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M Saijo

Publications and source records attributed to M Saijo.

At least 73 records · Page 4Linked to original sources
PubMed1995

The interactions of E2F with pRB and with p107 are regulated via the phosphorylation of pRB and p107 by a cyclin-dependent kinase.

It has been postulated that the product (pRB) of the retinoblastoma gene dissociates from the E2F-pRB complex upon phosphorylation by cyclin-dependent kinase(s) (cdk). However, there is no direct evident for the regulation of formation of the E2F-pRB complex via phosphorylation by purified cdk. Therefore, we investigated the regulation of formation of this complex by phosphorylation using pRB and purified cyclin A-cdk2, cyclin E-cdk2 or cyclin D1-cdk4. Purified pRB was incubated with nuclear extracts prepared from pRB-defective cells and then subjected to gel mobility shift assays. We confirmed that unphosphorylated pRB associated with various types of E2F but pRB has been phosphorylated by cyclin A-cdk2 did not. We found that E2F-pRB complexes were disrupted as a consequence of phosphorylation by cyclin A-cdk2, and the levels of the free forms of E2Fs increased. We also found that not only the E2F-pRB complexes but also the E2F-p107 complexes were disrupted upon phosphorylation by cyclin A-cdk2. Furthermore, E2F-pRB complexes were disrupted through phosphorylation by cyclin D1-cdk4 and cyclin E-cdk2, as well as by cyclin A-cdk2. These results clearly demonstrate that the phosphorylation of pRB and p107 by cdks regulates the formation of complexes between E2F and pRB or p107.

Base Sequence↗
PubMed1995

DNA repair protein XPA binds replication protein A (RPA).

XPA is a zinc finger DNA-binding protein, which is missing or altered in group A xeroderma pigmentosum cells and known to be involved in the damage-recognition step of the nucleotide excision repair (NER) processes. Using the yeast two-hybrid system to search for proteins that interact with XPA, we obtained the 34-kDa subunit of replication protein A (RPA, also known as HSSB and RFA). RPA is a stable complex of three polypeptides of 70, 34, 11 kDa and has been shown to be essential in the early steps of NER as well as in replication and recombination. We also demonstrate here that the RPA complex associates with XPA. These results suggest that RPA may cooperate with XPA in the early steps of the NER processes.

Base Sequence↗
PubMed1995

Expression of a transfected DNA repair gene (XPA) in xeroderma pigmentosum group A cells restores normal DNA repair and mutagenesis of UV-treated plasmids.

The XPA gene was initially cloned based on the ability of its cDNA to improve survival of cells from xeroderma pigmentosum complementation group A (XP-A) patients following irradiation of the cells with UV. We used plasmid host cell reactivation assays to compare UV mutagenesis and the proficiency of DNA repair in a cell line from an XP-A patient, XP2OS(SV40), two derivative cell lines stably expressing XPA cDNAs and in a DNA repair proficient human cell line. Expression of XPA protein in XP2OS cells allowed them to repair UV-treated plasmid pRSVCAT, increasing activity of the damaged CAT marker gene > 100-fold to levels produced by similarly damaged plasmids in normal cells. Expression of the XPA protein in XP2OS cells improved replication of the UV-treated shuttle vector pSP189, increasing plasmid survival and decreasing plasmid mutation frequency to the levels measured in normal cells. The sequence locations of most mutation hotspots in the plasmid marker gene were similar for the three cell lines and the differences did not correlate with the DNA repair status of the cells. This suggests that the location of mutation hotspots is not directly influenced by DNA repair. Expression of the XPA protein did cause a shift in the types of mutations seen in the plasmid gene. In the XP2OS cells > 95% of the plasmid mutations were G:C-->A:T transition mutations. In contrast, XP2OS cells expressing XPA produced other types of mutations: three times as many transversion mutations and a 12-fold increase in mutations at A:T base pairs. Furthermore, the distribution of these types of mutations was similar to the proportions measured in normal cells. Strikingly similar patterns of transition and transversion mutations were found by examination of reports of XP and non-XP skin carcinomas containing mutations in the p53 tumor suppressor gene, suggesting that the repair status of the cells influenced mutagenesis associated with these skin cancers. Our data suggest that loss of XPA gene function may be sufficient to effect the quantitative and qualitative changes in mutagenesis associated with the large increase in skin cancers seen in XP-A patients.

Base Sequence↗
PubMed1995

[Thrombocytopenic purpura associated with primary human herpesvirus-6 infection].

We report here a case of a 5-month-old infant with thrombocytopenic purpura associated with serologically confirmed primary human herpesvirus-6 (HHV-6) infection. His platelet count decreased to 17,000/mm3 on the 3rd day from the onset: Bone marrow aspiration showed that the megakaryocyte count was 129/mm3 and almost all of the them were of the non-platelet forming type. The IgG antibody to HHV-6 by indirect immunofluorescent assay in the sera in acute and convalescent phases were < 10x and 160x, respectively. IgM antibody to HHV-6 in the acute phase was positive. Platelet-associated IgG was also positive. We diagnosed this case as having thrombocytopenic purpura associated with primary HHV-6 infection. Steroid therapy was started for his thrombocytopenic purpura from the 4th day after the alleviation of his fever. His platelet count increased without side effects and became within the normal range without relapse despite discontinuation of steroid therapy. Thrombocytopenic purpura might be one of the complications of primary HHV-6 infection.

Antibodies, Viral↗
PubMed1995

[Respiratory tract infection and respiratory syncytial virus in young infants].

We investigated the role of respiratory syncytial virus (RSV) in respiratory tract infections of up-to-3-months old infants. This prospective study was carried out from April 1993 to March 1994. Detection of RSV antigen in nasopharyngeal specimens was done by enzyme immunoassay using TESTPACK RSV at our out patient clinic. During this study period, 65 young infants with respiratory tract infection visited our clinic. Seventeen patients (26%) were diagnosed as having RSV infection. Fifteen out of the 17 infants with RSV infection were observed in spring and winter. Fourteen out of the 17 infants with RSV infection had acute bronchiolitis or pneumonia. In contrast to RSV infection, only three patients out of the 48 infants with non-RSV infection had lower respiratory tract infection. In conclusion, the majority of young infants infected with RSV suffered from bronchiolitis or pneumonia.

Antigens, Viral↗
PubMed1995

[Comprehensive evaluation of pharmacokinetic and clinical studies on tazobactam/piperacillin in pediatric field].

To evaluate the pharmacokinetic and clinical effects of the newly developed combination antibiotic tazobactam/piperacillin (TAZ/PIPC, YP-14) on various infections in pediatric field, a study group was organized, and a joint research by 17 institutions and their related hospitals was conducted. Informed consents of subjects were obtained prior to the study. The results obtained in this study are as follows: 1. Blood concentration and urinary excretion Pharmacokinetics of TAZ/PIPC was studied in children at doses of 25 and 50 mg/kg through intravenous injection or intravenous drip infusion. With intravenous injection, maximum blood concentrations (Cmax's) of TAZ and PIPC were achieved 5 minutes after the administration. Cmax's of TAZ were 26.9 micrograms/ml with 25 mg/kg and 45.1 micrograms/ml with 50 mg/kg, and those of PIPC were 131.0 and 199.6 micrograms/ml, respectively. Values of the total area under the blood concentration curve (AUC's) of TAZ were 14.2 micrograms.hr/ml with 25 mg/kg and 26.1 micrograms.hr/ml with 50 mg/kg, and those of PIPC were 64.0 and 112.8 micrograms.hr/ml, respectively; thus dose dependency was observed with both TAZ and PIPC. The Cmax's of desethyl piperacillin (DEt-PIPC), the active metabolite of PIPC, achieved at 60 minutes after administration, were 1.2 and 2.0 micrograms/ml, respectively. The AUC's of DEt-PIPC were 2.6 and 4.2 micrograms.hr/ml, respectively. The half-lives (T 1/2's) of TAZ were 0.60 and 0.54 hour, respectively, and those of PIPC were 0.62 and 0.65 hour, respectively. In the first 6 hours after the initiation of administration, the cumulative recovery rates of TAZ in the urine were 46.7 and 56.0% respectively, those of PIPC were 46.1 and 57.2%, respectively, and those of DEt-PIPC were 5.9 and 3.0%, respectively. With intravenous drip infusion, the Cmax's of both TAZ and PIPC were achieved at the completion of drip; the Cmax's of TAZ were 12.1 micrograms/ml with 25 mg/kg and 28.9 micrograms/ml with 50 mg/kg, and those of PIPC were 54.6 and 137.9 micrograms/ml, respectively. The AUC's of TAZ were 11.6 micrograms.hr/ml with 25 mg/kg and 25.6 micrograms.hr/ml with 50 mg/kg, and those of PIPC were 49.0 and 117.2 micrograms.hr/ml, respectively; thus dose dependency was observed with both TAZ and PIPC. Cmax's of DEt-PIPC, achieved at 60 minutes after completion of drip, were 0.9 and 1.7 micrograms/ml, respectively. The AUC's of DEt-PIPC were 2.0 and 3.8 micrograms.hr/ml, respectively. The half-lives (T 1/2's) of TAZ were 0.59 and 0.62 hour, respectively, and those of PIPC were 0.58 and 0.57 hour, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)

Adolescent↗
PubMed1994

Identification of the nature of modification that causes the shift of DNA topoisomerase II beta to apparent higher molecular weight forms in the M phase.

Human cell lines express two genetically distinct isoforms of DNA topoisomerase (topo II) II: topo II alpha (p170) and topo II beta (p180). We detected a higher molecular weight form with an apparent molecular mass of about 190 kDa in M phase-arrested HeLa cells (Kimura, K., Saijo, M., Ui, M., and Enomoto, T. (1994) J. Biol. Chem. 269, 1173-1176). In this study we confirmed, using anti-topo II alpha and topo II beta monoclonal antibodies, that this higher molecular weight form is topo II beta and consists of doublet bands around 190 kDa. We confirmed that the doublet bands constituted an M phase-specific phenomenon and were not an artifact of the procedure used to accumulate mitotic cells. Digesting the immunoprecipitated materials from mitotic cell extracts with alkaline phosphatase resulted in the disappearance of the doublet bands and the appearance of the 180-kDa band with the concomitant disappearance of 32P label in the region of the doublet bands. Neither heat-inactivated alkaline phosphatase nor phosphodiesterase affected the doublet bands and the 32P label. Topo II beta in interphase cells was also phosphorylated, but the shift in apparent molecular weight was very slight after alkaline phosphatase digestion. Analysis of the labeled phosphoamino acids present in topo II beta from M phase and logarithmically growing cells indicated that phosphorylation occurred mainly on serine and fairly on threonine residues in both topo II beta isoforms. These results indicated that topo II beta is phosphorylated at specific sites in M phase, resulting in the formation of the doublet bands.

Amino Acid Sequence↗
PubMed1994

Inactivation of oncoprotein binding by a single Cys706-to-Tyr substitution in the retinoblastoma protein.

We previously found a new single amino acid substitution at codon 706 (Cys-to-Tyr) of the retinoblastoma (RB) gene in a sporadic retinoblastoma patient. The glutathione S-transferase-RB fused protein containing this mutation was here tested for binding to SV40 large T antigen and adenovirus E1A protein, and was shown to have lost its binding affinity. Thus, Tyr, as well as Phe, residues substituted for Cys706 were found to abolish the RB protein activity.

Adenovirus E1A Proteins↗
PubMed1994

Growth state- and cell cycle-dependent fluctuation in the expression of two forms of DNA topoisomerase II and possible specific modification of the higher molecular weight form in the M phase.

The fluctuations in the expression of two forms of DNA topoisomerase II, p170 and p180, were examined by immunoblotting during the transition of the growth state and the cell cycle in Swiss 3T3 cells. The level of p170 expression was higher during the exponentially growing phase than in the stationary phase, and the level of p180 did not appear to change when the cells changed the growth state. When quiescent cells were stimulated with serum, the p170 level began to increase at 12 h and reached a maximal level at 24-28 h, corresponding to the G2 phase. In contrast, the level of p180 was almost constant during the cell cycle, and the p180 band became obscure when the number of mitotic cells increased. Immunoblotting of the samples prepared from metaphase-synchronized HeLa cells did not have a p180 band, but did exhibit a band with a higher molecular weight. This band increased and p180 decreased in parallel with the increase in the number of mitotic cells. When the cells exited from the M phase, p180 reappeared and the higher molecular weight band disappeared.

3T3 Cells↗
PubMed1994

The role of respiratory syncytial virus in acute bronchiolitis in small children in northern Japan.

Respiratory syncytial virus (RSV) plays an important role in acute bronchiolitis, which is life threatening in some infants. We investigated the epidemiology of RSV acute bronchiolitis in children less than 3 years old in northern Japan. From April 1991 to March 1993, 162 infants with acute bronchiolitis were hospitalized in our pediatric wards. The diagnosis of RSV acute bronchiolitis was based on the typical clinical manifestations and the presence of RSV antigen in their nasopharyngeal specimens or the rise of the RSV antibody titer. 124 out of 162 patients (76.5%) were diagnosed as having RSV acute bronchiolitis. 43.5% of patients with RSV acute bronchiolitis were 6 months old or less. The epidemic of RSV acute bronchiolitis commenced in October, peaked in December and ended in summer. RSV is quite prevalent in infants with acute bronchiolitis in northern Japan.

Acute Disease↗
PubMed1994

[Epidemiological study on respiratory syncytial virus lower respiratory tract infections in northern Hokkaido, Japan].

We carried out an epidemiological study on lower respiratory tract infections, concerning the respiratory syncytial virus (RSV) infection, in northern Hokkaido, Japan, during the 2-year-study period from April 1991 to March 1993. There were 467 hospitalized patients with lower respiratory tract infections. We examined the presence of RSV antigen in their nasopharyngeal specimens of 456 out of 467 patients (97.6%) by enzyme immunoassay. The diagnosis of Mycoplasma pneumoniae infection was done by the rise of antibody titer, performed with complement fixation method, in the sera of the acute and convalescent phases. There were 133 patients with RSV infection (28.9%), 45 patients with Mycoplasma pneumoniae infection (9.8%) and 7 patients with measles virus infection (1.5%). RSV played an important role in the lower respiratory tract infections in northern Hokkaido, Japan. The epidemic of RSV infection was observed in spring and early winter in 1991 and only in early winter in 1992. The number of patients with RSV infection from April 1991 to March 1992 was greater than that from April 1992 to March 1993. In contrast with RSV infection, the epidemic of Mycoplasma pneumoniae infection was observed in 1992. We speculated that the RSV and Mycoplasma pneumoniae infections might interfere with each other.

Adolescent↗
PubMed1994

Complex formation between lamin A and the retinoblastoma gene product: identification of the domain on lamin A required for its interaction.

The retinoblastoma susceptibility gene product (pRB) has been known to function as a negative regulator of cell growth. Recent observations suggest that its biological activity might be modulated by an interaction with nuclear structures. By using in vitro binding assays, we have found that pRB can associate with lamin A, which has been known to be one of the major nuclear matrix proteins. A series of GST-lamin A deletion mutants was constructed to define the amino acid sequence required for binding to pRB. A GST-lamin A (247-355) contained an activity to associate with pRB, while the other constructs, such as GST-lamin A (37-244) or GST-lamin A (356-571), could not bind to pRB. Within the pRB-binding domain of lamin A, there exists the short amino acid sequence which is also present in the pRB-binding region of the transcription factor E2F-1. The similar experiments using a set of GST-RB deletion mutants revealed that a region containing the E1A-binding pocket B and the carboxy-terminal portion of pRB was responsible for binding to lamin A.

Amino Acid Sequence↗
PubMed1993

Acquisition of nonmaternal Enterobacteriaceae by infants delivered in hospitals.

To determine whether Escherichia coli strains that colonize the intestinal tract of newborn infants in hospitals are of maternal origin or come from the environment, plasmid profiles of E. coli strains isolated from the stools of infants were compared with those from the stools of their mothers. Twenty-nine mother-infant pairs were studied in three different hospitals. In only 4 of 29 pairs, plasmid profiles of E. coli or other Enterobacteriaceae were shared by infant and mother; vertical transmission seemed to be uncommon, unlike findings in previous reports. In one hospital, 8 of 10 infant fecal E. coli strains shared a single plasmid profile, strongly suggesting nosocomial acquisition. In another, 7 of 9 neonate strains also shared a unique profile, and additionally carried K1 capsular antigen, a known virulence factor. Two other infants from the latter nursery acquired a urinary tract infection with E. coli K1 carrying the same plasmid profile. This study indicates that nosocomial acquisition of hospital strains of E. coli by neonates may be common in some hospitals and that the clinical implications are potentially serious.

Antigens, Bacterial↗
PubMed1993

Respiratory syncytial virus infection in lower respiratory tract and asthma attack in hospitalized children in North Hokkaido, Japan.

Respiratory syncytial virus (RSV) infection is severe and life-threatening in some infants. To investigate the epidemiology of RSV infection in hospitalized children in North Hokkaido, Japan, we tried to detect RSV antigen in nasopharyngeal aspirates (NPA) from those children with lower respiratory tract infection (LRTI) and asthma attack. From April 1991 to March 1992, 317 patients were hospitalized in our pediatric ward for the treatment of LRTI and asthma attack. The presence of RSV antigen in NPA taken from 283 patients (89.3%) were examined by enzyme immunoassay. RSV antigen was detected in 88 patients (31.1%). RSV LRTI were noted throughout the year, and the epidemic peak was observed in November and December. There was no significant correlation between the RSV antigen positive rate and mean temperature. RSV played an important role in LTRI in children in North Hokkaido, Japan. RSV LRTI in North Hokkaido was not rare in summer, indicating that RSV was transmitted commonly among children throughout the year.

Adolescent↗
PubMed1993

[Pharmacokinetics and clinical studies on flomoxef in neonates and premature infants. A study of flomoxef in the perinatal collaboration research group].

We investigated pharmacokinetics and clinical effects of flomoxef sodium (6315-S, FMOX) in neonates and premature infants. These results are summarized as follows: 1. Pharmacokinetics (1) Plasma concentration (Ct) and half-lives (T1/2) were determined upon after intravenous one-shot injection (i.v.) of FMOX to neonates of different day-age groups (0-3 (n = 25), 4-7 (n = 18), 8-28 (n = 32) days of birth). At a dose of 10 mg/kg. i.v., mean C30 (30 minutes concentration) values were 21.2, 21.8 and 21.3 micrograms/ml, respectively, in the different groups mentioned above, and the mean T1/2 values were 3.37, 1.85 and 1.63 hours. At 20 mg/kg i.v., mean C15 (15 minutes concentration) values were 54.4, 51.4 and 50.7 micrograms/ml, and mean T1/2's were 2.99, 2.32 and 1.79 hours, respectively. At a dose of 40 mg/kg i.v., mean C15 values were 104.0, 95.9 and 99.2 micrograms/ml, and the mean T1/2's were 3.40, 1.20 and 1.80 hours, respectively. (2) Plasma concentrations and T1/2 after intravenous one-shot injection of FMOX in premature infants in group (0-3 (n = 14), 4-7 (n = 10), 8-28 (n = 13) days of birth). Mean C15's at doses of 10, 20 and 40 mg/kg in the different groups of infants were 24.0, 28.6, 21.7 and 54.0, 54.6, 55.5 and 98.2, 93.0, 106.0 micrograms/ml, and T1/2's were 4.10, 2.53, 2.57 and 4.28, 2.27, 3.02 and 4.66, 2.86, 2.09 hours, respectively. Mean Cmax values were clearly dose dependent, and mean T1/2 values tended to be longer in premature infants compared to neonates. (3) Urinary recovery rate of FMOX after intravenous injection in neonates and premature infants. Mean urinary recovery rates of FMOX in the first 6 hours after i.v. (one-shot) at doses of 10, 20 and 40 mg/kg to neonates and premature infants were 38.9-62.8% in the neonates and 30.7-61.5% in the premature infants. (4) Plasma concentrations and urinary recovery rates upon 1 hour drip infusion of 20 mg/kg in the neonate groups (or the premature infant groups) as follows: Mean C50 values were 31.0, 32.7 and 23.4 micrograms/ml, and T1/2 were 2.94, 3.68 and 2.25 hours, respectively. The recovery rates were 35.2-52.9% in the first 6 hours after administration. 2. Clinical studies The number of clinically evaluable cases in the FMOX treatment of premature infants was 199, in which the causative pathogens were identified in 71 cases (A group) and not identified in 128 cases (B group).(ABSTRACT TRUNCATED AT 400 WORDS)

Cephalosporins↗
PubMed1992

Growth state and cell cycle dependent phosphorylation of DNA topoisomerase II in Swiss 3T3 cells.

We have investigated the amount of DNA topoisomerase II and phosphorylation of the enzyme in Swiss 3T3 cells during the transition from cell quiescence to proliferation. A relatively high level of phosphorylation was observed with proliferating cells while no or a very low level of phosphorylation was observed with quiescent cells. Phosphoamino acid analysis of the phosphorylated topoisomerase II revealed that the phosphorylated aminoacyl residue was serine. When quiescent cells were stimulated to grow by the addition of serum, DNA synthesis began to increase at 9 h after serum addition, reaching a maximum at 15 h and then declining. The amount of topoisomerase II began to increase at 6 h and reached a maximum at 22-27 h, corresponding to the G2 phase. The phosphorylation of topoisomerase II measured by pulse-labeling gradually increased from 6 to 18 h and reached a maximum at 22 h when the amount of the enzyme was maximum. The level of phosphorylation measured by continuous-labeling increased gradually up to 12 h and markedly up to 28 h, and then declined. The increase in the rate of phosphorylation in the G2 phase was affected by inhibiting DNA synthesis, but the increase in the amount of the enzyme was not. Thus, it was suggested that the regulation of phosphorylation of topoisomerase II differs from that of the amount of the enzyme.

3T3 Cells↗
PubMed1992

Cranio-nasal median splitting for radical resection of craniopharyngioma.

A new surgical approach for radical resection of craniopharyngioma is presented. This approach (cranio-nasal median splitting) involves craniotomy in the centre of the frontal bone, removal of the median portion of the supraorbital bar that incorporates the nasal bone, and detachment of the medial canthal ligaments. The frontal lobes, the cribriform plates, the planum sphenoidale, and the upper nasal cavities are split in the midline. The extraventricular surface of the hypothalamus, the pituitary stalk, and the posterior portion of the Willis' arterial ring are well visualized through the midline infrachiasmatic route. The intraventricular surface of the hypothalamus is also visible in the same operative field through the lamina terminalis and/or the anterior portion of the corpus callosum. This excellent visualization is quite helpful for minimizing operative injury to the hypothalamus and the pituitary stalk whichever surface of the third ventricular floor the tumour is situated upon. Three cases of craniopharyngioma operated upon by this approach are presented. Discussions are focused not only on the indication, but on the advantages and disadvantages of this approach. The surgical techniques for reconstruction of the cranial base are also described, together with some precautions that should be taken to prevent possible postoperative complications.

Adult↗