Biomedical subjects

Kohei Shiota

Publications and source records attributed to Kohei Shiota.

31 records · Page 2Linked to original sources

PubMed2004

Organ culture of the fetal mouse palate for screening the developmental toxicity of chemicals: a validation study.

Using in vitro organ culture of the fetal mouse palate in a chemically defined serumless medium, the toxicity of 24 chemical compounds was investigated. Explanted palates of day-12.5 mouse fetuses were exposed for 72 h in vitro to various concentrations of each chemical, and the fusion rate and growth parameters were compared between the experimental group and respective controls. The average rate of palate fusion was 84% in vehicle controls. For compounds that are teratogenic in experimental animals in vivo, the fusion rates of palatal shelves decreased as the concentration of the test chemicals increased, showing a dose-dependent relationship. Palate fusion was inhibited by 11 of the 15 in vivo teratogens, and the predictability of in vivo developmental toxicity in this culture system was 73%. Cyclophosphamide itself did not inhibit the growth and fusion of explanted palates, but supplementation of hepatic S-9 fraction and cofactors for a monooxygenase system converted it to a toxic substance, as was shown in other in vitro systems. The 50% inhibitory concentration (IC50) value calculated based on the fusion rate was also found to be a useful parameter for evaluating the developmental toxicity of drugs. The teratogenic risk in the human fetus could be assessed by comparing the minimal toxic concentrations of the test compound on cultured palates with the maximal plasma level in pregnant women under therapeutic conditions and with the plasma concentrations when its minimal teratogenic dose is given to pregnant mice. This organ culture system of the fetal palate should be useful for screening the developmental toxicity of drugs and other environmental agents, and its value should increase when it is used in combination with other battery test systems.

Animals↗

PubMed2004

Terminal differentiation of palatal medial edge epithelial cells in vitro is not necessarily dependent on palatal shelf contact and midline epithelial seam formation.

During fusion of the mammalian secondary palate, it has been suggested that palatal medial edge epithelial (MEE) cells disappear by means of apoptosis, epithelial-mesenchymal transformation (EMT) and epithelial cell migration. However, it is widely believed that MEE cells never differentiate unless palatal shelves make contact and the midline epithelial seam is formed. In order to clarify the potential of MEE cells to differentiate, we cultured single (unpaired) palatal shelves of ICR mouse fetuses by using suspension and static culture methods with two kinds of gas-mixtures. We thereby found that MEE cells can disappear throughout the medial edge even without contact and adhesion to the opposing MEE in suspension culture with 95% O2/5% CO2. Careful examination of MEE cell behavior in the culture revealed that apoptosis, EMT, and epithelial cell migration all occurred at various stages of MEE cell disappearance, including the transient formation and disappearance of epithelial triangles and islets. In contrast, MEE cells showed poor differentiation in static culture in a CO2 incubator. Furthermore, mouse and human amniotic fluids were found to prevent MEE cell differentiation in the cultured single palatal shelf, although paired palatal shelves fused successfully even in the presence of amniotic fluid. We therefore conclude that terminal differentiation of MEE cells is not necessarily dependent on palatal shelf contact and midline epithelial seam formation, but such MEE cell differentiation appears to be prevented in utero by amniotic fluid unless palatal shelves make close contact and the midline epithelial seam is formed.

Amniotic Fluid↗

PubMed2004

Programmed cell death is not a necessary prerequisite for fusion of the fetal mouse palate.

It has been widely accepted that programmed cell death (PCD) is an essential event in palatogenesis and that its failure can result in cleft palate, one of the most common birth defects in the human. However, some conflicting results have been reported concerning the timing of cell death occurring in the fusing palate and therefore the role of PCD in palatal fusion is controversial. In order to clarify whether cell death is indispensable for mammalian palatogenesis, we cultivated the palates of day-13 mouse fetuses in vitro and prevented cell death by treating them with the inhibitors of caspases-1 and -3 or with aurintricarboxylic acid which inhibits the activity of caspase-activated DNase. Even when cell death was almost completely inhibited, palatal fusion took place successfully. Histological examination revealed that in the absence of apoptotic cell death, the medial edge epithelia of opposing palatal shelves adhered to each other and subsequently, the midline epithelial seam was disrupted and disappeared to bring about mesenchymal confluence across the palate. It seems that cell death is not a necessary prerequisite for palatal fusion but it may help to efficiently eliminate unnecessary cells which failed to migrate or differentiate properly.

Animals↗

PubMed2003

Spatial and temporal expression of folate-binding protein 1 (Fbp1) is closely associated with anterior neural tube closure in mice.

Periconceptional folate supplementation is widely believed to have significant preventive effects on the production of neural tube defects. Folate-binding protein 1 (FBP1) is one of the membrane proteins that mediate cellular uptake of folate. Although recent studies suggest that Fbp1 is essential for neural tube closure, the pattern of Fbp1 expression in embryonic tissues has not been examined in detail. To elucidate how Fbp1 contributes to neural tube closure, we examined the spatial and temporal expression patterns of Fbp1 in the developing neural folds and tube of mouse embryos by in situ hybridization. Fbp1 showed a distinct expression pattern in the neural folds, which preceded initiation of neural tube closure at the cervical region and the prosencephalic/mesencephalic boundary. Fbp1 expression was mainly localized to the most dorsal regions of the neural folds where fusion was to occur. With proceeding of neural fold fusion, Fbp1 expression extended to the adjacent unfused neural folds. In the rhombencephalon, robust expression of Fbp1 was observed in rhombomere2 (r2) and r6, suggesting its roles in development of neural crest cells. Fbp1 also showed intense expression in the yolk sac, indicating that FBP1 may mediate transferring maternal folate to embryos during neurulation. These findings indicate close association between Fbp1 and anterior neural tube closure.

Animals↗

PubMed2003

Stressor-like effects of cocaine on heat shock protein and stress-activated protein kinase expression in the rat hippocampus: interaction with ethanol and anti-toxicity drugs.

The present study examined the stressor-like effects of repeated (4 days) administration of cocaine hydrochloride(COC) (35 mg/kg, i.p.) on the expression of heat shock proteins (HSPs) (HSP27, HSP60, HSP70, HSC70) and stress-activated protein kinases (SAPKs) (SAPKalpha, SAPKbeta, SAPKgamma) in the rat hippocampus. The interactions with intraperitoneal ethanol and drugs known as antidotes against COC toxicity were also examined. Similar to the effects of a 10 min immobilization stress (IM) over 4 days, an early increase (5 h time point) in nerve cells immunoreactive for HSPs (HSP27, HSP60, HSP70, HSC70) and SAPKs (SAPKbeta, SAPKgamma) was observed in the COC group. At the 24 h time point, a recovery was observed only for SAPKs, which have been suggested to control the HSP levels. Before the 48 h time point, alterations in the number of HSP+cells as compared to the control group (increase for HSP27 and HSP70+cells, and attenuation for HSP60 and HSC70+cells) could still be observed. Stress-related, attenuated swimming behaviors in the forced swimming test were also the most severe at the 5 h time point. Ethanol (1.5 g/kg) cotreatment on each administration day, even at non-toxic and/or euphoric doses, enhanced these stressor-like alterations. On the other hand, the protective effects of daily coadministered drugs related to benzodiazepine (5 mg/kg Ro 15-4513), dopamine (0.5 mg/kg SCH 23390), muscarinic (0.25 mg/kg pirenzepine) and serotonin (5 mg/kg ketanserin) receptors could be observed on the number of HSP-immunoreactive (24 h) and SAPK-immunoreactive cells (5 h). Against the stressor-altered swimming behaviors, Ro 15-4513 and SCH 23390 were more effective as compared to pirenzepine and ketanserin.

Animals↗

PubMed2003

A quantitative study of the facial nerve in mice prenatally exposed to ethanol.

Pregnant ICR mice were given 20% ethanol intraperitoneally twice on day 13 of gestation and allowed to give birth to offspring. The offspring were killed at 56 days of age and the motor root of their facial nerve was examined histologically and morphometrically. The cross-sectional area of the facial nerve of mice prenatally exposed to ethanol was significantly smaller than that of the control mice. There was no significant difference in the total number of myelinated axons or the mean axonal diameter between control and ethanol-exposed mice, but the mean diameter of myelinated fibers (axon + myelin sheath) and the thickness of myelin sheath were significantly decreased in the treated group. These results suggest that prenatal exposure to ethanol disturbs myelination of the motor root of the facial nerve and may cause permanent neurological effects.

Animals↗

PubMed2002

Teratogenesis of holoprosencephaly.

Teratogenic causes of holoprosencephaly are critically assessed. A brief general review of holoprosencephaly is followed by four tables summarizing etiologic factors. Subjects evaluated here include: 1) maternal diabetes; 2) ethyl alcohol; 3) retinoic acid; 4) mutated genes and teratogens involving the sonic hedgehog signaling network and cholesterol biosynthesis; and 5) cholesterol trafficking, sterol adducts, target tissue response, and sterol sensing domain.

Abnormalities, Drug-Induced↗

PubMed2002

Methamphetamine induces apoptosis in seminiferous tubules in male mice testis.

We investigated whether methamphetamine (MAMP) induces apoptosis in seminiferous tubules in 10-week-old male ICR mice. Methamphetamine was dissolved in saline and injected ip at four doses (1, 5, 10, and 15 mg/kg). TUNEL-positive cells were detected in the seminiferous tubules in animals 24 h after a single treatment with 5, 10, or 15 mg/kg MAMP. The percentage of seminiferous tubules containing more than three TUNEL-positive cells (apoptotic tubules) was considered a reliable indicator for apoptotic changes. After a single treatment with 10 mg/kg MAMP, the percentage of apoptotic tubules increased with time, and it became significant at 24 h, at which time vacuolar changes in spermatogonia also peaked. Although the percentage of apoptotic tubules increased dose dependently from 5 to 15 mg/kg MAMP, the DNA ladder was detected in the testis of 15 mg/kg MAMP-treated mice. Thus, we have demonstrated that MAMP induces apoptosis in seminiferous tubules in male mice testis.

Animals↗

PubMed2002

Depletion of FGF acts as a lateral inhibitory factor in lung branching morphogenesis in vitro.

Previous studies have shown that the interaction of positive and inhibitory signals plays a crucial role during lung branching morphogenesis. We found that in mesenchyme-free conditions, the lung epithelium exerted a lateral inhibitory effect on the neighbouring epithelium via depletion of fibroblast growth factor 1 (FGF1). Contrary to previous suggestions, bone morphogenetic protein 4 could not substitute for the inhibitory effect. Based on of this observation, we used a reaction-diffusion model of the substrate-depletion type to represent the initial phase of in vitro branching morphogenesis of lung epithelium, with depletion of FGF playing the role of lateral inhibitor. The model was able to account for the effects of the FGF1 concentration, extracellular matrix degradation and different subtypes of FGF on morphogenesis of the lung bud epithelia. These results suggest that the depletion of FGF may be a key regulatory component in initial phase of branching morphogenesis of the lung bud epithelium in vitro.

Animals↗

PubMed2002

Comparative investigations of human and rat dermatoglyphics: palmar, plantar and digital pads and flexion creases.

The morphological features of the palmar, plantar and digital areas of the rat were studied and compared to the corresponding human traits. The location and the timing of appearance of the volar pads and flexion creases of human and rat fetuses were investigated to determine the feasibility of using rats as an experimental model for studying the factors influencing early development of the dermatoglyphics in humans. Comparisons between analogous developmental stages of human and rat fetuses demonstrate striking similarities in overall fetal development. However, marked differences between human and rat fetuses were found in the timing of developmental milestones and in some morphological features. Provided that these differences are taken into consideration, rats can serve as a useful experimental model in studies of the utility of the epidermal ridge configurations and flexion creases in medical disorders.

Animals↗

PubMed2002

Thioredoxin, an anti-oxidant protein, protects mouse embryos from oxidative stress-induced developmental anomalies.

During the early postimplantation period, rodent embryos survive in a relatively anaerobic environment in utero and are vulnerable to a high oxygen pressure. They become resistant to oxygen stress when they are exposed to a higher oxygen pressure after the uteroplacental circulation is established. However, it is unknown how embryos acquire such resistance against oxidative stress. This study was undertaken to examine whether an antioxidant protein thioredoxin (TRX) plays a significant role in the embryonic acquisition of the tolerance to oxidative stress. E7.5 embryos of C57BL/6 wild-type (WT) and human TRX (hTRX) inserted-transgenic (Tg) embryos were cultured under 10 or 25% O2 and their growth and morphological differentiation were evaluated. The TRX expression and the products of oxidative stress (8-hydroxy-2'-deoxy-guanosine and carbonylated proteins) in their tissues were also examined. When WT embryos were cultivated in vitro under 25% O2, their growth was significantly disturbed and various developmental abnormalities were induced, which did not occur in embryos grown under 10% O2. However, such embryotoxic effects of oxygen were significantly attenuated in the hTRX Tg embryos that continuously express hTRX. Accumulation of the products of oxidative stress was significantly reduced in hTRX Tg embryos as compared with that in WT embryos. The TRX transgene appears to provide the embryo with the resistance against oxidative stress and may play a crucial role in the redox regulation in embryos.

8-Hydroxy-2'-Deoxyguanosine↗

PubMed2001

Embryonic hydromyelia: cystic dilatation of the lumbosacral neural tube in human embryos.

In a large collection of human embryos (the Kyoto Collection of Human Embryos, Kyoto University), we encountered five cases with abnormal dilatation of the neural tube at the lumbosacral level. In these examples, the central canal was enlarged, and the roof plate of the neural tube was extremely thin and expanded. The mesenchymal tissue was scarce or lacking between the roof plate and the surface ectoderm. This type of anomaly was assumed to be formed after neural tube closure and may be an early form of spina bifida. In two of the cases, some abnormal cells were found ectopically between the thin roof plate and the surface ectoderm. Morphologically, these cells resembled those forming spinal ganglia and could be of the neural crest origin. Since neural crest cells are pluripotent and can differentiate into a variety of tissues, such ectopic cells might undergo abnormal differentiation into teratomatous tumors and/or lipomas, which are frequently associated with spina bifida. We also discuss the definition of spina bifida and the classification of neural tube defects from the embryological and pathogenic viewpoints and propose a new classification of neural tube defects.

Dilatation, Pathologic↗

PubMedDate not supplied

Microarray profile analysis of toxic cocaine-induced alterations in the expression of mouse brain gene sequences: a possible 'protective' effect of buprenorphine.

Using a mouse brain cDNA microarray consisting of 2688 gene sequences, which include unknown sequences with the empirical possibility of expression in the brain, the effects of repeated toxic doses of intraperitoneal (i.p.) cocaine (40 mg x kg(-1), 4 days) on the expression pro fi le of the cerebral genes were investigated. The modifications in this pro file caused by buprenorphine (BUP) (0.25 mg x kg(-1) i.p., 4 days), a protective drug against cocaine, were also examined. In the cocaine group, the expression levels reached the recommended increased levels (>or=2 times the control value in the saline-treated control group) in 24.0% of the genes but were equal to or less than the recommended attenuation levels ( 0.5 and <2 times the control value). Although statistically significant modifications in the expression of cocaine- or BUP-related brain-region-specific genes were not proved using whole cerebrums, including many unknown genes, our results suggest that the expression of genes related to neuronal cell damage, including non-peculiar genes related to the damage accompanying convulsive seizures and malignant tumors, were normalized and the genes related to the protection of neural cells were induced by BUP.

Animals↗