Immunohistochemical Characteristics of Spindle Cell Ameloblastic Carcinoma in a Horse.

A 2-year-old male Thoroughbred horse presented with a mass in the maxilla. The focally ulcerated mass, approximately 8 cm in diameter, covered the upper left intermediate and corner incisor teeth (nos. 602 and 603 according to the modified Triadan system) and radiographic examination revealed displacement and lysis of the incisors. Histologically, the tumour was composed of a dense proliferation of spindle-shaped cells and neoplastic odontogenic epithelial cells arranged in island, follicular, plexiform or sheetlike patterns. The spindle-shaped cells were immunopositive for cytokeratins AE1/AE3, 5/6, 14 and 19. The Ki-67 index was 32.6% in the spindle cell component. Based on the histological and immunohistochemical findings, the tumour was diagnosed as spindle cell ameloblastic carcinoma.

Harringtonine Ester Derivatives with Enhanced Antiproliferative Activities against HL-60 and HeLa Cells.

Harringtonine (HT), produced from Cephalotaxus species, is known to exhibit potent antiproliferative activity against myeloid leukemia cells by inhibiting protein synthesis. A previous study using acute promyelocytic leukemia (HL-60) cells raised the possibility that the C-5' methyl group of HT plays an important role in regulating leukemia cell line antiproliferative activity. In order to investigate the effect of hydrocarbon chains at C-5' on the resultant activity, the C-5' methyl group was replaced with various straight- and branched-chain hydrocarbons using the corresponding alcohols, and their antiproliferative activity against HL-60 and HeLa cells was investigated. As a result, 4'-n-heptyl-4'-demethylharringtonine (1f, n-heptyl derivative) showed the most potent cytotoxicity among the HT ester derivatives produced, with IC50 values of 9.4 nM and 0.4 µM for HL-60 and HeLa cells, respectively. Interestingly, the cytotoxicity of derivative 1f against HL-60 and HeLa cells respectively was ∼5 (IC50 = 50.5 nM) and ∼10 times (IC50 = 4.0 µM) those of HT and ∼2 (IC50 = 21.8 nM) and ∼4 times (IC50 = 1.7 µM) more than homoharringtonine (HHT). These results demonstrate the potential of the derivative 1f as a lead compound against leukemia.

A monoclonal antibody-based indirect competitive enzyme-linked immunosorbent assay to quantify swertiamarin and related compounds in Swertia japonica Makino.

INTRODUCTION: Swertia japonica Makino (S. japonica) has a long history of use as a folk medicine, and it is one of the three essential Japanese folk medicines. S.japonica has been reported to have various biological activities. The biologically active secoiridoid glycoside swertiamarin (SM) has been isolated from S. japonica. The efficacy of this plant is attributed to SM and related secoiridoid glycosides. To control the quality of S. japonica for medicinal use, a method for the determination of SM and other secoiridoid glycosides in the plant is needed. OBJECTIVE: To produce an anti-SM monoclonal antibody (MAb) and develop an indirect competitive enzyme-linked immunosorbent assay (icELISA) for S. japonica standardisation and quality control. METHODOLOGY: SM was conjugated to cationised bovine serum albumin (cBSA), and the SM-cBSA conjugate was used to immunise BALB/c mice. Splenocytes from the immunised mice were then fused with SP2/0 myeloma cells to produce hybridoma cells that expressed anti-SM MAb. RESULTS: The developed icELISA was sufficiently sensitive and had a quantitative range of 0.78 to 12.5 µg/mL. Coefficients of variation below 10% indicated good repeatability. Recoveries in a spike and recovery assay ranged from 91.84% to 115.50%, which confirmed that the icELISA was accurate. The SM content measured using the icELISA was in agreement with the results of a high-performance liquid chromatography-ultraviolet (HPLC-UV) assay. CONCLUSION: The icELISA is suitable for the high-throughput analysis of SM and other secoiridoid glycosides in S. japonica. The method is fast, economical, and reliable for S. japonica quality control.

Capillaria hepatica (Calodium hepaticum) infection in a horse: a case report.

BACKGROUND: Capillaria hepatica is a zoonotic parasite in humans and animals and has a worldwide distribution. However, infections in mammals apart from rodents, which are natural hosts of the parasite, have rarely been reported. This report describes the first known case of C. hepatica infection in a horse in Japan. CASE PRESENTATION: A 3-year-old filly without clinical signs was presented at a slaughterhouse in Japan. Gross examination revealed white to tan nodules 0.5 to 1.5 cm in diameter in the parenchyma of the liver. Histologically, the nodules had mature fibrous capsules and consisted of multifocal to coalescing granulomatous inflammations with numerous nematode eggs. The eggs were barrel shaped with an opercular plug on each end and double-layered shells; these findings are consistent with the features of C. hepatica eggs. CONCLUSIONS: To our knowledge, this is the first case of C. hepatica infection in a horse in Japan. The pathological findings confirmed the presence of this pathogen in this part of the world, and they highlight the importance of this nematode in the differential diagnosis of hepatic granulomatous lesions in horses.

Cardiac pathology and molecular epidemiology by avian leukosis viruses in Japan.

Epidemiological studies suggest that retroviruses, including human immunodeficiency virus type 1, are associated with cardiomyopathy and myocarditis, but a causal relationship remains to be established. We encountered unusual cardiomyocyte hypertrophy and mitosis in Japanese native fowls infected with subgroup A of the avian leukosis viruses (ALVs-A), which belong to the genus Alpharetrovirus of the family Retroviridae and mainly induce lymphoid neoplasm in chickens. The affected hearts were evaluated by histopathology and immunohistochemistry, viral isolation, viral genome sequencing and experimental infection. There was non-suppurative myocarditis in eighteen fowls and seven of them had abnormal cardiomyocytes, which were distributed predominantly in the left ventricular wall and showed hypertrophic cytoplasm and atypical large nuclei. Nuclear chains and mitosis were frequently noted in these cardiomyocytes and immunohistochemistry for proliferating cell nuclear antigen supported the enhancement of mitotic activity. ALVs were isolated from all affected cases and phylogenic analysis of envSU genes showed that the isolates were mainly classified into two different clusters, suggesting viral genome diversity. In ovo experimental infection with two of the isolates was demonstrated to cause myocarditis and cardiomyocyte hypertrophy similar to those in the naturally occurring lesions and cardiac hamartoma (rhabdomyoma) in a shorter period of time (at 70 days of age) than expected. These results indicate that ALVs cause myocarditis as well as cardiomyocyte abnormality in chickens, implying a pathogenetic mechanism different from insertional mutagenesis and the existence of retrovirus-induced heart disorder.

Epidemiological study of fowl glioma-inducing virus in chickens in Asia and Germany.

Fowl glioma-inducing virus (FGV), which belongs to avian leukosis virus (ALV) subgroup A, induces fowl glioma. This disease is characterized by multiple nodular gliomatous growths of astrocytes and has been previously reported in Europe, South Africa, Australia, the United States and Japan. FGV and FGV variants have spread to ornamental Japanese fowl, including Japanese bantams (Gallus gallus domesticus), in Japan. However, it is unclear how and where FGV emerged and whether FGV is related to the past fowl glioma in European countries. In this study, the prevalence of FGV in European, Asian and Japanese native chickens was examined. FGV could not be isolated from any chickens in Germany and Asian countries other than Japan. Eighty (26%) out of 307 chickens reared in Japan were positive by FGV-screening nested polymerase chain reaction and 11 FGV variants with an FGV-specific sequence in their 3' untranslated region were isolated. In addition, four other ALVs lacking the FGV-specific sequence were isolated from Japanese bantams with fowl glioma and/or cerebellar hypoplasia. These isolates were considered to be distinct recombinant viruses between FGV variants and endogenous/exogenous avian retroviruses. These results suggest that the variants as well as distinct recombinant ALVs are prevalent among Japanese native chickens in Japan and that FGV may have emerged by recombination among avian retroviruses in the chickens of this country.

Molecular characteristics and pathogenicity of an avian leukosis virus isolated from avian neurofibrosarcoma.

Peripheral nerve sheath tumors (PNSTs) are rare in chickens and their etiology remains to be elucidated. In this study, a naturally occurring PNST in a Japanese native fowl (Gallus gallus domesticus) was pathologically examined and the strain of avian leukosis virus (ALV) isolated from the neoplasm was characterized by molecular biological analysis. The fowl presented with a firm subcutaneous mass in the neck. The mass, connected to the adjacent spinal cord (C9-14), was microscopically composed of highly cellular tissue of spindle cells arranged in interlacing bundles, streams, and palisading patterns with Verocay bodies and less cellular tissue with abundant collagen. Immunohistochemically, neoplastic cells were divided into two types: perineurial cells positive for vimentin, glucose transporter 1 (GLUT1), and claudin1; and Schwann cells positive for vimentin, occasionally positive for S-100 alpha/beta but negative for GLUT1. Based on these findings, a diagnosis of neurofibrosarcoma was made. The complete nucleotide sequence of an ALV strain, CTS_5371, isolated from the neoplasm was determined and phylogenetic analysis indicated that the strain was a novel recombinant virus from avian leukosis/sarcoma viruses previously reported. Additionally, experimental infection revealed that CTS_5371 induced the proliferation of Schwann cells and perineurial cells. These results suggest that this ALV strain has the ability to induce PNSTs in chickens.

Pathogenicity of avian leukosis viruses related to fowl glioma-inducing virus.

Fowl glioma-inducing virus (FGV), which belongs to avian leukosis virus subgroup A, causes the so-called fowl glioma and cerebellar hypoplasia in chickens. In the present study, the complete nucleotide sequences of four isolates (Tym-43, U-1, Sp-40 and Sp-53) related to the FGV prototype were determined and their pathogenicity was investigated. Phylogenetic analysis showed that the 3'-long terminal repeat of all isolates grouped together in a cluster, while sequences of the surface (SU) proteins encoded by the env gene of these viruses had 85 to 96% identity with the corresponding region of FGV. The SU regions of Tym-43, U-1 and FGV grouped together in a cluster, but those of Sp-40 and Sp-53 formed a completely separate cluster. Next, C/O specific-pathogen-free chickens were inoculated in ovo with these isolates as well as the chimeric virus RCAS(A)-(FGVenvSU), constructed by substituting the SU region of FGV into the retroviral vector RCAS(A). The four variants induced fowl glioma and cerebellar hypoplasia and the birds inoculated with Sp-53 had the most severe lesions. In contrast, RCAS(A)-(FGVenvSU) provoked only mild non-suppurative inflammation. These results suggest that the ability to induce brain lesions similar to those of the FGV prototype is still preserved in these FGV variants.

A recombinant avian leukosis virus associated with fowl glioma in layer chickens in Japan.

Fowl glioma is characterized by multiple nodular growth of astrocytes, and fowl glioma-inducing virus belonging to avian leukosis virus has been isolated from Japanese bantam as a causal agent. Subcutaneous neoplasms of the head and neck have been reported in layer chickens since 2003 in Japan, and fowl glioma concurred in these affected layers. In the present study, the histopathology of 240 layers, including 18 layers with subcutaneous neoplasms and 222 layers kept with the affected layers, was performed to clarify the characteristics of fowl glioma in layers. Microscopically, 103 layers showed non-suppurative encephalitis, and four layers had locally extensive proliferation or multiple nodules of astrocytes. Gliomas concurred in 11 layers with subcutaneous neoplasms and occurred independently in three layers. In addition, two layers had locally extensive proliferation of small, round cells in the cerebrum. The fowl glioma-inducing virus genome was not detected in the affected brains by nested polymerase chain reaction. Ten isolates were obtained from the affected brains. By nucleotide sequencing of the env gene, SU coding regions of these isolates were most closely related to myeloblastosis-associated virus-like viruses, but TM regions showed the highest similarity to endogenous viral (ev) loci. The genome of one isolate mainly consisted of ev loci and contained several parts of other avian leukosis/sarcoma viruses. These results show that the causal avian leukosis virus of fowl glioma is not just fowl glioma-inducing virus and that different avian leukosis virus strains having oncogenicity in the central nervous system by recombination are spread in layers in Japan.