Biomedical subjects

Kyoji Hioki

Publications and source records attributed to Kyoji Hioki.

At least 19 recordsLinked to original sources

PubMed2006

The role of leptin in the development of the cortical neuron in mouse embryos.

Leptin is an obese gene product that decreases appetite and raises energy expenditure in adults. We previously reported that leptin was detected in the sera of mouse embryos and leptin receptors were expressed in the mouse embryonic cerebrum, suggesting that leptin plays a role in cerebral development. In this study, we injected leptin into the lateral ventricle of the cerebrum in leptin-deficient ob/ob mouse embryos to investigate the function of leptin in cerebral development. When leptin was injected on embryonic day (E) 14, the ratio of the number of cells in the cortical plate (CP) to that in the intermediate zone (IZ) was higher in leptin-injected than in vehicle-injected ob/ob embryos on E16, although there was no significant difference in the number of cells in the ventricular zone (VZ), IZ, or CP between these groups. The number of postmitotic BrdU-positive CP cells was larger in leptin-injected than in vehicle-injected ob/ob embryos on E16 and E17 when BrdU labeling and leptin injection were performed on E14. By in situ hybridization, NPY mRNA expression in CP neurons on E18 was weaker in leptin-injected than in vehicle-injected ob/ob embryos when leptin was injected on E16. These results suggest that leptin promotes the migration of neuronal lineage cells to CP and that the leptin-NPY axis in neurons works in the cerebral cortex on E16.

Age Factors↗

PubMed2006

The tyrosine kinase inhibitor imatinib [STI571] induces regression of xenografted canine mast cell tumors in SCID mice.

Canine mast cell tumors (MCTs) are the most common cutaneous tumors in the dog. They have a wide range of behaviour, which can make these tumors challenging to treat. Recently, mutations in c-kit proto-oncogene have been identified in several canine MCTs. Imatinib is the first member of a new class of agents that act by inhibiting particular tyrosin kinase enzymes, including KIT which is a product of the c-kit. In this study the efficacy of imatinib to reduce or abolish canine MCT [CMC-1] using xenografted MCT in severe combined immunodeficient [SCID] mice was evaluated. Imatinib was administered at doses of 200mg/kg and 100mg/kg once a day for one week. The antitumor responses in SCID mice with CMC-1 xenografts following treatment with imatinib were observed. Significant tumor regression occurred with 100mg/kg on days 7, 10, 14 and 21, and 200mg/kg on all days. Our results indicate that imatinib is effective against canine mast cell tumor in mouse xenograft models. Canine MCTs might be a potential target for imatinib therapy.

Animals↗

PubMed2006

Nine-week detection of six genotoxic lung carcinogens using the rasH2/BHT mouse model.

A 9-week in vivo rasH2/butylhydroxytoluene (BHT) model for the detection of genotoxic lung carcinogens was validated, using six potent positive test compounds, dimethylnitrosamine (DMN; 15 mg/kg, i.p.), diethylnitrosamine (DEN; 100 mg/kg, i.p.), ethylnitrosourea (ENU; 120 mg/kg, i.p.), 3-methylcholanthrene (MC; 100 mg/kg, i.p.), 7,12-dimethylbenz(a)anthracene (DMBA; 5 mg/kg, i.g.) and benzo(a)pyrene (B(a)P; 80 mg/kg, i.p.), each given to rasH2 mice of both genders by single administration for initiation followed by promoter BHT treatment. Statistically significant increase in the incidence and multiplicity of lung tumors was observed in rasH2 mice treated with BHT following exposure to all of the carcinogens tested. The data overall suggest the rasH2/BHT model to be a powerful screening tool for genotoxic lung carcinogens.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed2006

Phenotypic abnormalities observed in aged cloned mice from embryonic stem cells after long-term maintenance.

Somatic/embryonic stem cell cloning has made it possible to produce an individual genomically identical to another individual. However, the cloned animals have a variety of abnormalities caused by the aberrant gene modification, with insufficient reprogramming in cloning. We previously reported abnormalities in cloned mice at birth. In this study, we examined what abnormalities could be seen in cloned mice after long-term maintenance. The aged cloned mice showed multiple abnormalities: increase of body weight, some phenotypic abnormalities in the kidneys, testes and thymus, and lower urea nitrogen in their serum biochemical values. The kidneys of all cloned mice were hypertrophied, with a metamorphic or whitish appearance. The multiple lesions, including the enlarged renal pelvis and distension of the renal veins in histology, might be the result of urine accumulation by urinary tract obstruction. The testes of the cloned mice were atrophied, and showed no sperm formation in histology. In contrast, the thymus was rather hypertrophied, and a comparably increased number of lymphocytes were observed in the medulla, consisting mainly of T cells. By conducting a progeny test between the cloned mice, it was confirmed that these abnormalities in the aged cloned mice were not transmitted to their offspring, indicating that the incomplete reprogramming in clones might be in part responsible for the abnormalities detected in aged clones. These results indicate that the postnatal abnormalities observed in aged cloned mice are varied and can be restored through the germ line.

Aging↗

PubMed2005

The role of leptin in the development of the cerebral cortex in mouse embryos.

Leptin is detected in the sera, and leptin receptors are expressed in the cerebrum of mouse embryos, suggesting that leptin plays a role in cerebral development. Compared with the wild type, leptin-deficient (ob/ob) mice had fewer cells at embryonic day (E) 16 and E18 and had fewer 5-bromo-2'-deoxyuridine(+) cells at E14 and E16 in the neuroepithelium. Intracerebroventricular leptin injection in E14 ob/ob embryos increased the number of neuroepithelium cells at E16. In cultured neurosphere cells, leptin treatment increased Hes1 mRNA expression and maintained neural progenitors. Astrocyte differentiation was induced by low-dose (0.1 microg/ml) but not high-dose (1 microg/ml) leptin. High-dose leptin decreased Id mRNA and increased Ngn1 mRNA in neurosphere cells. The neuropeptide Y mRNA level in the cortical plate was lower in ob/ob than the wild type at E16 and E18. These results suggest that leptin maintains neural progenitors and is related to glial and neuronal development in embryos.

Animals↗

PubMed2005

Functional thalamocortical synapse reorganization from subplate to layer IV during postnatal development in the reeler-like mutant rat (shaking rat Kawasaki).

Transient synapse formation between thalamic axons and subplate neurons is thought to be important in thalamocortical targeting. Shaking rat Kawasaki (SRK), having reversed cortical layering similarly observed in reeler mouse, provides an interesting model system to test this idea. The spatial and temporal pattern of excitation was investigated using optical recording with voltage-sensitive dyes in thalamocortical slice preparations from SRK. At postnatal day 0 (P0), a strong optical response was elicited within the superplate of the SRK in the cell layer corresponding to subplate in wild-type (WT) rats. By P3, this response rapidly descended into deep cortical layers comprised of layer IV cells, as identified with 5-bromo-2'-deoxyuridine birthdating at embryonic day 17. During the first 3 postnatal days, both the subplate and cortical plate responses were present, but by P7, the subplate response was abolished. Tracing individual axons in SRK revealed that at P0-P3, a large number of thalamocortical axons reach the superplate, and by P7-P10, the ascending axons develop side branches into the lower or middle cortical layers. Synaptic currents were also demonstrated in WT subplate cells and in SRK superficial cortical cells using whole-cell recording. These currents were elicited monosynaptically, because partial AMPA current blockade did not modify the latencies. These results suggest that the general developmental pattern of synapse formation between thalamic axons and subplate (superplate) neurons in WT and SRK is very similar, and individual thalamic arbors in cortex are considerably remodeled during early postnatal development to find layer IV equivalent neurons.

Action Potentials↗

PubMed2005

A Kampo formula Juzen-taiho-to induces expression of metallothioneins in mice.

Juzen-taiho-to (JTX), one of the commonly prescribed traditional Japanese herbal medicines (Kampo), is indicated for adjunctive treatment of cancers and autoimmune diseases. To understand the mechanisms underlying the clinical effects of JTX, the effects of orally administered JTX on the expression of metallothioneins (MTs) were examined in the liver, spleen, small and large intestines of mice. In addition, the expression of MTs in specific pathogen free (SPF) mice was examined to understand the participation of intestinal bacteria in the expression of MTs. JTX enhanced expression of MT-I and -II significantly in the liver of SPF mice. Induction of MT-II expression was observed also in the small intestine. Intestinal bacteria appeared to have no effect on MTs expression. Neither expression of MT-III nor its induction was observed in any tissue. These findings strongly suggest that MTs should mediate at least some effects of JTX in mice.

Administration, Oral↗

PubMed2005

Estrous stage- and animal age-independent superovulation in the BrlHan:WIST@Jcl(GALAS) rat.

In most strains of rats, the effects of treatment for the induction of superovulation show major strain differences and are strongly influenced by the stage of the estrous cycle. This study demonstrated, however, that superovulation was easily induced in Wistar strain Brl Han:WIST@Jcl(GALAS) rats by PMSG and hCG administration. To confirm the effects of such treatment, we studied age differences in egg collection efficiency. After superovulation was induced by intraperitoneal administration of 150 IU/kg PMSG and 75 IU/kg hCG given 48 h apart, the mean numbers of oocytes obtained from rats at 4, 8, 12, 20 and 28 weeks of age were 38.9, 33.5, 46.1, 26.9 and 21.3, respectively. No differences caused by the estrous stage at the PMSG administration were observed. In an embryo transfer experiment, fertilized eggs obtained from superovulated rats at each week of age showed equivalent viability until full-term to those from untreated rats. These results suggest that estrous stage-independent superovulation is effective not only in the pre-pubertal stage but also in adult rats.

Aging↗

PubMed2004

Involvement of illegitimate V(D)J recombination or microhomology-mediated nonhomologous end-joining in the formation of intragenic deletions of the Notch1 gene in mouse thymic lymphomas.

Deregulated V(D)J recombination-mediated chromosomal rearrangements are implicated in the etiology of B- and T-cell lymphomagenesis. We describe three pathways for the formation of 5'-deletions of the Notch1 gene in thymic lymphomas of wild-type or V(D)J recombination-defective severe combined immune deficiency (scid) mice. A pair of recombination signal sequence-like sequences composed of heptamer- and nonamer-like motifs separated by 12- or 23-bp spacers (12- and 23-recombination signal sequence) were present in the vicinity of the deletion breakpoints in wild-type thymic lymphomas, accompanied by palindromic or nontemplated nucleotides at the junctions. In scid thymic lymphomas, the deletions at the recombination signal sequence-like sequences occurred at a significantly lower frequency than in wild-type mice, whereas the deletions did not occur in Rag2(-/-) thymocytes. These results show that the 5'-deletions are formed by Rag-mediated V(D)J recombination machinery at cryptic recombination signal sequences in the Notch1 locus. In contrast, one third of the deletions in radiation-induced scid thymic lymphomas had microhomology at both ends, indicating that in the absence of DNA-dependent protein kinase-dependent nonhomologous end-joining, the microhomology-mediated nonhomologous end-joining pathway functions as the main mechanism to produce deletions. Furthermore, the deletions were induced via a coupled pathway between Rag-mediated cleavage at a cryptic recombination signal sequence and microhomology-mediated end-joining in radiation-induced scid thymic lymphomas. As the deletions at cryptic recombination signal sequences occur spontaneously, microhomology-mediated pathways might participate mainly in radiation-induced lymphomagenesis. Recombination signal sequence-mediated deletions were present clonally in the thymocyte population, suggesting that thymocytes with a 5'-deletion of the Notch1 gene have a growth advantage and are involved in lymphomagenesis.

Animals↗

PubMed2004

PCR method for genotyping and zygosity-testing of RasH2 transgenic mice.

In short-term carcinogenicity testing using CB6F1-TgrasH2 mice, sibling nonTgrasH2 mice are used as a negative control. However, selection of TgrasH2 and nonTgrasH2 mice has been performed by PCR with only transgene specific primers by the conventional method. Therefore, the conventional method involves the risk of false negative results due to reaction failure, and contamination with TgrasH2 mice in the control mice group. Based on the nucleotide sequence information around the pre-integration site, we developed a genotyping method for distinguishing not only TgrasH2 mice (hemizygous for the Tg allele) but also nonTgrasH2 (homozygous for the nonTg allele) in a positive manner.

Animals↗

PubMed2003

Histological study of the hypertrophic placentas and open eyelids observed in cloned fetuses.

Mice cloned from somatic or ES cells showed signs of phenotypically various abnormalities. These abnormalities are now considered to result from aberrant gene expressions by epigenetic reprogramming errors but it is still unclear when these abnormalities occur and what histological changes occur during the gestation period. To address these issues, we histologically examined the hypertrophic placentas and open eyelids at 12.5, 17.5 and 19.5 days of the gestation period in ES-derived cloned mice that we have previously reported. In the placentas, the histology revealed that the hypertrophy had already occurred at 12.5 dpc and that the main change was the proliferation of trophoblast cells in the labyrinth layer. In the fetuses and placentas at 17.5 and 19.5 dpc, extensive proliferation of spongiotrophoblast and glycogen cells in the spongiotrophoblast layer and enlarged trophoblast giant cells were observed. Open eyelids in cloned mice were observed from 17.5 dpc, whereas the eyelids of the control mice had already been closed. The histology showed the malformation of eyelids where the formation of the stratum corneum and stratum granulosum in the epidermis was insufficient. Based on the histology described here, further comparative studies of the gene expression and histology of abnormalities seen in cloned mice and in gene-targeted and spontaneously mutated mice with similar phenotypic abnormalities could help illuminate these abnormalities and could contribute to the development of somatic cloning technology.

Animals↗

PubMed2003

Erythrocyte-replaced mouse model for Haemoparasite studies: comparison of NOD/shi-scid and C.B-17/Jcl-scid mouse upon acceptability of human erythrocytes.

The erythrocyte-exchanged chimera mouse model has become to be a significant tool for studying animal and human (hu) protozoan haemoparasites, though the usefulness of this model varies depending primarily on the acceptability of xenogeneic red blood cells (RBC). To find a superior recipient in comparison with C.B-17/Jcl mouse with severe combined immuno-deficiency (scid) mutation, we examined in this report the non-obese diabetes (NOD)/shi-scid mouse, a recently available strain of SCID. When 2.5 x 10(8) of fluorescent dye-labeled hu-RBCs were transfused, C.B-17scid mouse eliminated them logarithmically by a simple linear regression, while NOD-scid mouse eradicated hu-RBCs by a unique two-step fashion, i.e., a potent but only briefly functioning RBC eradication followed by a weak steadily functioning step. The means of regression line constance +/- their standard deviations (SD) of 205 C.B-17scid and of 213 NOD-scid mice for their short- and long-lasting steps were -0.73 +/- 0.63, -0.53 +/- 0.25 and -0.16 +/- 0.10, respectively. Hu-RBC half-lives determined from these means of C.B-17scid mice and of NOD-scid mice for the short- and long-living steps were 3.6, 4.9 and 16.3 hr, respectively. Higher hu-RBC acceptability of NOD-scid mouse, especially at their long-lasting step, was also demonstrated under at an activated state of mouse innate immunity. Treatment with 1.0 mg heat-killed Candida cells caused an acceleration of hu-RBC elimination in both mouse strains but the magnitudes for the short- and long-living steps of NOD-scid mice evaluated by "stimulation index" were only 1/2.6 and 1/7.6 of C.B-17scid mice, respectively.

Animals↗

PubMed2003

A genetic quality testing system for early stage embryos in the mouse.

We have established a genetic quality testing system for early stage embryos of the mouse. A method of preparation of template DNA for PCR was established using the lysis buffer (1 x PCR reaction buffer supplemented with proteinase K at a concentration of 40 microg/ml) developed by the authors. We demonstrated that two 8-cell embryos of an inbred strain provide sufficient volumes of template DNA for PCR to identify the strain of embryos using four microsatellite markers (D3Mit54, D5Mit18, D6Mit15 and D8Mit50) differentiating 13 inbred strains of mice. This system will be useful in embryo banks that have recently been established worldwide for demonstrating the genetic accuracy of a given strain prior to recovery of live animals.

Animals↗

PubMed2002

NOD/SCID/gamma(c)(null) mouse: an excellent recipient mouse model for engraftment of human cells.

To establish a more appropriate animal recipient for xenotransplantation, NOD/SCID/gamma(c)(null) mice double homozygous for the severe combined immunodeficiency (SCID) mutation and interleukin-2Rgamma (IL-2Rgamma) allelic mutation (gamma(c)(null)) were generated by 8 backcross matings of C57BL/6J-gamma(c)(null) mice and NOD/Shi-scid mice. When human CD34+ cells from umbilical cord blood were transplanted into this strain, the engraftment rate in the peripheral circulation, spleen, and bone marrow were significantly higher than that in NOD/Shi-scid mice treated with anti-asialo GM1 antibody or in the beta2-microglobulin-deficient NOD/LtSz-scid (NOD/SCID/beta2m(null)) mice, which were as completely defective in NK cell activity as NOD/SCID/gamma(c)(null) mice. The same high engraftment rate of human mature cells was observed in ascites when peripheral blood mononuclear cells were intraperitoneally transferred. In addition to the high engraftment rate, multilineage cell differentiation was also observed. Further, even 1 x 10(2) CD34+ cells could grow and differentiate in this strain. These results suggest that NOD/SCID/gamma(c)(null) mice were superior animal recipients for xenotransplantation and were especially valuable for human stem cell assay. To elucidate the mechanisms involved in the superior engraftment rate in NOD/SCID/gamma(c)(null) mice, cytokine production of spleen cells stimulated with Listeria monocytogenes antigens was compared among these 3 strains of mice. The interferon-gamma production from dendritic cells from the NOD/SCID/gamma(c)(null) mouse spleen was significantly suppressed in comparison with findings in 2 other strains of mice. It is suggested that multiple immunological dysfunctions, including cytokine production capability, in addition to functional incompetence of T, B, and NK cells, may lead to the high engraftment levels of xenograft in NOD/SCID/gamma(c)(null) mice.

Animals↗

PubMed2002

Abnormalities in cloned mice are not transmitted to the progeny.

Cloned animals suffer a wide range of severe fetal and placental malformations. Whether these malformations arise from insufficient epigenetic modifications or mutations has not yet been determined. To address this question, we examined siblings from both cloned XO and XY parents. These parents, which exhibited hypertrophic placentas, increased body weights, and open eyelids at birth, were created from the same ES cell sublines. The siblings from all three cloned pairs showed normal body and placenta weights and no open eyelids at birth. The results clearly showed that the phenotypic abnormalities seen in cloned mice were not transmitted to the progeny, a finding that suggests that abnormalities in cloned mice are responsible for insufficient epigenetic modifications/reprogramming.

Animals↗

PubMed2002

Transgene stability and features of rasH2 mice as an animal model for short-term carcinogenicity testing.

The transgenic mouse rasH2 line, in which the mouse carries the human c-Ha-ras gene under the control of its own enhancer and promoter, has been proposed as one of the alternative short-term models for carcinogenicity testing. To apply this purpose, we have produced a genetically homogeneous population as C57BL/6JJic-TgN(RASH2) (Tg-rasH2) by continuous backcrossing. In this study, we examined the transgene stability between different generations and the detailed transgene architecture of the integrated human c-Ha-ras gene. Fluorescence in situ hybridization analysis showed that the integrated human c-Ha-ras gene was stably located on chromosome 15E3 in Tg-rasH2 mice at generation number (N) 15 and 20. Southern and Northern blot analysis did not show any differences in the hybridized band pattern in each generation. Southern blot analyses showed that the Tg-rasH2 mouse contained three copies of the human c-Ha-ras gene arrayed in a head-to-tail configuration. We also determined the nucleotide sequence of the transgene in the Tg-rasH2 mouse at N20 and confirmed that the sequence of the coding region was perfectly matched with human c-Ha-ras cDNA. Cloning and sequencing of genome/transgene junctions revealed that integration of the microinjected human c-Ha-ras gene into mouse host genome resulted in a 1820-bp deletion in the rasH2 line. The deleted sequence did not have any sequence homologies with known functional genes. We assumed that either the deletion or the transgene insertion, or both, would not cause insertional mutation. In short-term carcinogenicity testing with a genetically engineered mouse model, confirmation of the transgene or modified gene stability at each generation is one of the important factors that affect the sensitivity to carcinogenic compounds in the same way as the genetic background, age and route of administration.

Animals↗

PubMed2002

Effects of cryopreservation of mouse embryos and in vitro fertilization on genotypic frequencies in colonies.

To evaluate the effects of cryopreservation and in vitro fertilization (IVF) on genotypic frequencies in mouse colonies, genotypic frequencies at 15 biochemical, 4 immunological and 20 microsatellite loci were examined in three colonies of MCH (ICR) mice derived from noncryopreserved embryos obtained by natural mating without the induction of superovulation, cryopreserved embryos obtained by natural mating with the induction of superovulation, and cryopreserved embryos obtained by the induction of superovulation and IVF. Three (Pgm-1, Ldr-1 and Hbb) out of the 15 biochemical loci, two (Thy-1 and H2K) out of four immunological loci and five (D5Mit18, D6Mit15, D12Mit5, D13Mit26, and D14Mit7) out of 20 microsatellite loci that showed polymorphisms in every colony were used for detection of genotypic frequencies. The genotypic frequencies of the loci in the three colonies did not differ from the predicted genotypic frequencies (P > 0.05). The results suggested that genetic drift does not occur among colonies established from treated and untreated embryos, and it was clear that the embryo banking by cryopreservation is suitable for preservation of outbred stock without genetic drift.

Animals↗

PubMed2002

The mouse rasH2/BHT model as an in vivo rapid assay for lung carcinogens.

We have demonstrated the utility of a 9-week in vivo two-stage assay for lung cancer initiating agents, using transgenic mice carrying the human prototype c-Ha-ras gene (rasH2 mice) and butylhydroxytoluene (BHT) as a potent lung promoter (rasH2/BHT model). In the present study, to ascertain appropriate conditions for BHT administration in this model, the effects of exposure on proliferation of alveolar type II cells in male rasH2 mice were examined. Additionally, use of BHT was validated for promotion of urethane (UR) carcinogenesis in male and female rasH2 mice. In a time-course study of a single intragastric administration of BHT at a dose of 400 mg/kg, increased bromodeoxyuridine-labeling index (LI) reached a maximum 3 days after treatment and was still observed after 7 days. In a dose-response study, effects were dose-dependent, the dose of 400 mg/kg causing eight-fold elevation as compared to the control. With repeated administration, whereas the LI was increased dramatically at first, effects gradually diminished with further exposure, and finally six BHT treatments failed to induce cell proliferation. In a two-stage model using UR as the initiator, although up to five consecutive doses of BHT were able to exert continued enhancing effects in terms of adenoma yield, no increment was evident with further treatments. The data overall indicate that a rasH2/BHT model with five weekly administrations of BHT at a dose of 400 mg/kg is most efficacious.

Animals↗