Carol B. Grindem

Blood cell markers.

Cytochemistry and immunocytochemistry are important adjunctive technologies to the morphologic characterization of blood cells and hematopoietic neoplasms. Although cytochemistry is briefly discussed, the emphasis of this article is on the clinical application of flow cytometry and leukocyte monoclonal antibodies in veterinary medicine. Classification and significance of immunophenotyping in canine and feline lymphomas and immunodeficiencies such as feline immunodeficiency virus and feline leukemia virus are discussed.

Cytology of bone marrow

Cytologic examination of bone marrow aspirates can provide a wealth of diagnostic information. Practitioners should not hesitate to perform bone marrow aspirates when indicated. This article is designed to assist the practitioner in the evaluation of bone marrow aspiration biopsies. The indications for marrow evaluation, methods of sample collection, sample preparation, and cytologic examination of bone marrow are discussed. Cases are provided to demonstrate accurate interpretation of bone marrow aspirates.

Thrombocytosis: a retrospective study of 165 dogs

BACKGROUND Thrombocytosis has been associated with various conditions, including inflammation, neoplasia, iron deficiency, splenectomy, and drug administration. OBJECTIVE The aim of this study was to characterize diseases and conditions associated with thrombocytosis in dogs. METHODS In this retrospective study, dogs with thrombocytosis (platelet count > 600 × 10(3) /μL) and complete medical records during a 1-year period were included, and breed, sex, age, CBC results, alkaline phosphatase and gamma-glutamyltransferase activities in some dogs, administration of glucocorticoids or vincristine, and primary diagnosis were evaluated. RESULTS Thrombocytosis was found in 240 of 5342 dogs (4.6%), and 165 (3.1%) met inclusion criteria. Thrombocytosis was secondary in all dogs, and underlying diseases and conditions (n,%) were neoplasia (56, 33.9%), inflammation (55, 33.3%), miscellaneous disorders (26, 15.8%), neoplasia plus a second disease (13, 7.9%), endocrine diseases (8, 4.8%), and multiple diseases (7, 4.2%). In dogs with neoplasia, carcinomas (24) and round cell neoplasms (20), especially lymphoma and mast cell tumor, were the most frequent tumors. Inflammatory disorders consisted of immune-mediated disorders (11), neurologic diseases (8), infectious diseases (6), allergic disease (5), orthopedic diseases (4), gastrointestinal diseases (4), and miscellaneous conditions (17). Of the 165 dogs, 73 (44.2%) had received glucocorticoids (55) or vincristine (18) Marked (850-969 × 10(3) platelets/μL) or extreme ( ≥ 970 × 10(3) platelets/μL) thrombocytosis occurred in 24 (14.5%) dogs; 12 (50.0%) had neoplasia. Thromboembolism occurred in 13 (7.9%) dogs. CONCLUSIONS Thrombocytosis in dogs occurred most frequently secondary to neoplastic and inflammatory diseases and was commonly associated with glucocorticoid and vincristine administration. Thromboembolic complications occurred in a small number of patients. Marked or extreme thrombocytosis was more likely to occur with neoplasia than with other conditions.

Bone Marrow Biopsy and Evaluation

Bone marrow evaluation provides valuable diagnostic and prognostic information about neoplastic, metabolic, and inflammatory diseases. Bone marrow biopsies should be done only after examination of peripheral blood, to avoid performing unnecessary biopsies. A blood sample should be taken at the time of the bone marrow biopsy, for complete hematopoietic evaluation. It is preferable to take both an aspiration and core biopsy simultaneously. A good sample is mandatory for accurate evaluation and interpretation. The method of evaluation should be systematic, complete, and cover the following points: adequacy of specimens; estimation of cellularity; identification of number, maturation pattern, and morphology of megakaryocytes, myeloid cells, and erythroid cells; estimation of M:E ratio; and identification of abnormal cells, cellular reactions, infectious agents, or abnormal stromal reactions. Bone marrow findings should be interpreted in conjunction with signalment, history, physical findings, and laboratory results. Reference or institutional laboratories should be contacted for proper handling of bone marrow specimens for special procedures, such as histopathology, cytochemistry, immunopathology, and electron microscopy.

Infection with Panola Mountain Ehrlichia sp. in a Dog with Atypical Lymphocytes and Clonal T-Cell Expansion

An 11-year-old, castrated male Scottish Terrier from Raleigh, NC that lived predominantly indoors and had no travel history was referred to the North Carolina State University Veterinary Health Complex (NCSU-VHC) in October 2012 for routine reevaluation of hepatobiliary disease. The dog’s previous 3-year medical history included biliary mucocele (2010); neutrophilic hepatitis (2011); recurrent Escherichia coli urinary tract infections, esophageal dysmotility, aspiration pneumonia, and transient thrombocytopenia (2012); and food responsive enteropathy (2009–2012). All of these medical problems were well controlled at the time of examination and no clinical abnormalities were reported by the dog’s owners. In the months before the present examination, ticks occasionally were noted and fleas were commonly found on the dog despite reported use of preventive therapies. On physical examination, the dog was obese (body condition score, 7 out of 9) and had mild hepatomegaly, both of which had been present for more than a year. Notable CBC findings included mild thrombocytopenia (platelet count, 143,000/lL; reference interval [RI], 190,000–468,000/lL) and an increased number of atypical lymphocytes (1,773/lL), with normal appearing lymphocytes within the laboratory reference range (1,854/lL; RI, 594–3,305/lL) and an otherwise normal differential cell count. A review of the blood smear by a pathologist identified a population of immature lymphocytes with angular nuclei and cell shape, multiple nucleoli, and deeply basophilic, vacuolated cytoplasm that had a tendency to mold into surrounding cells (Fig 1). These morphologic abnormalities raised the suspicion of possible lymphoid neoplasia, although these changes can be seen secondary to reactive processes such as chronic inflammatory or infectious diseases. Serum biochemical abnormalities included an increase in alkaline phosphatase activity (ALP; 817 IU/L; RI, 16–140 IU/L) and alanine aminotransferase activity (ALT; 60 IU/L; RI, 12–54). During the previous 12-month period, ALT activity varied between normal and 80 IU/L and ALP activity varied between 359 and 534 IU/L. Because these hematological and serum biochemical abnormalities were present 2 weeks later, abdominal ultrasound examination was repeated and identified mottled splenic parenchyma

Learning-Style Profiles of 150 Veterinary Medical Students

Awareness of student learning-style preferences is important for several reasons. Understanding differences in learning styles permits instructors to design course materials that allow all types of learners to absorb and process information. Students who know their own learning style are better able to help themselves in courses taught in a non-preferred method by developing study strategies in line with their preferred learning method. We used the Felder and Solomon Index of Learning Styles to assess the learning-style profiles of 150 veterinary students in three consecutive years. Students were predominantly active (56.7%), sensing (79.3%), visual (76.7%), and sequential (69.3%). Most were balanced on the active-reflective (59.3%) and global-sequential (50%) dimensions, and 61.3% and 54% were moderately to strongly sensing and visual, respectively. Small but significant numbers of students were moderately to strongly intuitive (8.7%), verbal (13%), and global (12%). The most common patterns were active-sensing-visual-sequential (26%), reflective-sensing-visual-sequential (19.3%), active-sensing-visual-global (8.7%), and active-sensing-verbal-sequential (8.7%). Although most students (65.3%) were balanced on one to two dimensions, 77.3% had one or more strong preferences. Our results show that although people have dominant learning-style preference and patterns, they have significant minor preferences and patterns across all dimensions with moderate to strong preferences on each scale. These results indicate that a balanced approach to teaching is essential to allow all students to learn optimally.

Hematologic Changes After Total Body Irradiation and Autologous Transplantation of Hematopoietic Peripheral Blood Progenitor Cells in Dogs With Lymphoma

Dogs with and without lymphoma have undergone hematopoietic cell transplantation in a research setting for decades. North Carolina State University is currently treating dogs with B- and T-cell lymphoma in a clinical setting with autologous peripheral blood progenitor cell transplants, using peripheral blood CD34+ progenitor cells harvested using an apheresis machine. Complete blood counts were performed daily for 15 to 19 days posttransplantation to monitor peripheral blood cell nadirs and subsequent CD34+ cell engraftment. This study documents the hematologic toxicities of total body irradiation in 10 dogs and the subsequent recovery of the affected cell lines after peripheral blood progenitor cell transplant, indicating successful CD34+ engraftment. All peripheral blood cell lines, excluding red blood cells, experienced grade 4 toxicities. All dogs had ≥ 500 neutrophils/μl by day 12, while thrombocytopenia persisted for many weeks. All dogs were clinically normal at discharge.

Human recombinant interleukin-2 induces maturation and activation signals for feline eosinophils in vivo.

Immunotherapy, with interleukin‐2 (IL‐2) or IL‐2 plus lymphokine‐activated killer (LAK) cells, has been used to treat cancer and acquired immunodeficiency syndrome (AIDS) in man. Similarities between feline leukemia virus (FeLV) infection in the cat and human immunodeficiency virus (HIV) infection in man have prompted immunotherapeutic studies in the cat. To develop baseline data on hematological responses to infused IL‐2, cats were given daily (1–14 days) i.v. injections of 5 × 104 U/kg of recombinant human IL‐2 (rHulL‐2). Complete blood cell (CBC) counts were done weekly. Red blood cell (RBC), neutrophil, and lymphocyte numbers did not change appreciably over the course of the study. In contrast, rHulL‐2 caused an eosinophilia in all but the 1 day treatment group. Treatment for 3 days generated a transient eosinophilia on day 7 that returned to baseline by 3 weeks. Five day and 7 day treatments generated an eosinophilia by day 7 that peaked on day 14 and returned to normal values by day 28. Treatment of cats for 14 days did not increase the magnitude or duration of the eosinophilia beyond the 5 or 7 day treatments. Bone marrow (BM) biopsies from rHulL‐2‐treated cats revealed a marked selective hyperplasia of eosinophil precursors. In the 5 day treatment group, all maturation stages of eosinophils were elevated by week 1 of treatment. By week 2, the early stages had returned to normal, whereas the late stage cells remained elevated, suggesting an ordered maturation response. Numbers of all eosinophil precursors approximated pretreatment numbers by weeks 3–4. Thus the BM hyperplasia preceded the blood eosinophilia by 1 week, suggesting that an enhanced maturation response of BM eosinophil precursors is a major contributor to the rHulL‐2‐induced blood eosinophilia. In addition to a maturation signal, rHulL‐2 induces a potent activation signal for eosinophils as measured by a decrease in density and an increase in longevity in culture. The significance of the activated eosinophil in the therapeutic or toxicologic response to rHulL‐2 infusion is discussed.

Comparison of manual polychromatophilic cell and automated reticulocyte quantification in evaluating regenerative response in anemic dogs

BACKGROUND Degree of polychromasia and reticulocyte counts are commonly used in veterinary medicine to evaluate response to anemia. The quantitative association between these 2 measurements has not been evaluated. OBJECTIVE The aim of this retrospective study was to compare automated reticulocyte percents and counts with those of polychromatophilic cells on blood films from dogs. METHODS Blood films and medical records from 30 clinically healthy dogs and 60 anemic dogs were evaluated. Manual percentage of polychromatophilic cells (MPP) was determined by counting cells in 10 1000× fields, and automated reticulocyte percentage (ARP) was measured using an ADVIA 120 hematology analyzer; absolute polychromatophilic cell and reticulocyte counts were then calculated. Degree of polychromasia, estimated as slight, mild, moderate, marked was documented. Percentages and absolute counts of polychromatophilic cells and reticulocytes were compared using least squares regression and Pearsons correlation coefficients. RESULTS The reference interval for MPP in healthy dogs was 0.21-0.26%. Correlation between MPP and ARP was highest when all levels were considered (n = 60, r = .98, P < .0001), strong when ARP was > 5% (n = 20, r = .91, P < .0001), and poor when ARP was 1-2% (n = 20, r = .35, P < .0001). Results were similar for comparisons of absolute reticulocyte and polychromatophilic cell counts, with the best correlation found when all levels of counts were analyzed together (r = .96) and for automated reticulocyte counts > 150,000/μL (r = .94). Correlation of estimates of polychromasia with ARP was good (r = .83). CONCLUSION In anemic dogs MPP can be used for assessment of regeneration, especially in dogs with higher levels of ARP. Standard quantification of reticulocytes is more accurate in dogs with lesser degrees of regeneration.

Extreme lymphocytosis with myelomonocytic morphology in a horse with diffuse large B-cell lymphoma.

An 11-year-old, 443-kg Haflinger mare was presented to the North Carolina State University Veterinary Teaching Hospital with a 2-week history of lethargy and a 3-day duration of anorexia, pyrexia, tachycardia, and ventral edema. Severe pitting edema, peripheral lymphadenopathy, and a caudal abdominal mass were noted on physical examination. An extreme leukocytosis (154.3 × 103 /μL) and microscopic hematologic findings suggestive of myelomonocytic leukemia were observed. Serum protein electrophoresis revealed a monoclonal gammopathy and urine protein electrophoresis revealed a monoclonal light chain proteinuria. Necropsy and histopathology confirmed widespread neoplastic infiltration in many organs with a heterogenous population of cells; there was no apparent evidence of bone marrow involvement. Immunohistochemistry confirmed presence of a majority of B cells with a limited antigen expression, admixed with a lower number of T cells. Molecular clonality analysis of IgH2, IgH3, and kappa-deleting element (KDE, B cell) on whole blood and KDE on infiltrated tissues revealed clonal rearrangements, and the KDE intron clones that amplified in blood and in infiltrated tissue were identical. In contrast, the clonality analysis of T-cell receptor γ revealed no clonality on blood cells and infiltrated tissues. In conjunction with the histopathologic changes, the lesion was interpreted to be composed of neoplastic B cells with a reactive T-cell population. Polymerase chain reaction testing for equine herpes virus 5 was negative. The final diagnosis was diffuse large B-cell lymphoma with a marked hematogenous component.