MR Imaging of the Pituitary Gland and Postsphenoid Ossification in Fetal Specimens.

BACKGROUND AND PURPOSE: A thorough knowledge of fetal growth and development is key to understanding both the normal and abnormal fetal MR imaging findings. We investigated the size and signal intensity of the normal pituitary gland and the intrasphenoidal ossification around the Rathke pouch in formalin-fixed fetuses on MR imaging. MATERIALS AND METHODS: Thirty-two fetuses with undamaged brains were included in this study (mean age, 19.93 weeks; age range, 12-31 weeks). Visual inspection of the pituitary and ossification around the Rathke pouch in the sphenoid bone or the postsphenoid ossification was conducted. The extent of pituitary and postsphenoid ossification, pituitary/pons signal ratio, and postsphenoidal ossification/sphenoid bone signal ratio was compared according to gestational age. RESULTS: The pituitary gland was identified as a hyperintense intrasellar structure in all cases, and postsphenoid ossification was identified as an intrasphenoidal hyperintense area in 27 of the 32 cases (84%). The mean pituitary/pons signal ratio was 1.13 ± 0.18 and correlated weakly with gestational age (R(2) = 0.243), while the mean postsphenoid ossification/sphenoid bone signal ratio was 2.14 ± 0.56 and did not show any increase with gestational age (R(2) = 0.05). No apparent change in the size of pituitary hyperintensity was seen with gestational age (R(2) = 0.001). Postsphenoid ossification showed an increase in size with gestational age (R(2) = 0.307). CONCLUSIONS: The fetal pituitary gland was hyperintense on T1-weighted images and the pituitary/pons ratio and extent of postsphenoid ossification correlated weakly with gestational age.

Three-dimensional morphology of the human embryonic brain.

The morphogenesis of the cerebral vesicles and ventricles was visualized in 3D movies using images derived from human embryo specimens between Carnegie stage 13 and 23 from the Kyoto Collection. These images were acquired with a magnetic resonance microscope equipped with a 2.35-T superconducting magnet. Three-dimensional images using the same scale demonstrated brain development and growth effectively. The non-uniform thickness of the brain tissue, which may indicate brain differentiation, was visualized with thickness-based surface color mapping. A closer view was obtained of the unique and complicated differentiation of the rhombencephalon, especially with regard to the internal view and thickening of the brain tissue. The present data contribute to a better understanding of brain and cerebral ventricle development.

Morphology and morphometry of the human embryonic brain: A three-dimensional analysis.

The three-dimensional dynamics and morphology of the human embryonic brain have not been previously analyzed using modern imaging techniques. The morphogenesis of the cerebral vesicles and ventricles was analyzed using images derived from human embryo specimens from the Kyoto Collection, which were acquired with a magnetic resonance microscope equipped with a 2.35-T superconducting magnet. A total of 101 embryos between Carnegie stages (CS) 13 and 23, without apparent morphological damage or torsion in the brain ventricles and axes, were studied. To estimate the uneven development of the cerebral vesicles, the volumes of the whole embryo and brain, prosencephalon, mesencephalon, and rhombencephalon with their respective ventricles were measured using image analyzing Amira™ software. The brain volume, excluding the ventricles (brain tissue), was 1.15 ± 0.43 mm(3) (mean ± SD) at CS13 and increased exponentially to 189.10 ± 36.91 mm(3) at CS23, a 164.4-fold increase, which is consistent with the observed morphological changes. The mean volume of the prosencephalon was 0.26 ± 0.15 mm(3) at CS13. The volume increased exponentially until CS23, when it reached 110.99 ± 27.58 mm(3). The mean volumes of the mesencephalon and rhombencephalon were 0.20 ± 0.07 mm(3) and 0.69 ± 0.23 mm(3) at CS13, respectively; the volumes reached 21.86 ± 3.30 mm(3) and 56.45 ± 7.64 mm(3) at CS23, respectively. The ratio of the cerebellum to the rhombencephalon was approximately 7.2% at CS20, and increased to 12.8% at CS23. The ratio of the volume of the cerebral vesicles to that of the whole embryo remained nearly constant between CS15 and CS23 (11.6-15.5%). The non-uniform thickness of the brain tissue during development, which may indicate the differentiation of the brain, was visualized with surface color mapping by thickness. At CS23, the basal regions of the prosencephalon and rhombencephalon were thicker than the corresponding dorsal regions. The brain was further studied by the serial digital subtraction of layers of tissue from both the external and internal surfaces to visualize the core region (COR) of the thickening brain tissue. The COR, associated with the development of nuclei, became apparent after CS16; this was particularly visible in the prosencephalon. The anatomical positions of the COR were mostly consistent with the formation of the basal ganglia, thalamus, and pyramidal tract. This was confirmed through comparisons with serial histological sections of the human embryonic brain. The approach used in this study may be suitable as a convenient alternative method for estimating the development and differentiation of the neural ganglia and tracts. These findings contribute to a better understanding of brain and cerebral ventricle development.

Three-dimensional reconstruction of rat knee joint using episcopic fluorescence image capture.

OBJECTIVE: Development of the knee joint was morphologically investigated, and the process of cavitation was analyzed by using episcopic fluorescence image capture (EFIC) to create spatial and temporal three-dimensional (3D) reconstructions. METHODS: Knee joints of Wister rat embryos between embryonic day (E)14 and E20 were investigated. Samples were sectioned and visualized using an EFIC. Then, two-dimensional image stacks were reconstructed using OsiriX software, and 3D reconstructions were generated using Amira software. RESULTS: Cavitations of the knee joint were constructed from five divided portions. Cavity formation initiated at multiple sites at E17; among them, the femoropatellar cavity (FPC) was the first. Cavitations of the medial side preceded those of the lateral side. Each cavity connected at E20 when cavitations around the anterior cruciate ligament (ACL) and posterior cruciate ligament (PCL) were completed. CONCLUSION: Cavity formation initiated from six portions. In each portion, development proceeded asymmetrically. These results concerning anatomical development of the knee joint using EFIC contribute to a better understanding of the structural feature of the knee joint.

Morphogenesis and three-dimensional movement of the stomach during the human embryonic period.

The stomach develops as the local widening of the foregut after Carnegie stage (CS) 13 that moves in a dramatic and dynamic manner during the embryonic period. Using the magnetic resonance images of 377 human embryos, we present the morphology, morphometry, and three-dimensional movement of the stomach during CS16 and CS23. The stomach morphology revealed stage-specific features. The angular incisura and the cardia were formed at CS18. The change in the angular incisura angle was approximately 90° during CS19 and CS20, and was <90° after CS 21. The prominent formations of the fundus and the pylorus differentiate at around CS20. Morphometry of the stomach revealed that the stomach gradually becomes "deflected" during development. The stomach may appear to move to the left laterally and caudally due to its deflection and differential growth. The track of the reference points in the stomach may reflect the visual three-dimensional movement. The movement of point M, representing the movement of the greater curvature, was different from that of points C (cardia) and P (pyloric antrum). The P and C were located just around the midsagittal plane in all the stages observed. Point M moved in the caudal-left lateral direction until CS22. Moreover, the vector CP does not rotate around the dorsoventral axis, as widely believed, but around the transverse axis. The plane CPM rotated mainly around the longitudinal axis. The data obtained will be useful for prenatal diagnosis in the near future.

The layered structure of the articular surface.

OBJECTIVE: Articular cartilage is roughly separated into three areas: the tangential, middle, and deep zones. The structure and molecular components of an additional important zone, the most superficial zone (MSZ), which directly faces the joint cavity, have yet to be conclusively elucidated. The purpose of the present study was to use multiple methods to study the MSZ in order to determine its structure. MATERIALS AND METHODS: Knees from 16 pigs (age, 6 months) were used. Full-thickness cartilage specimens were harvested from the femoral groove. The MSZ was observed using light microscopy, transmission electron microscopy (TEM), and scanning electron microscopy (SEM) in combination with histochemical and immunohistochemical methods. RESULTS: The combined findings from the three different observational methods indicate that the MSZ is subdivided into three layers. Among these three layers, collagen subtypes I, II, and III are present in the innermost (third) layer of the MSZ. Beneath the third layer, type II collagen is the predominant type, with small amounts of type III collagen. This layer beneath the third layer is considered to be the tangential layer. CONCLUSIONS: Our observations indicate that the MSZ is subdivided into three layers. Further analysis of the molecular components in each layer may improve our understanding of the structure of the articular surface.

Cell origin of pyothorax-associated lymphoma: a lymphoma strongly associated with Epstein-Barr virus infection.

Pyothorax-associated lymphoma (PAL) is an Epstein-Barr virus (EBV)-associated B cell lymphoma developing in the pleural cavity affected by chronic pyothorax. To clarify the cell origin of PAL, the expression of immunoglobulin heavy (IgH) and light chains in relation to somatic hypermutations (SHMs) of rearranged Ig heavy- and light-chain variable (IgV(H), IgV(L)) genes was examined using cell lines as well as clinical samples. SHMs without ongoing mutations of the IgV(H) gene were found in all PAL cell lines and clinical samples available for sequencing, indicating PAL to be derived from B cells at the postgerminal center (GC) stage of the differentiation process. They could be subdivided into post-GC cells with potentially productive IgV(H) genotypes (Group 1) and with sterile IgV(H) genotypes (Group 2). IgH expression was abrogated in Group 2 as expected and also in two cell lines in Group 1. DNA demethylation experiments with 5-aza-dC induced expression of IgH mRNA and protein in these cell lines. Most PAL cells were derived from crippled post-GC cells, which usually could not survive. Transformation of such B cells through EBV infection might provide a basis for the development of PAL with additional genetic changes.

Toxicity assessment of treated wastewater using cultured human cell lines.

Bioassay using cultured human cell lines was applied to an effluent of a wastewater treatment plant (WWTP) in Sapporo to assess their toxicity, and in order to investigate the fate of toxicity in the WWTP, bioassay of the water samples from several points in WWTP (influent, effluent, return flow from thickener, from dewatering process and from incineration process) was performed. We also applied bioassay to the mixture of the activated sludge from the investigated plant and artificial sewage. These results showed that the toxicity of the effluent was more intensive than the influent, and organic matter released from activated sludge bacteria during their decay process contributed to the increase of toxicity in the effluent.

Epigenetic regulation of the expression of the novel stem cell marker CDCP1 in cancer cells.

CDCP1 is a novel stem cell marker that is expressed in several types of cancer. The mechanisms by which CDCP1 expression is regulated, and the clinical implications of this marker, have not been clarified. In this report, we examine the epigenetic regulation of CDCP1 expression in cell lines and clinical samples from patients with breast cancer. Many CpG sequences were localized around the transcription initiation site of CDCP1. These CpG motifs were found to be poorly methylated in cell lines with high levels of CDCP1 expression and heavily methylated in cell lines with low levels of CDCP1 expression. The in vitro methylation of CpG sites decreased CDCP1 promoter activity, and the addition of a demethylating reagent restored activity. In 25 breast cancer samples, an inverse correlation was noted between the CDCP1 expression level and the proportion of methylated to non-methylated CpG sites. Tumours with high-level CDCP1 expression showed higher levels of proliferation, as revealed by immunohistochemical detection of the MIB-1 antigen, than tumours with low-level CDCP1 expression. These findings indicate that the expression of CDCP1 is regulated by methylation of its promoter region in tumours. CDCP1 expression may prove to be useful in the further characterization of cancers.

Toxicity assessment of the extract of compost as a final product from Bio-Toilet.

Bio-Toilet is the name of a dry closet or composting toilet using sawdust as an artificial soil matrix for bioconversion of human excrement into compost. Since feces and urine contain several chemicals such as pharmaceutical residues and endocrine disruptors and they may still remain in compost after biological reaction in the Bio-Toilet, it is required to examine the possibility of soil and/or groundwater pollution by applying compost to a soil system in farmland. In this study, toxicity of Bio-Toilet compost was evaluated by measuring the viability of human neuroblast (NB-1). The bio-assay was applied to the water extract of compost from the Bio-Toilets which are in practical use in Japan. The assay results showed that (1) the extract of feces showed no toxicity, and the extracts of unused sawdust had no or low level toxicity and (2) the extracts of composts had heavier toxicity than unused sawdust. These results implied that some chemicals that have toxicity were generated by biological reactions or accumulated in toilet system. The bioassay results with fractionated organic matter by its molecular weight showed that the small molecular weight fraction had stronger toxicity than other fractions. The effect of inorganic matter on toxicity was examined by comparing the dose-response relationship of the extracts of compost with positive control with 1M of sodium chloride solution. The comparison showed that sodium concentration in the extract was too low to develop the toxicity and the effect of inorganic matter could be neglected in this study.

Role of hydrophilic organic matter on developing toxicity in decay process of activated sludge.

It is known that the toxicity of effluent is more intensive than that of influent in the activated sludge process. In this study, we applied bioassay using cultured human cell lines to the decay process of activated sludge to evaluate the toxicity of organic matter generated and/or released from activated sludge bacteria. We also applied this bioassay to hydrophilic fraction of samples. The bioassay results showed that: (1) the variation in the dose-response relation obtained from assay with original samples was observed during decay; (2) on the other hand, the response curves of only hydrophilic fraction at each time show the same relationship between TOC and viability of MCF7 cells; (3) this trend was confirmed by plotting the time course of EC50. These results imply that: (1) the hydrophilic organic matter controlled for developing toxicity during decay process of activated sludge; and (2) the character of hydrophilic organic matter is not changed during the experimental period.

Organic matter released from activated sludge bacteria cells during their decay process.

Organic matter released from activated sludge bacteria is a considerable issue in the wastewater reclamation process. In this study, we focused 2-keto-3-deoxyoctulosonic acid in the Lipopolysaccharide existed in the gram-negative bacterial cell wall as an index of organic matter released from bacteria, and investigated the fate of 2-keto-3-deoxyoctulosonic acid in the aerated and ultrasonicated activated sludge samples. The results shows 1) 2-keto-3-deoxyoctulosonic acid concentration in the hydrolyzed sample was higher than non-hydrolyzed sample, and this implied that 2-keto-3-deoxyoctulosonic acid existed in the water phase as a monomer and also as a polymer such as Lipopolysaccharide form and their fragments; 2) the value of (2-keto-3-deoxyoctulosonic acid)/(dissolved organic carbon) ratio did not change in the sludge sonication process and was approximately 0.0006, on the other hand, in the bacteria decay process, the ratio varied from zero to approximately 0.0012; 3) the linear relationship was observed between the degraded heterotrophic biomass and the generated 2-keto-3-deoxyoctulosonic acid; and 4) 2-keto-3-deoxyoctulosonic acid might be considered as an index of organic matter originated from activated sludge bacteria cell.

Biological activity in the composting reactor of the bio-toilet system.

The bio-toilet is becoming commercially available and it is actually used in Japan in public parks, sightseeing areas, and households; however, the biological activity in the system during degradation of toilet wastes, particularly faeces, is unknown. Thus, in this study activity of microorganisms in the bio-toilet system during degradation of faeces was assessed through the quantification of reductions in total solids (TS), volatile solids (VS), and chemical oxygen demand (COD) during batch tests in laboratory-scale composting reactors. Additionally, the fate of nitrogen and its transformation processes in such reactors were evaluated. TS, VS, and COD reductions were on the order of 56%, 70%, and 75%, respectively, irrespective of the organic loading regarded. Total nitrogen (T-N) reductions quantified 94%, regardless of the organic loading. Furthermore, all T-N reductions observed during composting were equivalent to the NH(3)-N released from the reactor, i.e., 94% of ammonia was lost.

Mathematical model for describing reactions of residual chlorine with organic matter in reclaimed wastewater.

Among several applications of urban wastewater reuse, use of reclaimed wastewater to sustain stream flows has become attractive in the urban area. Since these rivers are used for recreational purposes and for restoring aquatic eco-system, the adequate control of residual chlorine is essential. Mathematical model for describing reactions between residual chlorine and organic matter in reclaimed wastewater has been developed. The model considers the effect of molecular weight distribution of organic matter on the reaction rate. Lab-scale experiments were performed to estimate reaction rates constants and to examine their temperature dependency. The experiments showed that 1) the smaller organic matter gave the larger reaction rate; 2) temperature effect on reaction rate was described by the Arrhenius formula; 3) decline of free chlorine had more temperature dependency than combined chlorine. The comparison of computed results with data from lab-scale experiments confirmed the validity of the model. We used the one-dimensional dispersion model with proposed reaction model and examined the seasonal variation of residual chlorine profile along the river sustained by reclaimed wastewater in Sapporo. Simulation showed that seasonal variation of nitrification performance in secondary treatment as well as change in temperature caused seasonal variation in residual chlorine profile along the river.

Onsite wastewater differential treatment system: modeling approach.

In this paper, the Onsite Wastewater Differential Treatment System (OWDTS), a new approach for improving the traditional onsite wastewater treatment systems (OWTS), is proposed based on differential management and treatment of household wastewater effluents. Three fractions of household wastewater have been differentiated, reduced-volume blackwater, higher-load graywater and lower-load graywater. Based on this differentiation, different treatment processes required for each fraction are discussed. The procedure adopted for treatment of toilet wastes (reduced-volume blackwater) is shown. In the case of graywater, a sketch of treatment processes is provided. The OWDTS seems to be a new approach with higher potential for improvement of traditional OWTS, dry ecological sanitation, recycling of resources (toilet wastes and water), conservation of water resources, etc. Aerobic biodegradation of toilet wastes by using sawdust as a matrix is an essential treatment process of the OWDTS. Membrane technology seems to be the most effective process to treat higher-load gray water. Natural biodegradation of lower-load gray water by soil bacteria needs to be deeply studied.

DNA sequence of immunoglobulin heavy chain variable region gene in thyroid lymphoma.

Patho-epidemiological studies have shown that thyroid lymphoma (TL) develops in thyroid affected by chronic lymphocytic thyroiditis (CLTH). CLTH is categorized as an organ-specific autoimmune disease, in which activated B-lymphocytes secrete a number of autoantibodies. Because antigenic stimulation might be involved in the pathogenesis of TL, the variable region in heavy chain (V(H)) genes was characterized in 13 cases with TL and 3 with CLTH. Clonal rearrangement of the V(H) gene was found in 11 cases of TL, and cloning study with sequencing of complimentary determining region (CDR) 3 revealed the presence of a major clone in 4. Three of the 4 cases used V(H) 3 gene, with the homologous germline gene of V3-30 in two cases and VH26 in one case. A biased usage of V(H) 3 and V(H) 4 genes with the homologous germline gene of VH26 in V(H) 3 gene was reported previously in cases with CLTH. A high level of somatic mutation (1-21%, average 12%) with non-random distribution of replacement and silent mutations was accumulated in all cases. The frequency of the occurrence of minor clones ranged from 29-44% per case, indicating the presence of on-going mutation. DNA sequencing of immunoglobulin V(H) gene suggests that TL develops among activated lymphoid cells in CLTH at the germinal center stage under antigen selection.

Reuse of heat energy in wastewater: implementation examples in Japan.

Sewage and treated water can be a heat source in urban area due to large heat capacity, thus recovery and reuse of its energy is one of the most desirable plans for the sewerage system. In this paper, characteristics of heat energy in wastewater, reuse plans, and some experiences in Japan are presented. Full-scale reuse projects for heating and cooling in the Tokyo Metropolitan Districts and project for melting snow in Sapporo City are discussed. The key factors found in experience of Tokyo were setting the heat pumps near the demand points and the technical developments of equipment to prevent system from clogging, corrosion, and decrease in the heat transfer efficiency. It was also found through the project for melting snow in Sapporo that the key factor in public acceptance was the multi-purpose use of the sewerage system both for melting snow in winter and retaining rain water in summer.

Primary pancreatic lymphoma: clinicopathological analysis of 19 cases from Japan and review of the literature.

Primary pancreatic lymphoma (PL) is an extremely rare disease, and large-scale studies are rarely performed even in Western countries, in which all cases of PL reported to date were of the B-cell type. Little information regarding PL is available in Japan. Nineteen cases of PL were collected through a nationwide study in Japan, and paraffin-embedded specimens were processed for staining with hematoxylin and eosin and by immunohistochemical procedures. Clinicopathological findings were reviewed and compared with those reported in Western countries. The patient population consisted of 13 men and 6 women, ranging in age from 46 to 84 (average 62) years. Abdominal pain was the most common presenting symptom. Tumors were located in the pancreatic head (12 cases), tail (4 cases) and body (2 cases), and ranged in size from 4 to 17 cm. Clinical stage was I(E) in 9 cases and II(E) in 10. Radical resection was performed in 10 cases and bypass operation in 1, followed by chemotherapy in 8. Immunophenotypically, 15 cases were B-cell and 4 were T-cell lymphomas. Seventy-three percent of B-cell tumors were diffuse large B-cell lymphomas. The 1-year actuarial survival rate for B-cell lymphomas (51.9%) was better than that of T-cell lymphomas (0%). However, in Japan the incidence of T-cell PLs was higher, and, partly as a consequence of this, prognosis was poorer than in Western countries.

Fas gene mutations in prostatic intraepithelial neoplasia and concurrent carcinoma: analysis of laser capture microdissected specimens.

Fas (Apo-1/CD95) is a cell-surface receptor involved in cell death signaling through binding of Fas ligand. Mutations of the Fas gene might be involved in proliferative diseases of the prostate by prolongation of programmed cell death of prostatic epithelial cells. Using the laser capture microdissection method, Fas gene mutations were examined on genomic DNA extracted from lesions with high-grade prostatic intraepithelial neoplasia (HGPIN), a possible precursor of prostatic cancer (PCA), and from PCA. A total of 193 lesions, 111 with HGPIN, 55 with PCA, and 27 benign glands, were microdissected from 27 patients with PCA. Polymerase chain reaction-amplified products were directly sequenced. Loss of heterozygosity (LOH) was examined at four sites of known polymorphisms. Fas gene mutations were detected in HGPIN: 4 of 27 (14.8%) cases or 4 of 111 (3.6%) lesions. All were point mutations: three missense and one nonsense in the death domain. Benign proliferative glands adjoining HGPIN and/or PCA, and PCA never showed mutations. LOH was found in 31.3% of PCA and 25% of HGPIN lesions, but was never found in benign glands. Exclusive occurrence of Fas mutations in HGPIN might underlie the development of these lesions. Occasional findings of LOH in HGPIN and PCA suggested that genetic instability might occur during the early phase of prostatic carcinogenesis.