Identification of potential plasma protein biomarkers for feline pancreatic carcinoma by liquid chromatography tandem mass spectrometry.

In both humans and cats, pancreatic carcinoma is an aggressive cancer with a grave prognosis. Proteomics techniques have successfully identified several blood-based biomarkers of human pancreatic neoplasia. Thus, this study aims to investigate whether similar biomarkers can be identified in the plasma of cats with FePAC by using liquid chromatography tandem mass spectrometry (LC-MS/MS). To facilitate evaluation of the low abundance plasma proteome, a human-based immunodepletion device (MARS-2) was first validated for use with feline plasma. Marked reduction and/or complete removal of albumin and immunoglobulins was confirmed by analysis of electrophoretograms and mass spectral data. Subsequently, plasma collected from 9 cats with pancreatic carcinoma (FePAC), 10 cats with symptomatic pancreatitis, and 10 healthy control cats was immunodepleted and subjected to LC-MS/MS. Thirty-seven plasma proteins were found to be differentially expressed (p < .05 in one-way ANOVA, FC >2 in fold change analysis). Among these proteins, ETS variant transcription factor 4 (p < .05) was overexpressed, while gelsolin (p < .01), tryptophan 2,3-dioxygenase (p < .05), serpin family F member 1 (p < .01), apolipoprotein A-IV (p < .01) and phosphatidylinositol-glycan-specific phospholipase D (p < .05) were down-regulated in cats with FePAC. Further studies on these potential biomarkers are needed to investigate their diagnostic value.

Comparison of Fine Arts- and Pathology-Based Observational Skills Training for Veterinary Students Learning Cytology.

Keen observational skills are essential for veterinarians; however, the development of these skills is not usually an explicit part of the veterinary curriculum. Fine arts-based (FAB) observation training has been shown to improve medical students' observational skills and might also improve veterinary students' observational skills. We compared FAB and pathology-based (PB) observation training in a veterinary cytology course. Students initially wrote a pre-test in which they described two cytology images and one art image, followed by participation in either FAB or PB observation training. Both groups completed a similar post-test immediately after training and a delayed post-test 4 weeks later following instruction in cytology. Differences between groups were noted only in the immediate post-test cytology descriptions. The PB group used significantly more specific vocabulary terms and significantly more accurate observations than the FAB group, suggesting an immediate benefit to the discipline-specific information gained in the PB observation training. In the delayed post-test, results for both groups were similar. The FAB group significantly increased their use of specific vocabulary terms and maintained but did not increase accurate observations following cytology instruction, while accurate observations decreased significantly for the PB group. The FAB group might have been able to generalize their observation skills to the discipline of cytology and to better retain these skills. Neither type of training resulted in both achievement and maintenance of the highest recorded scores for accurate observations. Both FAB and PB training led to improved observational skills, and explicit observation training may be useful for veterinary students.

Assessment of neutrophil function in canine cancer patients undergoing chemotherapy and correlation with neutrophil numbers.

Decreased neutrophil function following administration of chemotherapy has been reported in dogs with lymphoma. The first objective of our study was to determine if neutrophil oxidative burst and phagocytic activity are affected by chemotherapy 7 to 10 days following initiation of treatment in dogs with lymphoma and non-lymphoma malignancies. The second objective was to determine if there is a correlation between neutrophil numbers and neutrophil function before or after initiation of chemotherapy. Flow cytometric assessment of neutrophil oxidative burst and phagocytosis following stimulation with Escherichia coli was performed in 9 dogs diagnosed with lymphoma and 17 non-lymphoma tumor-bearing dogs pre- and post-chemotherapy, as well as 14 tumor-free control dogs. Spearman rank correlation was performed to determine if blood neutrophil numbers and neutrophil function were significantly correlated. Lymphoma patients showed significantly reduced percentage neutrophil oxidative burst post-chemotherapy compared to healthy controls as well as compared to pre-chemotherapy values (P = 0.0022 and P = 0.0020, respectively). Lymphoma patients also exhibited significantly reduced neutrophil phagocytosis activity post-chemotherapy compared to controls and pre-chemotherapy values (P = 0.0016 and P = 0.014, respectively). Dogs with non-lymphoma malignancies also showed a significant decrease in both percentage oxidative burst and phagocytosis post-chemotherapy compared to pre-chemotherapy values (P = 0.00040 and P = 0.029, respectively). Neutrophil numbers and function were not significantly correlated. The results of the study suggest that chemotherapeutic treatment decreases neutrophil oxidative burst and phagocytic activity 7 to 10 days post-treatment in dogs with various malignancies. Furthermore, neutrophil numbers cannot be used to predict neutrophil function.

Une diminution de la fonction des neutrophiles après l'administration d'une chimiothérapie a été rapportée chez des chiens atteints de lymphome. Le premier objectif de notre étude était de déterminer si la stimulation oxydative des neutrophiles et l'activité phagocytaire sont affectées par la chimiothérapie 7 à 10 jours après le début du traitement chez les chiens atteints de lymphomes et de tumeurs malignes non lymphomateuses. Le deuxième objectif était de déterminer s'il existe une corrélation entre les nombres de neutrophiles et la fonction des neutrophiles avant ou après le début de la chimiothérapie. L'évaluation par cytométrie en flux de la stimulation oxydative des neutrophiles et de la phagocytose après stimulation par Escherichia coli a été réalisée chez neuf chiens diagnostiqués avec un lymphome et 17 chiens avec des tumeurs non lymphomateuses avant et après la chimiothérapie, ainsi que 14 chiens témoins sans tumeur. Une corrélation des rangs de Spearman a été effectuée pour déterminer si les nombres de neutrophiles sanguins et la fonction des neutrophiles étaient significativement corrélés. Les patients atteints de lymphome ont montré un pourcentage significativement réduit de stimulation oxydative des neutrophiles après la chimiothérapie par rapport aux témoins sains ainsi que par rapport aux valeurs pré-chimiothérapie (P = 0,0022 et P = 0,0020, respectivement). Les patients atteints de lymphome ont également présenté une activité de phagocytose par les neutrophiles significativement réduite après la chimiothérapie par rapport aux témoins et aux valeurs pré-chimiothérapie (P = 0,0016 et P = 0,014, respectivement). Les chiens atteints de tumeurs malignes non lymphomateuses ont également montré une diminution significative du pourcentage de stimulation oxydative et de la phagocytose post-chimiothérapie par rapport aux valeurs pré-chimiothérapie (P = 0,00040 et P = 0,029, respectivement). Les nombres et la fonction des neutrophiles n'étaient pas significativement corrélés. Les résultats de l'étude suggèrent que le traitement chimiothérapeutique diminue la poussée oxydative des neutrophiles et l'activité phagocytaire 7 à 10 jours après le traitement chez les chiens atteints de diverses tumeurs malignes. De plus, les nombres de neutrophiles ne peuvent pas être utilisés pour prédire la fonction des neutrophiles.(Traduit par Docteur Serge Messier).

Tolerance of Ambiguity in Veterinary Students at the Beginning and End of a Second-year Clinical Pathology Course.

Tolerance of ambiguity (TOA) is essential for veterinarians because ambiguity and uncertainty are unavoidable aspects of veterinary practice. However, TOA has been little investigated in veterinarians or veterinary students. In this article, the 27-item Tolerance of Ambiguity of Veterinary Students (TAVS) scale, including eight additional clinical pathology-specific items, is used to evaluate TOA in veterinary students at the beginning and end of a clinical pathology course. Clinical pathology is often one of the first subjects in which students encounter ambiguity because real-life cases are used in teaching. The hypotheses are that TOA will increase across the course and that TOA will correlate with the final grade in the course. Analysis of the TAVS scale revealed very good inter-item reliability (α = 0.80) and a positive correlation between the original TAVS items and the new clinical pathology items (ρ = 0.63). Students demonstrated a significant increase in TOA across the course for TAVS items and a similar trend for clinical pathology items. Four items related to affinity for complexity and novice view showed significant increases in TOA. Two items related to discomfort from uncertainty showed significant decreases. There was no correlation between TOA and final grade in the course. Students rated their personal frustration with ambiguity in the course as low and did not think ambiguity in cases was problematic for teaching. The results suggest that the increased TOA at the end of the course might relate to students being taught-and learning how to cope with-ambiguity through the real-life cases used for teaching.

Using Fine Arts-Based Training to Develop Observational Skills in Veterinary Students Learning Cytology: A Pilot Study.

Arts-based training has been shown to improve medical students' observational skills. Veterinarians also need keen observational skills. Student veterinarians are expected to develop their observational skills; however, this training is usually not an explicit part of the veterinary curriculum. The impact of arts-based observation training has not been investigated in veterinary students learning cytology. In this pilot study, we compared student descriptions of art and cytology images before and immediately after receiving arts-based observation training. After 10 hours of cytology instruction, we again tested students' observational skills and asked for feedback via a survey. Pre-tests and post-tests were scored following a rubric based on expert descriptions of the images. Scores for art image descriptions were higher for both the immediate and delayed post-tests compared to the pre-test (p < .05). Scores for cytology image descriptions were higher for the immediate post-test than the pre-test, but this difference was not significant. Despite 10 hours of cytology instruction between post-tests, scores for cytology image descriptions were lower for the delayed post-test than the immediate post-test, but again, this difference was not significant. Student feedback on the arts-based observation training was positive. Overall, our results suggest that arts-based training may improve student observational skills, although context could be important, as the improvement in description was only significant for art images. Further investigation with a larger cohort of students and a control group that does not receive arts-based training would be valuable.

Intra- and inter-rater agreement for the detection of band neutrophils and toxic change in horses.

BACKGROUND: The detection of band neutrophils and toxic change via microscopic blood smear review is vitally important, as their presence indicates systemic inflammation. However, in-clinic evaluation of WBC morphology is often limited. OBJECTIVE: We aimed to determine the agreement between expert raters in the detection of bands and toxic change. METHODS: Three board-certified clinical pathologists each evaluated 109 blood smears from horses with acute disease, and 19 control smears from healthy horses. The pathologists determined if bands were present, and if so, the percentage of bands present. They also determined if toxic change was present, and if so, the grade of toxic change. Intra-rater agreement was evaluated using 12 duplicate blood smears. Agreement on the presence of bands between pathologists and an in-clinic hematology analyzer was evaluated. RESULTS: Intra-rater agreement was substantial to almost perfect. Agreement between pathologists for the detection of bands was moderate, but when pathologists agreed bands were present, there was excellent agreement on the percentage of bands and mature neutrophils. Agreement between pathologists for the detection of high-grade, clinically relevant toxic change was fair. When pathologists agreed high-grade toxic change was present, there was fair agreement on Döhle bodies and cytoplasmic basophilia and poor agreement on cytoplasmic vacuolation. Agreement between individual pathologists and the in-clinic hematology analyzer for the indication of bands was fair to moderate. CONCLUSIONS: Consistent identification of bands and toxic change is challenging, even for highly trained personnel. It is, thus,not surprising that in-clinic blood smear evaluation of WBC morphology by non-experts could be inadequate.

Do plasma protein:fibrinogen ratios in horses provide additional information compared with fibrinogen concentration alone?

The plasma protein:fibrinogen (PP:F) ratio was introduced to aid interpretation of hyperfibrinogenemia by accounting for dehydration. However, this ratio is inconsistently assessed in practice and its clinical value remains unknown. Our objective was to determine whether the PP:F ratio provides additional information in adult horses beyond fibrinogen concentration alone. Two databases were reviewed to identify 412 hyperfibrinogenemic horses. Plasma protein:fibrinogen ratios were calculated and their interpretation compared to the fibrinogen concentration. Ratios < 15 were supportive of inflammation. Albumin and total protein concentrations were evaluated when ratios were ≥ 15 to determine if inflammation was supported. Very good agreement (86%) was found on the presence of inflammation when PP:F ratios were compared to fibrinogen concentration. In 72% of cases in which PP:F ratios did not support inflammation, inflammation was considered likely based on albumin and total protein. These findings suggest that PP:F ratios do not provide additional information in horses over fibrinogen concentrations alone.

Est-ce que le ratio plasma protéine:fibrinogène chez les chevaux amène une information additionnelle par rapport à la seule concentration en fibrinogène? Le ratio plasma protéine: fibrinogène (PP:F) a été introduit pour aider à l'interprétation de l'hyperfibrinogénémie lors d'hémoconcentration. Toutefois, la valeur clinique de ce ratio est inconnue. Notre objectif était de déterminer si le ratio amène une information additionnelle comparé au fibrinogène. Deux bases de données ont été revues pour identifier 412 chevaux hyperfibrinogénémiques. Le ratio a été calculé et son interprétation comparée à celle du fibrinogène. Les ratios < 15 suggèrent l'inflammation. Pour les cas présentant un ratio ≥ 15, la concentration en albumine et protéines totales a été revue pour voir si l'inflammation était possible. Nos résultats indiquent que dans 86 % des cas, le ratio et le fibrinogène suggèrent la présence d'inflammation. Pour 72 % des cas où le ratio ne suggère pas l'inflammation, cette dernière est considérée probable étant donné la concentration en protéines totales et albumine. Ces résultats suggèrent que le ratio PP:F n'apporte pas d'information supplémentaire comparé au fibrinogène.(Traduit par les auteurs).

Changes in Student Perceptions and Study Strategies Over Time in a Veterinary Clinical Pathology Course Using Case-Based Instruction.

Veterinary students are challenged to develop new, nonlinear ways of thinking as they learn diagnostic reasoning skills. To support this process, we use real-life cases in our clinical pathology course. Changes in student perceptions regarding the use of cases and changes in study strategies over time have not been previously investigated or compared to student grades. Students participated in three voluntary online surveys that included 4-point Likert scale questions and open-ended questions on the helpfulness of cases for learning and study strategies used during the course. We used Friedman tests to detect any differences in perceptions over time; McNemar's test and Wilcoxon signed-rank tests were used to detect any differences in study strategies over time. Fisher's exact tests were used to examine the association between the Likert scale responses and grades in quartiles. Before beginning the course, 29% of students responded that cases were very helpful to their learning, with similar responses for helpfulness in applying course material and grasping important concepts. There was a significant trend of increasing positivity over the duration of the course, with 74% responding that cases were very helpful at the end of the course. The most-reported study strategy was working individually on cases before the midterm (74% of students), and the most helpful study strategy was attending class regularly (88% reported it as very helpful). Study strategies did not change significantly over time. Overall, perceptions and study strategies did not vary significantly with grades.

Quantifying Novice and Expert Differences in Visual Diagnostic Reasoning in Veterinary Pathology Using Eye-Tracking Technology.

Visual diagnostic reasoning is the cognitive process by which pathologists reach a diagnosis based on visual stimuli (cytologic, histopathologic, or gross imagery). Currently, there is little to no literature examining visual reasoning in veterinary pathology. The objective of the study was to use eye tracking to establish baseline quantitative and qualitative differences between the visual reasoning processes of novice and expert veterinary pathologists viewing cytology specimens. Novice and expert participants were each shown 10 cytology images and asked to formulate a diagnosis while wearing eye-tracking equipment (10 slides) and while concurrently verbalizing their thought processes using the think-aloud protocol (5 slides). Compared to novices, experts demonstrated significantly higher diagnostic accuracy (p <.017), shorter time to diagnosis (p <.017), and a higher percentage of time spent viewing areas of diagnostic interest (p <.017). Experts elicited more key diagnostic features in the think-aloud protocol and had more efficient patterns of eye movement. These findings suggest that experts' fast time to diagnosis, efficient eye-movement patterns, and preference for viewing areas of interest supports system 1 (pattern-recognition) reasoning and script-inductive knowledge structures with system 2 (analytic) reasoning to verify their diagnosis.

Evaluation of lymph node aspirates at diagnosis and relapse in dogs with high-grade multicentric lymphoma and comparison with survival time.

BACKGROUND: Canine high-grade multicentric lymphoma, a common disease with variable response to chemotherapy, is often diagnosed using cytology. OBJECTIVES: The purpose of the study was to compare cytologic features of canine peripheral lymph node aspirates collected at diagnosis and at relapse, and evaluate their usefulness in predicting survival. METHODS: Cytologic scoring based on a rubric and nuclear morphometry analyses were performed on cytologic smears collected at diagnosis and at relapse. Scores at diagnosis and relapse were compared by paired t-test and evaluated in relation to time from diagnosis to remission, remission to relapse, relapse to death, and total survival time, using the Cox proportional-hazards regression model. RESULTS: Number of mitoses and total cytologic score were significantly higher at relapse compared to diagnosis (P < .05). None of the nuclear morphometry measures were significantly different between diagnosis and relapse. The presence of binucleated or multinucleated cells at diagnosis was associated with a shorter remission and decreased total survival (P < .05). Increased mean nucleoli at relapse was associated with longer remission and total survival (P < .05). Increased minimum nuclear radius and diameter at diagnosis were associated with a decreased time from relapse to death (P < .05). Several nuclear morphometry measures at relapse were associated with a shorter time from diagnosis to remission (P < .05). CONCLUSIONS: Number of mitoses and total score were higher at relapse than at diagnosis in canine lymphoma. The presence of binucleated or multinucleated cells at diagnosis may be useful as indicator of a poor prognosis. Further studies including a larger number of cases are required to reinforce the prognostic values of these cytologic features.

Reliability of 400-cell and 5-field leukocyte differential counts for equine bronchoalveolar lavage fluid.

BACKGROUND: Reliable enumeration of mast cells and eosinophils in equine bronchoalveolar lavage (BAL) fluid is important because small increases in the percentages of these cells support the clinical diagnosis of inflammatory airway disease (IAD). Increases in BAL neutrophils also occur with IAD but are not specific due to overlap between IAD and recurrent airway obstruction (RAO). OBJECTIVES: The objectives of this study were to evaluate the reliability of a standard 400-cell leukocyte differential count and an alternate method evaluating 5 microscopic fields at 500× magnification in equine BAL fluid cytocentrifuged preparations. METHODS: BAL samples from 60 horses with and without pulmonary inflammation were evaluated using 400-cell and 5-field leukocyte differential counting methods. Reliability of enumeration of each leukocyte type was assessed by calculating and comparing intraclass correlation coefficients (ICC). Reliability of mast cell enumeration was further evaluated by comparing ICCs of slides with different cell densities. RESULTS: Reliability was higher for all cell types with the 5-field method; however, overall the difference between methods was not statistically significant. Neutrophil reliability was high (ICC > 0.90) with both methods. Adequate reliability (ICC > 0.85) for mast cells was achieved only with the 5-field method on slides with higher cell density. CONCLUSION: Enumeration of mast cells is unreliable when the standard 400-cell differential counting method is used, whereas the 5-field method on slides with higher cell density reached acceptable reproducibility. Neutrophil percentages were highly reliable with both methods.

What is your diagnosis? Ventral neck mass in a dog.

: A 14-year-old male Labrador Retriever was presented for lethargy and collapse. On physical examination, numerous abnormalities were found, including a large ventral neck mass (100 cm(3)) in the area of the thyroid gland. Fine-needle aspirates revealed 2 apparent populations of cells: one suspected to be a well-differentiated thyroid carcinoma, and the other consisting of large pleomorphic to spindloid cells suggestive of sarcoma. Two days later, the dog died at home. A full necropsy was not performed, but examination of the head and neck revealed a well-encapsulated mass adjacent to the cranial trachea and larynx. A section of the mass was evaluated histologically and a diagnosis of anaplastic thyroid carcinoma was made. Immunohistochemical evaluation with antibodies to thyroglobulin, cytokeratin, and vimentin confirmed distinct populations of malignant epithelial and malignant mesenchymal cells, and the diagnosis was amended to thyroid carcinosarcoma. Thyroid carcinosarcoma is a rare neoplasm in dogs in which the cell type comprising the mesenchymal component can vary. Immunochemistry to demonstrate the 2 cell types may be necessary to differentiate thyroid carcinosarcoma from anaplastic thyroid carcinoma.

Alkaline phosphatase: beyond the liver.

The alkaline phosphatases comprise a heterogeneous group of enzymes that are widely distributed in mammalian cells. They often are associated with cell membranes, but their exact physiologic function is unknown. Despite this, alkaline phosphatase activity is a very useful serum biochemical indicator of liver disease, particularly cholestatic disease. However, increases in the activity of alkaline phosphatase in serum and other body fluids may reflect physiologic or pathologic changes beyond those of hepatic origin. For example, nonhepatic increases in serum alkaline phosphatase activity are found in young animals, in pregnant and lactating females, and in association with high fat diets. Bone disease, endocrine disease, neoplasia, and other disorders can result in increased alkaline phosphatase activity. In addition, alkaline phosphatase activity may be increased due to induction by certain drugs such as glucocorticoids and anticonvulsants. In this article, we will review the physiologic and pathologic factors influencing the activity of alkaline phosphatase in serum and other body fluids, with an emphasis on disorders beyond liver disease.