Subleukaemic acute myeloid leukaemia with myelodysplasia in a horse

Acute myeloid leukaemia (AML) is rarely reported in horses and myelodysplastic syndrome (MDS) has been described only in one case. Acute myeloid leukaemia is defined as the presence of at least 20% blasts in the marrow or blood. On the other hand, MDS is characterised by morphologic abnormalities in one or more cell lineages with hypercellular marrow and peripheral cytopenias due to ineffective haematopoiesis. We report a case of acute myelomonocytic leukaemia with myelodysplasia-related features in a horse. A supposed diagnosis was based on abnormal morphology of circulating neoplastic cells and bone marrow cytology. A final diagnosis was made by using flow cytometry (FC) in conjunction with cytochemistry (CC) rarely reported in the haematopoietic neoplasms of the horse.     

Acute myelomonocytic leukemia in a horse

A 7-year-old Quarter Horse stallion with a myeloproliferative disorder was examined because of colic, and an enterolith was removed surgically. The horse experienced secondary complications after abdominal surgery, and leukopenia and thrombocytopenia were detected. Five months later, the horse was examined for recurrent peripheral edema and for repair of an abdominal incisional hernia. Acute myelomonocytic leukemia was diagnosed, and treatment with low-dose (noncytocidal) cytosine arabinoside was unsuccessful. Necropsy revealed neoplastic infiltrate in the spleen, liver, lung, adrenal gland, testes, and eye. The persistent hematologic abnormalities before the onset of overt leukemia may represent hematopoietic dysplasia or preleukemia.     

Detection of equine immunoglobulin-secreting cells by a plaque assay.

A protein A-hemolytic plaque assay was applied to detect immunoglobulin (Ig)-producing cells in horse peripheral blood, using pokeweed mitogen as a B lymphocyte activator. A maximum number of Ig-secreting cells was obtained when horse peripheral blood lymphocytes were cultured in a medium containing horse serum. The number of Ig-secreting cells in young horses (2 years old) was lower than that in adult horses (6 to 23 years old). In addition, the plaque formation was unchanged from blood samples kept at 4 degrees C for 24 hours, while blood samples kept for 72 hours did not yield plaques. These results indicate that the plaque assay is a reliable and useful method for detecting Ig-secreting cells in the peripheral blood of the horse.     

THE APPLICATION OF TECHNETIUM-99m HEXAMETHYLPROPYLENEAMINE OXIME (99mTc-HMPAO) LABELED WHITE BLOOD CELLS FOR THE DIAGNOSIS OF RIGHT DORSAL ULCERATIVE COLITIS IN TWO HORSES

The application of 99mTc-HMPAO labeled white blood cells to support the diagnosis of right dorsal ulcerative colitis was studied in two horses with a history and clinical signs consistent with phenylbutazone toxicity. These images were compared to a reference horse unaffected by right dorsal ulcerative colitis. Blood was collected aseptically in heparinized syringes from the patients for in vitro white blood cell (WBC) radiolabeling. The buffy coat was separated out and radiolabeled with 99mTc-HMPAO. The radiolabeled blood was re-injected i.v. and four images of the right and left side of the patient's abdomen were acquired at 4 hours and 20 hours post-injection. Results of the nuclear study revealed no abnormal findings in the abdomen at the four-hour post-injection images in any horse. Images obtained 20 hours post-injection revealed a linear uptake of radiolabeled WBCs in the right cranioventral abdomen in the region of the right dorsal colon in both horses with right dorsal ulcerative colitis. The reference horse had no radiopharmaceutical uptake in this region. This nuclear imaging study was a rapid, non-invasive method to identify right dorsal colon inflammation. These findings not only supported the diagnosis of right dorsal ulcerative colitis, but also facilitated appropriate medical management of each horse.     

Endocytosis of erythrocytes in vivo and particulate substances in vitro by feline neoplastic mast cells.

Clinical evidence for the phagocytic capability of neoplastic feline mast cells was provided by recognition of endocytosed erythrocytes in seven of 12 cytological smears of mast cell neoplasms, particularly in those cells collected from splenic tumors. The capability of these neoplastic mast cells to endocytose particulate substances was also studied in vitro. Evidence is presented that under cultural conditions, feline neoplastic mast cells are capable of endocytosing a variety of substances including polystyrene latex microspheres, zymosan particles, horse spleen ferritin, salmon sperm nuclei, horseradish peroxidase, and carbon particles.     

Pancytopenia Secondary to Lymphoid Leukemia in Three Horses

Pancytopenia was observed in two 3-year-old geldings and one 11-year-old mare. All horses had a brief history (2 days to 4 weeks) of fever, anorexia, and depression. One of the three horses had blast cells present on a peripheral blood smear. Examination of the bone marrow showed substantial infiltration with neoplastic lymphoid cells. At necropsy, neoplastic cells were restricted to the bone marrow in one horse, present in bone marrow, liver, and spleen in the second horse, and reported in multiple tissues in the third horse, including bone marrow, kidneys, lung, myocardium and lymph nodes. The value of a bone marrow aspirate and core biopsy in the investigation of pancytopenia is highlighted. (Journal of Veterinary Internal Medicine 1993; 7:360–363. Copyright © 1993 by the American College of Veterinary Internal Medicine.)     

Equine testicular interstitial cell tumors

Interstitial cell tumors from nine stallions were described. In all but one horse the tumors were found in undescended testes. Five animals had bilateral tumors. Two animals showed increased aggression. Tumors contained two cell types. The first type were large distinctly bordered eosinophilic cells interpreted to be hyperplastic and hypertrophic interstitial cells. They blended with pleomorphic often spindloid neoplastic cells which had fibrillar, vacuolated cytoplasm and indistinct cell borders. This latter cell population was arranged in nodules or broad sheets as endocrine-like packets or interweaving fascicles. Biologic behavior of the neoplasms could not be ascertained from histologic examination.     

Further Studies on the Cell Populations of an Intersex Horse

An intersex horse exhibiting cell types of different sex chromosome constitution was subjected to further studies in order to determine whether the house was a mosaic or a chimera. Cultures of gonadal tissue and peripheral blood revealed mainly 64/XX and 64/XY cells, the former predominating in both tissues. The frequency of drumstick-bearing poly-morphonuclear neutrophils in the intersex horse was similar to that noted in normal mares. Blood type analysis using 17 naturally occurring agglutinins and hemolysins revealed partial agglutinations with three antibodies for the factors of the A system (anti-A, anti-F, and anti-I), and partial hemolysis with anti-Fr₃ suggesting erythrocyte chimerism probably resulting from intrauterine interchange of blood cell precursors as noted in other domestic animals. On the other hand, the presence of XX and XY cells in cultures of gonads which in our intersex horse were apparently devoid of germ cells, would seem to indicate wholebody chimerism resulting from double fertilization or blastocyst fusion.     

The use of cytochemistry,immunophenotyping, flow cytometry,and in vitro differentiation to determine the ontogeny of a canine monoblastic leukemia

We evaluated the utility of cytochemistry, immunophenotyping, flow cytometry, and in vitro culture with forced differentiation of leukemic cells as diagnostic aids to identify the malignant cell ontogeny in a dog with leukemia. A tentative diagnosis of monoblastic leukemia was established by microscopic examination of Romanowsky-stained blood smears and bone marrow aspirate smears. This diagnosis also was supported by the light scatter signature that identified the blast cells as large, non-granular monocytic cells using a CellDyn 3500 automated hematology analyzer; as well as by the detection of N-butyrate esterase and the lack of choloroacetate esterase or leukocyte peroxidase by cytochemical staining. Subsequently, leukemic cells were isolated from the dog's peripheral blood and placed into tissue culture or cryopreserved. The leukemic cells grew in suspension cultures and proliferated spontaneously for up to 4 days. By day 7, proliferation was negligible. Upon culture with conditioned supernatant using mitogen-stimulated human T cells as a source of cytokines, an increased proportion of cells entered S phase by day 2 of culture; however, proliferation declined markedly by day 4, at which time the cells had apparently differentiated to adherent, vacuolated macrophages. The cytokine-stimulated leukemic cells were positive for the monocyte/macrophage specific markers alpha-1-antitrypsin, alpha-1-antichymotrypsin, lysozyme, CD14, MHC class II, and calprotectin, an antigen found in differentiated macrophages and granulocytes. Despite the strong tendency of the leukemic cells towards monocytic differentiation, our results suggested that they retained some features of a myelomonocytic precursor. These data show that cytochemistry, immunophenotyping, flow cytometry, and in vitro differentiation of canine leukemia cells are useful tools for confirming the lineage of malignant hematopoietic cells.     

畜禽体外培养细胞生长规律研究

本研究利用组织块贴壁培养法对鲁西黄牛、蒙古马、北京鸭和狼山鸡等4个品种体外培养细胞的生长、分化规律进行了研究。通过绘制其生长曲线和在激光共聚焦显微镜下拍摄细胞生长过程,初步分析成纤维细胞的生长、分化规律。试验结果表明,4个品种的细胞生长曲线均呈明显的“S”形,家畜和家禽的细胞潜伏期和停滞期的长短以及细胞群体倍增时间均有明显差异,蒙古马和狼山鸡体外培养细胞的单个细胞分裂期也有显著差异,蒙古马为97 min,而狼山鸡为85 min。本试验构建的4个品种成纤维细胞系不仅使畜禽遗传资源在细胞水平永久保存下来,而且为分子生物学研究提供宝贵材料。     

Identification of acute myeloid leukemia in dogs using flow cytometry with myeloperoxidase, MAC387, and a canine neutrophil-specific antibody

BACKGROUND: Flow cytometry may be used to determine immunophenotype or lineage of leukemic cells, but few antibodies are available that are specific for cells of monocytic and granulocytic lineage. OBJECTIVE: The purpose of this study was to evaluate the flow cytometric staining patterns of 3 commercial monoclonal antibodies for monocytes and granulocytes in clinically healthy dogs and in dogs with acute myeloid leukemia (AML). METHODS: Mouse antihuman macrophage antibody (MAC387), mouse anti-human myeloperoxidase (MPO), and a canine neutrophil-specific antibody (NSA) were evaluated using flow cytometry on blood from 6 clinically healthy control dogs, and on blood (n = 7) and/or bone marrow (n = 2) from 8 dogs with AML. A diagnosis of acute leukemia was confirmed by >30% blasts in bone marrow or >30% blasts in peripheral blood, together with bi- or pancytopenia, circulating CD34-positive blast cells, and clinical signs of disease. Leukemic samples also were evaluated using a wide panel of monoclonal antibodies. RESULTS: MAC387 stained neutrophils and monocytes from control dogs, although the staining profiles for the 2 cell types differed. MPO and NSA resulted in strong positive staining of neutrophils; MPO also stained monocytes weakly. Lymphocytes did not stain with any of the antibodies. One case was classified as AML of granulocytic lineage (AML-M1), 6 cases were classified as acute monocytic leukemia (AML-M5), and 1 case was classified as acute myelomonocytic leukemia (AML-M4). Neoplastic myeloblasts in the dog with granulocytic AML were positive for MPO, NSA, MAC387, and CD4. All monoblasts from the dogs with AML-M5 were positive for CD14, 5 of 6 were positive for MAC387, and 2 were positive for MPO. NSA staining was negative in the 2 dogs with AML-M5 in which it was evaluated. In the dog with AML-M4 variable percentages of blast cells were positive for CD14, MPO, MAC387, CD4, and NSA. CONCLUSIONS: Antigens identified by antibodies to MAC387, MPO, and NSA were expressed not just by normal mature neutrophils and monocytes, but also by neoplastic myeloblasts and monoblasts. These 3 antibodies may be useful as part of a wider panel for immunophenotyping AML in dogs.     

水牛外周血单核巨噬细胞的培养与鉴定

本试验旨在为阐明水牛感染大片吸虫的免疫机理提供单核巨噬细胞材料。经过密度梯度离心分离,贴壁法培养水牛外周血单核巨噬细胞,以MTT法检测培养4、24、48、72、96、120、144、168、192 h时贴壁单核巨噬细胞的相对存活数量,结果显示贴壁细胞体外培养72、96、120 h时,细胞数量相对稳定,组间差异不显著(P＜0.05)。贴壁72 h的细胞通过倒置显微镜、瑞氏染色观察,结果显示其具备单核巨噬细胞的典型形态特征;酸性磷酸酶、非特异性酯酶染色阳性率分别为(72.8±0.5)%和(84.6±0.4)%,显示其具备单核巨噬细胞酶化学特性;贴壁细胞表达单核巨噬细胞特异性表面抗原MHC Ⅱ;墨汁的吞噬率为(87.5±0.6)%,表明该细胞具有单核巨噬细胞的吞噬功能。结果表明贴壁法分离,5% BSA的DMEM培养水牛外周血单核巨噬细胞,培养72 h贴壁细胞具备单核巨噬细胞特征,在细胞数量相对稳定期(72、96、120 h)可以进行相应的试验研究。     

Multiple masses in a horse's tongue resulting from an atypical perineurial cell proliferative disorder

A 5-year-old National Show horse mare presented with a soft mass on the left dorsolateral aspect of the tongue. Over the next 2 years, the mare developed numerous, similar, coalescing masses that extended along the left dorsolateral aspect to the tip of the tongue. Microscopically, the bases for these masses were slender, fusiform, mesenchymal cells that formed compact whorls around myelinated and unmyelinated nerves. These cells were labeled by antibodies directed against vimentin but not by S-100. Ultrastructurally, multiple, concentrically arranged, long, slender cell processes, with discontinuous external laminae and many pinocytotic vesicles, helped to accurately phenotype the proliferative element. Whether this unusual perineurial cell proliferative disorder is neoplastic or not remained a matter of conjecture.     

A clinical case of acute myelomonocytic leukemia in a Holstein cow

A 2-year, 3-month-old Holstein cow presented with anorexia and enlarged superficial lymph nodes. Fine needle aspiration cytology of the superficial lymph nodes revealed large blast cells. Hematological examination revealed anemia, neutropenia, and blast cells in peripheral blood. Blast cells were the predominant cell type in bone marrow aspirates. Of the non-erythroid cells, 26%, 58%, and 18% were positive for myeloperoxidase, α-naphthyl acetate esterase, and naphthol AS-D chloroacetate esterase, respectively. Pathological examination revealed the proliferation of neoplastic cells, which were positive for monocytic markers, in the affected lymph nodes. The cow was diagnosed with acute myelomonocytic leukemia based on these findings. This report highlights the importance of performing bone marrow aspiration cytology and cytochemical staining when diagnosing bovine myeloid leukemia.     

Induction of lymphokine-activated killer cells of equine origin: specificity for equine target cells.

The in vitro stimulation of peripheral blood mononuclear cells (PBMC) with interleukin 2 (IL-2) results in the development of potent cytotoxic effector cells, referred to as lymphokine-activated killer (LAK) cells. LAK cells are capable of lysing a wide variety of autologous, allogeneic and xenogeneic tumor cells. The exact mechanism of target cell recognition by LAK cells remains unknown. LAK cell activity has been reported for a variety of domesticated species except the horse. We report here that IL-2-stimulated equine PBMC, which fail to lyse either human or murine tumor cell lines, exhibit potent cytolytic activity against an equine tumor cell line, EqT8888. Cytolytic activity against the EqT8888 cells required 3 days of incubation with IL-2, was mediated primarily by T-cells, and was not restricted by major histocompatibility complex antigens. Though LAK activity could only be demonstrated using equine-derived target cells, xenogeneic targets could be lysed in a lectin-mediated cytotoxicity assay. The xenogeneic targets also failed to block LAK cell-killing of the EqT8888 cells in a cold-target competition assay. These results indicate that LAK cells in the horse appear to utilize a species-specific recognition mechanism during target cell lysis.     

Acute myelomonocytic leukemia with neurologic manifestations in the dog

A dog was presented with mandibular paralysis, photophobia, and diffuse atrophy of the cranial skeletal muscles. Physical examination also revealed glossal paralysis, reduction of the swallowing reflex, reduction of the pupillary light response, and generalized lymphadenopathy. Cytologic and ultrastructural examinations of blood films, bone marrow, and lymph node aspirates were consistent with acute myelomonocytic leukemia. Post-mortem examination revealed extensive, multisystemic neoplastic infiltration with marked involvement of the central and peripheral nervous systems, especially the cranial and lumbar spinal nerves and associated ganglia. Neurologic manifestations are unusual in acute myelomonocytic leukemia in the dog.     

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.     

Lymphosarcoma and associated immune-mediated hemolytic anemia and thrombocytopenia in horses

Three horses with equine lymphosarcoma were examined because of clinical signs including chronic weight loss, respiratory distress, peripheral edema, and chronic colic. Clinicopathologic findings included evidence of an immune-mediated hemolytic anemia. Immune-mediated thrombocytopenia also was diagnosed in 1 of the horses and suspected in another. One horse died in spite of treatment, 1 died 5 hours after surgical removal of a tumor encircling the jejunum, and 1 was euthanatized because of deteriorating condition. Necropsy of each horse revealed extensive neoplastic infiltration of peripheral lymph nodes and abdominal or thoracic viscera with neoplastic lymphocytes.     

Pathological findings of two types of lymphoid malignancy in sheep inoculated with bovine leukemia virus

Different types of lymphoid malignancy were observed in two sheep inoculated with BLV-containing materials. Sheep 1 showed severe leukemic change in the peripheral blood and splenomegaly but lymphosarcoma in the lymph nodes was absent. Sheep 2 had lymphosarcoma in the lymph nodes and various organs. Neoplastic cells had B-cell marker in both cases and a few neoplastic cells contained intracytoplasmic IgM in sheep 2. It was presumed that B-cells might be transformed into neoplastic cells on the way of their differentiation. Some of neoplastic cells might have ability of immunoglobulin-production in sheep 2.