Biomedical subjects

Akira Kikuchi

Publications and source records attributed to Akira Kikuchi.

At least 19 recordsLinked to original sources

PubMed2007

Post-translational palmitoylation and glycosylation of Wnt-5a are necessary for its signalling.

Wnt-5a is a representative ligand that activates a beta-catenin-independent pathway in Wnt signalling. In the present paper, the roles of the post-translational modifications in the actions of Wnt-5a were investigated. We found that Wnt-5a is modified with palmitate at Cys104 and glycans at Asn114, Asn120, Asn311 and Asn325. The palmitoylation was not essential for the secretion of Wnt-5a, but was necessary for its ability to suppress Wnt-3a-dependent T-cell factor transcriptional activity and to stimulate cell migration. Wnt-5a activated focal adhesion kinase and this activation also required palmitoylation. Wild-type Wnt-5a induced the internalization of Fz (Frizzled) 5, but a Wnt-5a mutant that lacks the palmitoylation site did not. Furthermore, the binding of Wnt-5a to the extracellular domain of Fz5 required palmitoylation of Wnt-5a. These results indicate that palmitoylation of Wnt-5a is important for the triggering of signalling at the cell surface level and, therefore, that the lipid-unmodified form of Wnt-5a cannot activate intracellular signal cascades. In contrast, glycosylation was necessary for the secretion of Wnt-5a, but not essential for the actions of Wnt-5a. Thus the post-translational palmitoylation and glycosylation of Wnt-5a are important for the actions and secretion of Wnt-5a.

Amino Acid Sequence↗

PubMed2007

Moyamoya syndrome following childhood acute lymphoblastic leukemia.

BACKGROUND: Long-term survivors of childhood acute lymphoblastic leukemia (ALL) sometimes suffer from adverse long-term sequelae. We analyzed the incidence, clinical course and prognosis of moyamoya syndrome (MoS) following childhood ALL. PROCEDURE: A total of 1,846 ALL patients were treated with four consecutive TCCSG ALL protocols (L84-11, L89-12, L92-13, and L95-14) between 1984 and 1999. We surveyed the MoS cases among these patients in the follow-up studies. RESULTS: Six patients with MoS were identified: four boys and two girls whose ages ranged from 2 years and 1 month (abbreviated as "2y1m") to 14y 1 m at diagnosis of ALL. None of the patients had central nervous system (CNS) leukemia. All six patients received prophylactic cranial irradiation with a dosage of 18 or 24 Gy. Although one patient died of brain infarction due to MoS, no leukemic relapse was observed in the group. The cumulative incidence of MoS in our series was 0.46 +/- 0.02% at 8 years. Among several clinical characteristics, use of cranial irradiation was the only factor that appeared to be significantly related to the development of MoS. CONCLUSIONS: MoS occurs with increased frequency in children treated for ALL, and might be associated with cranial irradiation. Prophylactic cranial irradiation should be used cautiously in ALL patients who can be cured by other CNS-directed therapies.

Adolescent↗

PubMed2007

Phytochemical, pharmacological and ethnobotanical studies of Rhazya stricta Decne.

Rhazya stricta Decne. is an important medicinal species that is rich in alkaloids containing anticancer alkaloids also. This paper reviews phytochemical, pharmacological and ethnobotanical studies of R. stricta. Phytochemical and pharmacological aspects of the species have been thoroughly explored. Over 100 alkaloids have been isolated from R. stricta, but the pharmacological activities are known for only a few of these compounds. Very few ethnobotanical studies of R. stricta have been done, and there is a particular need for such studies in various regions of South Asia and in Middle Eastern countries where the species grows. These ethnobotanical studies should investigate herbal recipes of R. stricta and their use for curing diseases. The genetic diversity, chemotaxonomy and allelopathy of R. stricta have not yet been explored, and these studies are recommended, with the ultimate goal of benefiting local communities in regions where R. stricta grows. This should be achieved by sharing knowledge and through the manufacture of herbal drugs from R. stricta's active chemical constituents, while protecting intellectual property rights and following the World Health Organization's guidelines for safety and efficacy.

Animals↗

PubMed2007

Dvl regulates endo- and exocytotic processes through binding to synaptotagmin.

Dvl, an important component of the Wnt signalling pathway, is thought to be involved in synaptogenesis. In this study, we investigated whether Dvl regulates neurotransmitter release. Knockdown of Dvl in PC12 cells suppressed K(+)-induced dopamine release, and this phenotype was restored by expression of Dvl-1. We identified synaptotagmin (Syt) I, which is involved in neurotransmitter release, as a Dvl-binding protein. Dvl directly bound to the C2B domain of Syt I. Dvl colocalized with Syt I at the tip of neurites of differentiated PC12 cells and of neurons in the rat dorsal root ganglion. Dvl and Syt I was located in large dense-core vesicles, which contain dopamine. In addition, endocytosis of vesicles containing Syt I was suppressed in Dvl knockdown PC12 cells. Dvl inhibited the binding of Syt I to the complex consisting of syntaxin-1A and SNAP-25. Furthermore, micro2-adaptin of AP-2, which is known to play a role in endocytosis, formed a complex with Dvl and Syt I. Taken together, these results suggest that Dvl is involved in endo- and exocytotic processes through the binding to Syt I.

Adaptor Proteins, Signal Transducing↗

PubMed2006

Developmental stage-specific biphasic roles of Wnt/beta-catenin signaling in cardiomyogenesis and hematopoiesis.

Although Wingless (Wg)/Wnt signaling has been implicated in heart development of multiple organisms, conflicting results have been reported regarding the role of Wnt/beta-catenin pathway in cardiac myogenesis: Wg/armadillo signaling promotes heart development in Drosophila, whereas activation of Wnt/beta-catenin signaling inhibits heart formation in avians and amphibians. Using an in vitro system of mouse ES cell differentiation into cardiomyocytes, we show here that Wnt/beta-catenin signaling exhibits developmental stage-specific, biphasic, and antagonistic effects on cardiomyogenesis and hematopoiesis/vasculogenesis. Activation of the Wnt/beta-catenin pathway in the early phase during embryoid body (EB) formation enhances ES cell differentiation into cardiomyocytes while suppressing the differentiation into hematopoietic and vascular cell lineages. In contrast, activation of Wnt/beta-catenin signaling in the late phase after EB formation inhibits cardiomyocyte differentiation and enhances the expression of hematopoietic/vascular marker genes through suppression of bone morphogenetic protein signaling. Thus, Wnt/beta-catenin signaling exhibits biphasic and antagonistic effects on cardiomyogenesis and hematopoiesis/vasculogenesis, depending on the stage of development.

Animals↗

PubMed2006

Sprouty2 and Sprouty4 are essential for embryonic morphogenesis and regulation of FGF signaling.

Sprouty genes encode cytoplasmic membrane-associated proteins that inhibit receptor tyrosine kinase signaling. Four orthologs of Drosophila Sprouty (dSpry) (Sprouty1-4) have been identified in mammals. Physiological function of Sprouty1 and Sprouty2 has been investigated using gene targeting approaches, however to date detailed examination of Sprouty4 knockout (KO) mice has not been reported. In this study, Sprouty4 KO mice were generated and characterized. Although a significant fraction of Sprouty4 KO mice died shortly after birth due to mandible defects, the remainder were viable and fertile. Growth retardation was observed for most Sprouty4-deficient mice, with nearly all Sprouty4 KO mice having polysyndactyly. ERK activation was sustained in Sprouty4 KO mouse embryonic fibroblasts (MEFs) in response to FGF, but not to EGF. Sprouty2 and Sprouty4 double KO (DKO) mice were embryonic lethal and showed severe defects in craniofacial, limb, and lung morphogenesis. These findings suggest both redundant and non-redundant functions for Sprouty2 and Sprouty4 on embryonic development and FGF signaling.

Adaptor Proteins, Signal Transducing↗

PubMed2006

GSK-3beta-regulated interaction of BICD with dynein is involved in microtubule anchorage at centrosome.

Microtubule arrays direct intracellular organization and define cellular polarity. Here, we show a novel function of glycogen synthase kinase-3beta (GSK-3beta) in the organization of microtubule arrays through the interaction with Bicaudal-D (BICD). BICD is known to form a complex with dynein-dynactin and to function in the intracellular vesicle trafficking. Our data revealed that GSK-3beta is required for the binding of BICD to dynein but not to dynactin. Knockdown of GSK-3beta or BICD reduced centrosomally focused microtubules and induced the mislocalization of centrosomal proteins. The unfocused microtubules in GSK-3beta knockdown cells were rescued by the expression of the dynein intermediate chain-BICD fusion protein. Microtubule regrowth assays showed that GSK-3beta and BICD are required for the anchoring of microtubules to the centrosome. These results imply that GSK-3beta may function in transporting centrosomal proteins to the centrosome by stabilizing the BICD1 and dynein complex, resulting in the regulation of a focused microtubule organization.

Adaptor Proteins, Signal Transducing↗

PubMed2006

Multiplicity of the interactions of Wnt proteins and their receptors.

Wnts are secreted proteins that are essential for a wide array of developmental and physiological processes. They signal across the plasma membranes by interacting with serpentine receptors of the Frizzled (Fz) family and members of the low-density-lipoprotein receptor-related protein (LRP) family. Recent advances in the Wnt signaling field have revealed that Wnt-unrelated proteins activate or suppress Wnt signaling by binding to Fzs or LRP5/6 and that atypical receptor tyrosine kinases mediate Wnt signaling independently of Fz and/or function as a Fz co-receptor. This review highlights recent progress in our understanding of the multiplicity of Wnts and their receptors. We discuss how the interaction between the ligands and receptors activate distinct intracellular signaling pathways. We also discuss how intracellular trafficking of Wnt signaling components can regulate the sensitivity of cells to Wnts.

Animals↗

PubMed2006

Filopodia formation mediated by receptor tyrosine kinase Ror2 is required for Wnt5a-induced cell migration.

The receptor tyrosine kinase Ror2 plays important roles in developmental morphogenesis. It has recently been shown that Ror2 mediates Wnt5a-induced noncanonical Wnt signaling by activating the Wnt-JNK pathway and inhibiting the beta-catenin-TCF pathway. However, the function of Ror2 in noncanonical Wnt signaling leading to cell migration is largely unknown. We show, using genetically different or manipulated cultured cells, that Ror2 is critical for Wnt5a-induced, but not Wnt3a-induced, cell migration. Ror2-mediated cell migration requires the extracellular cysteine-rich domain (CRD), which is the binding site for Wnt5a, and the cytoplasmic proline-rich domain (PRD) of Ror2. Furthermore, Ror2 can mediate filopodia formation via actin reorganization, irrespective of Wnt5a, and this Ror2-mediated filopodia formation requires the actin-binding protein filamin A, which associates with the PRD of Ror2. Intriguingly, disruption of filopodia formation by suppressing the expression of either Ror2 or filamin A inhibits Wnt5a-induced cell migration, indicating that Ror2-mediated filopodia formation is essential for Wnt5a-induced cell migration.

Adaptor Proteins, Signal Transducing↗

PubMed2006

Expression of Wnt-5a is correlated with aggressiveness of gastric cancer by stimulating cell migration and invasion.

Wnt-5a is a representative ligand that activates a beta-catenin-independent pathway in the Wnt signaling. Although abnormal activation of beta-catenin-dependent pathway is often observed in human cancer, the relationship between beta-catenin-independent pathway and tumorigenesis is not clear. We sought to clarify how Wnt-5a is involved in aggressiveness of gastric cancer. Abnormal expression of Wnt-5a was observed in 71 of 237 gastric cancer cases by means of immunohistochemistry. The positivity of Wnt-5a expression was correlated with advanced stages and poor prognosis of gastric cancer. Wnt-5a had the abilities to stimulate cell migration and invasion in gastric cancer cells. Wnt-5a activated focal adhesion kinase and small GTP-binding protein Rac, both of which are known to play a role in cell migration. Cell migration, membrane ruffling, and turnover of paxillin were suppressed in Wnt-5a knockdown cells. Furthermore, anti-Wnt-5a antibody suppressed gastric cancer cell migration. These results suggest that Wnt-5a stimulates cell migration by regulating focal adhesion complexes and that Wnt-5a is not only a prognostic factor but also a good therapeutic target for gastric cancer.

Cell Movement↗

PubMed2006

Common gene expression signatures in t(8;21)- and inv(16)-acute myeloid leukaemia.

Human acute myeloid leukaemia (AML) involving a core-binding factor (CBF) transcription factor is called CBF leukaemia. In these leukaemias, AML1 (RUNX1, PEBP2alphaB, CBFalpha2)-MTG8 (ETO) and CBFbeta (PEBP2beta)-MYH11 chimaeric proteins are generated by t(8;21) and inv(16) respectively. We analysed gene expression profiles of leukaemic cells by microarray, and selected genes whose expression appeared to be modulated in association with t(8;21) and inv(16). In a pair-wise comparison, 15% of t(8;21)-associated transcripts exhibited high or low expression in inv(16)-AML, and 26% of inv(16)-associated transcripts did so equivalently in t(8;21)-AML. These common elements in gene expression profiles between t(8;21)- and inv(16)-AML probably reflect the situation that AML1-MTG8 and CBFbeta-MYH11 chimaeric proteins affect a common set of target genes in CBF leukaemic cells. On the other hand, 38% of t(8;21)-associated and 24% of inv(16)-associated transcripts were regulated in t(8;21)- and inv(16)-specific manners. These distinct features of t(8;21)- and inv(16)-associated genes correlate with the bimodular structures of the chimaeric proteins (CBF-related AML1 and CBFbeta portions, and CBF-unrelated MTG8 and MYH11 portions).

Acute Disease↗

PubMed2006

Human T-cell leukemia virus type 1 tax dysregulates beta-catenin signaling.

Dysregulation of beta-catenin signaling has been implicated in the malignant transformation of cells. However, the role of beta-catenin in the human T-cell leukemia virus type 1 (HTLV-1)-induced transformation of T cells is unknown. Here we found that beta-catenin protein was overexpressed in the nucleus and that beta-catenin-dependent transcription was significantly enhanced in Tax-positive HTLV-1-infected T-cell lines compared to that in Tax-negative HTLV-1-infected T-cell lines. Transfection with beta-catenin-specific small interfering RNA inhibited the growth of the Tax-positive HTLV-1-infected T-cell line HUT-102. Transient transfection of Tax appeared to enhance beta-catenin-dependent transcription by stabilizing the beta-catenin protein via activation of the cyclic AMP (cAMP) response element-binding protein. HTLV-1-infected T-cell lines overexpressing beta-catenin also showed increased Akt activity via Tax activation of the cAMP response element-binding protein, resulting in the phosphorylation and inactivation of glycogen synthase kinase 3beta, which phosphorylates beta-catenin for ubiquitination. The phosphatidylinositol 3-kinase inhibitor LY294002 reduced beta-catenin expression in Tax-positive T-cell lines, and inactivation of glycogen synthase kinase 3beta by lithium chloride restored beta-catenin expression in Tax-negative T-cell lines. Finally, we showed that dominant-negative Akt inhibited Tax-induced beta-catenin-dependent transcription. These results indicate that Tax activates beta-catenin through the Akt signaling pathway. Our findings suggest that activation of beta-catenin by Tax may be important in the transformation of T cells by HTLV-1 infection.

Base Sequence↗

PubMed2006

In vitro and in vivo evaluation of novel cationic liposomes utilized for cancer gene therapy.

Advanced peritoneal carcinomatoses is very difficult to treat. We have explored the potential therapeutic application of gene therapy using cationic liposomes in this disease. The lacZ gene was introduced in vitro into ovarian and endometrial cancer cells using cationic liposomes. The transfection efficiency was similar to that of commercially available liposomes in serum-free medium (11.0-20.9% vs. 5.4-26.0%). In serum-containing medium, the efficiency was 1.9-18.1%, which is comparable with the efficiency in serum-free medium. However, the efficiency of commercial liposomes decreased drastically to between 0.1% and 4.7% in the serum-containing medium. When cultured cells were transfected with the herpes simplex virus thymidine kinase (HSV-tk) gene and ganciclovir (GCV) was added, the anti-tumor effect of GCV was 47-640 times greater than when the same experiment was performed with lacZ gene. Evaluation of anti-tumor effect was performed with the MTT assay. In vivo, the HRA and mEIIL ascitic mice were treated with HSV-tk gene and GCV using the peritoneal route, a significant prolongation of the mean survival time was observed by Kaplan-Meier analysis (16-18 days and 15-30 days, respectively, p < 0.05). These results indicate a potential role for gene therapy in the treatment of advanced intraperitoneal carcinomatoses using the novel cationic liposomes.

Animals↗

PubMed2006

Oog1, an oocyte-specific protein, interacts with Ras and Ras-signaling proteins during early embryogenesis.

We previously identified an oocyte-specific gene, Oogenesin 1 (Oog1), that encodes 326 amino acids containing a leucine zipper structure and a leucine-rich repeat. In the present study, to identify the interacting proteins of Oog1, we performed a yeast two-hybrid screening using a GV-oocyte cDNA library and found that Ral guanine nucleotide dissociation stimulator (RalGDS) is the binding partner of Oog1. Coimmunoprecipitation assay confirmed the interaction between Oog1 and RalGDS proteins. Colocalization experiments provide the evidence that the nuclear localization of RalGDS depends on the expression of Oog1. Interestingly, RalGDS protein localized in the nucleus rather than the cytoplasm between late 1-cell and early 2-cell stages, the time when Oog1 localizes in the nucleus. We also examined the interaction between Oog1 and Ras by GST pull-down assay and revealed that Oog1 interacts with Ras in a GTP-dependent manner. These findings suggest a role of Oog1 as a Ras-binding protein.

Animals↗

PubMed2006

Regulation of Wnt signaling by protein-protein interaction and post-translational modifications.

The Wnt signaling pathway is conserved in various species from worms to mammals, and plays important roles in cellular proliferation, differentiation, and migration. Wnt stabilizes cytoplasmic beta-catenin and then the accumulated beta-catenin is translocated into the nucleus, where it activates the transcriptional factor T-cell factor (Tcf)/lymphoid enhancer factor (Lef), and thereby stimulates the expression of genes including c-myc, c-jun, fra-1, and cyclin D1. Tight regulation of this response involves post-translational modifications of the components of the Wnt signaling pathway. Phosphorylation, ubiquitination, and sumoylation have been shown to affect the half-life of beta-catenin and the transcriptional activity of Tcf/Lef. The precise spatio-temporal patterns of these multiple modifications determine the driving force of various cellular responses.

Animals↗

PubMed2006

Mutations of the PTPN11 and RAS genes in rhabdomyosarcoma and pediatric hematological malignancies.

PTPN11 has been identified as a causative gene in Noonan syndrome (NS), responsible for about 50% of cases of NS. Given the association between NS and an increased risk of some malignancies, notably leukemia and probably some solid tumors including neuroblastoma (NB) and rhabdomyosarcoma (RMS), recent studies have reported that gain-of-function somatic mutations in PTPN11 occur in some hematological malignancies, especially de novo juvenile myelomonocytic leukemia (JMML) and in some solid tumors such as NB, although at a low frequency. In a screen for mutations of PTPN11 in 7 cell lines and 30 fresh tumors of RMS and in 25 cell lines and 40 fresh tumors of NB, we identified a missense mutation (A72T) in an embryonal RMS patient. In the RMS samples, we also detected mutations of NRAS in 1 cell line and 1 patient; both mutations were in embryonal RMSs and had no PTPN11 mutations. No mutations of PTPN11 were detected in NB. In 95 leukemia cell lines and 261 fresh leukemia samples including 22 JMMLs, 9 kinds of missense mutations were detected in 17 leukemia samples, which included 11 (50.0%) mutations in JMML samples and lower frequencies in other hematological malignancies. Furthermore, we identified 4 (18.2%) NRAS mutations and 1 (4.5%) KRAS mutation in 5 JMML samples, 1 of which had a concomitant PTPN11 mutation. Our data suggest that mutations of PTPN11 as well as RAS play a role in the pathogenesis of not only myeloid hematological malignancies but also a subset of RMS malignancies.

Cell Line, Tumor↗

PubMed2006

Caveolin is necessary for Wnt-3a-dependent internalization of LRP6 and accumulation of beta-catenin.

beta-catenin-mediated Wnt signaling is critical in animal development and tumor progression. The single-span transmembrane Wnt receptor, low-density lipoprotein receptor-related protein 6 (LRP6), interacts with Axin to promote the Wnt-dependent accumulation of beta-catenin. However, the molecular mechanism of receptor internalization and its impact on signaling are unclear. Here, we present evidence that LRP6 is internalized with caveolin and that the components of this endocytic pathway are required not only for Wnt-3a-induced internalization of LRP6 but also for accumulation of beta-catenin. Overall, our data suggest that Wnt-3a triggers the interaction of LRP6 with caveolin and promotes recruitment of Axin to LRP6 phosphorylated by glycogen synthase kinase-3beta and that caveolin thereby inhibits the binding of beta-catenin to Axin. Thus, caveolin plays critical roles in inducing the internalization of LRP6 and activating the Wnt/beta-catenin pathway. We also discuss the idea that distinct endocytic pathways correlate with the specificity of Wnt signaling events.

Axin Protein↗

PubMed2006

Alteration of TH1 and TH2 cells by intracellular cytokine detection in patients with unexplained recurrent abortion before and after immunotherapy with the husband's mononuclear cells.

OBJECTIVE: To elucidate the possible mechanisms of immunotherapy for unexplained recurrent aborters using their husband's mononuclear cells. DESIGN: Prospective clinical study. SETTING: Institutional practice at the Outpatient Clinic for Infertility, Niigata University Medical Hospital. PATIENT(S): Fifty-two unexplained recurrent aborters were chosen as an experimental group. INTERVENTION(S): Each patient was injected with her husband's mononuclear cells as immunotherapy. Peripheral blood was obtained from the patients. MAIN OUTCOME MEASURE(S): The percentage of CD4-positive cells, TH1 cells, TH2 cells, and the TH1/TH2 ratio were analyzed in the patients before and after immunotherapy. The same analyses were performed in the successful and the unsuccessful group. RESULT(S): To date, 42 of the 52 patients have become newly pregnant. Of the 42, 34 patients have already delivered (successful group) and 3 are now pregnant, while the remaining 8 cases experienced repeated abortion (unsuccessful group). The percentage of TH2 cells significantly increased in the total patient population, while the TH1/TH2 ratio significantly decreased in the total patient population and in the successful group. CONCLUSION(S): These findings suggest that immunotherapy with the husband's mononuclear cells for unexplained recurrent abortion induces a dominant state of TH2 cells in the patients.

Abortion, Habitual↗