machine Learning-Based Approach for Classification of Focal Splenic Lesions Based on Their CT Features.

The aim of the study was to describe the CT features of focal splenic lesions (FSLs) in dogs in order to predict lesion histotype. Dogs that underwent a CT scan and had a FSL diagnosis by cytology or histopathology were retrospectively included in the study. For the statistical analysis the cases were divided into four groups, based on the results of cytopatholoy or hystopathology, namely: nodular hyperplasia (NH), other benign lesions (OBLs), sarcoma (SA), round cell tumour (RCT). Several qualitative and quantitative CT features were described for each case. The relationship occurring between each individual CT feature and the histopathological groups was explred by means of c chi-square test for the count data and by means of Kruskal-Wallis or ANOVA for the continuous data. Furthermore, the main features of each group were described using factorial discriminant analysis, and a decision tree for lesion classification was then developed. Sarcomas were characterised by large dimensions, a cystic appearance and an overall low post contrast-enhancement. NH and OBLs were characterised by small dimensions, a solid appearance and a high post-contrast enhancement. OBLs showed higher post-contrast values than NH. Lastly, RCTs did not exhibit any distinctive CT features. The proposed decision tree had a high accuracy for the classification of SA (0.89) and a moderate accuracy for the classification of OBLs and NH (0.79), whereas it was unable to classify RCTs. The results of the factorial analysis and the proposed decision tree could help the clinician in classifying FSLs based on their CT features. A definitive FSL diagnosis can only be obtained by microscopic examination of the spleen.

Hypermethylation-Mediated Silencing of CIDEA, MAL and PCDH17 Tumour Suppressor Genes in Canine DLBCL: From Multi-Omics Analyses to Mechanistic Studies.

Gene expression is controlled by epigenetic deregulation, a hallmark of cancer. The DNA methylome of canine diffuse large B-cell lymphoma (cDLBCL), the most frequent malignancy of B-lymphocytes in dog, has recently been investigated, suggesting that aberrant hypermethylation of CpG loci is associated with gene silencing. Here, we used a multi-omics approach (DNA methylome, transcriptome and copy number variations) combined with functional in vitro assays, to identify putative tumour suppressor genes subjected to DNA methylation in cDLBCL. Using four cDLBCL primary cell cultures and CLBL-1 cells, we found that CiDEA, MAL and PCDH17, which were significantly suppressed in DLBCL samples, were hypermethylated and also responsive (at the DNA, mRNA and protein level) to pharmacological unmasking with hypomethylating drugs and histone deacetylase inhibitors. The regulatory mechanism underneath the methylation-dependent inhibition of those target genes expression was then investigated through luciferase and in vitro methylation assays. In the most responsive CpG-rich regions, an in silico analysis allowed the prediction of putative transcription factor binding sites influenced by DNA methylation. Interestingly, regulatory elements for AP2, MZF1, NF-kB, PAX5 and SP1 were commonly identified in all three genes. This study provides a foundation for characterisation and experimental validation of novel epigenetically-dysregulated pathways in cDLBCL.

Corrigendum: Diagnostic Accuracy of Delayed Phase Post Contrast Computed Tomographic Images in the Diagnosis of Focal Liver Lesions in Dogs: 69 Cases.

[This corrects the article DOI: 10.3389/fvets.2021.611556.].

Undifferentiated laryngeal carcinoma with hyaline bodies in a cat.

BACKGROUND: Primary laryngeal neoplasms are rare in cats, with lymphoma and squamous cell carcinoma being the most commonly diagnosed tumour types. These tumours are usually highly aggressive, difficult to treat, and have a poor prognosis. Here an undifferentiated laryngeal carcinoma with hyaline bodies in a cat is reported. CASE PRESENTATION: A 13-year-old cat was presented for progressive respiratory signs. Diagnostic procedures revealed a partially obstructive laryngeal mass. Cytology was compatible with a poorly differentiated malignant tumour, with neoplastic cells frequently containing large intracytoplasmic hyaline bodies. After 1 month the patient was euthanised due to a worsening clinical condition and submitted for post-mortem examination, which confirmed the presence of two laryngeal masses. Histopathology confirmed the presence of an undifferentiated neoplasm with marked features of malignancy. Strong immunolabelling for pancytokeratin led to a diagnosis of undifferentiated carcinoma, however, histochemical and immunohistochemical investigations could not elucidate the origin of the large intracytoplasmic hyaline bodies observed in tumour cells, which appeared as non-membrane bound deposits of electron-dense material on transmission electron microscopy. CONCLUSION: This is the first report of primary undifferentiated laryngeal carcinoma in a cat. Our case confirms the clinical features and the short survival that have been reported in other studies describing feline laryngeal tumours. Moreover, for the first time in feline literature, we describe the presence of intracytoplasmic hyaline bodies in neoplastic cells that were compatible with the so-called hyaline granules reported in different human cancers and also in the dog.

Immunohistochemical Markers of Apoptotic and Hypoxic Damage Facilitate Evidence-Based Assessment in Pups with Neurological Disorders.

Seizures in puppies often present a diagnostic challenge in terms of identifying and treating the underlying cause. Dog breeds with mutations of the MDR1-gene are known to show adverse reactions to certain drugs, yet metabolic imbalance exacerbated by physiologically immature organs and other contributing pathologies require consideration before arriving at a diagnosis. This study analysed the brains of two male, 5-week-old Australian Shepherd siblings that died after displaying severe neurological symptoms upon administration of MilproVet® to treat severe intestinal helminth infection. Despite the initial symptoms being similar, their case histories varied in terms of the symptom duration, access to supportive therapy and post-mortem interval. Histopathology and immunohistochemistry were used to obtain more information about the phase of the pathological processes in the brain, employing protein markers associated with acute hypoxic damage (ß-amyloid precursor protein/APP) and apoptosis (diacylglycerolkinase-ζ/DGK-ζ, apoptotic protease activating factor 1/Apaf1, and B-cell lymphoma related protein 2/Bcl-2). The results seem to reflect the course of the animals' clinical deterioration, implicating that the hypoxic damage to the brains was incompatible with life, and suggesting the usefulness of the mentioned immunohistochemical markers in clarifying the cause of death in animals with acute neurological deficits.

Evaluation of TFR-1 Expression in Feline Mammary Cancer and In Vitro Antitumor Efficacy Study of Doxorubicin-Loaded H-Ferritin Nanocages.

The transferrin receptor 1 (TFR-1) has been found overexpressed in a broad range of solid tumors in humans and is, therefore, attracting great interest in clinical oncology for innovative targeted therapies, including nanomedicine. TFR-1 is recognized by H-Ferritin (HFn) and has been exploited to allow selective binding and drug internalization, applying an HFn nanocage loaded with doxorubicin (HFn(DOX)). In veterinary medicine, the role of TFR-1 in animal cancers remains poorly explored, and no attempts to use TFR-1 as a target for drug delivery have been conducted so far. In this study, we determined the TFR-1 expression both in feline mammary carcinomas during tumor progression, as compared to healthy tissue, and, in vitro, in a feline metastatic mammary cancer cell line. The efficacy of HFn(DOX) was compared to treatment with conventional doxorubicin in feline mammary cancer cells. Our results highlighted an increased TFR-1 expression associated with tumor metastatic progression, indicating a more aggressive behavior. Furthermore, it was demonstrated that the use of HFn(DOX) resulted in less proliferation of cells and increased apoptosis when compared to the drug alone. The results of this preliminary study suggest that the use of engineered bionanocages also offers unprecedented opportunities for selective targeted chemotherapy of solid tumors in veterinary medicine.

Diagnostic Accuracy of Delayed Phase Post Contrast Computed Tomographic Images in the Diagnosis of Focal Liver Lesions in Dogs: 69 Cases.

To describe the computed tomographic (CT) features of focal liver lesions (FLLs) in dogs, that could enable predicting lesion histotype. Dogs diagnosed with FLLs through both CT and cytopathology and/or histopathology were retrospectively collected. Ten qualitative and 6 quantitative CT features have been described for each case. Lastly, a machine learning-based decision tree was developed to predict the lesion histotype. Four categories of FLLs - hepatocellular carcinoma (HCC, n = 13), nodular hyperplasia (NH, n = 19), other benign lesions (OBL, n = 18), and other malignant lesions (OML, n = 19) - were evaluated in 69 dogs. Five of the observed qualitative CT features resulted to be statistically significant in the distinction between the 4 categories: surface, appearance, lymph-node appearance, capsule formation, and homogeneity of contrast medium distribution. Three of the observed quantitative CT features were significantly different between the 4 categories: the Hounsfield Units (HU) of the radiologically normal liver parenchyma during the pre-contrast scan, the maximum dimension, and the ellipsoid volume of the lesion. Using the machine learning-based decision tree, it was possible to correctly classify NHs, OBLs, HCCs, and OMLs with an accuracy of 0.74, 0.88, 0.87, and 0.75, respectively. The developed decision tree could be an easy-to-use tool to predict the histotype of different FLLs in dogs. Cytology and histology are necessary to obtain the final diagnosis of the lesions.

Contrast-enhanced ultrasound features of malignant focal liver masses in dogs.

A total of 185 cases (150 retrospectively and 35 prospectively) of malignant liver masses were collected. In the retrospectively collected cases hyperenhancement during wash-in was the most common feature in HCCs but there was a high percentage of cases showing no enhancement or hypo/isoenhancement. ICCs displayed a large variety of contrast enhancement patterns and, although statically significant differences between ICCs and HCCs were evident, no clear distinction between these two pathologies was possible based only on their CEUS appearance. Sarcomas displayed all the possible degrees of wash-in enhancement with non-enhancing being the most common appearance. Metastases displayed all the possible contrast-enhancement patterns, with the most common being hyperenhancement in the wash-in phase followed by hypoenhancement in the wash-out phase. A decision tree was developed based on the features of the retrospectively selected cases. Based on the developed decision tree 27/35 prospectively collected cases were correctly classified. Even if some significant differences among groups were evident, all the histotypes displayed all the possible patterns of contrast enhancement, and, therefore, the differentiation of liver masses in dogs based only on their CEUS features is not feasible and, therefore, cytology or histopathology is required.

Correlation between renal histopathology and renal ultrasound in dogs.

Fifty-three privately owned dogs were included in the study. Ultrasonography of the kidneys was performed ante mortem. All the dogs died or were euthanized for reasons unrelated to this study. Histopathology of both kidneys was performed, and a degeneration and an inflammation score ranging from zero to two was assigned by consensus between two pathologists. A numerical score based on a three level semi-quantitative scale (0, 0.5, 1) was assigned by consensus between two of the authors to the following ultrasonographic abnormalities: cortico-medullary definition, echogenicity of the renal cortex, echogenicity of the medulla, renal shape, cysts, scars, mineralizations, subcapsular perirenal fluid accumulation, pyelectasia. The scores deriving from the consensus were summed to create a summary index called renal ultrasound score (RUS). Statistically significant differences in cortico-medullary definition, echogenicity of the renal cortex, echogenicity of the medulla, renal shape, scars and pyelectasia were evident between the degeneration score groups. There were significantly different distributions of cortico-medullary definition, renal shape and scars between the inflammatory score groups. There were statistically significant differences in the RUS between the degenerative score groups (F = 24.154, p-value<.001). Post-hoc tests revealed significant differences between all groups. There were no significant differences in the RUS between the inflammatory score groups (F = 1.312, p-value = .264). Post-hoc tests revealed no significant differences between groups. The results of the present study suggest that the number and severity of the ultrasonographic abnormalities are correlated with the severity of the kidney degeneration. On the other hand, inflammation showed poor influence on the ultrasonographic appearance of the kidneys.

Preliminary investigation on feline coronavirus presence in the reproductive tract of the tom cat as a potential route of viral transmission.

OBJECTIVES: Feline infectious peritonitis (FIP) is an immune-mediated disease initiated by feline coronavirus (FCoV) infection. To date, the only proven route of transmission is the faecal-oral route, but a possible localisation of FCoV in the reproductive tract of tom cats is of concern, owing to the involvement of the male reproductive tract during FIP and to the presence of reproduction disorders in FCoV-endemic feline catteries. The aim of the study was to investigate the presence and localisation of FCoV in semen and/or in the reproductive tract of tom cats, and its possible association with seroconversion and viraemic phase. METHODS: Blood, serum, semen and/or testicle samples were obtained from 46 tom cats. Serology was performed on 38 serum samples, nested reverse transcriptase PCR (nRT-PCR) and reverse transcriptase quantitative PCR (RT-qPCR) were performed on 39 blood samples and on 17 semen samples, and histology, immunohistochemistry and nRT-PCR were performed on 39 testicles. RESULTS: Twenty-four of 38 serum samples were positive on serology. Semen samples were negative on RT-PCR and RT-qPCR for FCoV, while all blood samples were negative at both molecular methods, except for one sample positive at RT-qPCR with a very low viral load. All testicles were negative at immunohistochemistry, while six were positive at nRT-PCR for FCoV. Serology and blood PCR results suggest that the virus was present in the environment, stimulating transient seroconversion. FCoV seems not to localise in the semen of tom cats, making the venereal route as a way of transmission unlikely. Although viral RNA was found in some testicles, it could not be correlated with the viraemic phase. CONCLUSIONS AND RELEVANCE: In the light of these preliminary results, artificial insemination appears safer than natural mating as it eliminates the direct contact between animals, thus diminishing the probability of faecal-oral FCoV transmission.

Contrast-enhanced ultrasonography features of hepatobiliary neoplasms in cats.

BACKGROUND: Contrast-enhanced ultrasonography (CEUS) features of primary hepatobiliary neoplasms have been reported in dogs but no information is available in cats. METHODS: Qualitative and quantitative features of bile duct adenomas (BDAs, n=20), bile duct carcinomas (BDCs, n=16), and hepatocellular carcinomas (HCCs, n=8) are described in 44 cats. RESULTS: There was an overlap in CEUS qualitative features between different histotypes, both in wash-in and wash-out phases. Distinction between different neoplasms based only on the CEUS qualitative features was not possible. At peak of enhancement, the BDAs, BDCs and HCCs showed a large range of echogenicities, from hypoenhancement to hyperenhancement, in comparison to the liver parenchyma. Eight of 20 BDAs showed inhomogeneous hyperenhancement during wash-in, which is a feature reported as typical of malignant lesions in dogs. BDC had a significantly faster wash-in compared with both BDA and HCC but the diagnostic accuracy of all the included quantitative variables was only moderate. No significant differences in the wash-out quantitative features of BDA and BDC were evident. CONCLUSION: There is poor evidence that CEUS may be used to distinguish between different primary hepatobiliary neoplasms in cats.

Contrast-enhanced ultrasound features of hepatocellular carcinoma in dogs.

BACKGROUND: This study aimed to describe the contrast-enhanced ultrasound (CEUS) features of canine hepatocellular carcinoma (HCC) in relation to cellular differentiation and lesion size. METHODS: Sixty dogs with a cytological diagnosis of HCC and that underwent a CEUS examination were retrospectively selected. The wash-in and wash-out patterns of contrast enhancement, along with the time to wash-in and the time to wash-out, of each lesion were recorded. A dimensional cut-off value of 3 cm was adopted for classification. RESULTS: Cellular differentiation had a significant influence on both wash-in (chi-squared=16.99; P<0.001) and wash-out (chi-squared=10.9; P=0.004) patterns of contrast enhancement. Lesion size had a lower, but still significant, influence on both wash-in (chi-squared=12.7; P=0.005) and wash-out (chi-squared=7.42; P=0.024) patterns. A homogeneous hyperenhancement in the arterial phase followed by homogeneous wash-out were suggestive of a well-differentiated HCC. The cellular differentiation of lesions with inhomogeneous hyperenhancement or hypoenhancement/no enhancement as well as an inhomogeneous wash-out or no wash-out could not be inferred. CONCLUSIONS: No significant difference in the time to wash-in and the time to wash-out in relation to cellular differentiation or lesion size was evident. CEUS has the potential to improve efficiency in the diagnosis of HCCs in dogs.

Acute Phase Proteins in Marine Mammals: State of Art, Perspectives and Challenges.

The term "acute phase response" (APR) is referred to a nonspecific and complex reaction of an organism that occurs shortly after any tissue damage, such as infection, trauma, neoplasia, inflammation, and stress. The APR can be identified and monitored with some laboratory tests, such as the concentration of several plasma proteins, the acute phase proteins (APPs). The APPs are components of the non-specific innate immune response, and their plasma concentration is proportional to the severity and/or the extent of tissue damage. The evaluation of health status of marine mammals is difficult because the classical clinical signs of illness used for human and domestic animals are difficult to recognize and understand. For this reason, in the past years, several efforts were done to identify laboratory markers of disease in these animals. The APPs have demonstrated their role as early markers of inflammation in veterinary medicine, thus several APPs were tested in marine mammals, such as C-reactive protein (CRP), serum amyloid-A (SAA), and Haptoglobin (Hp). However, the difficulty to extrapolate the knowledge about APPs in one species to another, the lack of specie-specific reagents, the absence of data about negative APPs have hampered their extent use in marine mammals. Herein, the state of art of APPs in marine mammals is reviewed, with particular attention to pre-analytical and analytical factors that should be taken into account in validation and interpretation of APPs assays. Moreover, the current application, potential utility and the future developments of APPs in marine mammals is highlighted and discussed.

Relationship of diagnostic accuracy of renal cortical echogenicity with renal histopathology in dogs and cats, a quantitative study.

BACKGROUND: Renal cortical echogenicity is routinely evaluated during ultrasonographic investigation of the kidneys. Both in dog and cat previous ex-vivo studies have revealed a poor correlation between renal echogenicity and corresponding lesions. The aim of this study was to establish the in-vivo relationship between renal cortical echogenicity and renal histopathology. RESULTS: Thirty-eight dogs and fifteen cats euthanized for critical medical conditions were included in the study. Ultrasonographic images of both kidneys were acquired ante mortem at standardized ultrasonographic settings. The echogenicity was quantified by means of Mean Gray Value (MGV) of the renal cortex measured with ImageJ. A complete histopathological examination of both kidneys was performed. Five kidneys were excluded because histopathology revealed neoplastic lesions. Only samples affected by tubular atrophy showed statistically different values in dog, and histopathology explained 13% of the total variance. MGV was not correlated neither to the degeneration nor to the inflammation scores. However, significant differences were identified between mildly and severely degenerated samples. Overall, the classification efficiency of MGV to detect renal lesions was poor with a sensitivity of 39% and a specificity of 86%. In cats, samples affected by both tubular vacuolar degeneration and interstitial nephritis were statistically different and histopathology explained 44% of the total variance. A linear correlation was evident between degeneration and MGV, whereas no correlation with inflammation was found. Statistically significant differences were evident only between normal and severely degenerated samples with a sensitivity of 54.17% and a specificity of 83.3% and MGV resulted scarce to discriminate renal lesions in this species. CONCLUSIONS: Renal cortical echogenicity shows low relevance in detecting chronic renal disease in dog whereas it results worth to identify severe renal damage in cat.