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Biomedical subjects

T Shimokawa

Publications and source records attributed to T Shimokawa.

At least 109 records · Page 6Linked to original sources
PubMed1992

Recombinant human granulocyte-colony-stimulating factor in the treatment of patients with chronic benign granulocytopenia and congenital agranulocytosis (Kostmann's syndrome).

Seven patients with chronic benign granulocytopenia and nine patients with congenital agranulocytosis, received consecutive seven-day courses of recombinant human granulocyte-colony stimulating factor at a starting dose of 50 micrograms/m2/day, subcutaneously. If there was no response the doses were increased to 300 micrograms/m2. All patients with chronic benign granulocytopenia responded rapidly at the minimum dose within 1-3 days after administration. By contrast, only three of the nine patients with congenital agranulocytosis responded within 1-7 days at this dose. Four patients with congenital agranulocytosis showed a response between days 7-19 at a dose of granulocyte-colony-stimulating factor 100-200 micrograms/m2 but in the remaining two cases no response was obtained. The administration of granulocyte-colony-stimulating factor was shown to be safe and effective also in reducing infectious episodes in these patients. Previously it was reported that granulocyte-colony-stimulating factor 10-30 micrograms/kg/day was effective for patients with congenital agranulocytosis. These results indicate that patients with congenital agranulocytosis may require much higher doses of recombinant human granulocyte-colony-stimulating factor than patients with chronic benign granulocytopenia and that the response to ordinary doses of recombinant human granulocyte-colony-stimulating factor may be useful in differentiating between chronic benign granulocytopenia and congenital agranulocytosis.

Adolescent↗
PubMed1992

Lipid lowering effects of high linoleate and high alpha-linolenate diets in rats and mice. Consequence of long-term feedings.

Diets high in linoleate (safflower oil) or high in alpha-linolenate (perilla oil) were fed to rats for 11 months, and the effects of the diets on plasma and tissue lipids were compared. The plasma levels of total cholesterol (Cho), phospholipids (PL) and triacylglycerol (TG) were significantly lower in the high alpha-linolenate group than in the high linoleate group, the differences being more than 30% in the levels of total Cho and TG. The diets had differential effects on the lipid contents of major tissues: the TG level in muscle was higher but both the TG level in depot fat and the PL level in muscle were lower in the high alpha-linolenate group than in the high linoleate group. In order to clarify whether or not the hypolipidemic effect of the high alpha-linolenate diet was due to changes in the distribution of lipids among tissues, whole body lipids were estimated in mice fed these diets for 5 months. The whole body Cho content was significantly lower, by 28%, in the high alpha-linolenate group compared with the high linoleate group, but the total lipid content, PL and neutral lipids were similar between the groups. Our results indicate that the high alpha-linolenate diet has a more potent cholesterol lowering effect in plasma and body tissue than the high linoleate diet; interestingly, whole body TG levels are similar but tissue distributions of TG are different between the two dietary groups.

Animals↗
PubMed1991

Essential histidines of prostaglandin endoperoxide synthase. His-309 is involved in heme binding.

Prostaglandin endoperoxide (PGH) synthase has a single iron protoporphyrin IX which is required for both the cyclooxygenase and peroxidase activities of the enzyme. At room temperature, the heme iron is coordinated at the axial position by an imidazole, and about 20% of the heme iron is coordinated at the distal position by an imidazole. We have used site-directed mutagenesis to investigate which histidine residues are involved in PGH synthase catalysis and heme binding. Individual mutant cDNAs for ovine PGH synthases were prepared with amino acid substitutions at each of 13 conserved histidines. cos-1 cells were transfected with each of these cDNAs, and the cyclooxygenase and peroxidase activities of the resulting microsomal PGH synthases were measured. Mutant PGH synthases in which His-207, His-309, or His-388 was replaced with either glutamine or alanine lacked both activities. Gln-386 and Ala-386 PGH synthase mutants exhibited cyclooxygenase but not peroxidase activities. Other mutants exhibited both activities at varying levels. Because binding of heme renders native PGh synthase resistant to cleavage by trypsin, we examined the effects of heme on the relative sensitivities of native, Ala-204, Ala-207, Ala-309, Ala-386, and Ala-388 mutant PGH synthases to trypsin as a measure of the heme-protein interaction. The Ala-309 PGh synthase mutant was notably hypersensitive to tryptic cleavage, even in the presence of exogenous heme; in contrast, the native enzyme and the other alanine mutants exhibited similar, lower sensitivities toward trypsin and, except for the Ala-386 mutant, were partially protected from trypsin cleavage by heme. Preincubation of the native and each of the alanine mutant PGH synthases, including the Ala-309 mutant, with indomethacin protected the proteins from trypsin cleavage. Thus, all the mutant proteins retain sufficient three-dimensional structure to bind cyclooxygenase inhibitors. Our results suggest that His-309 is one of the heme ligands, probably the axial ligand, of PGH synthase. Two other histidines, His-207 and His-388, are essential for both PGH synthase activities suggesting that either His-207 or His-388 can serve as the distal heme ligand; however, the trypsin cleavage measurements imply that neither His-207 nor His-388 is required for heme binding. This is consistent with the fact that only 20% of the distal coordination position of the heme iron of PGH synthase is occupied by an imidazole side chain.

Base Sequence↗
PubMed1991

[Serum monitoring of methotrexate (MTX) and 7-hydroxymethotrexate concentrations in patients treated with MTX using high-pressure liquid chromatography (HPLC) and comparison of serum MTX levels between HPLC method and fluorescence polarization immunoassay (FPIA)].

In order to measure simultaneously the serum levels of methotrexate (MTX) and its major metabolite, 7-hydroxymethotrexate (7-OH-MTX), in samples obtained from patients treated with MTX, we have investigated the reversed-phase high-pressure liquid chromatographic assay using ion-pairing reagents. The mobile phase consisted of 77.5% solution of 0.005M tetrabutylammonium and 22.5% acetonitrile. SEP-PAK C18 Cartridges were used for the precolumn. The detectable range of MTX and 7-OH-MTX were 0.02-0.03 and 1.0 mumol/l respectively. A significant positive correlation was observed (r = 0.983) between FPIA and HPLC methods. Serum MTX levels with FPIA were significantly (p less than 0.05) higher than those of HPLC method. The serum 7-OH-MTX levels at 24 hr and 48 hr were 4.857 +/- 1.383 (n = 10) and 1.835 +/- 0.286 (n = 6) mumol/l respectively with the dosage of 400 mg/m2. The serum 7-OH-MTX levels at 48 hr were 6.254 +/- 3.053 mumol/l (n = 5) with the dosage of 3,000 mg. The serum half lives of MTX and 7-OH-MTX were 8.05 +/- 1.03 (n = 4) and 14.8 +/- 1.35 (n = 6) hours respectively between 24 hr and 48 hr after administration. The T1/2 7-OH-MTX/MTX ratio was 1.8. Percent cross-reactivity of 7-OH-MTX with concentrations ranging from 1-10 mumol/l were 0.6-2.0% by FPIA. However, patients' serum levels of 7-OH-MTX were 15-85 times (n = 21) higher than those of MTX. MTX levels of containing both MTX and 7-OH-MTX (7-OH-MTX/MTX ratio was 50/1) were significantly higher than those of containing MTX alone by FPIA.

Adult↗
PubMed1990

Tyrosine 385 of prostaglandin endoperoxide synthase is required for cyclooxygenase catalysis.

There are spectral and biochemical data suggesting that a tyrosine group(s) is involved in the cyclooxygenase reaction catalyzed by prostaglandin endoperoxide (PGH) synthase. Treatment with tetranitromethane, a reagent which nitrates tyrosine residues, abolishes cyclooxygenase activity, but this inactivation can be largely prevented by competitive cyclooxygenase inhibitors such as ibuprofen and indomethacin. To identify sites of nitration, native PGH synthase and indomethacin-pretreated PGH synthase were incubated with tetranitromethane, and the sequences of peptides containing nitrotyrosine were determined. Three unique tyrosines (Tyr-355, Tyr-385, and Tyr-417) were nitrated in the native enzyme but not in the indomethacin-treated PGH synthase. Using site-directed mutagenesis of sheep PGH synthase, each of these tyrosines, as well as two other tyrosine residues selected as controls (Tyr-254 and Tyr-262), were replaced with phenylalanine; cos-1 cells were transfected with constructs containing cDNAs coding for the native PGH synthase and each of the five phenylalanine mutants, and microsomes from these cells were assayed for cyclooxygenase and hydroperoxidase activities. The Phe-385 mutant of PGH synthase lacked cyclooxygenase activity but retained peroxidase activity; all other mutants expressed both enzyme activities. Our results establish that Tyr-385 is essential for the cyclooxygenase activity of PGH synthase and that nitration of this residue can be prevented by indomethacin. We conclude that Tyr-385 is at or near the cyclooxygenase active site of PGH synthase and could be the tyrosine residue proposed to be involved in the first step of the cyclooxygenase reaction, abstraction of the 13-proS hydrogen from arachidonate.

Animals↗
PubMed1990

Successful chemotherapy on a pregnant non-Hodgkin's lymphoma patient.

We report a case of a non-Hodgkin's lymphoma (NHL) patient treated successfully with combination chemotherapy during pregnancy who delivered a full-term baby. A 29 year-old patient with cervical and inguinal lymphadenopathy in the 27th week of gestation was referred to our hospital. The diagnosis of lymph node biopsy was NHL (diffuse, large cell type with B-cell phenotype). Three courses of CHOP regimen (adriamycin, cyclophosphamide, vincristine and prednisolone) were given before delivery. The patient has been in complete remission for three years and her baby has been in normal development. Our case supports previous reports that chemotherapy in the third trimester may be given safely on NHL patients.

Adult↗
PubMed1990

[Expansion of peripheral blood progenitors by recombinant human granulocyte colony-stimulating factor].

The levels of peripheral progenitor cells was measured serially after cancer chemotherapy in 4 patients with non-Hodgkin's lymphoma and one patient with rhabdomyosarcoma who received recombinant human granulocyte colony-stimulating factor (rG-CSF). This study was composed of two independent phases: in the first phase, patients received a course of cytotoxic chemotherapy only, and in the second phase, they received the same chemotherapy followed by subcutaneous injection of rG-CSF (2 micrograms/kg/day) for 10-14 days. In the control phase, the levels of granulocyte-macrophage colony-forming units (CFU-GM) and erythroid burst-forming units (BFU-E) per milliliter increased during the early recovery phase, but rG-CSF treatment increased the number of CFU-GM 3 to 18-folds, and the number of BFU-E increased 1.3 to 4.6-folds. An overshoot in the blood progenitor levels occurred at the day 8-10 of rG-CSF administration. And then, the peak of neutrophil count followed 3-5 days later. After the discontinuation of rG-CSF, the number of blood CFU-GM and BFU-E fell rapidly. This results suggest that in vivo expansion of circulating hemopoietic progenitors can be achieved by the administration of rG-CSF, and this approach might be clinically applicable to cancer patients who are a candidate of peripheral blood stem cell autotransplantation.

Adult↗
PubMed1990

[A successful case of congenital agranulocytosis treated with recombinant human granulocyte colony-stimulating factor].

A 1-month-old girl was admitted because of staphylococcal cellulitis of the buttock and the shoulder, and peripheral agranulocytosis. CBC on admission showed 10,100/microliters of WBC with 1% mature neutrophils, 5% monocytes, 6% eosinophils, 2% basophils and 86% lymphocytes. Bone marrow aspiration revealed maturation arrest of neutrophil precursors at the level of myelocyte. We treated the patient with subcutaneous rhG-CSF (Kirin-Amgen, Tokyo) for sequential 7-day course at the starting dose of 3 micrograms/kg, and increased weekly. The dose was escalated at the level of 18 micrograms/kg for 2 weeks subcutaneously, 8 days after effective dose of 18 micrograms/kg, the absolute neutrophil counts more than 1,000/microliters was attained, and bone marrow aspiration showed an increase of neutrophil precursors beyond the myelocyte level with maturation. Our case proved that both WBC and absolute neutrophil counts were increased parallel with the dose escalation of rhG-CSF. Shortly after the cessation of rhG-CSF, WBC and absolute neutrophil counts were decreased. No side effect was detected except for mild splenomegaly which was resolved after cessation of rhG-CSF. In methylcellulose culture with PHA-LCM, marrow cell of patient produced normal number of CFU-GM, and myeloid precursors could proliferate and differentiate to normal polymorphonuclear neutrophils, but rhG-CSF produced only small number of CFU-GM. Our case confirms that rhG-CSF is a new approach to control the life-threatening infection of congenital agranulocytosis.

Agranulocytosis↗
PubMed1990

[Successful treatment of a case of hepatitis-associated severe aplastic anemia by anti-lymphocyte globulin (ALG)].

A 7-year-old girl was admitted to our hospital because of fever and multiple petechiae following non-A non-B hepatitis. Peripheral blood on admission showed pancytopenia, and she was diagnosed to have hepatitis-associated severe aplastic anemia. She was treated with oxymetholone, prednisolone, 2 courses of bolus methylprednisolone, 2 courses of high dose gamma-globulin therapy and ALG (Pressimmun) without success. Nineteen months after diagnosis. ALG (Lymphoser) and bolus methylprednisolone followed by oxymetholone and prednisolone were tried. Hematologic conditions improved gradually, and have become normal except mild thrombocytopenia over a year. This case suggests that there is a difference in clinical efficacy between ALG preparations, and this observation gives the basis of repeating ALG therapy using a different preparation even if the preceding one has failed.

Anemia, Aplastic↗
PubMed1990

Structure-function relationships in sheep, mouse, and human prostaglandin endoperoxide G/H synthases.

Our studies are designed to determine which amino acid residues are involved in catalyzing the cyclooxygenase and hydroperoxidase activities of prostaglandin endoperoxide (PGG/H) synthase. We have deduced from complementary (c)DNAs the amino acid sequences of the sheep and mouse PGG/H synthases, and a portion of the human PGG/H synthase. These enzymes have amino acid sequences which are about 90% identical. Sequence similarities with putative heme binding regions of myeloperoxidase and thyroid peroxidase suggest that the sequence TI(L)WLREHNRV of PGG/H synthase contains the histidine (His309) which is the proximal heme ligand; the distal heme ligand may be His226 which is found in the sequence 222-KALGH-226. Using site-directed mutagenesis, we have replaced Ser530, the serine residue which is acetylated by aspirin, with Ala530 and with Asn530; the Ala530 mutant has both cyclooxygenase and hydroperoxidase activity, while the Asn530 mutant lacks cyclooxygenase activity but retains hydroperoxidase activity. These results establish that the hydroxyl group of Ser530 is not essential for catalysis or substrate binding and suggest that a bulky group at position 530, such as that introduced by aspirin acetylation, prevents arachidonate binding to the cyclooxygenase active site. Finally, we have found that tetranitromethane causes irreversible inactivation of cyclooxygenase activity and that the enzyme is protected from inactivation when ibuprofen is included in the reaction mixture. These results suggest that there is an essential tyrosine at the active site of PGG/H synthase.

Amino Acid Sequence↗
PubMed1990

Molecular basis for the inhibition of prostanoid biosynthesis by nonsteroidal anti-inflammatory agents.

Many nonsteroidal anti-inflammatory drugs exert their effects by inhibiting the synthesis of prostanoids. More specifically, these agents block the synthesis of prostaglandin endoperoxide G2 from arachidonic acid by competing with arachidonate for binding to the cyclooxygenase active site of prostaglandin endoperoxide synthase. Studies of the molecular biology of prostaglandin endoperoxide synthase indicate that there is a single gene for the enzyme. Thus, tissue-specific effects of nonsteroidal anti-inflammatory drugs probably result from differences in drug distribution and/or metabolism and not from the existence of tissue-specific prostaglandin endoperoxide synthase isozymes. Aspirin causes inactivation of prostaglandin endoperoxide synthase by first binding to the cyclooxygenase active site and then acetylating the protein at Ser530. Although the cyclooxygenase activity is inactivated, the hydroperoxidase activity of prostaglandin endoperoxide synthase is unaltered by Aspirin or other nonsteroidal anti-inflammatory drugs. Replacement of Ser530 of the native enzyme with an alanine residue by site-directed mutagenesis yields a prostaglandin endoperoxide synthase with unaltered catalytic and substrate binding activities. Thus, the hydroxyl group of Ser530 is not essential for enzyme activity. Instead, it appears likely that acetylation of prostaglandin endoperoxide synthase by Aspirin simply places a bulky acetyl group at or near the cyclooxygenase active site, thereby interfering with arachidonic acid binding.

Acetylation↗
PubMed1989

Sustained cytopenia after leukapheresis for collection of peripheral blood stem cells in small children.

Marked and persistent cytopenia was observed in two 3-year-old children with cancer, after lymphopheresis to collect peripheral blood stem cells (PBSC) for autotransplantation after high-dose chemotherapy. This finding suggests that PBSC are a major component of the hematopoietic cells in small children and, hence, that special care is necessary in the construction of a protocol for collection of stem cells from young children.

Child, Preschool↗
PubMed1989

Hypertension does not stimulate the development of hypercholesterolemia or fatty liver induced by a high cholesterol/cholate diet in rats.

A high cholesterol/cholate diet induced hypercholesterolemia and fatty liver in both spontaneously hypertensive rats (SHR) and normotensive control rats (WKY). However, in contrast to previous concepts, the levels of cholesterol ester, triacylglycerol and phosphatidylcholine in plasma as well as triacylglycerol in liver were higher in WKY than in SHR fed a normal diet. The high cholesterol/cholate diet elevated the levels of plasma cholesterol, plasma cholesterol ester and hepatic triacylglycerol, and the extent of elevation was significantly higher in WKY than in SHR. Increases both in monoene/saturated ratios, an indication of elevated delta 9-desaturase activity, and in linoleate/arachidonate ratios, a possible indication of impaired desaturation-elongation activity, were observed in hepatic and plasma lipids of both strains fed the high cholesterol/cholate diet. The increases in monoene/saturated ratios were similar in both strains, but the increases in the linoleate/arachidonate ratios were higher for the plasma cholesterol esters of WKY than of SHR. The n-6/n-3 ratios of plasma and hepatic lipids were higher in WKY than in SHR throughout the experiments. These diet-induced changes observed in hepatic and plasma lipids were not reflected in the aortic lipids. Thus, hypertension per se does not promote the development of hyperlipemia and fatty liver induced by a high cholesterol/cholate diet. Our results also suggest that the metabolism of polyenoic fatty acids is different between SHR and WKY.

Animals↗
PubMed1989

Surgical treatment of advanced carcinoma of the esophagus.

This report was done to evaluate the role of surgical treatment of locally advanced carcinoma of the esophagus. All 32 patients had histologically proved epidermoid carcinomas of the esophagus. The post-operative one and two year survival rates and median survival time were 34.6 and 17.3 per cent and 199 days for those in the group undergoing esophagectomy, zero per cent and 55 days for those in the group with bypass and zero per cent and 80 days for those undergoing formation of a stoma. Thirty day postoperative mortality rates were zero per cent for the esophagectomy group, 25.0 per cent for the stoma group and 33.3 per cent for the bypass group. When calculated from the date of diagnosis, the one and two year survival rates and the median survival time were 50.2 and 16.7 per cent and 296 days for the esophagectomy group, 16.7 and zero per cent and 196 days for the bypass group, 25.0 and zero per cent and 142 days for the stoma group and zero per cent and 142 days for the group who did not undergo surgical treatment. Complication related postoperative deaths occurred in 14.8 per cent of the 27 patients operated upon. For those in the esophagectomy group, it was 5.9 per cent, and for the bypass group, 50.0 per cent. Swallowing was improved in 58.8 per cent after esophagectomy, but in none of the patients was swallowing restored after the bypass procedures. When the invasion spared the aorta or major respiratory tract, postoperative survival time after esophagectomy was significantly better. Considering the short postoperative survival time and frequent complications, bypass procedure should not be the final resort after nonsurgical treatment.

Actuarial Analysis↗
PubMed1989

Quantitative analysis of the effect of colony-stimulating factors on human marrow progenitor growth in liquid-suspension cultures: application of limiting dilution assay.

Proliferation of human marrow progenitors in liquid cultures can be quantitated by limiting dilution clonal analysis (LDA) of progenitors in microwells. In this study, we have used LDA to study the effect of purified native or recombinant granulocyte colony-stimulating factors (G-CSFs) and recombinant granulocyte-macrophage CSF (GM-CSF) on progenitor growth. These results were compared to those of simultaneous cultures in methylcellulose. In LDA, single-hit kinetics were obtained with up to 500 U/ml of the recombinant preparation. In LDA with recombinant GM-CSF, progenitor growth conformed to single-hit kinetics from 100 to 2000 U/ml with maximum progenitor frequency at 500 U/ml. In simultaneous methylcellulose cultures with recombinant GM-CSF, colony formation reached a plateau at 100 U/ml. In LDA, purified native G-CSF was shown to support progenitor growth with single-hit kinetics at 100 U/ml, but at greater concentrations (greater than 150 U/ml), it suppressed progenitor growth with almost complete inhibition at a concentration of 200 U/ml. However, this dose-response effect was not observed in either simultaneous methylcellulose culture with G-CSF or in LDA with a purified recombinant preparation of the corresponding G-CSF. In methylcellulose cultures, colony formation reached a maximum at 100 U/ml and maintained a plateau up to 1000 U/ml. Hence, liquid culture allowed detection of contaminating suppressive activity in the G-CSF preparation that was not detected by methylcellulose assay. LDA may be more sensitive than methylcellulose culture for screening factors regulating human hematopoietic cell growth.

Bone Marrow↗
PubMed1989

Peripheral blood stem cell autotransplantation in treatment of childhood cancer.

The levels of circulating hematopoietic progenitor cells were measured sequentially in eight children receiving chemotherapy for acute leukemia or neuroblastoma. Significant increases in the progenitor levels (up to 50-fold in CFU-GM numbers) were observed during post-chemotherapy cytopenia in all cases, but differences among individuals in the kinetics of recovery of less committed progenitors (CFU-mix) contrasted with the synchronized-mode of expansion of committed progenitors (CFU-GM). Peripheral blood cells were collected by repeated continuous-flow leukaphereses from three of the children during post-chemotherapy expansion of the progenitor pool and were cryopreserved after fractionation procedures. Infusion of these stored cells into the patients after marrow-ablative chemotherapy established trilineage hematopoiesis. This use of stem cell rescue should be useful as an alternative to bone marrow transplantation and extends the application of cure-oriented salvage therapy to childhood cancers.

Blood Cell Count↗