5-fluorouracil-induced histopathological changes in the central nervous system of rat fetuses.

5-Fluorouracil (5-FU), a thymidylate synthesis inhibitor, has been well known to induce developmental anomalies in the craniofacial tissues and limb buds. Recently it was reported that microencephaly was also induced in rat neonates after 5-Fu-treatement in late phase of pregnancy (Kumar et al., 2006). In this study, pregnant rats were treated with 5-Fu (15, 30 or 50 mg/kg) on day 13 of gestation, and their fetuses were examined for histopathological changes, especially in the fetal central nervous system (CNS) at 12, 24 and 48 hours after treatment (HAT). At 12 HAT, an enhancement of pyknosis of neuronal progenitor cells and subsequent loss of dead cells were detected in the CNS in a dose-dependent manner. The severity of such histopathological changes in the CNS was most prominent in the telencephalon (middle and dorsal layers of the ventricular zone) and spinal cord (dorsal area). Pyknotic cells decreased towards 48 HAT in the brain while they increased towards 48 HAT in the spinal cord. Almost all of the nuclei of pyknotic cells were positively stained by TUNEL method and showed characteristics of apoptotic cells under electron microscopy. Therefore, these pyknotic cells were considered to be apoptotic ones. Enhanced apoptosis and reduced mitosis in neuronal progenitor cells in the telencephalon seem to be responsible for the later induction of microencephaly reported by Kumar et al. (2006).

Electron spin resonance of particulate soot samples from automobiles to help environmental studies.

The application of electron spin resonance (ESR) was studied for diesel soot samples and suspended particulate matter (SPM) from automobile engines. Soot samples or diesel exhaust particles (DEP) were recovered at various points: in the exhaust pipe of a diesel engine, at the dust sampler of a highway tunnel (standard DEP), on the soundproofing wall alongside a heavy traffic road, and on the filters of a dust sampler for SPM. The diesel soot samples apparently showed two ESR spectra: one was a broad spectrum at g=2.1 with a line width of ca. 80-120 mT and the other was a sharp signal of a carbon radical at g=2.003 with a line width of 0.4 mT. Annealing experiments with a DEP sample at 250 degrees C revealed drastic enhancement of the sharp ESR signal, which suggested a thermal process of carbonization of remnant organics. An oximetric study by ESR showed an enhancement of the broad signal in the diesel soot sample as well as in the sharp ESR signal. Therefore, the main part of the broad ESR signal would be attributed to carbon radicals, which form a different configuration, probably closely interacting aggregates. Enhancement of the sharp ESR signal was not observed in the standard DEP sample under vacuum condition, which suggested less adsorption sites on the surface of DEP samples.

Possibility of precipitated CaCO3 with vitamin C as a new dosimetric material.

Precipitated CaCO3 in the presence of vitamin C has been investigated as an ESR dosimeter. gamma-ray irradiation produced a sharp electron spin resonance (ESR) signal with a linewidth (0.015 mT) with high thermal stability. Required microwave power (0.02 mW) and magnetic field modulation (0.02 mT) for ESR measurement were about 250 and 30 times lower than those of DL-alpha-alanine, respectively. The lower detection limit was reduced from approximately 3 Gy for a commercial alanine dosimeter to approximately 20 mGy for gamma-rays for this material.

Loss of mRor1 enhances the heart and skeletal abnormalities in mRor2-deficient mice: redundant and pleiotropic functions of mRor1 and mRor2 receptor tyrosine kinases.

The mammalian Ror family of receptor tyrosine kinases consists of two structurally related proteins, Ror1 and Ror2. We have shown that mRor2-deficient mice exhibit widespread skeletal abnormalities, ventricular septal defects in the heart, and respiratory dysfunction, leading to neonatal lethality (S. Takeuchi, K. Takeda, I. Oishi, M. Nomi, M. Ikeya, K. Itoh, S. Tamura, T. Ueda, T. Hatta, H. Otani, T. Terashima, S. Takada, H. Yamamura, S. Akira, and Y. Minami, Genes Cells 5:71-78, 2000). Here we show that mRor1-deficient mice have no apparent skeletal or cardiac abnormalities, yet they also die soon after birth due to respiratory dysfunction. Interestingly, mRor1/mRor2 double mutant mice show markedly enhanced skeletal abnormalities compared with mRor2 mutant mice. Furthermore, double mutant mice also exhibit defects not observed in mRor2 mutant mice, including a sternal defect, dysplasia of the symphysis of the pubic bone, and complete transposition of the great arteries. These results indicate that mRor1 and mRor2 interact genetically in skeletal and cardiac development.

Expression of vinexin alpha in the dorsal half of the eye and in the cardiac outflow tract and atrioventricular canal.

Vinexin, a recently identified cytoskeletal protein, contains three SH3 domains and plays important roles in regulation of cytoskeletal organization and signal transduction. Using whole-mount in situ hybridization, we showed here that expression of vinexin alpha, the longer vinexin transcript, is strictly regulated, although the shorter transcript, vinexin beta, is expressed almost ubiquitously during embryonic development in mice. Expression of vinexin alpha was limited to within part of the eye and heart in 10.5 dpc embryos. Analysis of cryosections of 10.5 dpc embryos showed that vinexin alpha was expressed in a dorsal half of the retinal pigment epithelium and in the outflow tract and atrioventricular canal of the heart. Furthermore, we also found that vinexin alpha was expressed in the gonad and in a ventral part of the pons of 12.5 dpc embryos. These results indicated that the expression of vinexin alpha is strictly regulated in a temporally and spatially restricted manner.

Expression of the receptor tyrosine kinase genes, Ror1 and Ror2, during mouse development.

In mammals, the Ror-family receptor tyrosine kinases consist of two structurally related proteins, Ror1 and Ror2, characterized by the extracellular Frizzled-like cysteine-rich domain and membrane proximal kringle domains. As an attempt to gain insights into their roles in mouse development, expression patterns of Ror1 and Ror2 during early embryogenesis were examined and compared. Interestingly, at early stages, Ror1 and Ror2 exhibit similar expression patterns in the developing face, including the frontonasal process and pharyngeal arches, which are derived from cephalic neural crest cells. On the other hand, they exhibit different expression patterns in the developing limbs and brain, where the expression of Ror2 was detected broadly compared with that of Ror1. At a later stage, both genes are expressed in a similar fashion in the developing heart and lung, yet in a distinct manner in the brain and eye.

Wnt-3a is required for somite specification along the anteroposterior axis of the mouse embryo and for regulation of cdx-1 expression.

In vertebrates, each vertebra along the anteroposterior axis has a characteristic structure. It has recently been shown that several transcription factors and cell signaling molecules expressed in the primitive streak ectoderm and/or the tailbud play essential roles in establishing the correct anteroposterior specification of vertebrae during mouse development. Here, we report that Wnt-3a mutants exhibit homeotic transformations in the vertebrae along their entire body axis. In addition, reduced expression of cdx-1, the mutation of which results in an anterior transformation, as occurs in Wnt-3a mutants, was observed in the primitive streak and tail bud region of Wnt-3a mutant embryos. These results indicate that Wnt-3a is necessary for correct anteroposterior patterning of vertebra, and that cdx-1 may be one of the mediator genes of Wnt-3a signaling in this process.

Strategy for finding new materials for ESR dosimeters

The right strategy for finding a new ESR dosimetric material sensitive to radiation is to follow the orthodox procedures used in the development of thermoluminescence dosimeters (TLD) and phosphorescence studies. Modern procedures used in materials sciences, such as computer calculation of molecular orbitals (MO), should be employed to estimate the ESR and optical properties of prospective materials. Radiation effects in lithium and magnesium sulfates and metal salts of organic acids, such as lithium and magnesium lactates, have been investigated in search for tissue-equivalent dosimeter with a large G value.

Metal ion-organic compound for high sensitive ESR dosimetry

A systematic study to find a tissue equivalent and high sensitive dosimeter material has been made to stimulate the field of ESR dosimetry. Lithium acetate dihydrate (Li-Ac.2H2O:CH3COOLi.2H2O) and lithium phosphate (Li-phosphate:Li3PO4) were irradiated by gamma-rays to study radicals with (ESR) in addition to magnesium lactate (Mg-lactate (CH3CH(OH)COO)2Mg) doped with nominal pure lithium lactate (Mg(Li)-lactate) and lithium lactate (CH3CH(OH)COOLi) doped with Mg-lactate (Li(Mg)-lactate). A triplet spectrum with intensity ratio of 1:2:1 in Li-Ac.2H2O was ascribed to acetate radical which has g = 2.0031+/-0.0004 and hyperfine splitting of A/gbeta = 2.12+/-0.1 mT. The Li-phosphate spectrum shows splitting due to anisotropic g-factors of g(parallel) = 2.0190+/-0.0005 and g(perpendicular) = 1.9974+/-0.0004. Quartet spectra with the intensity ratio of 1:3:3:1 in Mg(Li)-lactate and Li(Mg)-lactate were ascribed to lactate radicals with g-factors of 2.0032+/-0.0004 and 2.0029+/-0.0004 and the intensity ratio of 1:3:3:1 and A/gbeta = 1.92+/-0.06 and 1.82+/-0.06 mT, respectively. The response to gamma-ray dose and the thermal stability as well as the effect of UV-illumination have been studied. The obtained number of free radicals per 100 eV (G-values) were 0.4+/-0.13, 1.02+/-0.31, 1.35+/-0.35 and 0.78+/-0 for Li-Ac.2H2O, Li-phosphate, Mg(Li)-lactate, and Li(Mg)-lactate, respectively. The lifetimes were estimated from Arrhenius plots to be approximately 2.0+/-0.6, 50.7+/-20 and 10+/-3.5 years for Li-phosphate, Mg(Li)-lactate and Li(Mg)-lactate, respectively. The lifetime for Li-Ac.2H2O cannot be estimated because of the decomposition by heating.

Radiation effects in dry ice: models for a peak on the Arrhenius curve

Dry ice (solid CO2) occurs in the polar caps of Mars, on the surface of Triton, and in places in the outer planets of our solar system. Radicals in gamma-irradiated solid CO2 have been studied by ESR for future applications of ESR dating on outer planets. The annealing curves for CO3- radical (ESR signal at g = 2.0126) can be described neither by the first-order nor the second-order decay kinetics. The peak observed in the Arrhenius plot can result from two parallel first-order kinetic processes. Radicals that provide overlapping signals are CO3- (g1 = 2.0057, g2 = 2.0126, g3 = 2.0161; activation energy E = 0.10 eV; frequency factor v0 = 4 x 10(1) s(-1)) and HO2 (g1 = 2.0040, g2 = 2.0055, g3 = 2.0360), which have E = 0.28 eV and v0 = 7 x 10(5) s(-1)). Hence, HO2 is more thermally stable, and use of HO2 is promising for ESR dating.

Mouse Ror2 receptor tyrosine kinase is required for the heart development and limb formation.

BACKGROUND: A mouse receptor tyrosine kinase (RTK), mRor2, which belongs to the Ror-family of RTKs consisting of at least two structurally related members, is primarily expressed in the heart and nervous system during mouse development. To elucidate the function of mRor2, we generated mice with a mutated mRor2 locus. RESULTS: Mice with a homozygous mutation in mRor2 died just after birth, exhibiting dwarfism, severe cyanosis, and short limbs and tails. Whole-mount in situ hybridization analysis showed that mRor2 was expressed in the branchial arches, heart and limb/tailbuds, in addition to the developing nervous system. The mutants had cardiac septal defects, mainly a ventricular septal defect. In addition, an examination of the skeletal systems revealed that the mutants had shorter limbs, vertebrae and facial structure, with a particular defect in their distal portions, and that almost no calcification was observed in their distal limbs. Histological examination showed abnormalities in the chondrocytes. CONCLUSIONS: Our findings suggest that mRor2 plays essential roles in the development of the heart and in limb/tail formation, in particular cardiac septal formation and ossification of distal portions of limbs and tails.

Wnt signaling from the dorsal neural tube is required for the formation of the medial dermomyotome.

Signals originating from tissues surrounding somites are involved in mediolateral and dorsoventral patterning of somites and in the differentiation of the myotome. Wnt-1 and Wnt-3a, which encode members of the Wnt family of cystein-rich secreted signaling molecules, are coexpressed at the dorsal midline of the developing neural tube, an area adjacent to the dorsomedial portion of the somite. Several lines of evidence indicate that Wnt-1 and Wnt-3a have the ability to induce the development of the medial and dorsal portion of somites, as well as to induce myogenesis. To address whether these Wnt signalings are really essential for the development of somites during normal embryogenesis, we investigated the development of somites in mouse embryos lacking both Wnt-1 and Wnt-3a. Here we demonstrate that the medial compartment of the dermomyotome is not formed and the expression of a lateral dermomyotome marker gene, Sim-1, is expanded more medially in the absence of these Wnt signalings. In addition, the expression of a myogenic gene, Myf-5, is decreased at 9.5 days post coitum whereas the level of expression of a number of myogenic genes in the later stage appeared normal. These results indicate that Wnt-1 and Wnt-3a signalings actually regulate the formation of the medial compartment of the dermomyotome and the early part of myogenesis.

Wnt signalling required for expansion of neural crest and CNS progenitors.

Interactions between cells help to elaborate pattern within the vertebrate central nervous system (CNS). The genes Wnt-1 and Wnt-3a, which encode members of the Wnt family of cysteine-rich secreted signals, are coexpressed at the dorsal midline of the developing neural tube, coincident with dorsal patterning. Each signal is essential for embryonic development, Wnt-1 for midbrain patterning, and Wnt-3a for formation of the paraxial mesoderm, but the absence of a dorsal neural-tube phenotype in each mutant suggests that Wnt signalling may be redundant. Here we demonstrate that in the absence of both Wnt- and Wnt-3a there is a marked deficiency in neural crest derivatives, which originate from the dorsal neural tube, and a pronounced reduction in dorsolateral neural precursors within the neural tube itself. These phenotypes do not seem to result from a disruption in the mechanisms responsible for establishing normal dorsoventral polarity. Rather, our results are consistent with a model in which local Wnt signalling regulates the expansion of dorsal neural precursors. Given the widespread expression of different Wnt genes in discrete areas of the mammalian neural tube, this may represent a general model for the action of Wnt signalling in the developing CNS.

Clinical, morphologic, and biochemical characteristics of Chediak-Higashi syndrome in fifty-six Japanese black cattle.

OBJECTIVE: To characterize Chediak-Higashi syndrome (C-HS) in Japanese Black cattle. ANIMALS: 56 of 200 cattle with a bleeding disorder and giant granules in leukocytes. PROCEDURE: Clinical observation, CBC, hemostatic screening test, platelet aggregometry, electron microscopy, platelet constituent analysis, and ophthalmoscopic examination were done. RESULTS: Affected Japanese Black cattle had increased bleeding tendency and abnormal granules in their leukocytes. Susceptibility to infection was not increased. Cutaneous albinism was evident in 6 new-born calves, but not in most affected cattle. In all affected cattle, the tapetal fundus was pale and the nontapetal fundus was almost devoid of pigment. By electron microscopy, a remarkable decrease in the number of dense granules in platelets was observed. Functionally, collagen-induced platelet aggregation was markedly reduced. CONCLUSIONS: This bleeding disorder was diagnosed as C-HS. With regard to susceptibility to infection, albinism, and mortality, clinical manifestations of C-HS in Japanese Black cattle were moderate, compared with C-HS in human beings and Hereford cattle. CLINICAL RELEVANCE: Because an autosomal recessive mode of inheritance was documented and recessive homozygotes could be easily detected, C-HS in Japanese Black cattle can be controlled.

[Thoracoscopic cutting needle biopsy].

We presented a new method of percutaneous cutting needle biopsy under the thoracoscope. Percutaneous cutting needles for lung tumor have been abandoned because the risk of hemorrhage, death, and pneumothorax is much greater with these instruments than the computed tomographic guided needle aspiration. On the other hand, thoracoscopic stapled lung wedge resection is easy to obtain a good sample for any lung disease, but should be needed two or three staples, and waste a time, in the case following lobectomy was needed for malignancy. This thoracoscopic cutting needle biopsy was easy to obtain a specimen for the frozened section diagnosis. Under the view of thoracoscope, percutaneous cutting needle biopsy is safety, saving the cost for staples and easy to control the bleeding from the lung. The indication of this thoracoscopic cutting needle biopsy is the case that could not obtain the precise diagnosis by the preoperative biopsy and imaging diagnosis might be suspicious a malignant tumor.

ESR dosimetry of a deceased radiation worker.

A case of overexposure of an industrial radiographer using 192Ir sources and having a filmbadge dosimeter record of 104 mSv has been examined with ESR dosimetry of postmortem tooth and bone specimens. ESR measurements of the tooth enamel showed an intense signal of CO2- and gave the equivalent dose (ED) of 14 Gy by the additive dose method using gamma-rays from a source of 60Co. The doses for a finger bone and humerus were 14.7 and 7.0 Gy, respectively. It was concluded that he had been exposed to radiation repeatedly over 10 yr and that ESR dosimetry can give a life-long cumulative dose for personnel using radiation.

Death of a classified worker probably caused by overexposure to gamma radiation.

This paper describes the case of an industrial radiographer who was seriously overexposed to gamma radiation. The exact circumstances of this exposure were not established but it was concluded that he was repeatedly irradiated probably to a total average whole body dose of at least 10 Gy over several years. Also, a much larger dose to a hand required its partial amputation. He developed myelodysplasia, which progressed to acute myeloid leukaemia from which he died. Karyotypic examination of the leukaemic blasts showed changes very similar to those associated with secondary leukaemia that may develop after radio or chemotherapy. The paper describes his medical case history, the investigation of his workplace, and the attempts to estimate his radiation dose by chromosomal analysis of blood lymphocytes and electron spin resonance of dental enamel and bone.

Applications of microwave scanning ESR microscope: human tooth with metal.

A microwave scanning ESR microscope has been developed to study the distribution of paramagnetic ions or radicals in various materials, especially in minerals. The microwave scanning microscope consists of a cavity with an aperture over which a sample is moved using mechanical stages and computer-controlled stepping motors. The EPR signal intensities at all positions are composed on a CRT display as an image. A new TE111-mode cylindrical cavity system was designed to obtain better sensitivity and resolution. ESR images of gamma-rayed human teeth were obtained to study the effect of secondary electrons from the adjacent metal on the defect formation in a tooth.