Prolonged holding time and sampling protocol affects viscoelastic coagulation parameters as measured by the VCM-Vet™ using fresh equine native whole blood.

OBJECTIVES: Determine the effect of sample holding time and single sample reuse on viscoelastic coagulation parameters when using fresh equine native whole blood. ANIMALS: 8 healthy adult horses from a university teaching herd. PROCEDURES: Blood collected by direct jugular venipuncture (18 ga needle, 3 mL syringe) was held at 37 °C for 2, 4, 6, or 8 minutes according to 1 of 2 protocols. Syringes were gently inverted twice, a small amount of blood was expressed, testing cartridges were filled, and placed within the VCM-Vet™ device (Entegrion Inc). Protocol A: samples were processed from a single syringe. Protocol B: 4 syringes were drawn through a single needle. VCM-Vet™ measures assessed included clot time (CT), clot formation time (CFT), alpha angle (AA), amplitude at 10/20 minutes (A10/A20), maximal clot firmness (MCF), and lysis index at 30/45 minutes (LI30/LI45). Differences over time were examined using the Friedman test and post hoc Wilcoxon Rank Sum Test with Bonferroni correction, P ≤ .05. RESULTS: Following Protocol A, there was a significant effect of holding time for CT (P = .02), CFT (P = .04), and AA (P = .05). CT and AA decreased over time, while CFT increased. Samples handled by Protocol B showed no significant difference over time for any of the VCM-Vet™ parameters. CLINICAL RELEVANCE: Sample holding time and handling protocol impact VCM-Vet™ testing results of fresh equine native whole blood. Viscoelastic coagulation samples tested using the VCM-Vet™ may be held unagitated for up to 8 minutes after collection while warm, but should not be reused.

Acute phase protein concentrations following serial procaine penicillin G injections in horses.

BACKGROUND: Acute phase protein (APP) measurement is used to detect inflammation. Intramuscular (IM) injections could cause tissue injury and induce an acute phase response (APR). OBJECTIVES: To evaluate the effects of IM procaine penicillin G (PPG) injections on APP concentrations in horses. STUDY DESIGN: Prospective longitudinal design. METHODS: PPG was administered intramuscularly to six horses, twice daily, for 5 days. Plasma fibrinogen (FIB), serum amyloid A (SAA), haptoglobin (HAP), creatine kinase (CK), and aspartate aminotransferase (AST) were quantified daily for 5 days before the first injection, during the course of administration, and for 4 days after the final dose. Analytes were quantified every other day for the remaining 16 days. Data were compared using a parametric or non-parametric repeated measures ANOVA and a Tukey's or Mann-Whitney rank sum test, respectively. Significance was set at p < 0.05. RESULTS: CK was increased over baseline (mean ± SD: 200 ± 74 IU/L) on Days 1-6 (p < 0.001 to p = 0.02, mean ± SD: 723-1177 ± 355-544 IU/L) and AST was increased above baseline (mean ± SD: 233 ± 58 IU/L) on Days 2-7 and 10 (p < 0.001 to p = 0.05, mean ± SD: 307-437 ± 79-146 IU/L). Increased FIB was noted over baseline (mean ± SD: 177 ± 30 mg/dl) on Days 6-8 and 10 (p = 0.02 to p = 0.03, mean ± SD: 234-252 ± 33-49 mg/dl). SAA was increased above baseline (mean ± SD: 4.7 ± 2.9) on Day 6 (p = 0.02, mean ± SD: 113 ± 186 µg/ml). There was no change in HAP. MAIN LIMITATIONS: Healthy horses were used, small sample size, and a lack of a negative control group. CONCLUSIONS: Serial intramuscular procaine penicillin G (IM PPG) injections may result in increased positive APP concentrations in horses and this must be considered when these test results are interpreted.

Total protein concentration as a predictor of neoplastic peritoneal and pleural effusions of dogs.

BACKGROUND: The diagnosis of neoplastic cavitary effusions requires the identification of neoplastic cells in effusions, yet the cytologic appearance of neoplastic effusions can be highly variable due to the varied mechanisms of formation. Additional parameters might aid in the interpretation of equivocal cytologic results. OBJECTIVES: Our goal was to evaluate whether total protein concentrations can be used to support the diagnosis of neoplasia in the peritoneal and pleural effusions of dogs with lower cellularities (≤5000 nucleated cells/µL). METHODS: Pleural and peritoneal fluid analyses from dogs presented to the University of Illinois Veterinary Teaching Hospital between 2014 and 2019 were evaluated retrospectively. Effusions were categorized as neoplastic or non-neoplastic based on histology or cytology. Non-neoplastic effusions were subcategorized according to mechanism: decreased oncotic pressure, increased hydrostatic pressure, increased vascular permeability, leakage of urine, and leakage of lymph. The TP and blood albumin to fluid TP ratio (Albblood :TPfluid ) were compared among groups. RESULTS: Twenty-seven neoplastic and 65 non-neoplastic cases were evaluated. TP was higher in the neoplastic group (P = .001) than in the non-neoplastic group. Neoplastic effusions had a lower Albblood :TPfluid than non-neoplastic (P = .001), and effusions with Albblood :TPfluid of ≤0.6 were 5.6 times more likely to be neoplastic (95% CI 1.69-17.36; P = .003). CONCLUSIONS: Fluid TP concentrations were significantly greater in neoplastic than non-neoplastic effusions; however, given the considerable overlap between groups, the diagnostic utility of this difference is low. A neoplastic etiology might be more likely in cases with an Albblood :TPfluid ≤0.6.

Utilizing feline oral squamous cell carcinoma patients to develop NQO1-targeted therapy.

Developing effective therapies for the treatment of advanced head-and-neck squamous cell carcinoma (HNSCC) remains a major challenge, and there is a limited landscape of effective targeted therapies on the horizon. NAD(P)H:quinone oxidoreductase 1 (NQO1) is a 2-electron reductase that is overexpressed in HNSCC and presents as a promising target for the treatment of HNSCC. Current NQO1-targeted drugs are hindered by their poor oxidative tolerability in human patients, underscoring a need for better preclinical screening for oxidative toxicities for NQO1-bioactivated small molecules. Herein, we describe our work to include felines and feline oral squamous cell carcinoma (FOSCC) patients in the preclinical assessment process to prioritize lead compounds with increased tolerability and efficacy prior to full human translation. Specifically, our data demonstrate that IB-DNQ, an NQO1-targeted small molecule, is well-tolerated in FOSCC patients and shows promising initial efficacy against FOSCC tumors in proof-of-concept single agent and radiotherapy combination cohorts. Furthermore, FOSCC tumors are amenable to evaluating a variety of target-inducible couplet hypotheses, evidenced herein with modulation of NQO1 levels with palliative radiotherapy. The use of felines and their naturally-occurring tumors provide an intriguing, often underutilized tool for preclinical drug development for NQO1-targeted approaches and has broader applications for the evaluation of other anticancer strategies.

Comparison of creatine kinase in cerebrospinal fluid collected from the cerebellomedullary and lumbar cisterna in 10 dogs with neurologic disease.

BACKGROUND: Analysis of cerebrospinal fluid CK (CSF-CK) might be useful as a prognostic indicator in dogs with neurologic disease. Previous studies have mostly analyzed CSF-CK collected from the cerebellomedullary (CM) cisterna, but CSF collection sites could affect its levels. OBJECTIVES: This is a pilot study aimed to evaluate differences in CSF-CK concentrations when collected from the CM or lumbar cisterna in dogs presenting with neurologic disease. METHODS: Ten dogs presenting for neurologic disease underwent magnetic resonance imaging and CSF collection from both the CM and lumbar cisterna. Cerebrospinal fluid CK was analyzed within 30 minutes. RESULTS: Ten dogs were prospectively recruited. Overall, there was no statistically significant difference between CSF-CK collected from the CM or lumbar cisterna (P = .31). When evaluated by neurolocalization, CSF-CK was different between sites in dogs with thoracolumbar myelopathy (P = .024), but not in dogs with intracranial or cervical neurolocalization (P = .93). All dogs with thoracolumbar myelopathy had equivocal or higher CK levels at the lumbar collection site compared with levels at the CM collection site. CONCLUSIONS: Cerebrospinal fluid CK values differed depending on the CSF site collection, especially in dogs with thoracolumbar myelopathy. In dogs with thoracolumbar myelopathy, CSF-CK was likely to be higher when CSF was taken from the lumbar cisterna compared with the CM cisterna. Collecting CSF from the thoracolumbar site could provide better prognostic information than if collected at the CM collection site.

Comparison of cerebellomedullary and lumbar cerebrospinal fluid analysis in dogs with neurological disease.

BACKGROUND: Cerebrospinal fluid (CSF) analysis aids in categorizing underlying disease processes in patients with neurologic disease. Convention suggests that CSF should be collected caudal to the lesion. However, little evidence exists to justify this assertion. HYPOTHESIS/OBJECTIVES: Evaluate the clinicopathologic differences between CSF collected from the cerebellomedullary (CM) and lumbar cisterns in dogs presented for evaluation of neurologic disease. ANIMALS: Fifty-one client-owned dogs undergoing magnetic resonance imaging (MRI) and CSF collection for investigation of neurologic disease. METHODS: Cerebrospinal fluid was prospectively collected from the CM and lumbar cisterns in all patients. The total protein (TP) concentration, red blood cell (RBC) count, and total nucleated cell count (TNCC) were analyzed within 30 minutes of collection. Results and cytology findings were interpreted by a single pathologist. RESULTS: Fifty-one paired samples were collected. The TNCC (P < .001), RBC (P < .001), and TP (P < .001) were different between collection sites. When grouped by neurolocalization, TP (intracranial, P < .001; cervical, P < .001; thoracolumbar, P < .001) and RBC (intracranial, P < .001; cervical, P ≤ .002; thoracolumbar, P = .006) counts were significantly different. The TNCC was significantly different in the cervical (P = .04) and thoracolumbar localizations (P = .004) but not for intracranial (P = .30) localizations. The pathologist's interpretation differed between sites in 66.7% of the cases (34/51). CONCLUSIONS: In dogs with lesions that neurolocalized to the brain or cervical spinal cord, there may be clinical benefit in collecting fluid from both the CM and lumbar cisterns. In dogs with thoracolumbar myelopathy, CSF collected from the CM cistern may not be representative of the underlying disease process.

Overexpression of prostate specific membrane antigen by canine hemangiosarcoma cells provides opportunity for the molecular detection of disease burdens within hemorrhagic body cavity effusions.

BACKGROUND: Canine hemangiosarcoma (cHSA) is a highly metastatic mesenchymal cancer that disseminates by hematogenous and direct implantation routes. Therapies for cHSA are generally ineffective, in part due to advanced clinical disease stage at the time of diagnosis. The validation of conventional molecular methods for detecting novel biomarkers preferentially expressed by cHSA could lead to more timely diagnosis, earlier therapeutic interventions, and improved outcomes. In humans, prostate-specific membrane antigen (PSMA) is a transmembrane protein overexpressed by prostate carcinoma and tumor-associated endothelium of various solid cancer histologies. Importantly, the preferential overexpression of PSMA by certain cancers has been leveraged for the development of diagnostic molecular imaging reagents and targeted therapeutics. Recently, PSMA has been qualitatively demonstrated to be expressed in cHSA cell lines, however, quantitative PSMA expressions and the potential utility of PSMA transcript identification in biologic fluids to support the presence of microscopic cHSA burden has not been reported. Therefore, this study sought to characterize the differential quantitative expressions of PSMA between cHSA and non-malignant tissues, and to determine the potential diagnostic utility of PCR-generated PSMA amplicons as a surrogate of rare cHSA cells dwelling within peritoneal and pericardial cavities. METHODS: Quantitative gene and protein expressions for PSMA were compared between one normal endothelial and six cHSA cell lines by RT-PCR, western blot analysis, and fluorescent microscopy. Additionally, gene and protein expressions of PSMA in normal canine tissues were characterized. Graded expressions of PSMA were determined in spontaneously-arising cHSA tumor samples and the feasibility of qualitative PCR as a molecular diagnostic to detect PSMA transcripts in whole blood from healthy dogs and hemorrhagic effusions from cHSA-bearing dogs were evaluated. RESULTS: PSMA gene and protein expressions were elevated (up to 6-fold) in cHSA cells compared with non-malignant endothelium. By immunohistochemistry, protein expressions of PSMA were detectable in all cHSA tissue samples evaluated. As predicted by human protein atlas data, PSMA's expression was comparably identified at substantial levels in select normal canine tissues including kidney, liver, and intestine. In young healthy pet dogs, PSMA amplicons could not be identified in circulating whole blood yet were detectable in hemorrhagic effusions collected from pet dogs with confirmed cHSA or PSMA-expressing cancer. CONCLUSIONS: PSMA is quantitatively overexpressed in cHSA compared to normal endothelium, but its protein expression is not restricted to only cHSA tumor tissues, as specific visceral organs also substantively express PSMA. Optimized qualitative PCR methods failed to amplify PSMA amplicons sufficiently for visible detection in circulating whole blood derived from healthy young dogs, yet PSMA transcripts were readily identifiable in hemorrhagic effusions collected from pet dogs with histologically confirmed cHSA or PSMA-expressing cancer. While preliminary, findings derived from a limited cohort of normal and diseased pet dogs provocatively raise the potential value of PSMA amplicon detection as an ancillary molecular diagnostic test for supporting the presence of microscopic cHSA disease burden within hemorrhagic body cavity effusions.

Cytology of Bone.

Cytology of bone is a useful diagnostic tool. Aspiration of lytic or proliferative lesions can assist with the diagnosis of inflammatory or neoplastic processes. Bacterial, fungal, and protozoal organisms can result in significant osteomyelitis, and these organisms can be identified on cytology. Neoplasms of bone including primary bone tumors such as osteosarcoma, chondrosarcoma, fibrosarcoma, synovial cell sarcoma, and histiocytic sarcoma and tumors of bone marrow including plasma cell neoplasia and lymphoma and metastatic neoplasia can result in significant bone lysis or proliferation and can be diagnosed effectively with cytology.

SYSTEMIC BLASTOMYCOSIS IN A CAPTIVE RED RUFFED LEMUR (VARECIA RUBRA).

A 5-yr-old, intact male red ruffed lemur ( Varecia rubra ) presented for evaluation as the result of a 1-wk history of lethargy and hyporexia. Physical examination findings included thin body condition, muffled heart sounds, harsh lung sounds, and liquid brown diarrhea. Complete blood count and serum biochemistry showed an inflammatory leukogram, mild hyponatremia, and mild hypochloremia. Orthogonal trunk radiographs revealed a severe alveolar pattern in the right cranial lung lobes with cardiac silhouette effacement. Thoracic ultrasound confirmed a large, hypoechoic mass in the right lung lobes. Fine-needle aspiration of the lung mass and cytology revealed fungal yeast organisms, consistent with Blastomyces dermatitidis. Blastomyces Quantitative EIA Test on urine was positive. Postmortem examination confirmed systemic blastomycosis involving the lung, tracheobronchial lymph nodes, spleen, kidney, liver, cerebrum, and eye. To the authors' knowledge, this is the first report of blastomycosis in a prosimian species.

Characterization of feline serum-cobalt binding.

BACKGROUND: Oxidative stress inhibits albumin's ability to complex with cobalt. Feline serum-cobalt binding has not been described. OBJECTIVES: The objective was to develop a cobalt binding test for use with feline serum, and correlate the results with other biochemical and cellular constituents in blood, and with clinical diseases of cats. METHODS: A colorimetric test of cobalt binding, based on the oxidation-reduction reaction of Co(+2) and dithiothreitol, was developed using feline serum. The test was used to measure cobalt binding in stored serum from 176 cats presented to the University of Illinois Veterinary Teaching Hospital for a variety of disease conditions. Time-matched hematology and biochemical data, and clinical information, were obtained from the medical record of each cat and correlated with the serum-cobalt binding results. RESULTS: Serial dilution of feline serum with phosphate-buffered saline resulted in a highly linear decrease in serum-cobalt binding (r(2)  = .9984). Serum-cobalt binding of the clinical samples also correlated with albumin concentrations in a stepwise linear regression model (r(2)  = .425), and both cobalt binding and albumin were significantly decreased in cases of inflammation. Albumin and cobalt binding also shared significant correlations with several erythron variables, and serum concentration of total calcium and bilirubin. CONCLUSIONS: The correlation of cobalt binding measured by a colorimetric test with albumin concentration in the clinical samples and with serum dilution is consistent with feline albumin-cobalt complex formation. Hypoalbuminemia is the likely cause of reduced serum-cobalt binding in inflammation and the correlations observed between cobalt binding and other variables.

Identification of Cytauxzoon felis infection in domestic cats from southern Illinois.

OBJECTIVES: The objective of this study was to document Cytauxzoon felis infection in domestic cats from southern Illinois. METHODS: Diagnosis of cytauxzoonosis was based upon clinical signs of illness and detection of piroplasms within erythrocytes on peripheral blood smears or schizonts in internal organs consistent with Cytauxzoon infection. Additionally, genomic DNA was extracted from histologic sections of splenic tissue from two cats. RESULTS: The internal transcribed spacer region-1 (ITS-1) and ITS-2 of the C felis genome were successfully sequenced, confirming infection with the organism. CONCLUSIONS AND RELEVANCE: Sequence analysis of C felis DNA isolated from histologic lesions in two domestic cats from southern Illinois show either mixed infection or possible heterozygosity (cytosine and thymine) in ITS-2 at the position equivalent to nucleotide 76 (thymine) in the most commonly isolated C felis ITS-2 sequence. Identification of C felis infection in domestic cats from southern Illinois is a critical finding that raises awareness of this often fatal disease process in an area of the USA where, previously, the disease was only anecdotally reported.

Vaginal fold histology reduces the variability introduced by vaginal exfoliative cytology in the classification of mouse estrous cycle stages.

Vaginal exfoliative cytology is commonly used in biomedical and toxicological research to classify the stages of the rodent estrous cycle. However, mouse vaginal exfoliative cytology is commonly used as a stand-alone tool and has not been evaluated in reference to vaginal histology and serum sex hormone levels. In this study, the direct and Giemsa-stained methods of vaginal exfoliative cytology were compared in reference to vaginal fold histology and serum sex hormone levels. Both methods predicted the estrous stages similarly with mean discordance rates of 55%, 77%, 46%, and 31%, for diestrus, proestrus, estrus, and metestrus, respectively. From these results, we conclude that vaginal exfoliative cytology may be used as a general guide to determine the desired estrous stage end point and that a definitive confirmation of the estrous stage should be obtained from evaluation of vaginal fold histology. Confirmation of the stage of the estrous cycle by vaginal fold histology will decrease the variability otherwise introduced by misclassification of estrous cycle stages with vaginal exfoliative cytology.

Dermatophilus congolensis in a feral cat.

A young adult feral cat presented to the Champaign County Humane Society with a subcutaneous mass near the stifle. The mass was aspirated. Chains of paired cocci organisms were identified, consistent with Dermatophilus congolensis. The identity of these organisms was confirmed by culture and polymerase chain reaction.

Cytologic identification of erythrophagocytic neoplasms in dogs.

BACKGROUND: Erythrophagia has been identified in multiple types of tumors in human medicine, but information in veterinary medicine is limited. In dogs and cats, erythrophagia has been reported to occur in mast cell and plasma cell tumors, lymphoma, megakaryocytic leukemia, and hemophagocytic histiocytic sarcoma. On the basis of clinical experience, we believed this to be an incomplete list that might bias veterinarians toward a diagnosis of histiocytic sarcoma when erythrophagia is observed in neoplastic spindle-shaped cells. OBJECTIVE: The objective of this retrospective study was to identify neoplasms in dogs that exhibit erythrophagia within the neoplastic cell population. METHODS: Reports of cytologic samples collected from dogs and submitted to the University of Illinois Veterinary Diagnostic Laboratory between January 2003 and May 2011 were searched for a description of erythrophagia. Only cases with a diagnosis confirmed by histopathologic examination using immunohistochemical analysis or by cytochemical staining were included. Slides were reviewed by one pathologist to confirm the presence of erythrophagocytic neoplastic cells. RESULTS: Of 20,016 reports searched, 897 contained the term "erythrophagia." In 55 reports, neoplastic cells exhibiting erythrophagia were identified. Of these, 21 cases were excluded because the cytologic diagnosis was not confirmed, and the remaining 34 cases included 12 hemangiosarcomas, 11 hemophagocytic histiocytic sarcomas, 9 osteosarcomas, one mast cell tumor, and one T-cell lymphoma. CONCLUSION: Erythrophagia can be found in many tumor types. When erythrophagia is observed in spindle-cell sarcomas, osteosarcoma and hemangiosarcoma should be considered in addition to hemophagocytic histiocytic sarcoma.

Neutropenia in dogs and cats: causes and consequences.

Neutropenia is a serious hematopathologic change that should not be ignored. In almost all patients, it is an important primary or secondary indicator of significant underlying disease. While in some neutropenic patients the diagnostic work up will be simple, in others it is challenging. The value of examining a blood smear for toxic changes in neutrophils cannot be overemphasized; it may indicate the presence of systemic inflammation, as well as providing clues about prognosis and the extent of treatment the patient may require.