Biomedical subjects
A Ito
Publications and source records attributed to A Ito.
Structural requirement for recognition of the precursor proteins by the mitochondrial processing peptidase.
We examined the structural characteristics of the extension peptides responsible for the recognition by the mitochondrial processing peptidase by using preadrenodoxin, which has a long extension peptide of 58 amino acid residues, as the substrate. The deletion of various parts of the extension peptide of pre-adrenodoxin indicated that more than 40 amino acid residues and the presence of basic amino acid residues in the distal portion (20-40 amino acid residues upstream of the cleavage site) were necessary for the recognition of the precursor by the peptidase. The processing of preadrenodoxin was strongly inhibited by the synthetic peptide corresponding to the middle portion of the extension peptide, whereas the peptide corresponding to the amino-terminal portion exhibited weak inhibition of the processing. The replacement of arginine residues in the middle portion of the extension peptide with neutral amino acids resulted in a great decrease in the processing. We conclude that basic amino acids at a position distal to the cleavage site are necessary for the recognition of the precursor proteins by the processing peptidase and that basic amino acids required for the mitochondrial targeting and those for the recognition by the peptidase are separately located in the extension peptide of pre-adrenodoxin.
Arginine residues in the extension peptide are required for cleavage of a precursor by mitochondrial processing peptidase. Demonstration using synthetic peptide as a substrate.
Mitochondrial processing peptidase (MPP) specifically recognizes a large variety of mitochondrial precursor proteins and correctly cleaves off the extension peptides. To determine the structure common to all the extension peptides that is required for specific recognition by MPP, we synthesized various oligopeptides of different chain lengths and amino acid sequences, based on the amino acid sequence of the extension peptide of pre-malate dehydrogenase, and determined kinetic parameters of the cleavage reactions. The minimal length of peptides for effective cleavage was 16 amino acid residues consisting of 11 and 5 residues from the cleavage site to the amino- and carboxyl-terminal sides, respectively. Two sets of basic amino acids in the peptide, the distal arginine residue at position -10 and the proximal ones at positions -3 and -2 relative to the cleavage site, were necessary for effective hydrolysis. Of these two, the residue at position -2 was more important for effective cleavage than the one at position -3 and could not be replaced by a lysine residue. The replacement of the distal arginine by lysine had no effect on the cleavage. Our study demonstrates that use of peptides with the proper length is essential for performing kinetic analyses on the cleavage reaction by MPP and that an arginine residue at position -2 to the cleavage site is necessary for the recognition and cleavage of the extension peptide.
[Diffuse slow washout pattern (DSWO) on exercise stress thallium-201 myocardial SPECT: correlative study with coronary arteriography and the related clinical factors].
DSWO was shown on bull's eye images of 98 of 1234 patients suspected of having coronary artery disease and examined with thallium-201 myocardial SPECT imaging. Fifty-eight of these 98 patients underwent coronary arteriography, and comparison studies were performed between the bull's eye SPECT images, the results of coronary arteriography and the laboratory data. DSWO was found in 11 of 58 cases (19.0%) of single vessel disease (1VD), 18 of 58 cases (31.0%) of double vessel disease (2VD) and 21 of 58 cases (36.2%) of triple vessel disease (3VD). Three of 58 cases (5.2%) of stenosis of a coronary artery less than 75% of its normal diameter (N group) also showed DSWO. DSWO was closely related with multiple vessel disease, as has been indicated by previous reports, but we also found another patient group that showed minor coronary arterial change (1VD and N group) and manifested DSWO. Based on the study of laboratory data, we clarified that this group of patients tended to show accompanying hypertension and hyperlipidemia as factors influencing the appearance of DSWO. DSWO was accompanied by hypertension and hyperlipidemia in 54.5% and 45.5% in 1VD. These values were higher than those in the 2VD and 3VD cases, which were 33.3% and 38.9% in 2VD, and 19.0% and 28.6% in 3VD, respectively. Hypertension and hyperlipidemia appeared to play an important role in causing DSWO by interfering with coronary circulation.
Suppression of interleukin 8 production by progesterone in rabbit uterine cervix.
Uterine cervical fibroblasts prepared from rabbits at 23 days of gestation were found to produce spontaneously the neutrophil chemotactic factor/interleukin 8 (IL-8). When the cells were treated with recombinant human interleukin 1 alpha and 1 beta (rhIL-1 alpha and -1 beta), both cytokines similarly enhanced the production of IL-8 in a dose-dependent manner. Recombinant tumour necrosis factor alpha also enhanced its production to a lesser extent, but interleukin 6 failed to modulate the production. Physiological concentrations of progesterone suppressed both the spontaneous and IL-1-mediated production of IL-8 in parallel with the decrease in the steady-state levels of its mRNA. These suppressive actions of progesterone were offset by co-treatment of cells with a progesterone antagonist, mifepristone (RU486). In conclusion, basal and IL-1-induced IL-8 production in rabbit uterine cervical fibroblasts is down-regulated by progesterone at the transcriptional level. These results obtained in vitro and our previous observations indicating that progesterone modulates the extra-cellular matrix breakdown via the suppression of production of matrix metalloproteinases and the augmentation of production matrix metalloproteinases and the augmentation of production of their specific inhibitors (TIMP-1) [Sato, Ito, Mori, Yamashita, Hayakawa and Nagase (1991) Biochem. J. 275, 645-650] may explain the mechanisms of the maintenance of pregnancy until parturition and the acceleration of uterine cervical ripening and dilatation at term.
Progesterone increases the production of tissue inhibitor of metalloproteinases-2 in rabbit uterine cervical fibroblasts.
Rabbit uterine cervical fibroblasts in culture produces tissue inhibitor of metalloproteinases (TIMP)-1 and TIMP-2. When cells were treated with physiological concentrations of progesterone, the production of two TIMPs increased, and essentially all TIMP-2 was found to be complexed with promatrix metalloproteinase 2 (proMMP-2)/progelatinase A. Progesterone did not modulate the production of proMMP-2 and resulted in the increased total amount of proMMP-2-TIMP-2 complex. These observations provide the first evidence that progesterone participates in maintaining the homeostasis of connective tissue matrix in uterine cervix by augmenting both TIMP-1 and TIMP-2 production along with the known suppressive effects on the proMMP-1 and proMMP-3 production.
Acceleration of desipramine-induced changes on the dopamine receptor-coupled adenylate cyclase system by pertussis toxin.
The response of adenylate cyclase to GTP and to dopamine (DA) was investigated in striatal membranes from desipramine (DMI)-treated rats (10 mg/kg, b.i.d., for 5 days). GPT exerted the same biphasic effect on basal and DA-stimulated enzyme activity in membranes from DMI-treated rats as on saline-treated rats. Rats were injected intraventricularly once with islet activating protein (IAP), pertussis toxin, and given extended treatment with DMI in order to study the effects on the inhibitory GTP-binding protein (Gi). Gi loses its function as a signal transducer on being ADP-ribosylated selectively by the IAP. D2 inhibition of adenylate cyclase by DA was attenuated by the IAP treatment in both DMI-and saline-treated rats; peak levels of DA plus GTP stimulation shifted from 1 microM to 100 microM GTP. D1 stimulation of adenylate cyclase by DA was also attenuated by the IAP in the DMI-treated rats. Since long-term treatment with DMI (15 mg/kg, once a day, for 3 weeks) resulted in suppression of D1 stimulation similar to that seen in the present findings, uncoupling between D2 receptors and Gi due to IAP treatment might accelerate DMI-induced adaptive changes of dual control of adenylate cyclase system by DA.
Role of laparoscopic cholecystectomy in treating gallbladder polyps.
Since the application of laparoscopic cholecystectomy (Lap C) to gallbladder polyps has not yet been fully evaluated, we performed Lap C on 26 patients with gallbladder polyps. Pathological examinations showed adenocarcinoma in three patients, adenoma in two, and cholesterol polyp in 21. Preoperative diagnoses of the cases with adenocarcinoma were a cholesterol polyp in one patient and an adenoma in two. Adenocarcinoma was confirmed to reside in the mucosa without any invasion of lymphatic ducts or small vessels in the three patients. This procedure was considered to be sufficient for this grade of cancer, and, therefore, no additional operations were performed. At present, our policy is to resect by Lap C a gallbladder polyp having a maximum size larger than 10 mm and a tendency to grow or presenting with suspicion of adenoma. When cancer is suspected by preoperative examinations, however, traditional surgery may be recommended.
Adoptive transfer of protection to Taenia taeniaeformis infection in rats with immune spleen cells.
The effector mechanism of the protection against re-infection with eggs of Taenia taeniaeformis was evaluated in rats by analysing protection dynamics against an ongoing primary infection. Immune spleen cells were prepared from F344 donor rats 10 days after oral inoculation with 200 eggs and used for adoptive transfer experiments. When F344 recipient rats were injected intravenously with immune spleen cells (2 x 10(8)) from 2 to 48 h after a primary egg inoculation, there was a statistically significant reduction in the number of metacestodes recovered at day 30 of the infection. The highest protection figures were obtained in rats given immune cells at 12 or 24 h of the primary infection. The maximum reduction induced with immune cells was 92% when they were injected at 12 h of the ongoing infection. It is strongly suggested that the vulnerable stage of the parasite damaged by immune spleen cells is the initial stage of infection (oncosphere and post-oncosphere stages in the liver within 2 days, especially between 12 and 24 h).
A study of CD45RO, CD45RA and CD29 antigen expression on human decidual T cells in an early stage of pregnancy.
The decidua is the place where the fertilized egg is implanted and where the immunocompetent cells of the mother come into direct contact with genetically disparate cells of the conceptus. Although the T cells in the decidua are exposed to fetal antigens, the fetus is not rejected by maternal immunocompetent cells. In the present study, we examined surface markers to determine whether the T cells in the human decidua are naive T cells without or memory T cells with a history of antigen stimulation. Although few T cells were present in the decidua, as compared to the peripheral blood, CD45RO+, CD29+ and CD45RA- CD4+ T cells as well as CD45RO+, CD29+ and CD45RA- CD8+ T cells, which are considered to be memory T cells, were in the majority, with only small numbers of CD45RO-, CD29- and CD45RA+ CD4+ and CD8+ cells, which are naive T cells, present. Also, the decidual mononuclear cells secreted IL-2 and IL-4. Since IL-4 is secreted only by memory T cells, it is suggested that in the decidua memory T cells increase in number and secrete cytokines, thereby in some way influencing the phenomenon of fertility.
Tumorigenicity study of phloxine (FR 104) in B6C3F1 mice.
The aim of this study was to examine the tumorigenicity of the food colouring phloxine (Food Red 104) in (C57BL/6N x C3H/HeN) F1 mice. Groups of male or female mice received 0, 0.1 and 0.4% phloxine in the diet starting at 6 wk of age for a maximum of 90 wk. Survivals of mice were more than 85% at 64 wk after the start of phloxine administration. Their body weights reached maximum at 48 wk and both male and female mice given 0.1 and 0.4% phloxine weighed significantly more than their respective controls. Tumours developed in 13 organs, the greatest incidence in males being of liver tumours (67.3%), followed by lung tumours (10.9%), lymphomas (9.7%), altered foci of liver (8.4%) and hepatic haemangioma (7.8%). In females, hepatic tumours (11.5%) had the greatest incidence followed by lymphomas (10.8%), pulmonary tumours (8.2%) and hepatic haemangioma (3.2%). In males, the incidence of altered foci and haemangioma in the liver was 19 and 14%, respectively, in the control groups, whereas only 2% occurred in the group given 0.4% phloxine (P < 0.05). It is concluded that average body weights increased significantly in female mice of groups given 0.1 or 0.4% phloxine, compared with body weights of the female controls, and that altered liver foci were significantly reduced in male mice given 0.4% phloxine. There was a significant increase in pituitary tumour incidence in the test groups compared with the control groups.
A sterol with an unusual side chain from Anoectochilus koshunensis.
A new sterol with a non-conventional side chain has been isolated from the whole plant of Anoectochilus koshunensis, together with four known sterols, a megastigmane glucoside and 2'-deoxyadenosine. The structure of the new sterol was elucidated as 26-methylstigmasta-5,22,25, (27)-trien-3 beta-ol based on chemical and detailed spectroscopic evidence.
Lignan glycosides from bark of Albizzia myriophylla.
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Cross protection against Taenia taeniaeformis in rats vaccinated with non-viable oncospheres of Asian Taenia or T. saginata.
It was determined to examine whether rats injected with non-viable oncospheres of Asian Taenia or Taenia saginata became resistant to challenge infection with eggs of Taenia taeniaeformis, since (a) metacestodes of Asian Taenia and T. taeniaeformis develop in the liver of pigs and rats, respectively, and (b) Asian Taenia and T. saginata have human origins. Rats injected intravenously or subcutaneously with complete Freund's adjuvant with non-viable oncospheres of Asian Taenia showed statistically significant resistance to challenge infection with eggs of T. taeniaeformis, whereas those injected with non-viable oncospheres of T. saginata did not show any resistance.
Antibody responses in the wild vole, Clethrionomys rufocanus bedfordiae, naturally infected with Echinococcus multilocularis by western blotting.
Antibody responses against crude antigens and the two (Em18 and Em16) epitopes of Echinococcus multilocularis were analysed by Western blotting using sera from the wild vole, Clethrionomys rufocanus bedfordiae (Crb), which were captured in Hokkaido, Japan and found to have been naturally infected with eggs, and compared with those in patients of AHD and mice experimentally infected with protoscoleces of this parasite. Antibody responses in the wild were demonstrated most clearly with anti-Crb-IgG antiserum but more faintly with anti-rat-IgG antiserum and poorly with anti-mouse-IgG or anti-human-IgG antisera or Protein G. Although only two serum samples of the wild vole found naturally infected were analysed, antibody responses against Em18 and Em16 in these voles appeared to be similar to those in AHD patients but differed from those in mice.
In vitro hatching of oncospheres of Taenia taeniaeformis using eggs isolated from fresh, frozen, formalin-fixed and ethanol-fixed segments.
In order to establish a simple means of handling eggs of taeniid cestodes under non-biohazardous conditions, gravid segments of Taenia taeniaeformis were fixed in ethanol or formalin or were frozen. In vitro hatching of oncospheres was carried out by the 0.5% sodium hypochlorite method using eggs isolated from the nonviable and viable segments. No oncospheres hatched from formalized eggs, whereas almost all oncospheres hatched from all other eggs but the number of oncospheres recovered was highly variable. Oncospheres hatched from eggs isolated from frozen segments were highly fragile. The highest figures for in vitro hatching of oncospheres were recorded when eggs isolated either from segments fixed in 70% ethanol or from viable segments were used.
Cytochrome b5-like hemoprotein/cytochrome b5 reductase complex in rat liver mitochondria has NADH-linked aquacobalamin reductase activity.
Rat liver mitochondrial NADH-linked aquacobalamin reductase was characterized to clarify its enzymological properties. Most of the enzyme was solubilized with 10 g/L Triton X-100 from rat liver mitochondrial membranes. The elution behavior of the solubilized enzyme was identical to that of NADH-cytochrome c reductase (b-type cytochromes/cytochrome b5 reductase complex) during DEAE-Sepharose Fast Flow column chromatography. By mixing both purified cytochrome b5-like hemoprotein (outer membrane-cytochrome b) and cytochrome b5 reductase, cob(II)alamin was formed from aquacobalamin and NADH. These results provide evidence that the outer membrane-cytochrome b/cytochrome b5 reductase complex has the activity of the NADH-linked aquacobalamin reductase in rat liver mitochondria. Some properties of the NADH-linked aquacobalamin reductase were studied using the function of rat liver mitochondrial membranes. The specific activity (109.5 +/- 14.3 nmol.min-1.mg protein-1) of the enzyme was shown under physiological conditions (pH 7.1 at 40 degrees C). The optimal pH and temperature for activity were 7.1 and 40 degrees C, respectively. The apparent Km values were 41.9 mumol/L for aquacobalamin in the presence of 0.2 mmol/L NADH and 14.4 mumol/L for NADH in the presence of 0.1 mmol/L aquacobalamin. The enzyme was specific for aquacobalamin, because cyanocobalamin could not be reduced by the enzyme.
Inhibition of mammary tumors by pretreatment with 17 beta-estradiol in F344 rats induced with N-methyl-N-nitrosourea.
The influence of 17 beta-estradiol (E2) and prolactin was studied on N-methyl-N-nitrosourea (MNU)-induced mammary carcinomas (MCAs) in rats. MNU was intravenously injected once into seven-week-old female. female F344 rats at a dose of 50 mg/kg body weight. Groups of rats also received either 2.5 mg of E2 or a continuous supply of prolactin and/or growth hormone via transplanted MtT/F84 (mammo-somatotropic pituitary tumor). Rats were observed for up to 36 weeks after MNU administration. Although simultaneous administration of MNU and E2 did not much affect the occurrence of MCAs as compared to administration of MNU alone, rats treated with 2.5 mg of E2 for two weeks before MNU administration had significantly reduced occurrence of MCAs compared to those given MNU alone. In contrast, rats with MNU plus MtT/F84 showed high incidence and shortened latency of MCAs and they also had a high incidence of clitorial gland hyperplasias. Average pituitary weights and serum prolactin levels in E2-treated rats were greatly increased compared to those of MNU-alone rats. Average serum E2 levels were about 100 ng/ml in E2-treated rats and 0.05 ng/ml in rats without E2 treatment. Serum prolactin levels were greatly increased in rats with MtT/F84. The results indicated that pretreatment with E2 before MNU administration was inhibitory while increased prolactin caused by grafting MtT/F84 after MNU injection was promotive for the occurrence of MCAs in female F344 rats.