Biomedical subjects

A Kubota

Publications and source records attributed to A Kubota.

At least 109 records · Page 6Linked to original sources

PubMed1996

Granulocyte colony-stimulating factor-induced mobilization of peripheral blood stem cells for autologous and allogeneic transplantation. Fukuoka Bone Marrow Transplantation Group.

Peripheral blood stem and progenitor cells (PBSC and PBPC), which circulate at very low levels during steady-state hematopoiesis, show a transient but marked increase during hematologic recovery from marrow-suppressive chemotherapy. To ensure rapid and sustained hematologic engraftment after autologous PBSC transplantation, sufficient PBSC or PBPC must be infused. To confirm the utility of granulocyte colony-stimulating factor (G-CSF) in chemotherapy-induced PBSC mobilization, we investigated the effect of G-CSF on PBSC mobilization in leukemia and lymphoma patients. The study design was such that PBSC mobilization with and without G-CSF was assessed in the same patients. The results indicate that PBSC mobilization can be enhanced significantly when G-CSF is given during the recovery phase postchemotherapy. Interestingly, progenitor cells of different lineages could be mobilized by G-CSF. We subsequently investigated the effect of increasing G-CSF dose on PBSC mobilization during steady-state hematopoiesis in healthy adult donors. The results indicate that not only committed but also primitive progenitor cells are mobilized into the circulation in a dose-and time-dependent manner when G-CSF at 5, 10, or 15 micrograms/kg was given on each of 5 days and leukapheresis was performed on day 6. From our data we estimate that sufficient PBSC for engraftment after allogeneic PBSC transplantation can be collected on day 5 of administration of G-CSF at 10 micrograms/kg and by 10-1 leukapheresis on days 5 and 6. Furthermore, we found that some G-CSF-mobilized PBSC retained their self-renewal capability. These observations suggest that hematopoietic stem cells for allogeneic PBSC transplantation can be mobilized by short-term administration of relatively high-dose G-CSF.

Animals↗

PubMed1996

Ultrastructural analysis of survival in cultured smooth muscle cells isolated from stroke-prone spontaneously hypertensive rats: effect of growth factors.

Our previous study in vivo suggested that vascular smooth muscle cells (VSMCs) in stroke-prone spontaneously hypertensive rats (SHRSP) were vulnerable when plasma components were deficient. Therefore, we cultured VSMCs isolated from normotensive and hypertensive rats to clarify the weakness of VSMCs isolated from hypertensive rats and maintained in plasma-deficient conditions by employing ultrastructural and biochemical analyses. VSMCs, obtained from normotensive rats and cultured without fetal bovine serum (FBS) for 1 week, were intact and well differentiated; without FBS for 2 weeks retained their original structures except for several degenerative changes. VSMCs, obtained from hypertensive rats and cultured without FBS for 2 weeks, were extensively damaged and lost their cell organelles. Apoptotic bodies were frequently observed. We also cultured VSMCs in medium containing a variety of growth factors. VSMCs obtained from normotensive rats and cultured with epidermal growth factor or insulin-like growth factor-1 for 2 weeks were almost intact, as were VSMCs from hypertensive rats, although some degenerative changes of cell organelles were observed. VSMCs from hypertensive rats, maintained with platelet-derived growth factor-BB or basic fibroblast growth factor, were generally in poor condition. Thus VSMCs from hypertensive rats have hereditary weaknesses in cell survival including apoptosis and require specific growth factors for their maintenance.

Animals↗

PubMed1996

Prenatal diagnosis of abdominal wall defects and their prognosis.

To determine whether associated anatomic features may be useful in predicting fetal prognosis, 43 fetuses who had ultrasonographic evaluation for abdominal wall defects during a 13-year period were reviewed. Thirty-one fetuses had omphalocele and were classified into three groups according to fetal ultrasonography results: ruptured omphalocele with exposed liver, giant omphalocele with exposed liver, and small omphalocele without liver herniation. Twelve fetuses had gastroschisis. Ten of the 12 fetuses with gastroschisis survived; one died in utero. Nine of the 12 with a small omphalocele survived. Ten of 12 fetuses with giant omphalocele survived, but six of the seven with ruptured omphalocele died of pulmonary hypoplasia and respiratory insufficiency. Ruptured omphalocele recognized in utero was accompanied by intrauterine growth retardation and liver herniation and frequently was associated with deformity of the spine, diaphragmatic defects, vesicointestinal fissure, and meningocele. These results indicated that, together with the deformity of the spine, rupture and absence of the covering membrane with an exposed liver (noted through fetal ultrasonography) may suggest a poor prognosis because of pulmonary hypoplasia.

Abdominal Muscles↗

PubMed1996

Effector coupling of somatostatin receptor subtypes on human endocrine tumors.

Effector coupling of somatostatin receptor subtypes sst1 and sst2 was examined in a reconstituted system. Forskolin-stimulated cyclic adenosine monophosphate (cAMP) formation was inhibited 66% by somatostatin (SRIF-14) in CHO cells expressing somatostatin receptor 1(sst1) (CHO-SR1), but not sst2, in a dose-dependent manner with an ED50 of 1 x 10(-9) mol/L SRIF-14. The inhibition was blocked by pertussis toxin (PTX), indicating that sst1 is coupled to adenylyl cyclase via PTX-sensitive Gi protein. In CHO cells, Gi alpha 2 and Gi alpha 3 mRNAs were detected. In adenylyl cyclase assays, 1 mumol/L SRIF-14 caused a 16% inhibition of forskolin-stimulated adenyly cyclase activity. Preincubation with Gi alpha 3, but not Gi alpha 1/Gi alpha 2, antiserum blocked this inhibition. By contrast, sst2 is coupled to adenylyl cyclase via Gi alpha 1. In cells expressing sst2 with Gi alpha 1(CHO-SR2G1), SRIF-14 significantly inhibited forskolin-stimulated cAMP formation by 53% and with an ED50 at 4 x 10(-9)mmol/L SRIF-14, which was completely blocked by PTX; ED50 values for sst1 and sst2 agree with the IC50 values in binding assays. In CHO-SR1, the rank of potency of agonists affecting adenyl cyclase was SRIF-14 = SRIF-28 > RC 160 > SMS 201-995. In CHO-SR2G1, the rank was RC-160 > SRIF-14 = SRIF-28 > SMS 201-995.

Adenylate Cyclase Toxin↗

PubMed1996

Generation of human natural killer cells from peripheral blood CD34+ cells mobilized by granulocyte colony-stimulating factor.

We studied the generation of human natural killer (NK) cells from CD34+ cells that were isolated from peripheral blood stem cells (PBSC) mobilized by granulocyte colony-stimulating factor (g-CSF). The isolated CD34+ cells were cultured in the presence of a combination of interleukin-1 (IL-1alpha), IL-2, and stem cell factor for 5 weeks without marrow stroma. We found that the CD34+ cells isolated from G-CSF-mobilized PBSC (G-CSF/PBSC) could differentiate into a population of NK cells which were CD56+(bright)/CD3- and showed morphologic characteristics of large granular lymphocytes. Immunophenotypic analysis of the NK cells thus generated showed that a small proportion of them expressed CD2, CD8 and CD16 surface markers and approximately half of them coexpressed CD7. This NK population exhibited cytotoxic activity against a NK-sensitive cell line, K562. These observations suggest that CD34+ cells from G-CSF/PBSC contain precursors of NK cells that can differentiate into functional NK cells.

Adult↗

PubMed1996

Immunophenotyping of peripheral blood mononuclear cells collected by cytotoxic and cytotoxic/G-CSF mobilization.

We investigated surface immunophenotypes of peripheral blood mononuclear cells (PBMC) collected by cytotoxic and cytotoxic/G-CSF mobilization of peripheral blood stem cells (PBSC) from 38 patients with haematological malignancies in complete remission who underwent consolidation chemotherapy. PBMC were collected by leucapheresis during the haematopoietic recovery phase after intensive chemotherapy. G-CSF was used for mobilization of PBSC in 19 cases. Surface immunophenotyping of frozen-thawed PBMC was performed by flow cytometry. Our findings showed that monocytes and T cells were the two major cell components of PBMC. There were very few B cells in PBMC. Expression of CD45RO and HLA-DR was elevated in lymphocytes, suggesting that T cells in PBMC were activated. The percentage of CD34 positive cells were significantly increased in PBMC collected by cytotoxic/G-CSF mobilization (group 1) compared with PBMC collected by cytotoxic mobilization (group 2). There were significantly higher percentages of CD14 and CD33 positive cells in group 1 than in group 2. The percentage of CD4 positive lymphocytes positive for HLA-DR was significantly higher in group 1 compared with group 2. These observations indicated that PBMC contained a large number of monocytes and activated T cells, especially in cytotoxic/G-CSF mobilization.

Adolescent↗

PubMed1996

Detection of an aberrant fragment in growth hormone gene cluster in a girl with congenital generalized lipodystrophy.

The etiology of congenital generalized lipodystrophy (CGL) is unknown, but one of the clinical features is an accelerated rate of growth. In order to determine whether this abnormality is related to an abnormality of the GH or the IGF-I gene, we studied the DNA of lymphocytes of a girl with CGL. The IGF-I gene autoradiographic patterns of this patient were found not to differ from those of normal subjects, but after BamHI digestion, this patient's GH gene cluster had an aberrant 5.7 kb fragment in addition to the constant 8.3, 6.7, 5.3, 3.8, 2.9 kg fragments. The new 5.7 kb fragment was not detected in any of 60 unrelated Japanese individuals. In conclusion, the CGL patient carries an abnormal GH gene cluster, but further studies are needed to clarify whether this abnormal fragment is specific to CGL or just a rare polymorphic fragment.

Autoradiography↗

PubMed1996

G-CSF-induced mobilization of peripheral blood stem cells from healthy adults for allogeneic transplantation.

We investigated a dose-escalation effect of G-CSF (5, 10, and 15 micrograms/kg) on mobilization of committed and primitive hemopoietic progenitor cells, including CFU-GM, BFU-E, and long-term culture-initiating cells (LTC-IC) in addition to CD34+ cells and yields of progenitor cells in PBSC harvests obtained by leukapheresis of healthy adult donors. Results indicate that the mobilization of these progenitor cells is both dose and time dependent. Despite the very small number of healthy donors studied, it is estimated from our data that a sufficient number of CD34+ cells for allogeneic PBSC transplant (PBSCT) could be collected using a 5 day administration of 10 micrograms/kg of G-CSF to normal adult donors. Adverse effects include general fatigue and bone pain in most of the donors and fever and headache in some. These symptoms were well tolerated in most instances. Laboratory test abnormalities, including transient thrombocytopenia, increased platelet aggregation, and increased serum levels of some liver enzymes, were induced by G-CSF administration, but all were reversible within a short time. These observations suggest that hemopoietic stem cells for allogeneic PBSCT can be mobilized by short-term administration of a relatively high-dose G-CSF.

Adult↗

PubMed1996

Identification of two missense mutations in the GIP receptor gene: a functional study and association analysis with NIDDM: no evidence of association with Japanese NIDDM subjects.

Gastric inhibitory polypeptide (GIP) potently stimulates insulin secretion from pancreatic islets in the presence of glucose as an incretin. Because the insulinotropic effect of GIP is reduced in NIDDM, it should be clarified whether defects in the GIP receptor gene contribute to the impaired insulin secretion in NIDDM. Using genomic DNA samples from Japanese NIDDM and non-NIDDM subjects, we have investigated the entire coding region of the GIP receptor gene by polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP). We have identified two missense mutations, Gly198-->Cys (Gly198Cys) in exon 7 and Glu354-->Gln (Glu354Gln) in exon 12. Investigation of the function of GIP receptor with either of these mutations reveals a half-maximal stimulation value of GIP-induced cAMP response in Chinese hamster ovary cells expressing the GIP receptor with Gly198Cys of 6.3 +/- 1.2 x 10(-10) mol/l (n = 3), which was considerably higher than that of the normal GIP receptor, 9.4 +/- 3.8 x 10(-12) mol/l GIP (n = 3), whereas that of the GIP receptor with Glu354Gln was not significantly different from that of the normal GIP receptor. To assess the possible role of the GIP receptor gene in genetic susceptibility to NIDDM, we have examined the allelic frequencies of Gly198Cys and Glu354Gln in NIDDM and control subjects. Association studies show no relationship between NIDDM and either of the two mutations.

Alleles↗

PubMed1996

E2011 a novel, selective and reversible inhibitor of monoamine oxidase type A.

E2011, (5R)-3-[2-((1S)-3-cyano-1hydroxypropyl)benzothiazol- 6-yl]-5-methoxymethyl-2-oxazolidine, is a novel inhibitor of monoamine oxidase type A (MAO-A). We have characterized the neurochemical and pharmacological profiles of E2011 and compared them with those of known inhibitors of MAO-A. E2011 potently inhibited MAO-A with more than 30,000 times higher selectivity for MAO-A relative to MAO-B in rat brain homogenate. E2011 did not affect putative neural receptors or reuptake of biogenic amines into synaptosomes of rat brain, which suggests that it is specific to monoaminergic systems. In vivo, E2011 at a dose of 0.3 mg/kg p.o. exhibited potent MAO-A inhibitory activity, whereas MAO-B inhibition was not observed even at 100 mg/kg p.o. E2011 inhibited monoamine metabolism in the rat brain, but the effect disappeared 24 h after administration. Like other reversible MAO-A inhibitors, E2011 did not show a cumulative inhibitory effect during repeated administration for 7 days. However, inhibition of MAO-A by E2011 in ex vivo experiments appeared to be less potent than that by moclobemide. The MAO-A inhibition by E2011 was partially but significantly reversed by dialysis at 4 degrees C for 24 h, which indicates that E2011 could be dissociated from the enzyme. These findings suggest that E2011 is a reversible and highly selective inhibitor of MAO-A. The potency of inhibition by highly reversible MAO-A inhibitors such as E2011 is likely to be underestimated in ex vivo studies because of dilution of the homogenate in the assay system.

Animals↗

PubMed1995

The 10-Hz rhythm in the sympathetic nerve activity of cats, rats and rabbits.

Since the 10-Hz rhythmic activity in the sympathetic nerves was reported only in cats, we examined whether the activity of the same frequency could be observed in rats and rabbits as well as in the cats. Histograms of inter-burst-peak intervals of discharges of the renal nerves revealed that the 100 ms interval activity, the reverse of the 10-Hz, was observed in all the three mammals, of which the baroreceptor afferents were intact or inactivated. Further, the same frequency activity could be evoked by intermittent electrical stimulation of the dorsolateral funiculus of the cervical cord in spinal animals. It was suggested that the 10-Hz rhythmic activity in the sympathetic nerves was a common phenomenon throughout mammals and this activity was produced in the spinal cord. The physiological significance of the 10-Hz activity in the sympathetic nerves was discussed.

Afferent Pathways↗

PubMed1995

The structures of the human calcium channel alpha 1 subunit (CACNL1A2) and beta subunit (CACNLB3) genes.

Calcium influx in pancreatic beta-cells is regulated mainly by L-type voltage-dependent calcium channels (VDCCs) and triggers insulin secretion. The alpha 1 subunit (CACN4) and the beta subunit (beta 3) of VDCCs, both of which are expressed in pancreatic islets, are major components for the VDCC activity, and so they may play a critical role in the regulation of insulin secretion. We have determined the structures of the human CACN4 (CACNL1A2) and the human beta 3 (CACNLB3) genes. The CACNL1A2 gene spans more than 155 kb and has 49 exons. Most of the positions interrupted by introns are well conserved between the CACNL1A2 gene and the previously reported L-type VDCC alpha 1 subunit, CACNL1A1, gene. On the other hand, the CACNLB3 gene distributes in approximately 8 kb and comprises 13 exons, most of which are located together within approximately 5 kb. Comparisons of the genomic sequences of CACNL1A2 with the previously reported cDNA sequences indicate that there are a number of polymorphisms in the human CACNL1A2 gene. In addition, the PCR-SSCP procedure of exon 1 of CACNL1A2 revealed a change from 7 to 8 ATG trinucleotide repeats in a patient with non-insulin-dependent diabetes mellitus (NIDDM), resulting in an addition of methionine at the amino-terminus of CACN4. The determination of the structures of the human CACNL1A2 and CACNLB3 genes should facilitate study of the role of these genes in the development of NIDDM and also other genetic diseases such as long QT syndrome.

Base Sequence↗

PubMed1995

Effects of TPN on brain, liver, and food intake in rats.

Postulating that total parenteral nutrition (TPN) has a direct metabolic effect in the brain which may be similar to that in the liver, we studied the effects of TPN on brain glycogen and triglyceride in rats. Control rats (n = 24) received normal saline via jugular catheter for 18 days; TPN rats (n = 24) had normal saline for 10 days and then TPN-100 for 4 days, followed by normal saline for 4 more days. TPN-100 (caloric ratio glucose: fat:amino acid = 50:30:20) provided 100% of the rat's daily caloric needs. Chow and water were available ad libitum during the study. Eight rats from each group were sacrificed after 1 and 4 days of TPN-100, and 4 days after stopping TPN-100. Glycogen in brain and liver glycogen, triglyceride, and glycogen synthetase and phosphorylase were measured, as were plasma glucose and insulin. Data were evaluated using Student's t test. With TPN, the following occurred: (a) an 85% decrease (P < 0.05) of spontaneous food intake; (b) an elevated plasma glucose; (c) a three- to fivefold increase (P < 0.01) in plasma insulin; (d) a 23% increase (P < 0.05) in whole brain glycogen but a 22-33% decrease (P < 0.05) in liver glycogen; (e) liver glycogen synthetase and phosphorylase activity were unchanged while whole brain glycogen synthetase activity decreased by 27% (P < 0.05) and phosphorylase activity increased by 10-16%; and (f) whole brain triglyceride content did not change, although there was a 155-241% increase (P < 0.01) in liver triglyceride.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

Early postoperative change of plasma levels of amino acids in neonates with perforative peritonitis and its prognostic significance.

As a possible prognostic index, plasma levels of amino acids were serially measured for 1 week in 22 neonates with perforative peritonitis including gastric rupture and necrotizing enterocolitis and 28 neonatal surgical patients without infection as a control, who were subjected to total parenteral nutrition with hypertonic glucose and amino acids formulated for neonates. Plasma levels of branched chain amino acids significantly increased on the first day after operation in peritonitis group, whereas plasma levels of phenylalanine and histidine increased on the third postoperative day and tyrosine increased seventh postoperative day. In the peritonitis group, 5 neonates died within 2 weeks after operation (early death group) because of cardiopulmonary failure and 5 neonates died with hyperbilirubinemia from 1 month to 6 months after operation (late death group). The early death group showed increased levels of total amino acids compared with those in the late death group and survivors. The late death group also showed higher levels of total amino acids than survivors. Plasma levels of tyrosine increased in the early and late death groups compared with survivors. These results indicated that serial measurements of plasma amino acids early after operation showed the existence of liver dysfunction in addition to the derangement of amino acid metabolism in the liver and muscle caused by septic insult, which predicted the prognosis of perforative peritonitis in neonates.

Amino Acids↗

PubMed1995

Analysis of c-kit expression of human erythroleukemia cell line, HEL: clonal variation and relationship with erythroid and megakaryocytic phenotype.

The proto-oncogene c-kit encodes the receptor for a stem cell factor. We examined HEL, a human erythroleukemia cell line, in order to clarify the correlation between the c-kit receptor (KR) expression and lineage-specific phenotype. Although HEL cells are known to express KR, we found two relatively distinct HEL cell populations in terms of KR expression. We then subcloned HEL cell lines with clone sorting on the basis of KR expression and compared their various characteristics. The highly KR-expressing subline, HEL-P1, expressed a high level of glycophorin A (GPA), a known erythroid lineage marker. HEL-N1, in which most of the cells were KR-negative, showed a higher megakaryocytic lineage marker CD41b expression than HEL-P1. However, the expression of granulomonocytic lineage markers were not significantly different between the two subclones. Cell growth rate and cell cycle analysis also did not detect significant differences between the sublines. HEL-P1 cells gradually lost their KR expression in serum-containing culture, while the percentage of KR-positive HEL-N1 cells increased in serum-free culture. These observations indicate that KR expression was associated with the synchronous expression of GPA and inversely correlated with CD41b, and reversible transitions between KR-positive cells and KR-negative cells exist. We suggest that KR plays an important part in commitment of erythroid and megakaryocytic precursor cells.

Antigens, CD↗

PubMed1995

Comparison of granisetron alone and granisetron plus hydroxyzine hydrochloride for prophylactic treatment of emesis induced by cisplatin chemotherapy.

The efficacy and safety of granisetron alone (group G) and granisetron plus hydroxyzine hydrochloride (group G/H) as prophylactic therapy for acute and delayed nausea and vomiting were evaluated in an open trial in head and neck cancer patients undergoing chemotherapy with cisplatin. The severity of nausea was significantly reduced on days 1 and 4 in patients receiving combination therapy, but the frequency of vomiting was not significantly different between the two groups. The only side-effect observed was headache in 1 patient from group G, and no drug-related laboratory test abnormalities were observed. These results suggest that the anti-emetic efficacy of granisetron can be augmented by hydroxyzine hydrochloride.

Acute Disease↗

PubMed1995

Quantitative analysis of AML1/ETO transcripts in peripheral blood stem cell harvests from patients with t(8;21) acute myelogenous leukaemia.

Peripheral blood stem cells (PBSC) have been used increasingly for haemopoietic reconstitution after marrow-ablative chemotherapy in patients with acute leukaemia because of the possibility that there is a lower risk of leukaemic contamination. We have developed a titration assay using a competitive reverse transcriptase polymerase chain reaction (RT-PCR) which is able to estimate the number of AML1/ETO transcripts so that minimal residual disease (MRD) can be monitored quantitatively in patients with t(8;21) acute myelogenous leukaemia (AML). Using a qualitative RT-PCR method, AML1/ETO transcripts could be detected in all samples from 15 first PBSC harvests and 11 second PBSC harvests obtained from 15 patients with t(8;21) AML. With our competitive RT-PCR assay, the number of AML1/ETO transcripts was found to be lower in the second PBSC harvest than that in the first in every individual. Furthermore, MRD in PBSC harvests was less than that in the corresponding bone marrow obtained on the day of PBSC collection in the individual patients studied. In 10 patients who received autologous blood stem cell transplantation (ABSCT), we could not find a relationship between the number of AML1/ETO transcripts in the infused PBSC harvests and the clinical outcome after ABSCT. The present study clearly indicates that although PBSC harvests collected after consolidation chemotherapy are contaminated by leukaemic cells, the degree of leukaemic contamination may decrease as chemotherapy is repeated. The mobilization of PBSC by repeated chemotherapy may provide an advantageous source of haemopoietic stem cells for ABSCT.

Adolescent↗

PubMed1995

High serum levels of granulocyte-macrophage colony-stimulating factor in patients with liver cirrhosis and granulocytopenia.

Leucopenia is often observed in patients with liver cirrhosis. We measured levels of serum granulocyte-macrophage colony-stimulating factor (GM-CSF) in patients with liver cirrhosis by a sensitive enzyme linked immunosorbent assay. Eight out of 22 patients with liver cirrhosis had detectable serum GM-CSF (range, 55 to 245 pg/ml:mean, 135 pg/ml). Serum GM-CSF was detected in all patients with a granulocyte count below 2.0 x 10(9)/l, but in only one patient with a granulocyte count above 2.0 x 10(9)/l. Haemoglobin concentration, platelet count, serum albumin, total bilirubin and aminotransferase levels did not correlate with serum GM-CSF levels. These findings may reflect a feedback mechanism between the number of circulating granulocytes and serum GM-CSF levels in patients with cirrhosis.

Adult↗