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Biomedical subjects

R Goitsuka

Publications and source records attributed to R Goitsuka.

At least 37 records · Page 2Linked to original sources
PubMed1996

Detection of apoptosis induced in peripheral blood lymphocytes from cats infected with feline immunodeficiency virus.

Peripheral blood lymphocytes (PBL) from cats infected with feline immunodeficiency virus (FIV) were examined for the occurrence of apoptosis after short-term culture. In the PBL from FIV-infected cats, changes in flow-cytometry scattergram, morphological characteristics of apoptosis and nucleosomal DNA fragmentation were observed. Percentages of apoptotic cells by flowcytometry analysis in PBL from FIV-infected cats (22.4% +/- 9.4%) were significantly higher than those in PBL from uninfected control cats (9.2% +/- 3.5%). The lymphocytes which underwent apoptosis included CD5+, CD4+ and sIgM+ cells, indicating that induction of apoptosis was not restricted to a special subset of lymphocytes. These findings provide evidence of the apoptotic state of PBL in cats with FIV infection.

Animals↗
PubMed1996

Molecular cloning of bovine mb-1 cDNA.

Ig-alpha of the B-cell antigen receptor complex forms a heterodimeric structure with Ig-beta on the plasma membrane of B-lymphocytes and is apparently involved in signal transduction during the activation of B-cells. Bovine leukemia virus (BLV) is predominantly a B-cell tropic retrovirus, which induces persistent lymphocytosis and leukemia/lymphoma of B-cell lineage in cattle. To understand the mechanisms of proliferation and tumorigenesis of bovine B-cells that are associated with BLV infection, we investigated the B-cell antigen receptor complex, especially bovine mb-1 encoding the bovine Ig-alpha protein. We isolated a full-length bovine mb-1 cDNA clone encoding 223 amino acid residues. The deduced amino acid sequence of the bovine mb-1 showed extensive homology with those of human and murine mb-1. The cytoplasmic tail of the bovine mb-1 also contained a consensus motif (D/E-X7-D/E-X2-L/I-X7-Y-X2-L/I) that may interact with the SH2 domain of src-type kinase. Interestingly, a similar consensus sequence motif was found in the BLV gp30env, although the overall sequence similarity between bovine mb-1 and BLVgp30 was not significant. Furthermore, elevated levels of mb-1 transcript were detected in various bovine leukemia/lymphoma cell lines. These results indicated that the proliferation of B-cells associated with BLV-infection may be related to abnormal signal transduction through the B-cell antigen receptor complex.

Amino Acid Sequence↗
PubMed1996

Elevation of serum G-CSF level in horses with transportation-induced fever.

Levels of granulocyte-colony stimulating factor (G-CSF) in the blood of horses were measured before and after a long-distance transportation to clarify the pathogenesis of transportation-induced fever. The serum G-CSF level was measured by its ability to stimulate growth in a mouse myeloblastic cell line, NFS-60. Of 26 horses transported for a long distance, 9 had fever more than 39.0 degrees C during or after transportation. After transportation, the serum G-CSF level significantly increased in horses with transportation-induced fever but not in those without fever, and the serum G-CSF level correlated positively with the peak body temperature and with an increase in peripheral white blood cell count. These data indicate that microbial infection, which is closely related to the elevation of the serum G-CSF levels, is the causative factor of transportation-induced fever.

Animals↗
PubMed1995

Molecular cloning of equine interleukin-1 alpha and -beta cDNAs.

Equine interleukin-1 alpha (IL-1 alpha) and IL-1 beta were molecularly cloned to establish a basis for research on inflammatory and immune responses in the horse. Equine peripheral blood mononuclear cells (PBMC) were stimulated with lipopolysaccharide (LPS), and cDNA clones of equine IL-1 alpha and IL-1 beta covering the whole coding sequences were isolated from them. These equine IL-1 alpha and IL-1 beta clones contained open reading frames encoding 271 and 269 amino acids, respectively. The deduced amino acid sequence of equine IL-1 alpha showed 71.6% and 60.2% similarity with that of human and murine IL-1 alpha, respectively. Similarly, the amino acid sequence of equine IL-1 beta showed 66.7% and 61.8% similarity with that of human and murine IL-1 beta, respectively. In both equine IL-1 alpha and IL-1 beta, amino acids at miristoylation sites were well conserved. Dot blot analysis indicated that the expression of IL-1 beta was predominant to that of IL-1 alpha in equine PBMC stimulated with LPS or phorbol myristate acetate.

Amino Acid Sequence↗
PubMed1995

Erythroleukemia in two cats naturally infected with feline leukemia virus in the same household.

Erythroleukemia was observed in two unrelated cats infected with feline leukemia virus (FeLV) from the same household. Case 1, a 1-year-old neutered male cat developed erythroleukemia (M6) after a diagnosis of myelodysplastic syndrome (MDS-Er) on the criteria of FAB classification of acute leukemias. Case 2, a 1-year-old neutered female cat, which had close contact with Case 1, also developed erythroleukemia (M6Er). In both cases, marked proliferation of erythroid progenitor cells with disproportionally large numbers of immature forms was observed in the bone marrow. In Case 1, neoplastic proliferation of myeloid cells in the bone marrow was also noted at the terminal stage. Combination chemotherapy with daunomycin was partially effective for treatment of these erythroid neoplasias, but did not induce complete remission. Southern blot analysis using exogenous FeLV-specific probes indicated the clonal origin of these hematopoietic tumor cells. Furthermore, the erythroid and myeloid tumor cells in Case 1 were shown to be derived from independent transformed clones. A variant FeLV was shown to be integrated into the tumor cells in Case 1, while a full-length FeLV was found in both cases. Because these erythroid neoplastic diseases occurred in two unrelated cats kept in the same household and these diseases are rare, they may both have been associated with the same FeLV strain.

Animals↗
PubMed1994

Two forms of the mb-1 gene transcript in cattle.

We have isolated two forms of bovine mb-1 cDNA clones from a cDNA library of a bovine leukemia virus-infected B-lymphoblastoid cell line. The nucleotide sequence analysis of these cDNA clones indicated that the shorter form of cDNA had a 119-bp deletion that precisely corresponded to the exon III domain of the human and murine mb-1 genes. This deletion would result in a frame shift, producing a premature translation termination leading to the lack of the entire transmembrane and cytoplasmic domains of the normal mb-1 molecule. The existence of two species of mRNA transcripts corresponding to the isolated cDNA clones were confirmed in all of the bovine leukemia cell lines examined by PCR with the use of the primers flanking the deletion site. Furthermore, the shorter mb-1 transcript was induced in normal PBMC by stimulation with mitogens or culture supernatants of bovine leukemia virus-infected cell lines. These findings indicate the presence of the truncated mb-1 gene product which may have some function different from that of the normal mb-1 gene product.

Alternative Splicing↗
PubMed1994

Cloning of feline p53 tumor-suppressor gene and its aberration in hematopoietic tumors.

Alterations of the p53 tumor-suppressor gene have been observed in a variety of human and mouse tumors. For investigation of the role of this gene in tumors of cats, feline p53 cDNA was molecularly cloned from a feline lymph-node cDNA library. The cloned cDNA (FF53) contained the whole open reading frame of p53 gene encoding 386 amino acids. The amino-acid sequence of the feline p53 gene showed 82.1% and 74.9% similarities with those of the human and mouse counterparts, respectively, and had structural characteristics in common with the p53 genes of several other species. Aberrations of the p53 gene were investigated by RT-PCR and single-strand conformation polymorphism analyses. Of 10 primary hematopoietic tumors and 3 lymphoma cell lines examined, one lymphoma and one lymphoma cell line had a point mutation of the p53 gene, resulting in single amino-acid substitutions.

Amino Acid Sequence↗
PubMed1994

Induction of apoptosis in a T lymphoblastoid cell line infected with feline immunodeficiency virus.

The mechanism of cell death induced by feline immunodeficiency virus (FIV) infection was investigated in an interleukin 2(IL-2)-dependent T-lymphoblastoid cell line (MYA-1). DNA extracted from FIV-infected MYA-1 cells showed a ladder of nucleosomal DNA, indicating that the cytopathic effect (CPE) observed in these cells was due to apoptosis. Infection of MYA-1 cells with FIV was associated with suppression of the proliferative response of the cells to exogenous IL-2 prior to DNA fragmentation. These findings suggest that FIV-induced CPE in these T-lymphoblastoid cells is associated with apoptosis possibly due to a defect in the IL-2 signal transduction pathway.

Animals↗
PubMed1994

Molecular cloning and expression of canine interleukin 8 cDNA.

Molecular cloning of canine interleukin-8 (IL-8) was performed to establish a basis for its investigation in the canine immune system. From a cDNA pool constructed from LPS-stimulated popliteal lymph node cells, canine IL-8 cDNA covering the whole coding region was amplified by polymerase chain reaction. The nucleotide sequence of a canine IL-8 clone, designated pcIL-8#38, was highly similar to those of human, rabbit and porcine IL-8, and comprised 353 bp with an open reading frame that encoded 101 amino acids. Analysis of the deduced amino acid sequence of insert DNA in pcIL-8#38 showed 76.5, 80.2, and 87.0% similarities with human, rabbit and porcine IL-8 proteins, respectively. Insert DNA of pcIL-8#38 was transferred to a mammalian expression vector, pcDL-SR alpha 296, and transfected into Cos7 cells. The supernatant of the transfectant had neutrophil chemotactic activity when it was examined by the neutrophil migration assay, suggesting that our cloned cDNA was biologically active. The cloned canine IL-8 cDNA will be useful for canine inflammatory disease and comparative immunology research.

Amino Acid Sequence↗
PubMed1993

Detection of feline immunodeficiency virus proviral DNA in feline peripheral blood mononuclear cells by the nested two-step polymerase chain reaction.

The polymerase chain reaction (PCR) was applied to detect feline immunodeficiency virus (FIV) proviral DNAs in primary peripheral blood mononuclear cells (PBMC). Suitable conditions for PCR amplification were examined to obtain highly sensitive and specific results by simple staining in agarose gel. Specific amplification of FIV proviral DNA in PBMC DNA of FIV-infected cats was achieved by a nested two-step PCR that amplified the DNA first with outer primers and then with inner primers nested within the first primers. PCR amplification using different primers indicated that those based on the gag sequence of the FIV/TM2 strain isolated in Japan were suitable for the detection of FIV genomes in naturally infected Japanese pet cats. By the nested two-step PCR with mixed gag primers of TM2 and Petaluma, isolated in the USA, we could detect FIV genomes in all 11 primary PBMC samples from FIV-seropositive cats tested. The PCR protocol developed here is sensitive and specific for molecular detection of FIV infection in cats.

Animals↗
PubMed1993

Apoptosis induced by tumor necrosis factor in cells chronically infected with feline immunodeficiency virus.

Tumor necrosis factor alpha (TNF-alpha) induced morphologic changes such as chromatin condensation and cell shrinkage in a feline fibroblastic cell line (CRFK) chronically infected with feline immunodeficiency virus (FIV) but not in uninfected CRFK cells. DNA extracted from TNF-alpha-treated CRFK cells infected with FIV showed a ladder of nucleosomal DNA, indicating that this cytocidal effect by TNF-alpha was due to programmed cell death, or apoptosis. These findings may have implications for understanding the pathogenesis of FIV infection and for the design of specific therapeutic strategies for AIDS in humans as well as cats.

Animals↗
PubMed1993

Malignant histiocytosis with multiple skin lesions in a dog.

A 7-year-old male Yorkshire terrier was examined for multiple plaques and nodules on the skin. The clinical, cytological and histopathological features indicated a malignant histiocytosis. Cytoreductive chemotherapy produced moderate clinical improvement, but died at day 90 after the first admission. Pathological examination revealed the neoplastic histiocytes in the skin as well as in the myocardium and skeletal muscles.

Animals↗
PubMed1993

Rearrangements of immunoglobulin and T-cell receptor genes in canine lymphoma/leukemia cells.

Tumor cells from 15 canine lymphoma/leukemia cases were examined for genetic rearrangements of immunoglobulin and T-cell receptor (TCR) genes in parallel with cell surface antigens. Ten of these 15 cases showed rearrangements of the immunoglobulin heavy chain (IgH) gene, while 4 cases displayed TCR beta-chain gene rearrangements on Southern blot analysis. All the cases with IgH gene rearrangements had multicentric form lymphoma, and 6 of the 10 cases were cell surface immunoglobulin-positive. On the other hand, the cases with TCR gene rearrangements included atypical lymphoma/leukemia cases, and 3 of the 4 cases were Thy-1 antigen-positive. Although the tumor cell lineage of a considerable number of lymphoma/leukemia cases could not be determined by phenotypic analysis, examination of the IgH and TCR gene rearrangements disclosed the lineages of 14 of 15 cases. Genetic analysis demonstrated that the tumor cells in most canine multicentric lymphomas were formed by clonal expansion of B-lymphocyte. These findings show that studies on the rearrangements of immunoglobulin and TCR genes are very useful for understanding the cellular origin, clonality and hierarchy of canine lymphoma/leukemia cells.

Animals↗
PubMed1993

Molecular cloning and chromosomal mapping of feline p53 tumor suppressor gene.

Alterations of the p53 tumor suppressor gene have been observed in a variety of human and mouse tumors. For investigation of the role of this gene in tumors in cats, feline p53 cDNA was molecularly cloned by PCR amplifications using primers based on the sequences conserved among several species. The cloned cDNA appeared to cover approximately 90% of the open reading frame of the feline p53 gene and had characteristic structures in common with the p53 genes of several other species. The amino acid sequence similarities of the feline p53 with the human, mouse, rat and chicken counterparts were 82.9%, 75.6%, 76.5% and 57.2% respectively. Moreover, using a panel of feline x rodent somatic cell hybrids, the feline p53 gene was assigned to feline chromosome E1. These data will be useful for determining the role of the p53 tumor suppressor gene in feline tumors.

Amino Acid Sequence↗
PubMed1993

Establishment and characterization of a feline large granular lymphoma cell line expressing interleukin 2 receptor alpha-chain.

A feline large granular lymphocyte (LGL) cell line was established from a cat with an alimentary-form lymphoma. This cell line, designated as FGL, had many large azurophilic granules in the cytoplasm, which were typical to LGL cells. Proviral genome of feline leukemia virus was detected in the chromosomal DNA of FGL cells, and reverse transcriptase activity was also demonstrated in the culture supernatant. Furthermore, we found expression of interleukin 2 (IL-2) receptor alpha-chain on the cell surface of FGL and its natural killer activity against human erythroblastic leukemia cell line, K562.

Animals↗
PubMed1993

Molecular cloning of feline interleukin-6 cDNA.

We cloned and sequenced feline interleukin-6 (IL-6) cDNA using the polymerase chain reaction (PCR) technique to obtain information on regulation of the IL-6 gene. The feline IL-6 cDNA cloned in this study was 752 bp long and included all the amino acid coding region. At the amino acid level, the overall identities of the feline IL-6 to the human and murine counterparts were 58% and 39%, respectively. After transfection with the expression vector containing the cDNA, CRFK cells produced biologically active proteins that showed hybridoma growth promoting activity.

Amino Acid Sequence↗
PubMed1992

Anti-human interleukin-6 receptor antibody inhibits human myeloma growth in vivo.

Myeloma is one of the interleukin (IL)-6-related diseases to which abnormal expression of IL-6 has been reported to be linked. We examined the in vivo inhibitory effect of anti-human IL-6 receptor (IL-6R) antibody on human myeloma cell growth in mice. SCID mice were subcutaneously inoculated with solid tumor of the myeloma cell line S6B45 in which human IL-6 was acting as an autocrine growth factor. Ten intraperitoneal administrations of 100 micrograms of the anti-human IL-6R antibody PM1 at 48-h intervals strongly inhibited the growth of S6B45 cells when the administration started 24 h after tumor inoculation. The tumor growth inhibition in vivo was also observed by administration of the anti-human IL-6 antibody MH166 using the same procedure as for PM1. The inhibitory effect of PM1 was not significant when the administration started 5 or more days after tumor inoculation. This work indicates that anti-human IL-6R antibody, as well as anti-human IL-6 antibody inhibits human myeloma growth in vivo, and provides an animal model for testing the therapeutic value of agents such as antibodies to human IL-6, IL-6R and gp130, an IL-6R-associated signal transducer, in the treatment of human myelomas.

Animals↗