Biomedical subjects
Publications and source records attributed to T Ohno.
The extent of restriction fragment length polymorphism in genes for the arginine deiminase enzyme among 28 species of mycoplasmas was assessed by Southern blot analysis of DNA digested with EcoRI or TaqI nuclease probed with a 725-bp internal fragment of the arginine deiminase gene from Mycoplasma arginini. The results indicated unexpected heterogeneity among species of a single genus.
For the multiplication of RNA viruses, specific host factors are considered essential, but as of yet little is known about this aspect of virus multiplication. To identify such host factors, we previously isolated PD114, a mutant of Arabidopsis thaliana, in which the accumulation of the coat protein of tobacco mosaic virus (TMV) in uninoculated leaves of an infected plant was reduced to low levels. The causal mutation, designated tom1, was single, nuclear, and recessive. Here, we demonstrate that the tom1 mutation affects the amplification of TMV-related RNAs in a single cell. When protoplasts were inoculated with TMV RNA by electroporation, the percentage of TMV-positive protoplasts (detected by indirect immunofluorescence staining with anti-TMV antibodies) was lower (about 1/5 to 1/10) among PD114 protoplasts than among wild-type protoplasts. In TMV-positive PD114 protoplasts, the amounts of the positive-strand RNAs (the genomic RNA and subgenomic mRNAs) and coat protein reached levels similar to, or slightly lower than, those reached in TMV-positive wild-type protoplasts, but the accumulation of the positive-strand RNAs and coat protein occurred more slowly than with the wild-type protoplasts. The parallel decrease in the amounts of the coat protein and its mRNA suggests that the coat protein is translated from its mRNA with normal efficiency. These observations support the idea that the TOM1 gene encodes a host factor necessary for the efficient amplification of TMV RNA in an infected cell. Furthermore, we show that TMV multiplication in PD114 protoplasts is severely affected by the coinoculation of cucumber mosaic virus (CMV) RNA. When PD114 protoplasts were inoculated with a mixture of TMV and CMV RNAs by electroporation, the accumulation of TMV-related molecules was approximately one-fifth of that in PD114 protoplasts inoculated with TMV RNA alone. No such reduction in the accumulation of TMV-related molecules was observed when wild-type protoplasts were inoculated with a mixture of TMV and CMV RNAs or when wild-type and PD114 protoplasts were inoculated with a mixture of TMV and turnip crinkle virus RNAs. These observations are compatible with a hypothetical model in which a gene(s) that is distinct from the TOM1 gene is involved in both TMV and CMV multiplication.
Effects of carperitide (alpha-human atrial natriuretic peptide) on hemodynamics and renal function in dogs with congestive heart failure (CHF) produced by volume expansion and ligation of the left anterior descending coronary artery were compared with those of various anti-heart failure agents (cardiotonic, vasodilator and diuretic). Carperitide (0.1-1 microgram/kg/min) dose-dependently decreased the elevated left ventricular end-diastolic pressure (LVEDP). No significant changes in cardiac contractility (LV dP/dtmax) and heart rate (HR) were noted, although cardiac output (CO) tended to reduce during the infusion of carperitide. Nitroglycerin (NG; 3 micrograms/kg/min) and furosemide (1 mg/kg) also decreased LVEDP, but the potency was less than that of carperitide. Sodium nitroprusside (SNP; 10 micrograms/kg/min) and dobutamine (10 micrograms/kg/min) caused a reduction in LVEDP and increased CO with an increase in HR. Hydralazine (H; 100 micrograms/kg/min) increased CO without reduction in LVEDP and induced a pronounced increase in HR. Double product (systolic blood pressure x HR), an index of myocardial oxygen consumption, was significantly reduced by carperitide, but significantly increased by DB and H. Carperitide, unlike NG, SNP, H and DB, increased urine volume and urinary electrolyte excretion. These results suggest that carperitide will be an useful therapeutic agent for the treatment of CHF.
The cardiovascular and diuretic actions of carperitide were studied in experimental animals. Carperitide relaxed various canine arteries and veins that were contracted by high K+ or norepinephrine. Carperitide stimulated particulate guanylate cyclase from rat thoracic aortas. Carperitide had almost no effect on coronary perfusion pressure or heart rate, but caused a slight decrease in contractile force in isolated guinea pig hearts. Carperitide tended to decrease isoproterenol-induced renin release from isolated rat kidney slices and elicited decreases in angiotensin II-induced aldosterone release from bovine zona glomerulosa cells. Intravenous injection of carperitide elicited decreases in arterial blood pressure and total peripheral resistance in the anesthetized and conscious dogs. Carperitide also elicited transient increases in cardiac output and coronary blood flow followed by slight decreases in them. Intravenous infusion of carperitide elicited decreases in pulmonary capillary wedge pressure, pulmonary pressure and right atrial pressure in association with elevating plasma carperitide (ANP like immuno-reactivity) level in dogs with heart failure induced by coronary artery occlusion and saline loading. These results suggest that carperitide decreases both preload and afterload and can improve the untoward hemodynamic alterations in animals with acute experimental heart failure.
Saccharomyces cerevisiae strains carrying the mutant allele skt5 are resistant to the killer toxin of Kluyveromyces lactis and are defective in protoplast regeneration. The DNA sequence analysis of the cloned mutant skt5 gene showed a nucleotide substitution (causing a glycine-to-glutamic acid substitution) and also a nucleotide insertion (causing a frameshift at the extreme carboxyterminal region) in the structural region from its wild-type gene.
Remission of diabetes, i.e. significant amelioration from absolute insulin-dependency, has been sometimes observed in diabetic BB/Wor//Tky rats which were treated with insulin. In remission BB/Wor//Tky rats, plasma glucose levels improved to near normal level and insulin content was also preserved as much as that between diabetic and non-diabetic rats. In this process, we hypothesized that autoimmune insulitis was suppressed and remaining islet B-cells was restored from severe destruction by recovering in number and/or function. While, recently, a novel regenerating (reg) gene, identified in the regenerating pancreatic islets of surgical models, is reported to be related to the replication of pancreatic B-cells in vitro. Based on these findings, we histologically investigated whether the reg protein could be actually expressed or not in the islets from remission BB/Wor//Tky rats. As expected, reg protein was observed in the islets from remission BB/Wor//Tky rats mainly in accordance with pancreatic B-cells. Thus, the present findings suggested that the regeneration of pancreatic B-cells represented by the expression of reg protein might be, at least in part, relevant to remission induced by insulin therapy in spontaneously occurring Type 1 diabetes in BB/Wor//Tky rats.
It has been reported that the urine of patients with prolidase deficiency contains various iminodipeptides with a carboxyl-terminal proline (hydroxyproline). These iminodipeptides have hitherto been detected indirectly by acid hydrolysis or enzymatic digestion, followed by amino acid analysis. In the present study, it was shown that X-Pro could be distinguished from Pro-X when the iminodipeptides were analysed directly by liquid chromatography coupled with atmospheric pressure ionization mass spectrometry (LC/API-MS), with scanning of the protonated molecule ions ([M+H]+). The same procedure also successfully quantified urinary iminodipeptides from patients with prolidase deficiency. A quantitative investigation of two siblings with prolidase deficiency revealed that the patient with severe clinical symptoms excreted more iminodipeptides than the other who did not have serious symptoms. LC/API-MS also revealed iminodipeptides (Gly-Hyp and Pro-Hyp) in the urine of the mother of the patients and in normal volunteers. Patients excreted much more Pro-Hyp than normal volunteers, whereas no quantitative differences were found between the mother and controls. In patients, the excretion of large quantities of X-Pro is due to their very low prolidase activity towards this type of substrate. In the erythrocytes of patients, prolidase activity towards X-Hyp was extremely low; even in the mother and normal volunteers, it was remarkably low in comparison with the activity against X-Pro.
Uterine endometrial carcinoma has been reported to synthesize and secrete a putative peptide mitogen that elicits a potent proliferative response in endometrial fibroblasts. The extract from endometrial carcinoma stimulated [3H]thymidine incorporation into human endometrial fibroblasts in a dose-dependent manner. Concomitant exposure of the fibroblasts to prolactin (PRL) led to a remarkable inhibition of the extract-stimulated mitogenic activity of the fibroblasts. This inhibition was dependent on PRL dose, and maximal effect occurred at 1 microM of PRL. PRL (10 nM) suppressed an apparent maximal activity of the extract by 50%, and the half-maximal stimulated effect of the extract on thymidine incorporation was observed at the same concentration in the absence or presence of PRL. This noncompetitive manner may imply that PRL acts at a stage after the interaction of the mitogen in the extract with the specific receptor for mitogen. When the fibroblasts were first exposed to the extract for 12 hr and then to PRL, PRL suppressed the mitogenic activity with no lag. The rapid growth-inhibitory effect of PRL was mimicked by prostaglandin E1, but the combination of both types of ligand was not additive in the inhibitory action on growth. PRL and prostaglandin E1 may inhibit a similar mitogenic signaling cascade. Specific receptor sites for PRL were detected in the endometrial fibroblasts, showing high binding affinity (Kd = 16.1 nM) and low binding capacity (Bmax = 1.59 pmol/mg protein). Treatment of the fibroblasts with the endometrial carcinoma extract induced no changes in the level of PRL receptor, excluding the possibility that PRL competes for the binding sites with the mitogen. These findings would suggest that PRL may block the mitogenic activity of the fibroblasts stimulated by the endometrial carcinoma-derived mitogen via a PRL receptor-mediated mechanism, perhaps prostaglandin production.
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To study the relationship between aging and development of diabetic nephropathy, we studied the time until the development of microalbuminuria in old onset (> 50 years old, n = 21) and young onset (< 40 years old, n = 26), normotensive NIDDM patients. Microalbuminuria which is associated with the early stage of diabetic nephropathy was defined as urinary albumin index (UAI; mg/g.creatinine) of more than 10mg/g.creatinine, using timed overnight urine. In these two groups, there were no significant differences in duration of diabetes, observation periods, glycemic control, systolic diastolic blood pressure, body mass index and creatinine clearance at the time of the last observation. Mean UAI +/- standard deviation of the two groups were 37.5 +/- 78.2 mg/g.cr and 93.0 +/- 127.2 mg/g.cr in the young onset group and the old onset group, and prevalences of microalbuminuria were 38% and 76% in the young onset and old onset group, respectively. Thus, UAI and prevalence of microalbuminuria in the old onset group are significantly higher than those of the young onset group (P < 0.05). These results suggest that aging, in itself, is one of the significant risk factors for the development of diabetic nephropathy in NIDDM patients.
Twelve cases of sinonasal 'T'-cell lymphoma were studied immunohistologically with a panel of antibodies that included two antibodies to CD3, Leu4 and rabbit anti-human CD3 epsilon. Leu4 was positive in five cases, whereas rabbit anti-human CD3 epsilon was positive in 11. This discordant reaction of the antibodies, suggestive of cytoplasmic CD3 epsilon +, CD3 delta-, and CD3 gamma-, has recently been demonstrated in activated adult natural killer (NK) cells also. Leu4-negative cases had the phenotype CD2+, CD5-, CD7+/-, T-cell receptor (TCR)-, and NK cell markers+, and had non-rearranged TCR genes. These findings suggest that a large group of sinonasal 'T'-cell lymphomas is derived from NK cells.
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Three of the ets oncogene superfamily members v-ets, Spi-1/PU.1 and Fli-1, have been shown to be directly involved in retroviral-mediated acute erythroleukemias. The Fli-1 gene was found to be rearranged in 75% of the erythroleukemias induced by Friend murine leukemia virus (F-MuLV), suggesting that it could play a key role in cellular transformation. We have previously isolated and characterized the human Fli-1 gene and have found it to be highly homologous (80%) to the human erg-2 gene. Human Fli-1 was also shown to be rearranged in Ewing's sarcoma cases, in which the amino-terminal region of the Fli-1 gene was replaced with a novel coding region of a putative RNA-binding protein, EWS. In this report, we show that the recombinant Fli-1 protein expressed in bacteria binds to DNA in a sequence-specific manner. It appears that Fli-1 and erg proteins fall into the category of ets proteins that recognize limited ets target sequences, unlike c-ets-1, ets-2 and Elk-1. The Fli-1 gene was found to activate the transcription of the reporter gene that was linked to Fli-1 target sequences, suggesting that Fli-1 is a sequence-specific transcriptional activator. Deletion analysis revealed the presence of two autonomous transcriptional activation domains, one at the amino-terminal region (amino-terminal transcriptional activation domain, ATA) and the other at the carboxy-terminal region (carboxy-terminal transcriptional activation domain, CTA). Secondary structural analysis of ATA and CTA domains revealed the presence of helix-loop-helix (H-L-H) and/or turn-loop-turn (T-L-T) regions. From these results it appears that a portion of the Fli-1 ATA domain (H-L-H region) was replaced by the amino-terminal domain of EWS gene in Ewing's sarcoma cases. Therefore alteration in the transcriptional activation function of Fli-1 may be responsible for human malignancies such as sarcomas, leukemias and lymphomas in which this gene is rearranged.
We describe the case of a patient with peripheral gamma/delta T-cell lymphoma (T-ML) with hepatosplenomegaly, generalized lymphadenopathy, and bone marrow involvement. A 44-year-old man had lymphoma, which became clinically apparent 2 months after the onset of myositis and insulin-dependent diabetes mellitus. A cervical lymph node biopsy specimen showed diffuse infiltration by large neoplastic cells with vascular proliferation. The neoplastic cells expressed the T-cell receptor (TCR)delta chain detected by TCR delta 1 and delta-TCS1, CD3, CD30, CD45RO, and epithelial membrane antigen, but not the TCR beta chain detected by beta F1, CD1a, CD2, CD4, CD5, CD7, CD8, CD25, HLA-DR, and terminal deoxynucleotidyl transferase. The cells had a clonal rearrangement of TCR gamma chain gene and a germ-line configuration of immunoglobulin heavy chain gene and TCR beta chain gene. Despite chemotherapy, the patient died of refractory lymphoma 4 months after diagnosis. Examination at autopsy revealed that the main hepatic and splenic neoplastic infiltration sites were the portal area and white pulp, respectively. Our patient differed from those with gamma/delta T-ML with hepatosplenic involvement reported previously with respect to the hepatic and splenic neoplastic infiltration patterns and the presence of lymphadenopathy.
We report a case of intermediate lymphocytic lymphoma (ILL) with multiple lymphomatous polyposis. A 56-year-old man presented with general fatigue and bloody stool. Physical examination showed cervical and axillary lymphadenopathy, bilateral tonsillar hypertrophy, and moderate splenomegaly. Leukocyte count was 9,570/microliters with 11% abnormal cells, infiltration of which was observed in the bone marrow too. Examinations of the gastrointestinal tract revealed diffuse small polypoid lesions throughout the stomach and the entire large bowel. The biopsied specimens from both the stomach and large bowel showed diffuse infiltration of medium-sized lymphoid cells in the submucosa and the lamina propria. Lymph node biopsies showed ILL (mantle zone lymphoma). The phenotype of lymphoma cells was CD5 (+) CD10 (-) CD19 (+) CD20 (+) CD21 (+), and sIg mu delta-lambda. The patient was initially given the multiple agent chemotherapy, which did not improve the peripheral blood findings and was switched to the regimen that comprised of etoposide and prednisolone. The patient's lymphoma is well controlled by this regimen 35 months after diagnosis.
The characteristics of correlation between the right-to-left ventricular systolic pressure ratios (RVp/LVp) and the thallium-201 right-to-left ventricular (201Tl R/L) count ratios was investigated in children with various congenital heart diseases. High-resolution three-headed SPECT system equipped with either parallel-hole or fan-beam collimators was used. In a total of 102 patients, the correlation between RVp/LVp and 201Tl R/L average count ratios was good in both planar (r = 0.89, p = 0.0001) and SPECT studies (r = 0.80, p = 0.0001). Quantitative analysis of myocardial uptake by SPECT demonstrated the characteristic pattern of each disease as well as the differences in the right ventricular overload types. When the linear regression analysis was performed in each heart disease, ventricular septal defect showed most excellent correlation. Complex heart anomalies also showed positive correlation (r = 0.51, p = 0.05) with RVp/LVp, and it can be used to estimate right ventricular pressure. After surgical treatment of tetralogy of Fallot and pulmonary stenosis, the decrease of 201Tl R/L count ratio was in accordance with improvement of right ventricular overload. We conclude that 201Tl SPECT study can be a good indicator for estimation of right ventricular pressure.
We analyzed the pre-S and S region of hepatitis B virus by polymerase chain reaction in 60 chronic carriers. In 10 hepatitis B e antigen-positive chronic carriers, a single DNA band was detected at the expected size, whereas additional shorter DNA bands were observed in some cases with chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. The smaller-sized DNA bands from chronic hepatitis patients had the deletion of 183 bp in the pre-S gene, and the deletion of 492 bp in S gene. The reasons of the appearance of these mutants are unknown, but it may be interesting to study in relationship to the mechanism of host immune response.
We have compared the physico-chemical behaviors alone and in the presence of a synthetic bilayer membrane, in aqueous solution and the bioavailability after intraduodenal administration to rabbits, of the two heparin diamine salts ITF-300 and ITF-331 with those of the heparin-amine salt ITF-1175. The three salts have similar structures but different characteristics of compounds tend to form aggregates in solution, but at different critical concentrations. The compounds induce fusion of single-walled vesicles of a synthetic peptide lipid into multi-walled lamellae. The minimal concentrations of the compounds required for the formation of such lamellae differ. This behavior in solution explains the differences in absorption in the animal model. This makes it possible to correlate enhanced heparin bioavailability with the structural nature of the diamine counter-ions used to prepare heparin salts.