Search PubMed⌕ Search

Biomedical subjects

A Kumagai

Publications and source records attributed to A Kumagai.

At least 19 recordsLinked to original sources
PubMed2001

Inhibition of green tea catechins against the growth of cancerous human colon and hepatic epithelial cells.

The ability of (-)-epicatechin (EC), (-)-epigallocatechin (EGC) and (-)-epigallocatechin gallate (EGCG) to inhibit the growth of HCT 116 colorectal and Hep G2 hepatocellular carcinoma cells was examined by MTT and clonogenic assays (CA). The respective catechins inhibited the growth of HCT 116 more strongly than Hep G2. In MTT assay, IC(50) values of EGC and EGCG against HCT 116 grew smaller on prolongation of the exposure times of the cells to the catechins. In CA, however, these two catechins had IC(50) values ranging between 7.6+/-0.4 and 11.2+/-0.5 microM against the same cells regardless of the exposure times. EC showed much weaker growth inhibitions relative to the two aforementioned catechins.

Beverages↗
PubMed2001

Protacs: chimeric molecules that target proteins to the Skp1-Cullin-F box complex for ubiquitination and degradation.

The intracellular levels of many proteins are regulated by ubiquitin-dependent proteolysis. One of the best-characterized enzymes that catalyzes the attachment of ubiquitin to proteins is a ubiquitin ligase complex, Skp1-Cullin-F box complex containing Hrt1 (SCF). We sought to artificially target a protein to the SCF complex for ubiquitination and degradation. To this end, we tested methionine aminopeptidase-2 (MetAP-2), which covalently binds the angiogenesis inhibitor ovalicin. A chimeric compound, protein-targeting chimeric molecule 1 (Protac-1), was synthesized to recruit MetAP-2 to SCF. One domain of Protac-1 contains the I kappa B alpha phosphopeptide that is recognized by the F-box protein beta-TRCP, whereas the other domain is composed of ovalicin. We show that MetAP-2 can be tethered to SCF(beta-TRCP), ubiquitinated, and degraded in a Protac-1-dependent manner. In the future, this approach may be useful for conditional inactivation of proteins, and for targeting disease-causing proteins for destruction.

Aminopeptidases↗
PubMed2001

Regulation of Op18 during spindle assembly in Xenopus egg extracts.

Oncoprotein 18 (Op18) is a microtubule-destabilizing protein that is negatively regulated by phosphorylation. To evaluate the role of the three Op18 phosphorylation sites in Xenopus (Ser 16, 25, and 39), we added wild-type Op18, a nonphosphorylatable triple Ser to Ala mutant (Op18-AAA), and to mimic phosphorylation, a triple Ser to Glu mutant (Op18-EEE) to egg extracts and monitored spindle assembly. Op18-AAA dramatically decreased microtubule length and density, while Op18-EEE did not significantly affect spindle microtubules. Affinity chromatography with these proteins revealed that the microtubule-destabilizing activity correlated with the ability of Op18 to bind tubulin. Since hyperphosphorylation of Op18 is observed upon addition of mitotic chromatin to extracts, we reasoned that chromatin-associated proteins might play a role in Op18 regulation. We have performed a preliminary characterization of the chromatin proteins recruited to DNA beads, and identified the Xenopus polo-like kinase Plx1 as a chromatin-associated kinase that regulates Op18 phosphorylation. Depletion of Plx1 inhibits chromatin-induced Op18 hyperphosphorylation and spindle assembly in extracts. Therefore, Plx1 may promote microtubule stabilization and spindle assembly by inhibiting Op18.

Animals↗
PubMed2001

Design, synthesis, and biological evaluation of anti-HIV double-drugs. conjugates of HIV protease inhibitors with a reverse transcriptase inhibitor through spontaneously cleavable linkers.

Based on the prodrug concept as well as the combination of two different classes of anti-HIV agents, we designed and synthesized a series of anti-HIV double-drugs consisting of HIV protease inhibitors conjugated with a nucleoside reverse transcriptase inhibitor in an effort to enhance the antiviral activity. For the conjugation, a series of linkers that conjoins the two different classes of inhibitors has been investigated. Double-drugs using a succinyl amino acid linker were shown to release the parent drugs via spontaneous imide formation at a faster rate compared to compounds using a glutaryl amino acid linker, as expected from the energetically favorable cyclization to the five-membered ring. Among the double-drugs, KNI-1039 (3b) with a glutarylglycine linker exhibited extremely potent anti-HIV activity compared with that of the individual components. Double-drug 3b was relatively stable in culture medium, whereas it regenerated active species in cell homogenate. These results suggested that the synergistic enhancement of anti-HIV activities of 3b may be due to their ability to penetrate into the target cell and subsequent regeneration of two different classes of anti-HIV agents in the cytoplasm.

Animals↗
PubMed2001

Attributable risk of H. pylori in peptic ulcer disease: does declining prevalence of infection in general population explain increasing frequency of non-H. pylori ulcers?

Recent reports in the United States have found that fewer peptic ulcers are due to Helicobacter pylori than previously believed. The aim of this study is to determine if the declining prevalence of H. pylori infection in the general population can account for the apparent increase in the frequency of non-H. pylori ulcers. A total of 396 patients with peptic ulcer or ulcer scar were enrolled in this study. The pre-1950 population consisted of 149 patients with gastric ulcers and with 44 duodenal ulcers. The post-1950 population consisted of 96 patients with gastric ulcers and 107 with duodenal ulcers. The frequency of H. pylori-negative gastric ulcers was 5.4% in patients born before 1950 and 4.2% in patients born after 1950, and the frequency of H. pylori-negative duodenal ulcers was 0% and 1.9%, respectively. There are no statistical differences between the two populations in gastric and duodenal ulcers. H. pylori seropositivity was 74.9% in asymptomatic volunteers born before 1950 and 20.7% in those born after 1950 (P < 0.01) in the general population. The attributable risk of H. pylori infection in peptic ulcer diseases was not affected by the prevalence of H. pylori infection in the general population in Japan. This suggests that the apparent increase in frequency of non-H. pylori ulcers in the United States is not simply due to the declining prevalence of infection. Other explanations for non-H. pylori ulcers should be sought.

Adult↗
PubMed2001

Positive regulation of Wee1 by Chk1 and 14-3-3 proteins.

Wee1 inactivates the Cdc2-cyclin B complex during interphase by phosphorylating Cdc2 on Tyr-15. The activity of Wee1 is highly regulated during the cell cycle. In frog egg extracts, it has been established previously that Xenopus Wee1 (Xwee1) is present in a hypophosphorylated, active form during interphase and undergoes down-regulation by extensive phosphorylation at M-phase. We report that Xwee1 is also regulated by association with 14-3-3 proteins. Binding of 14-3-3 to Xwee1 occurs during interphase, but not M-phase, and requires phosphorylation of Xwee1 on Ser-549. A mutant of Xwee1 (S549A) that cannot bind 14-3-3 is substantially less active than wild-type Xwee1 in its ability to phosphorylate Cdc2. This mutation also affects the intranuclear distribution of Xwee1. In cell-free kinase assays, Xchk1 phosphorylates Xwee1 on Ser-549. The results of experiments in which Xwee1, Xchk1, or both were immunodepleted from Xenopus egg extracts suggested that these two enzymes are involved in a common pathway in the DNA replication checkpoint response. Replacement of endogenous Xwee1 with recombinant Xwee1-S549A in egg extracts attenuated the cell cycle delay induced by addition of excess recombinant Xchk1. Taken together, these results suggest that Xchk1 and 14-3-3 proteins act together as positive regulators of Xwee1.

14-3-3 Proteins↗
PubMed2001

Expression of Fas and Fas ligand mRNA in rat and human preimplantation embryos.

The Fas-Fas ligand (L) system is one of the major signalling pathways to induce apoptosis in various cells and tissues. The aim of this study was to investigate the expression of the Fas-Fas L system in rat and human oocytes and preimplantation embryos. We determined the expression of Fas and Fas L mRNA of rat oocytes and embryos up to the blastocyst stage, and of human embryos at the 2- or 4-cell stage, using reverse transcription polymerase chain reaction (PCR) and nested PCR techniques. Moreover, we investigated the expression of Fas mRNA in human fragmented embryos. In rat embryos, Fas mRNA was expressed at the 2-cell stage only, whereas Fas L mRNA was expressed in oocytes, and at the pronuclear (1-cell) and 2-cell stages. In human embryos, Fas mRNA was expressed at the 4-cell stage only, whereas Fas L mRNA was expressed at both 2- and 4-cell stages. Human fragmented embryos expressed both Fas and Fas L mRNA. Because simultaneous expression of Fas and Fas L mRNA occurred in 2-cell rat embryos and in 4-cell human embryos, the Fas-Fas L system might be involved in the apoptotic pathway in the early embryos of these species.

Animals↗
PubMed2000

Requirement for Atr in phosphorylation of Chk1 and cell cycle regulation in response to DNA replication blocks and UV-damaged DNA in Xenopus egg extracts.

The checkpoint kinase Xchk1 becomes phosphorylated in Xenopus egg extracts in response to DNA replication blocks or UV-damaged DNA. Xchk1 is also required for the cell cycle delay that is induced by unreplicated or UV-damaged DNA. In this report, we have removed the Xenopus homolog of ATR (Xatr) from egg extracts by immunodepletion. In Xatr-depleted extracts, the checkpoint-associated phosphorylation of Xchk1 is abolished, and the cell cycle delay induced by replication blocks is strongly compromised. Xatr from egg extracts phosphorylated recombinant Xchk1 in vitro, but not a mutant form of Xchk1 (Xchk1-4AQ) containing nonphosphorylatable residues in its four conserved SQ/TQ motifs. Recombinant human ATR, but not a kinase-inactive mutant, phosphorylated the same sites in Xchk1. Furthermore, the Xchk1-4AQ mutant was found to be defective in mediating a checkpoint response in egg extracts. These findings suggest that Xchk1 is a functionally important target of Xatr during a checkpoint response to unreplicated or UV-damaged DNA.

Amino Acid Motifs↗
PubMed2000

Claspin, a novel protein required for the activation of Chk1 during a DNA replication checkpoint response in Xenopus egg extracts.

We have identified Claspin, a novel protein that binds to Xenopus Chk1 (Xchk1). Binding of Claspin to Xchk1 is highly elevated in the presence of DNA templates that trigger a checkpoint arrest of the cell cycle in Xenopus egg extracts. Xchk1 becomes phosphorylated during a checkpoint response, and we demonstrate directly that this phosphorylation results in the activation of Xchk1. Immunodepletion of Claspin from egg extracts abolishes both the phosphorylation and activation of Xchk1. Furthermore, Claspin-depleted extracts are unable to arrest the cell cycle in response to DNA replication blocks. Taken together, these findings indicate that Claspin is an essential upstream regulator of Xchk1.

Adaptor Proteins, Signal Transducing↗
PubMed2000

Chaperonin 10 in the rat oocytes and early embryos: its expression and activity for early pregnancy factor.

PROBLEM: The aim of this study was to investigate the expression of chaperonin (cpn) 10 and cpn 60 mRNA in oocytes or embryos, and to further explore the possibility that early pregnancy factor (EPF) is identical with cpn 10. METHOD OF STUDY: The expressions of cpn 10 and cpn 60 mRNA in oocytes and embryos at the different stages (1-cell, 2-cell, 8-cell, and morula) were examined by polymerase chain reaction techniques. The EPF activity of native rat cpn 10 isolated from rat livers was evaluated by the rosette inhibition test. RESULTS: Similar levels of mRNA of cpn 10 and cpn 60 were detected in oocytes and embryos at every stage. There were no detectable EPF activities in the native cpn 10. Immunoprecipitation using polyclonal antibodies against cpn 10 did not affect the activity of EPF in the pregnant rat serum. CONCLUSION: Our results do not support the hypothesis that cpn 10 is identical with EPF.

Animals↗
PubMed1999

The xenopus Suc1/Cks protein promotes the phosphorylation of G(2)/M regulators.

The entry into mitosis is controlled by Cdc2/cyclin B, also known as maturation or M-phase promoting factor (MPF). In Xenopus egg extracts, the inhibitory phosphorylations of Cdc2 on Tyr-15 and Thr-14 are controlled by the phosphatase Cdc25 and the kinases Myt1 and Wee1. At mitosis, Cdc25 is activated and Myt1 and Wee1 are inactivated through phosphorylation by multiple kinases, including Cdc2 itself. The Cdc2-associated Suc1/Cks1 protein (p9) is also essential for entry of egg extracts into mitosis, but the molecular basis of this requirement has been unknown. We find that p9 strongly stimulates the regulatory phosphorylations of Cdc25, Myt1, and Wee1 that are carried out by the Cdc2/cyclin B complex. Overexpression of the prolyl isomerase Pin1, which binds to the hyperphosphorylated forms of Cdc25, Myt1, and Wee1 found at M-phase, is known to block the initiation of mitosis in egg extracts. We have observed that Pin1 specifically antagonizes the stimulatory effect of p9 on phosphorylation of Cdc25 by Cdc2/cyclin B. This observation could explain why overexpression of Pin1 inhibits mitotic initiation. These findings suggest that p9 promotes the entry into mitosis by facilitating phosphorylation of the key upstream regulators of Cdc2.

Adaptor Proteins, Signal Transducing↗
PubMed1999

Binding of 14-3-3 proteins and nuclear export control the intracellular localization of the mitotic inducer Cdc25.

Binding of 14-3-3 proteins near the nuclear localization sequence of Xenopus Cdc25 suppresses its ability to induce entry into mitosis. We have examined the intracellular localization of green fluorescent protein (GFP)-tagged wild-type Cdc25 or a mutant (S287A) that cannot bind 14-3-3 proteins. Upon coexpression with Myc-14-3-3epsilon, GFP-Cdc25-WT was predominantly cytoplasmic, whereas GFP-Cdc25-S287A was exclusively nuclear. Leptomycin B, an inhibitor of nuclear export, elicited a prompt redistribution of GFP-Cdc25-WT to the nucleus. Mutagenesis experiments demonstrated that Cdc25 contains multiple nuclear export sequences. These studies indicate that the binding of 14-3-3 proteins and nuclear export regulate the intracellular localization of Cdc25.

14-3-3 Proteins↗
PubMed1999

Non-traumatic renal arteriopelvic fistula.

PURPOSE: In the present paper, we report on a 34-year-old female with macroscopic hematuria due to a nontraumatic renal arteriopelvic fistula (APF). The patient initially presented at another hospital with asymptomatic macroscopic hematuria. Following abdominal ultrasonography, computed tomography (CT) and laboratory data, no abnormal findings were seen. Therefore, the patient was referred to Teine Keijinkai Hospital for a more precise evaluation of the urinary tract and vascular abnormality. METHODS/RESULTS: Endoscopically, there was bleeding from the right ureteral orifice, so the patient was admitted for further examination. No abnormal findings were seen on urinary cytology and following an intravenous pyelogram. A selective right lower polar renal arteriogram revealed arterial extravasation directly into the pelvis before the venous phase, so APF of the kidney was diagnosed. The patient had no history of urinary tract trauma, so the APF was thought to be idiopathic. After transcatheter arterial embolization (TAE) with a gelatine sponge, macroscopic and microscopic hematuria disappeared and a low-density area was seen in the middle pole of the right kidney in an abdominal CT scan 4 days after TAE. This was thought to be renal infarction due to TAE. CONCLUSIONS: After discharge, the patient had no further hematuria.

Adult↗
PubMed1998

The Xenopus Chk1 protein kinase mediates a caffeine-sensitive pathway of checkpoint control in cell-free extracts.

We have analyzed the role of the protein kinase Chk1 in checkpoint control by using cell-free extracts from Xenopus eggs. Recombinant Xenopus Chk1 (Xchk1) phosphorylates the mitotic inducer Cdc25 in vitro on multiple sites including Ser-287. The Xchk1-catalyzed phosphorylation of Cdc25 on Ser-287 is sufficient to confer the binding of 14-3-3 proteins. Egg extracts from which Xchk1 has been removed by immunodepletion are strongly but not totally compromised in their ability to undergo a cell cycle delay in response to the presence of unreplicated DNA. Cdc25 in Xchk1-depleted extracts remains bound to 14-3-3 due to the action of a distinct Ser-287-specific kinase in addition to Xchk1. Xchk1 is highly phosphorylated in the presence of unreplicated or damaged DNA, and this phosphorylation is abolished by caffeine, an agent which attenuates checkpoint control. The checkpoint response to unreplicated DNA in this system involves both caffeine-sensitive and caffeine-insensitive steps. Our results indicate that caffeine disrupts the checkpoint pathway containing Xchk1.

14-3-3 Proteins↗
PubMed1998

14-3-3 proteins act as negative regulators of the mitotic inducer Cdc25 in Xenopus egg extracts.

Cdc25, the dual-specificity phosphatase that dephosphorylates the Cdc2-cyclin B complex at mitosis, is highly regulated during the cell cycle. In Xenopus egg extracts, Cdc25 is associated with two isoforms of the 14-3-3 protein. Cdc25 is complexed primarily with 14-3-3epsilon and to a lesser extent with 14-3-3zeta. The association of these 14-3-3 proteins with Cdc25 varies dramatically during the cell cycle: binding is high during interphase but virtually absent at mitosis. Interaction with 14-3-3 is mediated by phosphorylation of Xenopus Cdc25 at Ser-287, which resides in a consensus 14-3-3 binding site. Recombinant Cdc25 with a point mutation at this residue (Cdc25-S287A) is incapable of binding to 14-3-3. Addition of the Cdc25-S287A mutant to Xenopus egg extracts accelerates mitosis and overrides checkpoint-mediated arrests of mitotic entry due to the presence of unreplicated and damaged DNA. These findings indicate that 14-3-3 proteins act as negative regulators of Cdc25 in controlling the G2-M transition.

14-3-3 Proteins↗
PubMed1998

[Experience of nephron sparing surgery for renal mass].

BACKGROUND: In this paper, we reported nephron sparing surgery for renal mass in our hospital. METHODS: From March, 1994 to October, 1997, nephron sparing surgeries were done for 19 (11 males and 8 females, whose age were ranged from 38 to 75 years old) renal mass in our hospital. The sites of lesion were 12 in right side and 7 in left side. The tumor size were from 8 to 40/mm. Almost all cases were discovered incidentally. RESULTS: In all 19 cases, 6 cases were resultingly renal cell carcinoma (RCC). There was no difference in age, sex, tumor size and computed tomography findings between RCC and benign lesion. In magnetic resonance imaging (MRI) findings, iso intensity lesions of T2-weighted image were more frequent in RCC significantly. Postoperative 24 hours creatinine clearance and serum creatinine were not changed significantly compared with preoperative data. CONCLUSION: In MRI findings, iso intensity of T2-weighted image was useful for differential diagnosis between RCC and benign lesion.

Adult↗
PubMed1998

[Pain-free injection of propofol].

Pain on injection is one of the well-known side effects of propofol. Previous studies have shown several methods to alleviate this discomfort. We employed all these methods together to clarity whether pain-free injection of propofol was possible. Sixty adult patients premedicated with midazolam were studied. Control group patients (n = 20) received an induction dose of propofol via a vein on the dorsum of the hand at a slow injection speed with carrier i.v. fluid. Study group patients (n = 40) received i.v. fentanyl 0.1 mg, followed by bolus injection of cold propofol premixed with lidocaine (final concentration of lidocaine was 0.2%) in a forearm vein without carrier i.v. fluid. Eighteen patients (90%) in the control group experienced injection pain. In the study group, however, no patients complained of pain or discomfort. In conclusion, pain-free injection of propofol was possible when prior-administration of fentanyl, premixing of lidocaine, cooling to 4 degrees C, and rapid injection via a forearm vein without carrier i.v. fluid was the adopted precedure.

Adult↗