Biomedical subjects
Publications and source records attributed to S Izumo.
We identified a novel AvaI polymorphism within 3' non-coding region within exon 5 of the human rhodopsin gene and determined the allele frequency in a Japanese population. The polymorphism was found to be due to A/G transversion at nucleotide 5510 of the gene.
Leukocyte adhesion molecules to endothelium plays an important role in the pathogenesis of inflammatory diseases, including HTLV-I-associated myelopathy (HAM)/tropical spastic paraparesis (TSP). To help define the role of adhesion molecules in HAM/TSP, we studied the expression of lymphocyte function-associated antigen-1 (LFA-1), Mac-1, very late antigen-4 (VLA-4), Sialyl Lewisx (SLex), intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), endothelial leukocyte adhesion molecule-1 (ELAM-1) and monocyte chemoattractant protein-1 (MCP-1) in the spinal cord lesions of HAM/TSP. The results indicate that spinal cord lesions of HAM/TSP have greater VCAM-1 expression on endothelium compared with those of controls. Infiltrating mononuclear cells, especially perivascular lesions, expressed VLA-4. Although the expression of ICAM-1 in the spinal cords was not distinctive between HAM/TSP and controls, infiltrating mononuclear cells in the spinal cords of HAM/TSP strongly expressed LFA-1 and Mac-1. ELAM-1 was expressed on endothelium in the inactive-chronic lesions from three of five HAM/HAM/TSP, but was not detectable in the spinal cords of controls. SLex reactions was detectable on occasional perivascular cells in the spinal cord of HAM/TSP, but not in those controls. MCP-1 was detectable on perivascular infiltrating cells and vascular endothelium in active-chronic lesions. This study suggest that VLA-4/VCAM-1 interaction may play an important role for lymphocyte migration into the central nervous system (CNS) and MCP-1 may also be involved in inflammatory cell recruitment to the CNS in HAM/TSP.
We have analyzed muscle biopsy specimens from polymyositis patients who are also positive for human T cell lymphotropic virus type I (HTLV-I) using both immunohistochemistry for surface antigens of lymphocytes and macrophages and in situ polymerase chain reaction for HTLV-I proviral DNA on the same sections. We found HTLV-I in CD4+ cells but not in macrophages. This finding suggests that most of the HTLV-I-containing CD4+ cells are not macrophages but lymphocytes.
Five T-cell clones reactive to autologous HTLV-I-infected T-cells (KODA-TV) were established from peripheral blood lymphocytes of a HAM/TSP patient (KODA) by the limiting dilution method. All the clones showed CD3+, CD4+ and CD25+ surface markers and expressed alpha beta+ T-cell receptors to recognize KODA-TV antigens. One of the five T-cell clones (KODA-408) was infected with HTLV-I but the remaining four clones (KODA-400, 404, 405 and 409) were free of HTLV-I infection. KODA-408 recognized both KODA-TV and spinal cord antigens, the latter being extracted from autopsy tissues of a HTLV-I seronegative donor. KODA-408 did not recognize either alloantigens of peripheral blood mononuclear cells extracted from unrelated HTLV-I seronegative donors or purified human myelin basic protein. KODA-408 T-cell clone produced a considerable amount of TNF-alpha, IFN-gamma, and IL-6. The CDR3 motif of KODA-408 T-cell receptor showed a unique sequence CASSAGQS of v beta 8-D beta-J beta 1.5. These results indicated that HAM/TSP CD4+ T-cells were polyclonally activated by HTLV-I infection and antigenic stimulation. The T-cell repertoire shaped by HTLV-I infection included T-cells which recognized HTLV-I-infected T-cell antigens as well as spinal cord antigen in particular.
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Neurologic diseases associated with human T-cell lymphotropic virus type I (HTLV-I) infection have a clinical spectrum that includes myelopathy (HTLV-I-associated myelopathy/tropical spastic paraparesis, HAM/TSP) as the central manifestation. Many clinical signs of involvement outside the central nervous system (CNS) have been described in some patients with HAM/TSP and have triggered and advanced the discovery of some HTLV-I-associated concepts in HTLV-I-infected individuals without signs of CNS involvement. Most of these HTLV-I-associated diseases exhibit common viroimmunologic characteristics that include a distributional bias of HTLV-I activation between the blood flow and the affected lesions and accumulated cellular immune responses in the lesions. These facts suggest that the vulnerable tissue(s) in some HTLV-I-infected individuals may not be defined by an exclusive tissue specificity, but that common steps of HTLV-I-versus-host interactions may have an important role in the pathologic process(es) in these diseases. This review summarizes the recent perspectives of the clinical spectrum and the pathogenesis of HAM/TSP in Japan. Furthermore, the feasible pathogenic involvement of cellular interactions between infected cells and responding immunocompetent cells in the affected tissues is emphasized.
INTRODUCTION: In HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP), we correlated human leukocyte antigen (HLA) haplotypes to the fine specificities of intrathecally synthesized IgG antibodies against HTLV-1. PATIENTS AND METHODS: HLA haplotypes of HAM/TSP patients were determined by the standard NIH microcytotoxicity test and family HLA studies. IgG antibodies against HTLV-1 synthetic peptides in paired CSF and serum were measured by enzyme immunoassay, and intrathecal synthesis of antibodies was evaluated. RESULTS: HAM/TSP patients with particular HLA haplotypes (A24Cw7B7DR1DQ5, A2Cw7B7DR1DQ5, A24Cw-B52DR15DQ6, A11Cw1B54DR4DQ4, and A24Cw1B54DR4DQ4) showed more frequently intrathecal synthesis of antibodies against HTLV-1 synthetic peptides, especially against HTLV-1 env gp21 synthetic peptides. CONCLUSION: In HAM/TSP, a retrovirus-induced human chronic inflammatory disease of the CNS, this is the first report to provide evidence that the intrathecal antiviral immune response is influenced by immunogenetic factors of the HLA complex.
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A double-blind, multi-center study was performed on patients with HTLV-I-associated myelopathy (HAM) to evaluate the therapeutic effect of treatment with natural interferon-alpha (HLBI). Forty-eight HAM patients were enrolled and treated with either 0.3 MU (n = 15), 1.0 MU (n = 17), or 3.0 MU (n = 16) of HLBI for 28 days. Clinical evaluation included motor dysfunction, urinary disturbances, and changes of neurologic signs. The frequency of therapeutic response judged as excellent to good 4 weeks after starting therapy and 4 weeks after completion of therapy were 7.1% (1 of 14) and 8.3% (1 of 12) in the 0.3-MU group, 23.5% (4 of 17) and 26.7% (4 of 15) for the 1.0-MU group, and 66.7% (10 of 15) and 61.5% (8 of 13) for the 3.0-MU group. The therapeutic benefit in the 3.0-MU group was significantly higher than in the 0.3-MU group. There was no significant difference in the incidence of symptomatic side effects between groups. Abnormal laboratory data were obtained for some patients in the 1.0-MU and 3.0-MU groups; however, the treatment schedule could be continued in most patients. These results suggest that HAM patients may be safely treated with HLBI 3.0 MU every day for 4 weeks with favorable clinical effects.
To create muscle cell lines that conditionally differentiate in vitro we introduced a temperature-sensitive SV40 T antigen by retroviral infection into rat aortic smooth muscle cells (SMCs) and neonatal heart-derived cells. After G418 selection cell lines isolated were characterized at permissive (33 degrees C) and non-permissive (39 degrees C) temperatures. [3H]Thymidine uptake showed tht progression through the cell cycle is greatly reduced at 39 degrees C. Cytoskeletal proteins, such as actins and vimentin did not change significantly after temperature shift, while the number of desmin-positive SMCs significantly increased when cells were switched to 39 degrees C. Heart-derived muscle cells showed sarcomeric myosin heavy chain reactivity only when grown at 39 degrees C. After thrombin stimulation intracellular calcium in both cell types increased severalfold in 39 degrees C-cells but not in 33 degrees C-cells. Whole cell patch-clamp recordings of SMCs and heart-derived cells revealed a strong increase in nicardipine-sensitive Ca2+ current when cells were switched to 39 degrees C. Nicardipine-insensitive Ca2+ current also increased in both cell types at the non-permissive temperature. Na+ current in SMCs was large at 33 degrees C and small or not detectable at 39 degrees C and absent in heart-derived cells. Using a cDNA probe specific for the alpha 1 subunit of the dihydropyridine-sensitive Ca2+ channel we demonstrate a temperature-sensitive expression of the dihydropyridine receptor mRNA in smooth muscle-derived cells but not in heart-derived H10 cells. Our results suggest that upon downregulation of SV40 T antigen these cells become quiescent and exhibit a more differentiated phenotype. These cell lines may provide a useful tool to investigate ion channel- and receptor signal transduction, as well as cell cycle control in smooth and possibly cardiac muscle cell differentiation.
Endothelium exposed to fluid shear stress (FSS) undergoes cell shape change, alignment and microfilament network remodeling in the direction of flow by an unknown mechanism. In this study we explore the role of tyrosine kinase (TK) activity, intracellular calcium ([Ca2+]i), mechanosensitive channels and cytoskeleton in the mechanism of cell shape change and actin stress fiber induction in bovine aortic endothelium (BAE). We report that FSS induces beta-actin mRNA in a time- and magnitude-dependent fashion. Treatment with quin2-AM to chelate intracellular calcium release and herbimycin A to inhibit TK activity abolished BAE shape change and actin stress fiber induction by FSS, while inhibition of protein kinase C with chelerythrine had no effect. Altering intermediate filament structure with acrylamide did not affect alignment or F-actin induction by FSS. Examining the role of the BAE cytoskeleton revealed a critical role for microtubules (MT). MT disruption with nocodazole blocked both FSS-induced morphological change and actin stress fiber induction. In contrast, MT hyperpolymerization with taxol attenuated the cell shape change but did not prevent actin stress fiber induction under flow. Mechanosensitive channels were found not to be involved in the FSS-induced shape change. Blocking the shear-activated current (IK.S) with barium and the stretch-activated cation channels (ISA) with gadolinium had no effect on the shear-induced changes in morphology and cytoskeleton. In summary, FSS has a profound effect on endothelial shape and F-actin network by a mechanism which depends on TK activity, intracellular calcium, and an intact microtubule network, but is independent of protein kinase C, intermediate filaments and shear- and stretch-activated mechanosensitive channels.
The genomic sequence of human T-cell leukemia virus type 1 (HTLV-1) is highly conserved, although minor sequence variations enable classification of the isolates into several subgroups. We previously reported, however, that the Tax-coding sequence of HTLV-1 genome is highly variable in a random fashion within individuals with HAM/TSP and asymptomatic carriers. Here, we describe frequent base substitutions in the LTR sequence similarly to those in Tax-coding sequence. These observations indicate that frequent mutations are not unique to the sequence encoding the most effective antigen for cytotoxic T lymphocytes, but also seen in the LTR, a non-coding sequence. Thus, frequent mutations seem to occur during the viral replication process rather than the selection of rare mutants by immune surveillance.
We report here the results of therapeutic trials in 200 patients with HTLV-I-associated myelopathy (HAM)/tropical spastic paraparesis (TSP) conducted in our department between 1986 and 1993. Motor disability grades were improved by more than one grade in 69.5% (91/131) of patients by oral administration of prednisolone, 50% (3/6) by eperisone hydrochloride only, 43.8% (7/16) by blood purification (lymphocytapheresis and plasmapheresis), 40.0% (2/5) by intrathecal injection of hydrocortisone, 30.0% (3/10) by intravenous injection of high-dose methylprednisolone, 23.3% (10/43) by interferon-alpha (intramuscular injection and inhalation), 22.2% (2/9) by azathioprine, 20.0% (4/20) by high-dose vitamin C, 16.0% (4/25) by erythromycin, 12.5% (3/24) by salazosulfapyridine, 11.8% (2/17) by mizoribine, 7.1% (1/14) by fosfomycin, and 6.3% (1/16) by thyrotropin releasing hormone. No critical side effects of these therapies were seen with the exceptions of one patient with adult respiratory distress syndrome due to cytomegalovirus infection and one patient with drug-induced hepatitis/hepatic failure. Selection of these treatments for patients with HAM/ TSP must be considered on the basis of age, sex, disease severity and complications to reduce adverse events and to improve quality of life. Although the results were a synopsis of different treatments given to 200 patients with HAM/ TSP as an open trial, we consider this the first report of a large-scale therapeutic trial in patients with HAM/TSP. The results of this study indicate that immunomodulatory therapies have some beneficial effects in HAM/TSP, and the functions of these agents are related to the pathophysiology of this disease.
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OBJECTIVE: To elucidate the nature of ataxia observed in 3 cats spanning 2 generations. DESIGN: Experimental breeding was attempted to confirm heritability of the disease and establish the mode of inheritance; the original 3 cats and their offspring were studied. ANIMALS: Seven diseased cats spanning 3 generations and 11 neurologically normal cats. PROCEDURE: Cats were examined by use of the following methods: clinical observation, hematologic and serum biochemical examinations, neurologic examination, electrodiagnostics, magnetic resonance imaging, lysosomal enzyme activity assay, horizontal transmission test, and virologic and pathologic examinations. RESULTS: All kittens (1 male and 3 females) obtained by backcrosses developed pure cerebellar dysfunction from the age of 7 to 8 weeks onward. It became progressively worse, but not fatal, between 1 and 2.5 months. Prenatal or perinatal infection with feline panleukopenia virus, inherited lysosomal storage diseases, including gangliosidosis and mannosidosis, and feline hereditary neuroaxonal dystrophy were excluded. Magnetic resonance imaging indicated that size of the cerebellum of diseased cats was markedly reduced. Cerebellar cortical degeneration, especially with extensive destruction of Purkinje cells, was observed microscopically. CONCLUSION: The disease was concluded to be cerebellar degeneration of a new clinical form in cats having an autosomal recessive mode of inheritance. CLINICAL RELEVANCE: When cerebellar dysfunction is diagnosed in a cat, hereditary cerebellar degeneration of this type should be considered in the differential diagnosis.
Fibrovascular tissues obtained at therapeutic vitrectomy from 22 patients with proliferative diabetic retinopathy were studied for a mutation of tRNALEU(UUR) at nucleotide position (np) 3243 of mitochondrial DNA (mtDNA). We found a mutation of mtDNA in the vitreous sample of one patient, a 44-year-old woman who had maternally inherited, familial, non-insulin-dependent diabetes mellitus of 25-year duration with recurrent vitreous hemorrhages due to proliferative retinopathy. Heteroduplex analysis also detected the mutant-type mtDNA in the vitreous sample of this patient. The abnormality was, however, not observed in the peripheral blood sample from either this patient orr her family members with diabetes mellitus. The other 21 patients were negative for the mutation in vitreous specimens as well as in peripheral blood. Although a firm conclusion cannot be drawn from a single case report, our observations suggest that this case was an example of tissue-specific expression of mtDNA mutation. Further studies of a larger number of patients with familial diabetes mellitus seem justified.
BACKGROUND: Csx/Nkx2.5, a murine nonclustered homeobox gene expressed primarily in the heart, has significant sequence similarity to the Drosophila tinman gene. Tinman is essential for heart and gut formation in Drosophila. Targeted mutation in the mouse gene, Csx/Nkx2.5, arrests cardiac development during early embryonic stages, suggesting an evolutionary conservation in cardiogenesis. MATERIALS AND METHODS: We have isolated and characterized a human homolog, hCsx, from an adult cardiac cDNA library. Northern blotting and ribonuclease protection was used to define the pattern of expression during normal development and in disease states. Chromosomal localization of the gene was determined by somatic cell hybrid analysis and fluorescent in situ hybridization. RESULTS: The predicted amino acid sequence of hCsx has 87% overall homology to the murine gene with 100% identity in the homeodomain. The homeodomain sequence of hCsx is 95% identical to its Xenopus homolog, and 65% to tinman. hCsx mRNA was detected exclusively in the heart. hCsx transcript was detected at 12 weeks in human embryonic heart, the earliest time point examined, and was up-regulated 5-fold between 12 and 19 weeks. There was no significant alteration of hCsx message level in the myocardium of 14 patients with end stage heart failure compared to a normal control. The human gene mapped to the distal portion of chromosome 5, the 5q34-q35 region. This defines a new synteny region between human chromosome 5q and the t-locus of mouse chromosome 17, where the mouse Csx gene is located. CONCLUSIONS: hCsx, the human homolog of Drosophila tinman, is expressed in heart in a tissue restricted manner. Distal 5q trisomies produce several phenotypic abnormalities, including a high incidence of congenital heart disease. Isolation of the hCsx gene will allow further studies of mutations in this gene and their potential associations with some forms of congenital heart disease in humans.
Autopsy specimens from 3 patients with human T-lymphotropic virus (HTLV-I)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) were examined for the presence of HTLV-I in the central nervous system (CNS). In situ hybridization using an HTLV-I tax RNA probe detected cells containing HTLV-I RNA in spinal cord and cerebellar sections. HTLV-I infected cells were located within the white matter and, in particular, within the anterior and lateral funiculi of the spinal cord. Consistent with previously described HAM/TSP pathology, there were perivascular infiltrates in these CNS specimens. Significantly, HTLV-I RNA was not localized to these infiltrates but was detected deeper within the neural tissue. Furthermore, phenotypic analysis demonstrated that at least some of the infected cells were astrocytes. While previous polymerase chain reaction studies have demonstrated the presence of proviral HTLV-I in CNS specimens, here we provide evidence for the in situ expression of HTLV-I RNA in the CNS of HAM/TSP patients.