Megakaryocytes with intracytoplasmic blood cells

The phenomenon of megakaryocytes containing intracytoplasmic blood cells was frequently observed in the bone marrow of rats. The frequency appeared to increase in inflammatory or neoplastic conditions. The types of blood cells engulfed were mainly neutrophils. For the first time, megakaryocytes also were found to be engulfed.     

Type of smear may influence thrombopoietic cell counts in the bone marrow of clinically healthy dogs

BACKGROUND: The expected number of thrombopoietic cells in normal canine bone marrow is poorly defined and there is no consensus on the most appropriate way to prepare cytologic smears to evaluate these cells nor on the optimum method for their quantification. OBJECTIVE: The purpose of this study was to determine total and differential counts of thrombopoietic cells in the bone marrow of clinically healthy Beagle dogs by comparing 4 different smear types and bone marrow core biopsies. METHODS: Twenty-two clinically healthy, male Beagle dogs, 10 to 12 months old, were used in the study. Following bone marrow aspiration and core biopsy from the iliac crest, Giemsa-stained smears were prepared by 4 techniques: drop-squash, particle-squash, buffy coat, and fat-layer smears. Thrombopoietic cells were counted in up to 100 low-power fields (LPF, X10 objective) in the aspiration smears and in all possible high-power fields (HPF, X40 objective) in H&E-stained biopsy sections. RESULTS: Mean total thrombopoietic cell counts were 2.76 cells/LPF (drop-squash), 1.55 cells/LPF (particle-squash), 8.05 cells/LPF (buffy coat), and 3.08 cells/LPF (fat-layer). Core biopsies yielded 5.31 cells/HPF but frequently failed to provide interpretable specimens. There was a significant difference in cell counts among the 4 smear types (P <.001). Based on evaluation of buffy coat smears, thrombopoietic cells included 1.23% megakaryoblasts, 8.77% promegakaryocytes, and 90% megakaryocytes, with a mean maturation index of 0.11. CONCLUSION: Thrombopoietic cell counts in canine bone marrow are influenced by the smear technique. Buffy coat and fat-layer smears may be useful to obtain cellular smears in hemodiluted or small aspirate samples.     

Ultrastructural analysis of platelets and megakaryocytes from a dog with probable essential thrombocythemia

Blood platelets and bone marrow megakaryocytes from a dog with essential thrombocythemia were analyzed by transmission electron microscopy and compared with those of control dogs. Platelets varied in size and shape and contained enlarged and dilated open canalicular systems. Megakaryocytes were dysmorphic and had evidence of abnormal maturation, with large numbers of megakaryoblasts containing alpha granules and poorly organized and reduplicated demarcation membranes. The fewer, more mature megakaryocytes lacked normal cytoplasmic organization. Most notable was the absence of well-demarcated "platelet fields," due to the excessive and disorderly array of demarcation membranes.     

Scanning electron microscopic studies of megakaryocytes and platelet formation in the dog and rat

Megakaryocyte morphology and platelet formation in canine and murine bone marrows were studied by scanning electron microscopy. In situ-fixed bone marrow preparations and cell suspensions of bone marrow provided complementary information for the 2 species (dogs and rats). Cylindrical processes (proplatelets) of variable length and thickness, originating from the megakaryocyte surface, were in the larger marrow sinusoids and the central vein. Regional constrictions along the length of proplatelets, particularly near their apical region, and the presence of fragments of such processes supported the concept of platelet formation through segmentation of proplatelets. Megakaryocytes presented varied morphology. Surface features resembling platelets were observed on megakaryocytes, indicating that platelets may have been released through surface budding. In conclusion, megakaryocytes formed long proplatelet processes that actively migrated to venous sinusoids to release platelets by fragmentation. Scanning electron microscopy analysis revealed a complex and variable megakaryocyte surface topography. The platelet-like structures on megakaryocyte surfaces may represent platelet release by a budding mechanism. The similarity between murine platelet release and canine platelet release demonstrates that data from rodent models may be applicable to nonrodents.     

Thrombocytosis and central nervous system involvement in a case of canine acute megakaryoblastic leukemia

This case report presents a 14‐month‐old female Poodle mix with acute megakaryoblastic leukemia based on a marked thrombocytosis, abnormal platelet morphology, circulating dwarf megakaryocytes, and blast cells in the blood. Bone marrow abnormalities included dysmegakaryopoiesis dygranulopoiesis, and an increased number of blast cells was observed in the blood. Extensive leukemic involvement was also found in the liver, spleen, lymph nodes, lungs, kidneys, and brain. The cytopathologic features of the abnormal circulating cells were highly suggestive of being megakaryocytic in origin, which was supported by negative myeloperoxidase staining and positive von Willebrand factor staining on immunocytochemistry (ICC). The neoplastic cells were also CD61 positive and had variable von Willebrand factor expression on ICC. Although there were only 25% blast cells in the bone marrow, which theoretically supported myelodysplastic syndrome, the hypothesis that this case represented acute myeloid leukemia of megakaryoblastic origin was confirmed by the continuous increase in circulating blast cell numbers during follow‐up visits and the extensive leukemic involvement of parenchymal organs.     

Acute megakaryoblastic leukemia with erythrophagocytosis and thrombosis in a dog

A 7-year-old, intact male Dachshund was presented to the Lyon veterinary school for lethargy and anorexia of several weeks duration. The main clinical signs were pale and icteric mucous membranes, hepatomegaly, splenomegaly, and lymphadenopathy. Results of a CBC and plasma biochemistry tests revealed severe nonregenerative anemia, thrombocytopenia, and increased alanine aminotransferase and alkaline phosphatase activities. Blood smear evaluation and cytologic examination of lymph node and bone marrow aspirate specimens revealed a large population of poorly differentiated blast cells with morphologic features suggesting megakaryocytic lineage. A low number of well-differentiated but dysplastic megakaryocytes also were observed in lymph node and bone marrow smears. A few blast cells were erythrophagocytic. Blast cells were positive for glycoprotein IIIa, factor VIII-related antigen, and factor XIII using immunocytochemistry. The dog was euthanized and necropsied. Histologic findings consisted of diffuse, massive infiltration of lymph nodes, liver, and spleen by megakaryoblasts and atypical megakaryocytes, with widespread thrombosis. This case confirms the usefulness of immunochemistry, including for factor XIII, in the diagnosis of megakaryoblastic leukemia, and demonstrates the unique features of tumor cell erythrophagocytosis and marked fibrinous thrombosis, which have not been reported previously in dogs.     

不同周龄SPF级SD大鼠胰腺自发性病变的组织学观察

研究不同周龄SPF级SD大鼠胰腺自发性病变的种类及其病变发生率,为药物安全性评价提供背景资料。收集3年安评试验中11、19、31周龄试验对照组大鼠胰腺制作病理切片,光学显微镜下观察SD大鼠胰腺病变的种类及组织形态学特点,并统计其病变发生率。结果显示,大鼠胰腺主要出现了以下病变:①单核细胞浸润:11周龄大鼠该病变的总体发生率为0.6%,其中雌性为0,雄性为1.25%;19周龄大鼠该病变的总体发生率为1.0%,其中雌性为1.0%,雄性为1.0%;31周龄大鼠该病变的总体发生率为1.0%,其中雌性为0,雄性为1.96%。②腺泡细胞空泡变性:11周龄大鼠未观察到该病变的发生;19周龄大鼠该病变的总体发生率为1.0%,其中雌性为0,雄性为2.0%;31周龄大鼠该病变的总体发生率为3%,其中雌性为2.1%,雄性为3.9%。③腺泡细胞萎缩、腺管增生:11周龄大鼠未观察到该病变的发生;19周龄大鼠该病变的总体发生率为0.5%,其中雌性为0,雄性为1.0%;31周龄大鼠该病变的总体发生率为3.0%,其中雌性为1.0%,雄性为4.9%。结果表明,SPF级大鼠胰腺可发生单核细胞浸润、腺泡细胞空泡变性、腺泡细胞萎缩及腺管增生等自发性病变,且病变发生率可随着年龄的增加而增加。     

Scanning electron microscope study of platelet release by canine megakaryocytes in vitro

Megakaryocytes were isolated from bone marrow from healthy dogs, using a combination of density-gradient centrifugation and polysucrose-velocity sedimentation techniques. The 2-step separation technique resulted in a preparation comprising 30% to 35% megakaryocytes of total nucleated cells. Accessibility to large numbers of viable canine megakaryocytes allowed investigation of platelet release by these cells in short-term cultures. Megakaryocytes were observed to form long cytoplasmic processes that gradually developed segmental constrictions and subsequently fragmented into platelet-sized pieces. Some platelet-sized cytoplasmic pieces of megakaryocytes presumably underwent discoid transformation.     

Bone marrow failure in a dog

A 7-month old Boxer bitch with lethargy and inappetence of several days' duration was found to have pancytopenia. A bone marrow aspirate contained many lymphocytes and immature myeloid cells but few erythrocyte precursors; marrow phagocytes appeared active and megakaryocytes were immature. Circumstantial evidence suggested that the cause of marrow failure was prior administration of thiacetarsamide, an organic arsenical. Recovery was spontaneous and within four weeks the haemo-gram was normal, except that platelet numbers were not fully restored.
The bitch was examined 6 months later because of a recurrence of signs, with several syncopic episodes during the preceding week. A severe non-regenerative anaemia was present, with absence of erythroid precursors from bone marrow. Neutrophil and platelet counts were normal. The cause of the erythrocyte aplasia was not determined. The dog was given blood transfusions, oxymetholone and prednisolone but died after one month. A post-mortem marrow sample contained many erythroid cells, some with morphological abnormalities suggesting dyserythropoiesis.     

Canine distemper virus-induced thrombocytopenia

Effects of canine distemper virus (CDV) infection on circulating platelet values were studied in gnotobiotic dogs inoculated with R252-CDV. Thrombocytopenia (less than 200,000 platelets/microliter) was present on postinoculation day (PID) 5 and persisted through PID 15. Peak thrombocytopenia occurred on PID 10 (less than 85,000 platelets/microliter). Thrombocytopenia was accompanied by lymphopenia, neutropenia, and monocytopenia. Platelet membrane-bound CDV antigen and IgG were present from PID 7 onward; neither the third component of complement nor IgM was detected on platelets from CDV-inoculated dogs. The mean number of megakaryocytes per unit of bone marrow surface area was unchanged. Megakaryocyte infection was present in dogs euthanatized on PID 4 (0.33%), increased slightly in dogs euthanatized on PID 8 (3%), and increased sharply in dogs euthanatized on PID 9 and 10 to 17.8% and 8.3%, respectively. Phagocytosis of platelets by stellate reticuloendothelial (Kupffer's) cells in the liver was prominent in dogs euthanatized from PID 5 onward. Seemingly, CDV-induced thrombocytopenia was mediated by virus-antibody immune complexes on platelet membranes. Decreased platelet production after PID 8 resulting from direct viral infection of megakaryocytes was a likely contributing factor and occurred against a background of profound virus-induced dysfunctions of all hematopoietic cellular elements.     

The influence of dietary fat and non-specific immunotherapy on carcinogen-induced rat mammary adenocarcinoma

The incidence of mammary adenocarcinoma in Sprague-Dawley female rats, caused by the carcinogen 7,12-dimethylbenz(alpha)anthracene, was influenced by the level of dietary fat fed after exposure to carcinogen. Carcinogen was given by stomach tube to 50-day-old rats, and tumors were evaluated when rats were 9 months old. Rats on diets containing 20% unsaturated fat had a tumor incidence of 97%, while rats changed to a low-fat diet (2% unsaturated fat) three or four weeks after exposure to the carcinogen had an incidence of 45%. Some rats on each diet were given two treatments with the methanol extraction residue of Bacillus Calmette-Guerin, either three and five weeks after carcinogen or four and six weeks after carcinogen. Tumor incidence in the treated group and the untreated group was the same when rats were maintained on the high-fat diet, but tumors in the treated group were larger and the disease was more severe by histological criteria. These tumors were more anaplastic and many were extensively infiltrated with lymphocytes compared to the untreated group. Tumor incidence was significantly lower in rats changed to the low-fat diet (45%) than in those on the high-fat diet (97%), and tumor incidence was reduced to 20% when rats changed to the low-fat diet were treated with methanol extraction residue. The treated group had less severe disease than the untreated group on the low-fat diet. Only half the tumor-bearing rats in this group had malignant tumors, and none were invasive. Methanol extraction residue protected most rats on the low-fat diet against mammary adenocarcinoma, and reduced the severity of disease in those rats that did develop tumors. Methanol extraction residue treatment provided no protection, and even increased the severity of disease, in rats on the high-fat diet.     

Electron microscopy of a spiral-shaped bacterium in the blood and bone marrow of a rhinoceros iguana.

Spiral shaped bacteria present in blood smears and bone marrow of a sick rhinoceros iguana were examined by light and electron microscopy. The organisms averaged 10 micron in length and had at least three spiral turns. The cell contained nuclear areas, vacuoles and ribosomes, except at the poles where there was a virtual absence of organelles. The bacterium was found by a cell membrane, cell wall and enveloping sheath. "Blebs" were present with regularity on the cell surface. About 14 flagella were present at each pole, and at these sites there was a specialized thickening of the cell membrane. Organisms were present within a phagocytic vacuole of macrophages in the blood and bone marrow, and often these engulfed organisms were degenerated. The taxonomic position of the bacterium is unknown.     

Corneal endothelial cell morphology of normal dogs in different ages

Endothelial cell function is essential to maintain corneal transparency, but unfortunately the regenerative capacity of the endothelium is limited. There are only a few reports describing the effect of age on morphologic appearance of corneal endothelial cells of dogs. Studies of normal corneal endothelial cells in humans and dogs have shown a decrease in endothelial cell density (ECD) and an increase in pleomorphism and polymegethism with advancing age. The purpose of this study was to investigate the effect of age on ECD and endothelial cell morphology in dogs. A total of 30 dogs were divided into three groups (10 dogs/group) based on age: group 1 (2-12 months old), group 2 (24-72 months old), and group 3 (84 months or older). Corneas were processed for light and scanning electron microscopy. Results showed only difference in cell density between group 1 and groups 2 and 3, showing an initial decrease in cell density as the animal matured. Whereas there was significantly greater variation in cell size within the dogs in group 3 than there was within the other two groups, suggesting that there was increased polymegethism and pleomorphism with advancing age.     

Gold-induced immune thrombocytopenia in the dog

In two seven-year studies with gold compounds in dogs of both sexes, thrombocytopenia was observed after 45 to 72 months of dosing in three of 14 and two of 14 dogs in high-dose groups that received 2.4 to 3.6 mg/kg of auranofin per day orally or 0.5 to 2.0 mg/kg of gold sodium thiomalate intramuscularly once every three days, respectively. An immune basis for the disorder was suggested by the apparent consumptive nature of the thrombocytopenia (increased bone marrow megakaryocytes and large peripheral blood platelets), the response to corticosteroid therapy and the demonstration of increased platelet-associated immunoglobulin. The latter was demonstrated with a solid phase radioimmunoassay and by electron microscopy using a staphylococcal protein A-colloidal gold conjugate. Platelet-associated immunoglobulin decreased as the platelet counts rose, and in one dog monitored over periods of steroid-induced remissions and subsequent relapses, the amount of platelet-associated immunoglobulin G correlated inversely with the platelet count (r = 0.82). These findings suggest that the long-term administration of gold compounds in dogs is associated with a dose-dependent incidence of thrombocytopenia, which is immune-mediated and similar to that associated with parenteral chrysotherapy in man. The application of tests for platelet-associated immunoglobulin to canine patients with immune thrombocytopenia should be useful in the diagnosis of the disorder in clinical practice.     

高氟低碘对老年大鼠肺脏细胞DNA损伤的影响

研究高氟低碘对老年大鼠肺脏细胞DNA损伤的影响。健康离乳Wistar大鼠21只,随机分为4组：对照组（5只）,饲喂大鼠标准日粮,饮自来水;高氟组（5只）,饲喂大鼠对照日粮,饮100 mg/L氟化钠（NaF）去离子水;低碘组（5只）,饲喂低碘日粮（定做）,饮去离子水;高氟低碘组（6只）,饲喂低碘日粮,饮100 mg/L氟化钠（NaF）去离子水。在大鼠20月龄时,处死采取肺脏组织,检查其DNA的损伤情况。高氟组、低碘组和高氟低碘组的老年大鼠肺细胞拖尾率比对照组显著增高（P＜0.01）,高氟低碘组大鼠肺细胞拖尾率比高氟组、低碘组高,并且均有差异显著性（分别为P＜0.01和P＜0.05）;高氟组、低碘组和高氟低碘组老年大鼠肺细胞的拖尾长度比对照组显著增长,显著性差异分别为P＜0.01、P＜0.05 和P＜0.01。高氟低碘组大鼠肺细胞的拖尾长度比高氟组及低碘组增长,与低碘组有差异显著性（P＜0.01）,但与高氟组差异不显著（P＞0.05）;高氟组和高氟低碘组老年大鼠肺细胞DNA损伤主要是Ⅲ级损伤（Ⅲ级分别为70.00%和79.31%）。低碘组的肺细胞DNA损伤进入Ⅱ级和Ⅲ级范围的比较多,并且Ⅲ级占有率较高（Ⅱ级为31.25%;Ⅲ级为40.62%）。结果表明,高氟、低碘、高氟低碘都影响了大鼠肺脏细胞的DNA损伤。     

Detection of feline immunodeficiency virus infection in bone marrow of cats.

Natural or experimental feline immunodeficiency virus (FIV) infection in cats is often associated with hematologic abnormalities which are similar to those observed in human immunodeficiency virus (HIV) infected patients. To determine if cells in bone marrow are infected with FIV and whether severity of hematopoietic disorder is correlated with the level of viral infection, bone marrow tissues from ten experimentally and two naturally FIV infected cats were examined by in situ hybridization for presence of FIV RNA. Seven of the 12 FIV infected cats were also naturally or experimentally coinfected with feline leukemia virus (FeLV). FIV RNA was detected mainly in megakaryocytes and unidentified mononuclear cells in the bone marrow of cats that were sick and had marrow hypercellularity and immaturity. These included all cats in the acute phase of FIV infection and two of seven long term FIV infected cats. One long term FIV infected cat with lymphosarcoma was also positive for FIV RNA in bone marrow cells. The other four long term FIV infected cats were relatively healthy, with normal bone marrow morphology, and were negative for FIV infected cells. Bone marrow from three non-infected and two cats infected with FeLV alone were also negative for FIV RNA by in situ hybridization. We concluded that megakaryocytes and mononuclear cells were targets of the viral infection and that the presence of FIV RNA in cells of the bone marrow correlated with marrow hypercellularity and immaturity, and severity of illness.     

The morphological effects of vincristine sulfate on canine bone marrow cells

This study documents the morphologic changes observed in the bone marrow aspirate biopsies from dogs 6 and 24 hours after receiving a single therapeutic dose (0.025 mg/kg) of vincristine sulfate (Oncovin: Eli Lilly & Co., Indianapolis, Ind.) intravenously. The most striking cytologic changes were observed in the erythroid cell line. Abnormalities included increased numbers of mitotic figures, abnormal nuclear configurations, and fragmented nuclei. Erythroid cells in metaphase were prominent in marrow samples collected 6 hours post-vincristine, accounting for a mean of 27% of all erythroid precursors. Fragmented nuclei and atypical nuclear configurations were seen in low numbers (mean = 7%) of erythroid cells from these animals. In contrast, marrow collected from dogs 24 hours post-vincristine exhibited low numbers (mean = 1%) of erythroid cells in metaphase, but erythroid cells with atypical nuclear configurations and fragmented nuctei accounted for a mean of 41% of the erythroid cells present. Less dramatic increases in the number of mitotic non-erythroid cells were seen 6 hours post-vincristine (mean = 5% of non-erythroid cells) and 24 hours post-vincristine (mean = 1% of non-erythroid cells). Only rare nuclear fragmentation was observed in these cell lines. Significant alterations in megakaryocytes and myeloid to erythroid (M:E) ratios were not observed in samples taken 6 hours post-vincristine; however, M:E ratios were considerably higher in three of the four samples taken from dogs 24 hours post-vincristine. Similar time-related changes were observed in four clinical cases in which bone marrow aspirates were performed after vincristine administration.     

Epididymal interstitial (Leydig) cell tumors in B6C3F1 mice

Six primary interstitial cell tumors of the epididymis were identified from 46,752 male B6C3F1 mice used in chronic toxicity and carcinogenicity studies. Five of the tumors occurred at the end of 2-year studies; none were attributed to treatment. None of the mice with epididymal tumors had a primary testicular tumor. Histologically, tumors were characterized by a nodular or diffuse proliferation of tumor cells in the epididymal interstitium. Most cells were polygonal with highly vacuolated cytoplasm (vacuolated cells) or eosinophilic cytoplasm (eosinophilic cells). Smaller hyperchromatic cells with scant basophilic cytoplasm (basophilic cells) and cells with yellow-brown pigment characteristic of lipofuscin (pigmented cells) were less common. In each tumor two or more cell types were present. Extension of these tumors through the capsule, invasion of the testis, or metastasis did not occur. By electron microscopy both eosinophilic and vacuolated cell types had a large round or oval nucleus with sparse heterochromatin, abundant mitochondria with tubulovesicular cristae, and frequent desmosome structures between cell membranes. Vacuolated cells contained numerous lipid droplets. Morphological features of the epididymal tumors are similar to those of the testicular interstitial (Leydig) cell tumor in mice and rats.     

Cytologic evaluation of primary and secondary myelodysplastic syndromes in the dog

Myelodysplastic syndromes are a heterogeneous group of acquired primary and secondary alterations of hematopoietic stem cells that result in cytopenias in blood and cytologic features of dysplasia in blood and/or bone marrow. To better understand the cytologic features that would permit differentiation of primary and secondary forms of myelodysplasia, we reviewed 267 consecutive bone marrow reports from dogs. These reports indicated that 34 dogs (12.7%) had dysgranulopoiesis, dyserythropoiesis, and/or dysthrombopoiesis in >10% of granulopoietic cells, erythroid cells, and/or megakaryocytes, respectively. Thirteen dogs had primary myelodysplastic syndromes, and 21 had secondary myelodysplastic syndromes. Of the 13 dogs with primary myelodysplasia, 4 were subclassified as myelodysplastic syndrome with refractory anemia (MDS-RA), and 9 were subclassified as myelodysplastic syndrome with excess blasts (MDS-EB). Secondary conditions associated with dysplasia in the bone marrow included malignant lymphoma (n = 5), myelofibrosis (n = 3), immune-mediated thrombocytopenia (n = 4), immune-mediated hemolytic anemia (n = 5), multiple myeloma with melphalan administration (n = 1), pyometra with estrogen administration (n = 1), polycythemia vera (n = 1), and thrombopathia (n = 1). MDS-RA was characterized by <5% myeloblasts in bone marrow, normal granulocyte maturation ratio, increased erythroid maturation ratio, and dysplastic changes in >15% of erythroid cells. MSD-EB was characterized by >/=5% myeloblasts in bone marrow, high granulocyte maturation and erythroid maturation ratios, >/=32% dysplastic granulocytes, and the presence of small atypical immature myeloid cells. Secondary myelodysplastic syndromes were characterized by <5% myeloblasts in bone marrow, variable granulocyte maturation and erythroid maturation ratios, and variable dysplastic features. These results indicate that morphology alone cannot be used to distinguish primary and secondary myelodysplastic syndromes in dogs.     

Surface parasitism of the fish mycoplasma Mycoplasma mobile 163 K on tracheal epithelial cells

The interaction of Mycoplasma mobile 163 K with epithelial cells was investigated in tracheal organ cultures of gnotobiotic rats by electron microscopy. Ultrathin sections showed that the epithelial cells were heavily colonized and damaged by the mycoplasmas 2 days after infection. Numerous organisms were attached to fragments of the cytoplasmic membranes; others entered the injured cells and adhered to cell structures. Organisms were interconnected by fibrillary material supporting their adherence to the cell surface. Non-infected tracheal rings remained viable, differentiated and well organized up to 28 days of cultivation. The present investigation characterizes M. mobile 163 K as a surface parasite.