Biomedical subjects

Nobuharu Fujii

Publications and source records attributed to Nobuharu Fujii.

At least 37 records · Page 2Linked to original sources

PubMed2004

Possible involvement of CS-ACS1 and ethylene in auxin-induced peg formation of cucumber seedlings.

BACKGROUND AND AIMS: Cucumber (Cucumis sativus) seedlings develop a peg on the concave side of the gravitropically bending transition zone between the hypocotyl and the root after seed germination. Peg initiation occurs in response to auxin when its levels in the concave side of the transition zone exceed a particular threshold through the graviresponse. Ethylene also plays an important role in peg formation, but its relationship to auxin in this event is not understood. Here, the role ethylene plays in auxin-induced peg formation is studied. METHODS: Peg formation of cucumber seedlings exposed to ethylene at different stages of growth or during exogenous auxin treatment was observed. In addition, ethylene evolution from the concave and convex sides of the transition zone was compared and their transcription of CS-ACS (1-aminocyclopropane-1-carboxylic acid synthase) genes was analysed by RT-PCR and in situ hybridization. KEY RESULTS: Seedlings treated with ethylene after peg initiation produced an enlarged peg, whereas ethylene treatment before peg initiation inhibited peg formation. Ethylene also promoted the development of the peg in the auxin-treated seedlings. Furthermore, the concave side of the transition zone at peg initiation produced more ethylene and CS-ACS1 mRNA than the convex side. CONCLUSIONS: Since CS-ACS1 is an auxin-inducible gene, the greater abundance of auxin in the concave side of the transition zone causes peg initiation and increases CS-ACS1-mediated ethylene biosynthesis, which then facilitates peg development.

Amino Acids, Cyclic↗

PubMed2004

Isolation of cucumber CsARF cDNAs and expression of the corresponding mRNAs during gravity-regulated morphogenesis of cucumber seedlings.

Cucumber seedlings show positive gravitropism and bend in the transition zone between the hypocotyl and the root. The peg, a specialized protuberance, develops on the concave side of the bending transition zone. Auxin and the mRNA of an auxin-inducible gene (CsIAA1) isolated from cucumber are differentially accumulated across the transition zone during the gravity-regulated peg formation. In this study, five cDNAs of Auxin Response Factors (ARFs) from cucumber were isolated and their mRNA accumulation was compared with that of CsIAA1. The tissue specificity of CsARF2 mRNA accumulation was similar to that of CsIAA1. Because the structural character of CsARF2 predicts that it is a transcriptional activator, CsARF2 may be involved in the activation of CsIAA1 transcription, which plays a role in gravity-regulated peg formation. Neither gravity nor auxin affected mRNA accumulation of five CsARFs including CsARF2, suggesting that CsARF2 may be regulated at a post-transcriptional level to induce the asymmetric expression of the CsIAA1 gene in response to gravistimulation and auxin in cucumber seedlings.

Amino Acid Sequence↗

PubMed2004

cDNA cloning and functional characterization of a novel splice variant of c-Cbl-associated protein from mouse skeletal muscle.

c-Cbl-associated protein (CAP) is an SH3-containing adapter protein that binds to the proto-oncogene c-Cbl. Recent work suggests that signaling through these molecules is involved in the regulation of insulin-stimulated glucose uptake in 3T3-L1 adipocytes. Skeletal muscle is the major site of insulin-stimulated glucose disposal but there have been no reports of CAP function in this tissue. Using RT-PCR of mouse skeletal muscle RNA, we discovered a novel splice variant of CAP (CAPSM; GenBank Accession No. AF521593) that is different from the adipocyte form by inclusion of a novel 168 bp fragment. This fragment encodes a peptide sequence that shows very high similarity with exon 25 of the human homologue of CAP (SORBS1). To understand the function of CAPSM in glucose uptake regulation, L6 myotubes were transfected with either CAPSM or a truncated CAPSM devoid of all three SH3-binding domains (CAPDeltaSH3), which prevents CAP association with c-Cbl. Transfection with CAPDeltaSH3 decreased insulin-stimulated 2-deoxyglucose (2-DG) uptake and reduced c-Cbl phosphorylation. In contrast, transfection of L6 myotubes with CAPDeltaSH3 had no effect on dinitrophenol (DNP)- or hypoxia-stimulated glucose uptake, stimuli that work through insulin-independent mechanisms for the regulation of glucose uptake. These data demonstrate the existence of a novel CAP isoform expressed in skeletal muscle, and suggest the involvement of the CAP/Cbl pathway in the regulation of insulin-stimulated glucose uptake in L6 myotubes.

Alternative Splicing↗

PubMed2004

Overexpression or ablation of JNK in skeletal muscle has no effect on glycogen synthase activity.

c-Jun NH(2)-terminal kinase (JNK) is highly expressed in skeletal muscle and is robustly activated in response to muscle contraction. Little is known about the biological functions of JNK signaling in terminally differentiated muscle cells, although this protein has been proposed to regulate insulin-stimulated glycogen synthase activity in mouse skeletal muscle. To determine whether JNK signaling regulates contraction-stimulated glycogen synthase activation, we applied an electroporation technique to induce JNK overexpression (O/E) in mouse skeletal muscle. Ten days after electroporation, in situ muscle contraction increased JNK activity 2.6-fold in control muscles and 15-fold in the JNK O/E muscles. Despite the enormous activation of JNK activity in JNK O/E muscles, contraction resulted in similar increases in glycogen synthase activity in control and JNK O/E muscles. Consistent with these findings, basal and contraction-induced glycogen synthase activity was normal in muscles of both JNK1- and JNK2-deficient mice. JNK overexpression in muscle resulted in significant alterations in the basal phosphorylation state of several signaling proteins, such as extracellular signal-regulated kinase 1/2, p90 S6 kinase, glycogen synthase kinase 3, protein kinase B/Akt, and p70 S6 kinase, in the absence of changes in the expression of these proteins. These data suggest that JNK signaling regulates the phosphorylation state of several kinases in skeletal muscle. JNK activation is unlikely to be the major mechanism by which contractile activity increases glycogen synthase activity in skeletal muscle.

Animals↗

PubMed2004

Regulation of glucose transport by the AMP-activated protein kinase.

The AMP-activated protein kinase (AMPK) is an energy-sensing enzyme that is activated during exercise and muscle contraction as a result of acute decreases in ATP:AMP and phosphocreatine:creatine. Physical exercise increases muscle glucose uptake, enhances insulin sensitivity and leads to fatty acid oxidation in muscle. An important issue in muscle biology is to understand whether AMPK plays a role in mediating these metabolic processes. AMPK has also been implicated in regulating gene transcription and, therefore, may function in some of the cellular adaptations to training exercise. Recent studies have shown that the magnitude of AMPK activation and associated metabolic responses are affected by factors such as glycogen content, exercise training and fibre type. There have also been conflicting reports as to whether AMPK activity is necessary for contraction-stimulated glucose transport. Thus, during the next several years considerably more research will be necessary in order to fully understand the role of AMPK in regulating glucose transport in skeletal muscle.

Adenylate Kinase↗

PubMed2004

Peripheral blood circulating immature cell counts predict CD34+ cell yields in G-CSF-induced PBPC mobilization in healthy donors.

BACKGROUND: It has been previously reported that the number of circulating immature cells (CIC) in peripheral blood (PB) estimates the number of CD34+ cells collected in G-CSF plus chemotherapy-induced PBPC mobilization. The correlation of CIC counts in PB with CD34+ cell yield and its usefulness was evaluated in G-CSF-induced PBPC mobilization for healthy donors. STUDY DESIGN AND METHODS: CIC counts in PB and CD34+ cell counts in the apheresis product from 122 collections were assessed, and the relationship between these two variables was evaluated with the Pearson rank correlation analysis, the chi-squared test, and the U-test. RESULTS: CIC counts were correlated weakly with the number of CD34+ cells per L of blood processed in the apheresis product (Pearson rank correlation analysis; r=0.357, p<0.0001). When a level of 1.7 x 10(9) CICs per L was selected as a cutoff value, the sensitivity and specificity for collecting more than 20 x 10(6) CD34+ cells per L of blood processed were 63.6 and 77.5 percent, respectively. CONCLUSION: The present study suggests that the number of CICs in PB may estimate the number of CD34+ cells collected. The data indicate that CIC counts above 1.7 x 10(9) per L can be used as a good predictor for PBPC collections containing more than 20 x 10(6) CD34+ cells per L of blood processed in a single apheresis procedure.

Adolescent↗

PubMed2004

[Clinical evaluation of cefozopran as treatment for febrile neutropenia].

Clinical effects and safety of cefozopran (CZOP) were evaluated by the Okayama Bone Marrow Transplantation Group. Twenty-five patients expected to experience febrile neutropenia during induction chemotherapy or consolidation chemotherapy of acute leukemia were enrolled between July 2000 and November 2002. CZOP was administrated by drip infusion at 4g/day bid for a minimum of 3 days. The clinical effects and safety were evaluated in 20 patients with fever of 37.5 degrees C or more from a clinically suspected infection. The underlying disease was acute myeloid leukemia in 17 patients, acute lymphoid leukemia in 1 and acute promyelogeneous leukemia in 1. The complicating infections were sepsis and suspected sepsis. Clinical efficacy was excellent in 11 patients, good in 1, fair in 2 and poor in 6, with an efficacy rate of 60.0%. The efficacy rate in patients whose albumin levels before therapy were less than 3.8 g/dl was 37.5%, whereas the rate in patients whose albumin levels before therapy were between 3.8 g/dl and 5.3 g/dl was 80.0%. The efficacy rate in patients whose neutrophil counts before therapy were less than 100/microliter was 50.0%, whereas the rate in patients whose neutrophil counts after therapy were less than 100/microliter was 53.8%. The efficacy rate in patients whose neutrophil counts both before and after therapy were less than 100/microliter was 37.5%. Side effect of exanthema was observed in 1 patient. These results indicate that CZOP is an effective and safe antibiotic for the treatment of febrile neutropenia in patients with hematological malignancies.

Acute Disease↗

PubMed2003

p38gamma MAPK regulation of glucose transporter expression and glucose uptake in L6 myotubes and mouse skeletal muscle.

Skeletal muscle expresses at least three p38 MAPKs (alpha, beta, gamma). However, no studies have examined the potential regulation of glucose uptake by p38gamma, the isoform predominantly expressed in skeletal muscle and highly regulated by exercise. L6 myotubes were transfected with empty vector (pCAGGS), activating MKK6 (MKK6CA), or p38gamma-specific siRNA. MKK6CA-transfected cells had higher rates of basal 2-deoxy-d-[3H]glucose (2-DG) uptake (P < 0.05) but lower rates of 2,4-dinitrophenol (DNP)-stimulated glucose uptake, an uncoupler of oxidative phosphorylation that operates through an insulin-independent mechanism (P < 0.05). These effects were reversed when MKK6CA cells were cotransfected with p38gamma-specific siRNA. To determine whether the p38gamma isoform is involved in the regulation of contraction-stimulated glucose uptake in adult skeletal muscle, the tibialis anterior muscles of mice were injected with pCAGGS or wild-type p38gamma (p38gammaWT) followed by intramuscular electroporation. Basal and contraction-stimulated 2-DG uptake in vivo was determined 14 days later. Overexpression of p38gammaWT resulted in higher basal rates of glucose uptake compared with pCAGGS (P < 0.05). Muscles overexpressing p38gammaWT showed a trend for lower in situ contraction-mediated glucose uptake (P = 0.08) and significantly lower total GLUT4 levels (P < 0.05). These data suggest that p38gamma increases basal glucose uptake and decreases DNP- and contraction-stimulated glucose uptake, partially by affecting levels of glucose transporter expression in skeletal muscle. These findings are consistent with the hypothesis that activation of stress kinases such as p38 are negative regulators of stimulated glucose uptake in peripheral tissues.

2,4-Dinitrophenol↗

PubMed2003

Cloning and characterization of mouse 5'-AMP-activated protein kinase gamma3 subunit.

Naturally occurring mutations in the regulatory gamma-subunit of 5'-AMP-activated protein kinase (AMPK) can result in pronounced pathological changes that may stem from increases in muscle glycogen levels, making it critical to understand the role(s) of the gamma-subunit in AMPK function. In this study we cloned the mouse AMPKgamma3 subunit and revealed that there are two transcription start sites, which result in a long form, gamma3L (AF525500) and a short form, gamma3S (AF525501). AMPKgamma3L is the predominant form in mouse and is specifically expressed in mouse skeletal muscle at the protein level. In skeletal muscle, AMPKgamma3 shows higher levels of expression in fast-twitch white glycolytic muscle (type IIb) compared with fast-twitch red oxidative glycolytic muscle (type IIa), whereas gamma3 is undetectable in soleus muscle, a slow-twitch oxidative muscle with predominantly type I fibers. AMPKgamma3 can coimmunoprecipititate with both alpha and beta AMPK subunits. Overexpression of gamma3S and gamma3L in mouse tibialis anterior muscle in vivo has no effect on alpha1 and alpha2 subunit expression and does not alter AMPKalpha2 catalytic activity. However, gamma3S and gamma3L overexpression significantly increases AMPKalpha1 phosphorylation and activity by approximately 50%. The increase in AMPKalpha1 activity is not associated with alterations in glycogen accumulation or glycogen synthase expression. In conclusion, the gamma3 subunit of AMPK is highly expressed in fast-twitch glycolytic skeletal muscle, and wild-type gamma3 functions in the regulation of alpha1 catalytic activity, but it is not associated with changes in muscle glycogen concentrations.

AMP-Activated Protein Kinases↗

PubMed2003

Gravity-induced modification of auxin transport and distribution for peg formation in cucumber seedlings: possible roles for CS-AUX1 and CS-PIN1.

Cucurbit seedlings potentially develop a peg on each side of the transition zone between the hypocotyl and root. Seedlings grown in a horizontal position suppress the development of the peg on the upper side of the transition zone in response to gravity. It is suggested that this suppression occurs due to a reduction in auxin levels to below the threshold value. We show in this study that the free indole-3-acetic acid (IAA) content is low, while IAA conjugates are significantly more abundant in the upper side of the transition zone of gravistimulated seedlings, compared to the lower side. A transient increase in mRNA of the auxin-inducible gene, CS-IAA1, was observed in the excised transition zone. The result suggests that the transition zone is a source of auxin. Cucumber seedlings treated with auxin-transport inhibitors exhibited agravitropic growth and developed a peg on each side of the transition zone. Auxin-transport inhibitors additionally caused an increase in CS-IAA1 mRNA accumulation at the transition zone, indicating a rise in intracellular auxin concentrations due to a block of auxin efflux. To study the involvement of the auxin transport system in peg formation, we isolated the cDNAs of a putative auxin influx carrier, CS-AUX1, and putative efflux carrier, CS-PIN1, from cucumber (Cucumis sativus L.) plants. Both genes (CS-AUX1 in particular) were auxin-inducible. Accumulation of CS-AUX1 and CS-PIN1 mRNAs was observed in vascular tissue, cortex and epidermis of the transition zone. A reduced level of CS-AUX1 mRNA was observed in the upper side of the gravistimulated transition zone, compared with the lower side. It is therefore possible that a balance in the activities of auxin influx and efflux carriers controls intracellular auxin concentration at the transition zone, which results in lateral placement of a peg in cucumber seedlings.

Arabidopsis Proteins↗

PubMed2003

Glucose metabolism and energy homeostasis in mouse hearts overexpressing dominant negative alpha2 subunit of AMP-activated protein kinase.

AMP-activated protein kinase (AMPK) is an energy-sensing enzyme that plays a pivotal role in regulating cellular metabolism for sustaining energy homeostasis under stress conditions. Activation of AMPK has been observed in the heart during acute and chronic stresses, but its functional role has not been completely understood because of the lack of effective activators and inhibitors of this kinase in the heart. We generated transgenic mice (TG) with cardiac-specific overexpression of a dominant negative mutant of the AMPK alpha2 catalytic subunit to clarify the functional role of this kinase in myocardial ischemia. In isolated perfused hearts subjected to a 10-min ischemia, AMPK alpha2 activity in wild type (WT) increased substantially (by 4.5-fold), whereas AMPK alpha2 activity in TG was similar to the level of WT at base line. Basal AMPK alpha1 activity was unchanged in TG and increased normally during ischemia. Ischemia stimulated a 2.5-fold increase in 2-deoxyglucose uptake over base line in WT, whereas the inactivation of AMPK alpha2 in TG significantly blunted this response. Using 31P NMR spectroscopy, we found that ATP depletion was accelerated in TG hearts during no-flow ischemia, and these hearts developed left ventricular dysfunction manifested by an early and more rapid increase in left ventricular end-diastolic pressure. The exacerbated ATP depletion could not be attributed to impaired glycolytic ATP synthesis because TG hearts consumed slightly more glycogen during this period of no-flow ischemia. Thus, AMPK alpha2 is necessary for maintaining myocardial energy homeostasis during ischemia. It is likely that the functional role of AMPK in myocardial energy metabolism resides both in energy supply and utilization.

AMP-Activated Protein Kinases↗

PubMed2003

Successful treatment with cyclosporin A of myelodysplastic syndrome with erythroid hypoplasia associated with t(6;8)(q15;q22).

We report a t(6;8)(q15;q22) in a patient with myelodysplastic syndrome (MDS) with erythroid hypoplasia. The patient was successfully treated with an immunosuppressive treatment with cyclosporin A, while the translocation was repeatedly detected as the sole anomaly with the percentages of positive cells ranging from 5% to 70%. To our knowledge, the t(6:8) has never been described in MDS.

Aged↗

PubMed2003

Potential limitations in using minor histocompatibility antigen-specific cytotoxic T cells for targeting solid tumor cells.

We have shown previously that KIAA0223, a gene encoding a minor histocompatibility antigen, HA-1, whose expression was believed to be restricted to the hematopoietic cells, is aberrantly expressed in some solid tumor cell lines. However, its significance in tumor immunity needs to be determined. Cytotoxic activity of HA-1(H)-specific cytotoxic T lymphocytes (CTLs) was assessed against solid tumor cell lines expressing KIAA0223 using (51)Cr release assays. Five of seven cell lines were lysed when HLA-A*0201 was adequately expressed. One of the two CTL-resistant cell lines became susceptible after treatment with IFN-gamma and TNF-alpha, while the other was lysed only after pulsing with HA-1(H) peptide. In most cell lines tested, HA-1(H) peptide was properly generated and presented for recognition by the CTL. However, impaired antigen processing and presentation observed in this study may result in escape from CTL recognition in vivo, as well as in vitro, as observed in this study.

Gene Expression Regulation, Neoplastic↗

PubMed2003

A novel fusion variant of the MORF and CBP genes detected in therapy-related myelodysplastic syndrome with t(10;16)(q22;p13).

We report a case of therapy-related myelodysplastic syndrome (t-MDS) with t(10;16)(q22;p13), in which novel fusion transcripts of the MORF and CBP genes were detected. In one MORF-CBP fusion transcript, exon 15 of the MORF gene was fused in frame with exon 5 of the CBP gene. In a reciprocal CBP-MORF transcript, exon 4 of the CBP gene was fused in frame with exon 16 of the MORF gene. This is the first reported case of t-MDS associated with t(10;16), and provides molecular evidence that the novel MORF-CBP and/or CBP-MORF fusion protein(s) might play an important role in the development of t-MDS.

Acetyltransferases↗

PubMed2003

Human herpes virus-8-negative primary effusion lymphoma in a patient with common variable immunodeficiency.

Primary effusion lymphoma (PEL) is a newly described high-grade B cell lymphoma developing in association with human herpes virus type 8 (HHV-8) in human immunodeficiency virus (HIV)-infecting individuals. Common variable immunodeficiency (CVID) is a primary immunodeficiency disease characterized by reduced serum immunoglobulin and heterogeneous clinical features. The risk of cancer in CVID patients is increased. Here, we describe a PEL that developed in the pleural and pericardial cavities of an HIV-negative and HHV-8-negative patient with CVID.

Common Variable Immunodeficiency↗

PubMed2003

Cardiac hypertrophy by hypertension and exercise training exhibits different gene expression of enzymes in energy metabolism.

Hypertension-induced pathological cardiac hypertrophy (hypertensive heart) and exercise training-induced physiological cardiac hypertrophy (athletic heart) have differences in cardiac properties. We hypothesized that gene expression of energy metabolic enzymes differs between these two types of cardiac hypertrophy. To investigate whether mRNA expression of key enzymes in the long-chain fatty acid (FA), glucose, and lactic acid metabolic pathways differs between these two types of cardiac hypertrophy, we used the hearts of spontaneously hypertensive rats (SHR; 19 weeks old) as a model of the hypertensive heart, swim-trained rats (Trained; 19 weeks old, swimming training for 15 weeks) as a model of the athletic heart, and sedentary Wistar-Kyoto rats (Control; 19 weeks old). SHR developed hypertensive cardiac hypertrophy, of which cardiac function was deteriorated, whereas Trained rats developed an athletic heart, of which cardiac function was enhanced. The mRNA expression of CD36, which involved in uptake of long-chain FA, in the heart was almost never detected in the SHR group. Furthermore, the mRNA expression of key enzymes in the long-chain FA metabolic pathway (acyl CoA synthase [ACoAS], carnitine palmitoyl transferase [CPT]-I, CPT-II, and isocitrate dehydrogenase [ISCD]) in the heart was significantly higher in the SHR group compared with the Control group. The mRNA expression of ACoAS, CPT-I, ISCD, and CD36 in the heart did not differ between Trained group and Control group, whereas that of CPT-II in the Trained group was significantly higher compared with the Control group. The mRNA expression of key enzymes (phosphofructokinase and lactate dehydrogenase) in glycolytic metabolic pathway in the heart was markedly higher in the SHR group compared with the Control group, whereas these mRNA expressions did not differ between Trained group and Control group. These findings suggest that the molecular phenotypes in the energy metabolic system differ in hypertension-induced pathological and exercise training-induced physiological cardiac hypertrophy, and these differences may participate in the differences in cardiac function.

Animals↗

PubMed2003

[Auxin-mediated response of cucumber seedlings to gravity].

Gravity regulates peg formation because cucumber seedlings grown in a horizontal position develop a peg on the lower side of the transition zone (TR zone) but not on the upper side. Studies on peg formation have suggested the regulation of peg formation by gravity as follows. Cucumber seedlings potentially develop a peg on both the lower and upper sides of the TR zone. The development of the peg on upper side of the TR zone is suppressed in response to gravity. A phytohormone, auxin, induces peg formation. Upon gravistimulation the auxin concentration on the upper side of the TR zone is reduced to a level below the threshold value necessary for peg formation. The unequally distributed auxin across TR zone is caused by a change in accumulation of auxin influx carrier (CsAUX1) protein and auxin efflux carrier (CsPIN1) protein in response to gravity. In addition, TR zone before peg initiation expresses both CsARF2 (putative activator of auxin response factor) and CsIAA1 (putative repressor of auxin-inducible gene expression), by which TR zone could respond the auxin gradient regulated by gravity.

Cucumis sativus↗

PubMed2003

The effect of light on the gravimorphogenesis of cucumber seedlings.

Light affects the responses of plants to gravity. For example, light is inhibitory to the gravimorphogenesis of cucumber seedlings, peg formation. We have suggested that auxin regulates peg formation by stimulating the expression of auxin-inducible genes including CsACS1 that encodes key enzyme in ethylene synthesis. We also have hypothesized that ethylene promotes the development of peg in cucumber seedlings. To understand how light inhibits peg formation, we examined the effects of light on the auxin-induced peg formation, the expression of auxin-inducible genes and ethylene production in cucumber seedlings. Number of seedlings that produced a peg in response to applied IAA was significantly decreased by exposure to light compared with that in the dark. In addition, exposure of seedlings to light decreased mRNA accumulation of CsACS1 and ethylene production. However ethylene application did not compensate the inhibition of peg formation by light. These results suggest that light blocks auxin-mediated step(s) including CsACS1 expression for the initiation of peg formation.

Cucumis sativus↗