[Serum carboxy-terminal propeptide of human type 1 procollagen (P1CP)].
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Biomarkers↗
Biomedical subjects
Y Fujiwara
Publications and source records attributed to Y Fujiwara.
Detection of K-ras gene mutations in plasma DNA of patients with pancreatic adenocarcinoma: correlation with clinicopathological features.
We investigated the presence of K-ras gene mutation in plasma DNA and assessed its clinical value in patients with pancreatic adenocarcinoma. Mutations in codon 12 of the K-ras gene were examined by mutant allele-specific amplification method using DNA extracted from surgical specimens and plasma samples of 21 patients with pancreatic adenocarcinoma. K-ras gene mutation was detected in 15 of 21 (71%) primary tumors. In 9 of 15 (60%) patients with K-ras gene mutation-positive tumors, an identical mutation was detected in the plasma DNA. None of four patients with chronic pancreatitis or five healthy subjects had such mutations in plasma DNA. Tumors positive for K-ras gene mutation in plasma DNA were significantly larger (P = 0.04) and less likely to result in a curative cure after surgical resection (P = 0.09) than those negative for the mutation. Other clinicopathological features, including age, sex, histological type, mode of invasion, and metastasis, did not correlate with K-ras gene mutations in plasma DNA. Treatment resulted in disappearance of K-ras gene mutations in plasma DNA in six of nine (67%) patients. Three patients with a persistently positive K-ras gene mutation in pre- and post-treatment plasma samples were likely to show early recurrence or have a progressive disease. Our findings suggest that K-ras gene mutation can be detected in plasma DNA of patients with pancreatic adenocarcinoma. Detection of K-ras mutations in plasma may be clinically useful for evaluating tumor burden and efficacy of treatment.
Adenocarcinoma↗
[Complete disappearance of metastatic pulmonary tumors in a case of hepatocellular carcinoma treated with UFT].
The disappearance of multiple pulmonary metastatic lesions in hepatocellular carcinoma patient following the oral administration of UFT is reported. Pulmonary and pelvic metastases were detected in a 71-year-old female hepatocellular carcinoma patient treated by TAE (Trans-Arterial Embolization with epirubicin, mitomycin C, lipiodol and Gelform) three times in one year. Four months after beginning oral administration of UFT-E granule 1.5 g/day (tegafur 300 mg/day, uracil 672 mg/day), three pulmonary metastatic lesions markedly reduced in size, and disappeared completely 6 months after beginning UFT. A pelvic metastatic tumor markedly reduced in size by the combined effect of radiation, TAE (epirubicin, mitomycin C, lipiodol, Gelform) and oral UFT. We consider UFT was effective in the disappearance of multiple pulmonary metastatic lesions, because three TAE administrations of epirubicin and mitomycin C could not prevent systemic spreading of hepatocellular carcinoma.
Administration, Oral↗
Ulcer recurrence: cytokines and inflammatory response-dependent process.
H. pylori and nonsteroidal antiinflammatory drugs (NSAIDs) are important factors in the recurrence of peptic ulcer diseases. However, H. pylori-negative recurring ulcers can also be found in nonusers of NSAIDs. The aim of this paper is to review recent data pertaining to mechanisms of ulcer recurrence. Prostaglandin E2 generation is impaired in the tissues of the ulcer scar site and prostaglandin depletion induced by administration of indomethacin during the healing of experimental gastric ulcer predisposes to future ulcer recurrence. Therefore, the prostaglandin deficiency may impair the quality of ulcer healing and thus increase the likelihood of future ulcer recurrence. Persistent infiltration of polymorphonuclear cells is the most prominent finding in the gastric ulcer scar in rats treated with indomethacin. Concomitant administration of prostaglandin E1-analog with indomethacin attenuates inflammatory infiltration and reduces future ulcer recurrence. Therefore, the inflammatory responses at the ulcer scar site may be a key to the quality of ulcer healing. Recent clinical findings suggest a close relationship between the quality of ulcer healing, infiltration of neutrophils and mononuclear cells, and future ulcer recurrence. Gastroprotective drugs such as prostaglandin analogs and prostaglandin inducers improve the quality of ulcer healing and reduce future recurrence. Production of inflammatory cytokines is stimulated by ulcerogenic factors such as NSAIDs, stress, and H. pylori infection. Inflammatory cytokines such as interleukin-1beta and tumor necrosis factor-alpha cause recurrence of healed ulcer. Synthetic prostaglandin E2 inhibits recurrence as well as the production of the cytokines.
Anti-Inflammatory Agents, Non-Steroidal↗
Protective effect of rebamipide against ammonia-induced gastric mucosal lesions.
We investigated the protective effect of rebamipide against ammonia-induced gastric mucosal lesions. Participation of prostaglandin E2 and nitric oxide in the action of rebamipide was also examined. Rebamipide was administered intraperitoneally (10-100 mg/kg) to male Wistar/ST rats (150-325 g) fasted for 24 hr. Thirty minutes later, 1% NH4OH (1 ml) solution was given intragastrically. One hour later, the length of the mucosal lesions was measured (lesion index), and prostaglandin E2 (PGE2) was determined by radioimmunoassay. A 1% NH4OH solution caused gastric mucosal lesions with hemorrhagic necrosis and submucosal edema. PGE2 synthesis was not affected by NH4OH but was significantly increased by rebamipide. Rebamipide decreased the severity of NH4OH-induced gastric mucosal lesions in a dose-dependent manner. Pretreatment with indomethacin (5 mg/kg, subcutaneously) did not affect the protective effect of rebamipide; however, pretreatment with N(omega)-nitro-L-arginine (L-NNA, 1-10 mg/kg, intravenously), an inhibitor of nitric oxide synthase, attenuated the protective effect of rebamipide in a dose-dependent manner. Simultaneous administration of L-arginine (100 mg/kg) and L-NNA completely restored the protective effect of rebamipide, whereas D-arginine was inactive. These results suggest that nitric oxide contributes significantly to the protective effect of rebamipide against ammonia-induced gastric mucosal lesions.
Alanine↗
Rebamipide prevents recurrence of gastric ulcers without affecting Helicobacter pylori status.
Rebamipide, a gastroprotective drug developed in Japan, accelerates ulcer healing and reduces recurrence of experimental gastric ulcers. We examined the effects of rebamipide, given during healing of human gastric ulcers infected with Helicobacter pylori, on the quality of ulcer healing and ulcer recurrence. Sixty H. pylori-positive patients with gastric ulcers were randomly allocated to three treatment groups: group O (N = 20) received 20 mg of omeprazole every day for eight weeks, group OR (N = 20) received the same dose of omeprazole and 300 mg of rebamipide three times a day for eight weeks, and group OA (N = 20) received the same dose of omeprazole for eight weeks and 1500 mg of amoxicillin three times a day for the first two weeks. After this treatment was completed no other medication was given. Endoscopic examinations were performed at the end of therapy (for healing rate), one month later (for rate of H. pylori eradication) and every three months for follow-up (for ulcer recurrence rate). At the end of therapy, biopsy specimens were taken from the gastric ulcer scar and examined under the microscope for neutrophil and mononuclear cell infiltration. The ulcer healing rate of the three groups was almost the same; H. pylori in group OA was 65% and that of the other two groups was 0%. The number of patients with a flat ulcer scar pattern (good quality of ulcer healing) was increased and the neutrophil infiltration was significantly improved in groups OR and OA compared to group O. The ulcer recurrence rate was significantly lower in group OA and group OR than in group O. In conclusion, rebamipide is almost equipotent to amoxicillin plus omeprazole for the reduction of ulcer recurrence. The decreased recurrence rate by rebamipide may be due to improvement of the quality of ulcer healing, reflected as in the suppression of inflammatory cell infiltration in the scar, which results from either cure of H. pylori infection and/or treatment with a gastroprotective drug such as rebamipide.
Aged↗
Increased mRNA levels of transforming growth factor-beta1 and monocyte chemoattractant protein-1 in ulcer relapse caused by interleukin-1beta in rats.
This study investigated the mRNA expression of transforming growth factor-beta1 (TGF-beta1) and monocyte chemoattractant protein-1 (MCP-1) in rat gastric tissues in which ulcers had relapsed due to interleukin-1beta (IL-1beta) administration. Rats with healed ulcers were administered IL-1beta (1 microg/kg) and killed after 0, 12, 24, or 48 hr. Both TGF-beta1 and MCP-1 mRNA levels were increased in the scarred gastric tissues at 24 hr (fourfold), when ulcers had not relapsed. Furthermore, the expression of these genes also increased in the ulcerated gastric tissues at 48 hr (fivefold), when 90% of healed ulcers had relapsed. On the other hand, the number of macrophages that had infiltrated the scarred gastric tissues at 24 hr was two times higher than that at 0 hr. At 48 hr, the number of macrophages that had infiltrated gastric tissues in which ulcers had relapsed was similar to that at 24 hr. Thus, TGF-beta1 and MCP-1 may be implicated in the macrophage infiltration, thereby leading to ulcer relapse due to IL-1beta.
Animals↗
[Gastrointestinal lesions in liver cirrhosis].
The gastrointestinal bleeding commonly observed in patients with liver cirrhosis is usually from esophageal and gastric varices, gastroduodenal ulcer, and congestive gastropathy. Portal hypertension is the major causative factor of pathogenesis of GI lesions. In the present review, we focus in gastric mucosal defense and Helicobacter pylori infection in liver cirrhosis. Gastric mucosal defense is reduced in liver cirrhosis, especially prostaglandins which play a role in the gastric mucosal defense decreased in the gastric mucosal of patients with liver cirrhosis and rat portal hypertension model. Although H. pylori is strongly associated with peptic ulcer disease and chronic gastritis, several studies showed no relationship between H. pylori infection and gastroduodenal ulcer or the infection and congestive gastropathy in liver cirrhosis. Reduced gastric mucosal defense may account for the pathogenesis of GI lesions in liver cirrhosis.
Animals↗
A rare case of a thyrotropin-secreting pituitary tumor which responded to methimazole.
A 52-year-old Japanese male complained of palpitations and excessive sweating. He showed evidence of hyperthyroidism, but without suppression of the serum thyroid-stimulating hormone (TSH). On admission, the serum level of the alpha-subunit of TSH was elevated, but tests for thyroid autoantibodies were negative. The TSH response to thyrotropin-releasing hormone (TRH) was blunted. Imaging studies revealed a pituitary tumor 2 cm in diameter. Administration of methimazole lowered the serum levels of thyroid hormones, and elevated the serum level of TSH. These findings led to the diagnosis of a thyrotropin-secreting pituitary tumor. Immunohistochemical analysis and electron microscopic findings verified the diagnosis.
Antithyroid Agents↗
The LIM-domain binding protein Ldb1 and its partner LMO2 act as negative regulators of erythroid differentiation.
The nuclear LIM domain protein LMO2, a T cell oncoprotein, is essential for embryonic erythropoiesis. LIM-only proteins are presumed to act primarily through protein-protein interactions. We, and others, have identified a widely expressed protein, Ldb1, whose C-terminal 76-residues are sufficient to mediate interaction with LMO2. In murine erythroleukemia cells, the endogenous Lbd1 and LMO2 proteins exist in a stable complex, whose binding affinity appears greater than that between LMO2 and the bHLH transcription factor SCL. However, Ldb1, LMO2, and SCL/E12 can assemble as a multiprotein complex on a consensus SCL binding site. Like LMO2, the Ldb1 gene is expressed in fetal liver and erythroid cell lines. Forced expression of Ldb1 in G1ER proerythroblast cells inhibited cellular maturation, a finding compatible with the decrease in Ldb1 gene expression that normally occurs during erythroid differentiation. Overexpression of the LMO2 gene also inhibited erythroid differentiation. Our studies demonstrate a function for Ldb1 in hemopoietic cells and suggest that one role of the Ldb1/LMO2 complex is to maintain erythroid precursors in an immature state.
Adaptor Proteins, Signal Transducing↗
Epiregulin stimulates proliferation of rabbit gastric cells in primary culture through autophosphorylation of the epidermal growth factor receptor.
Epiregulin, a growth factor of the epidermal growth factor (EGF) family, was recently purified from conditioned medium of a mouse fibroblast-derived tumor cell line. It was reported that epiregulin exhibited bifunctional properties in the regulation of cell growth. However, the effect of epiregulin on gastric cell proliferation is not known. The aims of this study were to determine whether: (1) epiregulin affects proliferation of rabbit cultured gastric cells, (2) epiregulin-induced stimulation of cell proliferation is mediated by the tyrosine kinase pathway, and (3) epiregulin stimulates autophosphorylation of EGF-receptors. Epiregulin stimulated cell proliferation to a significant extent. This effect was completely blocked by treatment with genistein. Epiregulin stimulated tyrosine phosphorylation of a 170 kDa protein, which represents the EGF receptor, in a dose-dependent fashion. These findings suggest that epiregulin has mitogenic effects on rabbit gastric cultured cells, possibly mediated via the tyrosine kinase pathway through autophosphorylation of EGF receptors.
Animals↗
Identification of active sites in amidase: evolutionary relationship between amide bond- and peptide bond-cleaving enzymes.
Mainly based on various inhibitor studies previously performed, amidases came to be regarded as sulfhydryl enzymes. Not completely satisfied with this generally accepted interpretation, we performed a series of site-directed mutagenesis studies on one particular amidase of Rhodococcus rhodochrous J1 that was involved in its nitrile metabolism. For these experiments, the recombinant amidase was produced as the inclusion body in Escherichia coli to greatly facilitate its recovery and subsequent purification. With regard to the presumptive active site residue Cys203, a Cys203 --> Ala mutant enzyme still retained 11.5% of the original specific activity. In sharp contrast, substitutions in certain other positions in the neighborhood of Cys203 had a far more dramatic effect on the amidase. Glutamic acid substitution of Asp191 reduced the specific activity of the mutant enzyme to 1.33% of the wild-type activity. Furthermore, Asp191 --> Asn substitution as well as Ser195 --> Ala substitution completely abolished the specific activity. It would thus appear that, among various conserved residues residing within the so-called signature sequence common to all amidases, the real active site residues are Asp191 and Ser195 rather than Cys203. Inasmuch as an amide bond (CO-NH2) in the amide substrate is not too far structurally removed from a peptide bond (CO-NH-), the signature sequences of various amidases were compared with the active site sequences of various types of proteases. It was found that aspartic acid and serine residues corresponding to Asp191 and Ser195 of the Rhodococcus amidase are present within the active site sequences of aspartic proteinases, thus suggesting the evolutionary relationship between the two.
Amides↗
An upstream, DNase I hypersensitive region of the hematopoietic-expressed transcription factor GATA-1 gene confers developmental specificity in transgenic mice.
The transcription factor GATA-1, which is expressed in several hematopoietic lineages and multipotential progenitors, is required for the development of red blood cells and platelets. To identify control elements of the mouse GATA-1 gene, we analyzed DNase I hypersensitivity of the locus in erythroid chromatin and the expression of GATA-1/Escherichia coli beta-galactosidase (lacZ) transgenes in mice. Transgenes with 2.7 kb of promoter sequences are expressed infrequently and only within adult (definitive) erythroid cells. We show that inclusion of an upstream hypersensitive site (HS I) markedly enhances the frequency of expressing transgenic lines and activates expression in primitive erythroid cells. This pattern recapitulates the proper pattern of GATA-1 expression during development. By breeding a GATA-1/lacZ transgene into a GATA-1(-) background, we also have shown that the activation or maintenance of GATA-1 expression does not require the presence of GATA-1 itself, thereby excluding simple models of positive autoregulation. The transgene cassette reported here should be useful in directing expression of foreign sequences at the onset of hematopoiesis in the embryo and may assist in the identification of upstream regulators of the GATA-1 gene.
3T3 Cells↗
Overcoming CPT-11 resistance by using a biscoclaurine alkaloid, cepharanthine, to modulate plasma trans-membrane potential.
Irinotecan, 7-ethyl-10-[4-(1-piperidino)-1-piperidino] carbonyloxycamptothecin, (CPT-11) resistance was overcome by using a biscoclaurine alkaloid, cepharanthine, in CPT-11- and multidrug-resistant 50MT-1 cells. 50MT-1 cells were established from a mouse breast-cancer cell line, FM3A, by subjecting the cells to a low dose of CPT-11 continuously. 50MT-1 cells exhibited resistance to CPT-11 (40-fold in colony-formation assay) and to other drugs such as doxorubicin (11.7-fold) and etoposide (VP-16) (16.8-fold). The plasma trans-membrane potential was lower in 50MT-1 cells than in FM3A cells, although there were no differences in expressions of P-glycoprotein and of DNA topoisomerase-I and -II proteins. The lower membrane potential in 50MT-1 cells was augmented by co-treatment with a non-toxic dose of cepharanthine. CPT-11 resistance in 50MT-1 cells was overcome (5.0- to 1.4-fold, 6-hr exposure) by the co-treatment with cepharanthine through increasing intracellular accumulation of CPT-11. Resistance to doxorubicin and VP-16 was also overcome by cepharanthine treatment (2.5- to 0.69-fold and 4.2- to 1.4-fold respectively). We conclude that the modification of plasma trans-membrane potential by cepharanthine should be effective in overcoming CPT-11 and multidrug resistance in 50MT-1 cells.
Alkaloids↗
Yolk sac angiogenic defect and intra-embryonic apoptosis in mice lacking the Ets-related factor TEL.
The TEL gene, which is frequently rearranged in human leukemias of both myeloid and lymphoid origin, encodes a member of the Ets family of transcription factors. The TEL gene is widely expressed throughout embryonic development and in the adult. To determine the requirement for the TEL gene product in development we generated TEL knockout mice (TEL-/-) by gene targeting in embryonic stem cells. TEL-/- mice are embryonic lethal and die between E10.5-11.5 with defective yolk sac angiogenesis and intra-embryonic apoptosis of mesenchymal and neural cells. Two-thirds of TEL-deficient yolk sacs at E9.5 lack vitelline vessels, yet possess capillaries, indicative of normal vasculogenesis. Vitelline vessels regress by E10.5 in the remaining TEL-/- yolk sacs. Hematopoiesis at the yolk sac stage, however, appears unaffected in TEL-/- embryos. Our findings demonstrate that TEL is required for maintenance of the developing vascular network in the yolk sac and for survival of selected cell types within the embryo proper.
Animals↗
FOG, a multitype zinc finger protein, acts as a cofactor for transcription factor GATA-1 in erythroid and megakaryocytic differentiation.
The hematopoietic transcription factor GATA-1 is essential for development of the erythroid and megakaryocytic lineages. Using the conserved zinc finger DNA-binding domain of GATA-1 in the yeast two-hybrid system, we have identified a novel, multitype zinc finger protein, Friend of GATA-1 (FOG), which binds GATA-1 but not a functionally inactive mutant lacking the amino (N) finger. FOG is coexpressed with GATA-1 during embryonic development and in erythroid and megakaryocytic cells. Furthermore, FOG and GATA-1 synergistically activate transcription from a hematopoietic-specific regulatory region and cooperate during both erythroid and megakaryocytic cell differentiation. These findings indicate that FOG acts as a cofactor for GATA-1 and provide a paradigm for the regulation of cell type-specific gene expression by GATA transcription factors.
3T3 Cells↗
A lineage-selective knockout establishes the critical role of transcription factor GATA-1 in megakaryocyte growth and platelet development.
Transcription factor GATA-1 is essential for red blood cell maturation and, therefore, for survival of developing mouse embryos. GATA-1 is also expressed in megakaryocytes, mast cells, eosinophils, multipotential hematopoietic progenitors and Sertoli cells of the testis, where its functions have been elusive. Indeed, interpretation of gene function in conventional knockout mice is often limited by embryonic lethality or absence of mature cells of interest, creating the need for alternate methods to assess gene function in selected cell lineages. Emerging strategies for conditional gene inactivation through site-specific recombinases rely on the availability of mouse strains with high fidelity of transgene expression and efficient, tissue-restricted DNA excision. In an alternate approach, we modified sequences upstream of the GATA-1 locus in embryonic stem cells, including a DNase I-hypersensitive region. This resulted in generation of mice with selective loss of megakaryocyte GATA-1 expression, yet sufficient erythroid cell levels to avoid lethal anemia. The mutant mice have markedly reduced platelet numbers, associated with deregulated megakaryocyte proliferation and severely impaired cytoplasmic maturation. These findings reveal a critical role for GATA-1 in megakaryocyte growth regulation and platelet biogenesis, and illustrate how targeted mutation of cis-elements can generate lineage-specific knockout mice.
Animals↗
A "knockdown" mutation created by cis-element gene targeting reveals the dependence of erythroid cell maturation on the level of transcription factor GATA-1.
The hematopoietic-restricted transcription factor GATA-1 is required for both mammalian erythroid cell and megakaryocyte differentiation. To define the mechanisms governing its transcriptional regulation, we replaced upstream sequences including a DNase I hypersensitive (HS) region with a neomycin-resistance cassette by homologous recombination in mouse embryonic stem cells and generated mice either harboring this mutation (neoDeltaHS) or lacking the selection cassette (DeltaneoDeltaHS). Studies of the consequences of these targeted mutations provide novel insights into GATA-1 function in erythroid cells. First, the neoDeltaHS mutation leads to a marked impairment in the rate or efficiency of erythroid cell maturation due to a modest (4- to 5-fold) decrease in GATA-1 expression. Hence, erythroid differentiation is dose-dependent with respect to GATA-1. Second, since expression of GATA-1 from the DeltaneoDeltaHS allele in erythroid cells is largely restored, transcription interference imposed by the introduced cassette must account for the "knockdown" effect of the mutation. Finally, despite the potency of the upstream sequences in conferring high-level, developmentally appropriate expression of transgenes in mice, other cis-regulatory elements within the GATA-1 compensate for its absence in erythroid cells. Our work illustrates the usefulness of targeted mutations to create knockdown mutations that may uncover important quantitative contributions of gene function not revealed by conventional knockouts.
Alleles↗