PURPOSE: The purposes of this study was to evaluates shear bond strength between zirconia core and veneer-ceramic in order to examine the clinical practice of colored zirconia block fabricated by infiltration method into the metal chloride solution. MATERIAL AND METHODS: CNU block and Everest(R) ZS blank were used. VITA In-Ceram(R)2000 YZ Coloring liquid (LL1) and 3 aqueous metal chloride solutions containing chromium and molybdenum ingredients were used. 40 zirconia specimens were prepared into cuboid shape (5 x 5 x 10 mm). All specimens were divided into 5 groups by infiltrating into the coloring liquids. After that, porcelain was build up into the shape of 5 x 5 x 4 mm3, followed by sintering. The maximum loading and shear bond strength was measured. Failure patterns and failure sites were examined. RESULTS: 1. There were no statistical differences in shear bond strength between zirconia blocks (P > .05). 2. There were no statistically significant differences in shear bond strength between non-colored and colored zirconia blocks, while shear bond strength of non-colored zirconia blocks is higher than that of colored specimen (P > .05). 3. In the comparison with shear bond strength among colored zirconia blocks, there were no statistical differences according to kinds of coloring liquid (P > .05). 4. Mixed failure patterns were mainly observed in the failure between zirconia and veneering ceramic. The veneering ceramic failure of all specimens was observed in either interface of zirconia or veneering ceramic. CONCLUSION: Shear bond strength between colored zirconia and veneering ceramic shows lower tendency than non-colored zirconia, but there was clinically allowable value.
Ceramics ; Chromium ; Dental Porcelain ; Ethylnitrosourea ; Molybdenum ; Zirconium

PURPOSE: The purpose of this study is to fabricate the new zirconia block (CNU block) and to evaluate fit of core and porcelain veneered zirconia crown. MATERIAL AND METHODS: The experimental blocks were fabricated from the commercial ytrria-stabilized zirconia powder (KZ-3YE Type A). The powder was uniaxial pressing and the green bodies were conducted using the Cold Isostatic Pressing. The zirconia blocks were presintered at 1040degrees C and the final sintering was performed at 1450degrees C. The Kavo Everest ZS blank(R) (KaVo, Biberach/Ri beta.) was used as a control group. The linear shrinkage of CNU block and Kavo block were compared. Twenty-one cores for porcelain veneered crowns were fabricated with CAD/CAM system (Everest(R), Biberach/Ribeta.). Group I: seven cores fabricated from Kavo blocks, Group II: seven cores fabricated from CNU blocks, Group III: seven cores from CNU blocks and porcelain veneering for crowns. All specimens were cemented and sectioned into two planes: diagonal and bucco-lingual. The measurement of the marginal, internal, and occlusal fit was carried out using SEM (S-4800(R)) at 30 x. The results were analyzed by one-way ANOVA test. RESULTS: The linear shrinkage of the CNU block and the KaVo block was 19.00% and 20.09%. The marginal gap of cores (29.67 +/- 6.58 micrometer) fabricated from CNU blocks showed significantly smaller than that of the cores of Kavo blocks (36.84 +/- 7.18 micrometer) (P < .05). The internal gaps of the porcelain veneered crowns (32.23 +/- 6.33 micrometer) were larger than those of the other two groups (37.57 +/- 6.81 micrometer and 38.14 +/- 6.81 micrometer). CONCLUSION: No statistically significant difference was found in between experimental groups and control group. The experimental groups in marginal gap showed significantly smaller than the control group.
Cold Temperature ; Crowns ; Dental Porcelain ; Ethylnitrosourea ; Zirconium

OBJECTIVETo screen and identify zebrafish mutants with erythropoiesis defects by N-ethyl-N-nitrosourea (ENU) mutagenesis and large-scale forward genetic screening using beta e 1 as the marker.

METHODSThe chemical mutagen ENU was used to treat healthy wild-type male fish (AB strain, F0). The surviving ENU-treated fish were mated with wild-type female fish to generate F1, and further F2 family was generated by F1 family intercross. The adult F2 fish were intercrossed within each F2 family and the resulting F3 embryos from each crossing were subjected to whole mount in situ hybridization (WISH) with the beta e 1 probe. Mutagenesis was performed by treating the male zebrafish with ENU to induce mutations in pre-meiotic germ cells to generate the founders, which were outcrossed to obtained the F1 fish. The F1 fish from different founders were mated to generate the F2 families. F3 embryos from the sibling cross in the F2 family were examined by whole mount in situ hybridization using beta e 1-globin probe. The putative mutants were then characterized with different hematopoiesis markers.

RESULTS AND CONCLUSIONWe identified 4 beta e 1-deficient mutants with erythropoiesis defects, including two with specific erythiod lineage defects and two with concurrent lymphopoiesis defects.

Animals ; Erythropoiesis ; genetics ; Ethylnitrosourea ; Female ; Gene Expression Regulation, Developmental ; Male ; Mutagenesis, Insertional ; Mutation ; Zebrafish ; genetics

OBJECTIVE: Tumor angiogenesis is an important factor for tumor growth, treatment response and prognosis. Noninvasive imaging methods for the evaluation of tumor angiogenesis have been studied, but a method for the quantification of tumor angiogenesis has not been established. This study was designed to evaluate tumor angiogenesis in a rat breast tumor model by the use of a contrast-enhanced ultrasound (US) examination with a second-generation US contrast agent. MATERIALS AND METHODS: The alkylating agent 19N-ethyl-N-nitrosourea (ENU) was injected into the intraperitoneal cavity of 30-day-old female Sprague-Dawley rats. Three to four months later, breast tumors were detected along the mammary lines of the rats. A total of 17 breast tumors larger than 1 cm in nine rats were evaluated by gray-scale US, color Doppler US and contrast-enhanced US using SonoVue. The results were recorded as digital video images; time-intensity curves and hemodynamic parameters were analyzed. Pathological breast tumor specimens were obtained just after the US examinations. The tumor specimens were stained with hematoxylin and eosin (H & E) and the expression of CD31, an endothelial cell marker, was determined by immunohistochemical staining. We also evaluated the pathological diagnosis of the tumors and the microvessel density (MVD). Spearman's correlation and the Kruskal-Wallis test were used for the analysis. RESULTS: The pathological diagnoses were 11 invasive ductal carcinomas and six benign intraductal epithelial proliferations. The MVD did not correlate with the pathological diagnosis. However, blood volume (BV) showed a statistically significant correlation with MVD (Spearman's correlation, p < 0.05). CONCLUSION: Contrast-enhanced US using a second-generation US contrast material was useful for the evaluation of tumor angiogenesis of breast tumors in the rat.
Animals ; Contrast Media ; Ethylnitrosourea ; Female ; Hemodynamics ; Image Enhancement ; Mammary Neoplasms, Experimental/chemically induced/pathology/*ultrasonography ; Neovascularization, Pathologic/*ultrasonography ; Rats ; Rats, Sprague-Dawley

BACKGROUND/AIMS: Hepatocytes on the hepatic lobule mipate from portal zone to centrilobular mea as the DNA synthesis within it. And also, the xenobiotic reactions reveal characteristic differences associated with zone specific metabolism in the liver acinus. In this study, the zonal distribution of ethylnitrosourea (ENU)-induced hepatic precancerous lesion was stereologically investigated. METHODS: Nine B6C3F1 mices were given I.p. injection of ENU (60 ug/pn body weight) when the pups were 15 days old prior to sacrifices at 8 weeks of life. All the 150 consecutive sections, 3 p m in thickness, were stained with hematoxylin and eosin and identified the basophilic precancerous lesions with 80-165 p m diameter in equatorial plane by the Zeiss microprojector. And then the distances from the center of selected foci to terminal hepatic vein or portal vein branches were estimated under the microscopic fields. As a control group, the same estimations were performed from the random points by the appointments of random digit table. RESULTS: Mean distance between ENU-induced 52 hepatocellular foci and the nearest terminal hepytic vein was 181.15+112.39 p m (Mean+ SD), but that of randomly selected 104 points was 291.73+157.98pm (Mean+5D) (Students t-test, p<0.0005). Substantially, 52.7% of ENU-induced 52 hepatocellular foci were within 300 p m from the terminal hepatic vein, but randomly selected 104 points were only 50.9% (Shapiro Wilk W test, w=0.819857, p=0.048038). Mean distance from ENU-induced 52 foci to portal vein was 398.85+149.98pm (Mean+SD), but that from the randomly selected 104 points was 315.87+145.79 pm (Mean+SD)(Students t-test, p<0.0005). CONCLUSION: Stereologically, ENU-induced mice liver cell foci distribute non-randomly to Zone III, centrilobular zone of mouse hepatic acini where promote invasion toward terminal hepatic veins.
Animals ; Appointments and Schedules ; Basophils ; Cholestasis ; DNA ; Eosine Yellowish-(YS) ; Ethylnitrosourea ; Fluconazole ; Hematoxylin ; Hepatic Veins ; Hepatocytes ; Liver ; Metabolism ; Mice* ; Portal Vein ; Veins

N-ethyl-N-nitrosourea (ENU) is a potent mutagen in a mouse model by inducing point mutation in a random manner and, in particular, causing heritable base substitutions in spermatogonia. In this study, systematic development of phenotype-driven mutant mice with large scale was carried out by using ENU. Nine-week-old male mice of C57BL/6J received intraperitoneal injection at three times with 100 mg/kg of ENU at weekly intervals for three weeks. After injections with ENU, the changes of body weight, fatality, recovery of fertile period, and breeding record were measured in these mice. Body weight lost as a result of ENU treatments was reversed after the last ENU injection. Live fertile male mice recovered from infertility from 104 to 165 days after ENU treatments were mated with C57BL/6J female mice for generation of G1 offspring. An average birth rate was 5.9 mice from 1 pair of paternal and maternal mice. All of 231 G1 offspring mice were analyzed by modified-SHIRPA with standard procedure at nine weeks of age. Among G1 mice, 166 mice were identified as mutagenic phenotypes in 20 test items. The changes in mutagenic phenotypes after ENU treatments, for instance, pattern in the region with a different color, touch escape, changes in head morphology, pupil, and teeth, and negative geotaxis etc., were found in these mice. Taken together, these results indicate that ENU may be a trans-generational mutagen in C57BL/6J mice.
Animals ; Birth Rate ; Body Weight ; Breeding* ; Ethylnitrosourea* ; Female ; Fertile Period* ; Head ; Humans ; Infertility ; Injections, Intraperitoneal ; Male ; Mice* ; Phenotype ; Point Mutation ; Pupil ; Spermatogonia ; Tooth ; United Nations

OBJECTIVETo perform the genetic identification of cloche(172) mutant zebrafish.

METHODSThe chemical mutagen N-ethyl-N-nitrosourea (ENU) was used to treat the AB stain male fish. Large-scale forward genetic screening was carried out to search for lyC-deficient zebrafish mutant by WISH. The morphology changes of the embryos at 3 days postfertilization (3dpf) stage were observed and the cloche(172) gene was identified by mapping and complementation test.

RESULTSWe selected 4 lyC-deficient zebrafish by WISH. cloche(172) mutant showed morphological changes similar to cloche mutant in 3dpf stage. One fourth of the embryos showed cloche phenotype as found in complementation test, and the cloche(172) gene was mapped on the telomere of zebrafish 13 chromosome where cloche gene was located. Numerous red blood cells were observed in the cloche(172) mutant, while only a few cells were found in the cloche mutant in the tail region by o-dianisdine staining.

CONCLUSIONcloche(172) gene which is responsible for the phenotype of cloche mutant may be a novel point mutation allele of the cloche mutant.

Alleles ; Animals ; Chromosome Mapping ; Cloning, Molecular ; Embryo, Nonmammalian ; embryology ; metabolism ; Ethylnitrosourea ; toxicity ; Genetic Complementation Test ; Male ; Muramidase ; genetics ; Mutation ; Zebrafish ; embryology ; genetics ; Zebrafish Proteins ; genetics

OBJECTIVETo explore the molecular spectra and mechanism of human hypoxanthine guanine phosphoribosyl transferase (HPRT) gene mutation induced by ethyluitrosourea (ENU).

METHODSSingle cell cloning culture, two-way screening, multiple PCR amplification and electrophoresis technique were used.

RESULTSWith dose of ENU increasing, cell plating efficiency reduced (in the group with 100-200 micro g/ml doses, P < 0.01) and mutation frequency increased (in the group with 12.5-200.0 micro g/ml doses, P < 0.05) significantly. There was no all exons deletion in spontaneous mutations, and only 7.7% of them were detected as single exon deletion. But, deletion was found in 79.7% of ENU-induced mutations (62.5%-89.4%, P < 0.01), and deletion mutations in all nine exons of HPRT gene. Most of ENU-induced mutations were chain deletion with multiple exons (88.1%).

CONCLUSIONSThe spectra in spontaneous mutations differed completely from ENU-induced ones. ENU was liable to cause bigger changes in genetic structure, which suggested a stronger ENU's mutagenesis.

Alkylating Agents ; pharmacology ; Ethylnitrosourea ; pharmacology ; HL-60 Cells ; Humans ; Hypoxanthine Phosphoribosyltransferase ; genetics ; metabolism ; Leukemia, Myeloid ; pathology ; Mutation ; drug effects ; Tumor Stem Cell Assay

PURPOSE: Genes of the HoxD cluster play a major role in vertebrate limb development, and changes that modify the Hoxd12 locus affect other genes also, suggesting that HoxD function is coordinated by a control mechanism involving multiple genes during limb morphogenesis. In this study, mutant phenotypes were produced by treatment of mice with a chemical mutagen, N-ethyl-N-nitrosourea (ENU). We analyzed mutant mice exhibiting the specific microdactyly phenotype and examined the genes affected. MATERIALS AND METHODS: We focused on phenotype characteristics including size, bone formation, and digit morphology of ENU-induced microdactyly mice. The expressions of several molecules were analyzed by genome-wide screening and quantitative real-time PCR to define the affected genes. RESULTS: We report on limb phenotypes of an ENU-induced A-to-C mutation in the Hoxd12 gene, resulting in alanine-to-serine conversion. Microdactyly mice exhibited growth defects in the zeugopod and autopod, shortening of digits, a missing tip of digit I, limb growth affected, and dramatic increases in the expressions of Fgf4 and Lmx1b. However, the expression level of Shh was not changed in Hoxd12 point mutated mice. CONCLUSION: These results suggest that point mutation rather than the entire deletion of Hoxd12, such as in knockout and transgenic mice, causes the abnormal limb phenotype in microdactyly mice. The precise nature of the spectrum of differences requires further investigation.
Animals ; Base Sequence ; DNA/genetics ; DNA Primers/genetics ; Ethylnitrosourea/toxicity ; Genes, Homeobox ; Homeodomain Proteins/*genetics ; Limb Deformities, Congenital/genetics ; Male ; Mice ; Mice, Inbred BALB C ; Mutagens/toxicity ; *Point Mutation ; Transcription Factors/*genetics

Striatal-enriched protein tyrosine phosphatase (STEP) is abundantly expressed in the striatum, which strongly expresses dopamine and opioid receptors and mediates the effects of many drugs of abuse. However, little is known about the role of STEP in opioid receptor function. In the present study, we generated STEP-targeted mice carrying a nonsense mutation (C230X) in the kinase interaction domain of STEP by screening the N-ethyl-N-nitrosourea (ENU)-driven mutant mouse genomic DNA library and subsequent in vitro fertilization. It was confirmed that the C230X nonsense mutation completely abolished functional STEP protein expression in the brain. STEP(C230X−/−) mice showed attenuated acute morphine-induced psychomotor activity and withdrawal symptoms, whereas morphine-induced analgesia, tolerance and reward behaviors were unaffected. STEP(C230X−/−) mice displayed reduced hyperlocomotion in response to intrastriatal injection of the μ-opioid receptor agonist DAMGO, but the behavioral responses to δ- and κ-opioid receptor agonists remained intact. These results suggest that STEP has a key role in the regulation of psychomotor action and physical dependency to morphine. These data suggest that STEP inhibition may be a critical target for the treatment of withdrawal symptoms associated with morphine.
Analgesia* ; Animals ; Brain ; Codon, Nonsense ; Dopamine ; Enkephalin, Ala(2)-MePhe(4)-Gly(5)- ; Ethylnitrosourea ; Fertilization in Vitro ; Gene Library ; Mass Screening ; Mice ; Morphine* ; Phosphotransferases ; Protein Tyrosine Phosphatases ; Receptors, Opioid ; Reward* ; Street Drugs ; Substance Withdrawal Syndrome*