9 results on '"Structural Research"'
Search Results
2. Establishment of an organ culture system to induce Sertoli cell differentiation from undifferentiated mouse gonads.
- Author
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Hasegawa C, Yokoyama T, Umemura Y, Kawanishi K, Miura Y, Takada N, Ohno S, Onaru K, Omotehara T, Hirano T, Mantani Y, Miki T, and Hoshi N
- Subjects
- Animals, Anti-Mullerian Hormone metabolism, Embryo, Mammalian, Female, Male, Mice, Inbred ICR, RNA, Messenger, SOX9 Transcription Factor metabolism, Sex Differentiation, Sex-Determining Region Y Protein genetics, Sex-Determining Region Y Protein metabolism, Testis cytology, Cell Differentiation, Organ Culture Techniques veterinary, Sertoli Cells, Testis embryology
- Abstract
Organ culture systems are useful for elucidating the process of testicular differentiation from mammalian undifferentiated genetically male gonads, as they permit various experiments, including experiments involving the control of gene expression. However, without addition of testicular differentiation-related factors, it is difficult to induce the formation of testis cord from immature gonads by a time point earlier 12 tail somites (ts) that corresponding to 11.0 days post coitum (dpc). In this study, we attempted to establish an organ culture system that induces testis formation from immature gonads (around 8 ts: 10.5 dpc) just before Sry (sex-determining region of the Y chromosome) expression. A paired gonad-mesonephros complex of around 8 ts was placed in the groove of an agarose gel block and put the semi-cylindrical piece of agarose gel to maintain the gonad morphology. The gonads were cultured in the gas phase for 96 hr. As a result, testis cord-like structures appeared in many genetically male gonads. Cells expressing the Sertoli cell markers Sox9 (SRY-box 9) and Amh (anti-Müllerian hormone) were observed, while granulosa cell marker Foxl2 (forkhead box L2) was not detected. In addition, Sox9- and Amh-expressing cells were observed throughout the entire gonad in many individuals. Amh mRNA expression was also upregulated. Surprisingly, formation of a partial testicular structure was observed from more immature gonads (6 ts). These results show that our gonadal organ culture system is useful for elucidating the regulation mechanism of Sry expression in undifferentiated bipotential gonads.
- Published
- 2020
- Full Text
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3. Immunotoxicity evaluation by subchronic oral administration of clothianidin in Sprague-Dawley rats.
- Author
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Onaru K, Ohno S, Kubo S, Nakanishi S, Hirano T, Mantani Y, Yokoyama T, and Hoshi N
- Subjects
- Administration, Oral, Animals, Gastrointestinal Microbiome drug effects, Granuloma chemically induced, Guanidines administration & dosage, Hypersensitivity, Delayed chemically induced, Immunohistochemistry, Insecticides administration & dosage, Male, Neonicotinoids administration & dosage, Nicotinic Agonists, Organ Size drug effects, Rats, Sprague-Dawley, Thiazoles administration & dosage, Thymus Gland drug effects, Guanidines toxicity, Immune System drug effects, Insecticides toxicity, Neonicotinoids toxicity, Thiazoles toxicity
- Abstract
Neonicotinoid pesticides (NNs) act as agonists on nicotinic acetylcholine receptors (nAChRs) of insects, and there have been concerns about the effects of NNs on the health of mammals. Since nAChRs are expressed in immune cells, it is possible that NNs disturb the immune system. However, few reports have examined the immunotoxicity of clothianidin (CLO), a widely-used NN. Here, we report the effects of CLO on immune organs and type IV allergic reactions in ear auricles. We orally administered CLO at 0, 30 and 300 mg/kg/day (CLO-0, 30 and 300) to Sprague-Dawley rats for 28 days. The effects were evaluated by organ and body weights, histopathology, and immunohistochemistry (TCRαβ, CD4, CD8, CD11b, CD68, CD103). In addition, some cecal contents were subjected to preliminary gut microbiota analysis, because microbiota contribute to host homeostasis, including the immunity. Our results showed loose stool, suppression of body weight gain, significant changes in organ weights (thymus: decreased; liver: increased) and changes of the gut microbiota in the CLO-300 group. There were no obvious histopathological changes in immune organs. Granulomas of the ear auricles were found in one rat of each of the CLO-30 and 300 groups, but CLO had no apparent effect on the thickness or immunohistochemistry in the ear auricles. We present new evidence that CLO affects the thymus and intestine, and might enhance the local inflammatory response. These findings should contribute to the appropriate evaluation of the safety of NNs in the future.
- Published
- 2020
- Full Text
- View/download PDF
4. Combined exposure to dinotefuran and chronic mild stress counteracts the change of the emotional and monoaminergic neuronal activity induced by either exposure singly despite corticosterone elevation in mice.
- Author
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Takada T, Yoneda N, Hirano T, Onaru K, Mantani Y, Yokoyama T, Kitagawa H, Tabuchi Y, Nimako C, Ishizuka M, Ikenaka Y, and Hoshi N
- Subjects
- Animals, Behavior, Animal drug effects, Corticosterone blood, Emotions physiology, Exploratory Behavior drug effects, Guanidines blood, Hindlimb Suspension, Insecticides toxicity, Male, Mice, Inbred C57BL, Neonicotinoids blood, Nitro Compounds blood, Serotonin metabolism, Swimming, Tryptophan Hydroxylase metabolism, Tyrosine 3-Monooxygenase metabolism, Brain metabolism, Guanidines toxicity, Neonicotinoids toxicity, Nitro Compounds toxicity, Stress, Psychological
- Abstract
Dinotefuran (DIN) belongs to the neonicotinoids (NNs), a class of globally applied pesticides originally developed to exhibit selective toxicity in insects. However, several reports have suggested that NNs also exert neurotoxic effects in mammals. We previously demonstrated neurobehavioral effects of DIN on mice under non-stressful conditions. For further toxicity assessments in the present study, we investigated the effects of DIN on mice exposed to stressful conditions. After subacutely administering a no-observed-effect-level (NOEL) dose of DIN and/or chronic unpredictable mild stress (CUMS) to mice, we conducted three behavioral tests (i.e., open field test [OFT], tail suspension test [TST] and forced swimming test [FST]). In addition, serotonin (5-HT) and tryptophan hydroxylase 2 (TPH2) of the dorsal raphe nuclei (DRN) and median raphe nuclei (MRN) and tyrosine hydroxylase (TH) of the ventral tegmental area and substantia nigra (SN) were evaluated immunohistochemically. A NOEL dose of DIN or CUMS alone increased of the total distance in OFT, decreased or increased the immobility time in TST or FST, respectively, and increased the positive intensity of 5-HT and TPH2 in the DRN/MRN, and TH in the SN. These changes were suppressed under the conditions of combined exposure to DIN and CUMS, though the blood corticosterone level was increased depending on the blood DIN values and the presence of CUMS. The present study suggests the multifaceted toxicity of the neurotoxin DIN.
- Published
- 2020
- Full Text
- View/download PDF
5. Genetic differences between C57BL/6 substrains affect the process of testis differentiation in Y POS mice.
- Author
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Yokoyama T, Miura Y, Yamamoto A, Hasegawa C, Kawanishi K, Takada N, Omotehara T, Hirano T, Mantani Y, Miki T, and Hoshi N
- Subjects
- Animals, Crosses, Genetic, Female, Male, Mice, Inbred C57BL, Ovary cytology, Sex Differentiation genetics, Species Specificity, Sex Determination Processes genetics, Testis cytology, Y Chromosome
- Abstract
C57BL/6J-XY
POS (B6J-XYPOS ) mice, which have the Y chromosome derived from Mus musculus poschiavinus on a B6J genetic background, form ovotestes or ovaries. Previously, we replaced the genetic background of B6J-XYPOS mice with B6N and found that individuals with testes also appeared in addition to those with ovaries or ovotestes. To investigate the effect of the B6J genetic sequence on the testis differentiation, the genetic background of B6N-XYPOS mice was replaced with B6J again. The recovery of the B6J genetic background significantly decreased the incidence of testes; only ovaries developed. These results indicate that the testicular differentiation process tends to be perturbed especially in the B6J substrain. This shows the importance of substrain differences in mice usually treated as B6 collectively.- Published
- 2019
- Full Text
- View/download PDF
6. Identification of reference genes for quantitative PCR analyses in developing mouse gonads.
- Author
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Yokoyama T, Omotehara T, Hirano T, Kubota N, Yanai S, Hasegawa C, Takada T, Mantani Y, and Hoshi N
- Subjects
- Algorithms, Animals, Female, Genes, Reporter, Male, Mice, Mice, Inbred C57BL, Reference Standards, Genes, Developmental, Gonads growth & development, Real-Time Polymerase Chain Reaction
- Abstract
Stable reference genes are important for gene expression analyses such as quantitative PCR. The stability of 15 candidate reference genes that can be used to developing mouse gonads was thoroughly verified using combinations of multiple algorithms. The expression of these genes fluctuated greatly depending on the analysis period and/or gender. Peptidylprolyl isomerase A (Ppia) and polymerase (RNA) II (DNA directed) polypeptide A (Polr2a) were the reference genes that were used stably for a wide analysis period in developing mouse gonads. Furthermore, the stable reference genes corresponding to the analysis period and/or gender were ranked. These results are useful for the selection of the optimal reference gene required for high-precision measurements.
- Published
- 2018
- Full Text
- View/download PDF
7. Verification of the causal relationship between subchronic exposures to dinotefuran and depression-related phenotype in juvenile mice.
- Author
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Takada T, Yoneda N, Hirano T, Yanai S, Yamamoto A, Mantani Y, Yokoyama T, Kitagawa H, Tabuchi Y, and Hoshi N
- Subjects
- Animals, Brain Chemistry drug effects, Guanidines administration & dosage, Hindlimb Suspension veterinary, Male, Mice, Neonicotinoids administration & dosage, Nitro Compounds administration & dosage, Serotonin analysis, Depression chemically induced, Guanidines adverse effects, Insecticides adverse effects, Neonicotinoids adverse effects, Nitro Compounds adverse effects
- Abstract
It has been suggested that an increase in the use of pesticides affects neurodevelopment, but there has been no animal experiment showing a causal relation between neonicotinoid pesticides (NNs) and depression. We examined whether dinotefuran (DIN), the most widely used NN in Japan, induces depression. Male mice were administered DIN between 3 and 8 weeks of age, referring to the no-observed-effect level (NOEL). The mice were then subjected to a tail suspension test (TST) and a forced swimming test (FST). After these tests, their brains were dissected for immunohistochemical analyses of serotonin (5-HT). Antidepressant activity in TST and no decrease in 5-HT-positive cells were observed. The subchronic exposure to DIN alone in juvenile male mice may not cause depression-like indication.
- Published
- 2018
- Full Text
- View/download PDF
8. The mechanisms underlying the effects of AMH on Müllerian duct regression in male mice.
- Author
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Yamamoto A, Omotehara T, Miura Y, Takada T, Yoneda N, Hirano T, Mantani Y, Kitagawa H, Yokoyama T, and Hoshi N
- Subjects
- Animals, Anti-Mullerian Hormone genetics, Female, Hermaphroditic Organisms, Male, Mice, Receptors, Transforming Growth Factor beta, Sex Differentiation, Anti-Mullerian Hormone metabolism, Gene Expression Regulation, Developmental physiology, Mullerian Ducts embryology, Testis embryology, Testis metabolism
- Abstract
Anti-Müllerian hormone (AMH) produced in the developing testis induces the regression of the Müllerian duct, which develops into the oviducts, uterus and upper vagina. In our true hermaphrodite mouse with an ovary on one side and a testis on the other (O/T), the oviduct and uterus are present only on the ovary side, and nothing derived from the Müllerian duct is present on the testis side. Here, we investigate the mechanism underlying the unilateral Müllerian duct regression and the mode of AMH signaling, by performing immunohistology, Western blotting, and organ culture analyses. The histological analysis revealed that during the start of the Müllerian duct regression, the duct in the O/T mice was clearly regressed on the AMH-positive testis side compared to the AMH-negative ovary side. The immunohistochemistry showed a diffuse immunoreaction of AMH in the interstitium surrounding the testis cord and boundary region between the testis and mesonephros, especially in the cranial portion. Western blotting revealed that the amount of AMH in the cranial half of the mesonephros was larger than that in the caudal half. AMH injected into the gonads in organ culture induced the regression of the Müllerian duct via the interstitium of the organ. These results suggest that AMH acts on the Müllerian duct in male mice by exuding into the interstitium surrounding the testis cord and infiltrating through the cranial region from the testis to the mesonephros.
- Published
- 2018
- Full Text
- View/download PDF
9. Peripubertal exposure to the neonicotinoid pesticide dinotefuran affects dopaminergic neurons and causes hyperactivity in male mice.
- Author
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Yoneda N, Takada T, Hirano T, Yanai S, Yamamoto A, Mantani Y, Yokoyama T, Kitagawa H, Tabuchi Y, and Hoshi N
- Subjects
- Animals, Guanidines administration & dosage, Insecticides administration & dosage, Male, Mice, Neonicotinoids administration & dosage, Nitro Compounds administration & dosage, Sexual Maturation, Dopaminergic Neurons drug effects, Guanidines toxicity, Hyperkinesis chemically induced, Insecticides toxicity, Neonicotinoids toxicity, Nitro Compounds toxicity
- Abstract
Although neonicotinoid pesticides are expected to have harmful influence on mammals, there is little animal experimental data to support the effect and mechanisms. Since acetylcholine causes the release of dopamine, neonicotinoids may confer a risk of developmental disorders via a disturbance in the monoamine systems. Male mice were peripubertally administered dinotefuran (DIN) referring to no observed effect level (NOEL) and performed behavioral and immunohistological analyses. In an open field test, the total locomotor activity was increased in a dose-dependent manner. The immunoreactivity of tyrosine hydroxylase in the substantia nigra was increased in DIN-exposed mice. These results suggest that exposure to DIN in peripubertal male mice causes hyperactivity and a disturbance of dopaminergic signaling.
- Published
- 2018
- Full Text
- View/download PDF
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