The Fat-Dachsous signaling pathway regulates growth of horns in Trypoxylus dichotomus, but does not affect horn allometry.

Males of the Asian rhinoceros beetle, Trypoxylus dichotomus, possess exaggerated head and thoracic horns that scale dramatically out of proportion to body size. While studies of insulin signaling suggest that this pathway regulates nutrition-dependent growth including exaggerated horns, what regulates disproportionate growth has yet to be identified. The Fat signaling pathway is a potential candidate for regulating disproportionate growth of sexually-selected traits, a hypothesis we advanced in a previous paper (Gotoh et al., 2015). To investigate the role of Fat signaling in the growth and scaling of the sexually dimorphic, condition-dependent traits of the in the Asian rhinoceros beetle T. dichotomus, we used RNA interference to knock down expression of fat and its co-receptor dachsous. Knockdown of fat, and to a lesser degree dachsous, caused shortening and widening of appendages, including the head and thoracic horns. However, scaling of horns to body size was not affected. Our results show that Fat signaling regulates horn growth in T. dichotomus as it does in appendage growth in other insects. However, we provide evidence that Fat signaling does not mediate the disproportionate, positive allometric growth of horns in T. dichotomus.

Endocrine Control of Exaggerated Trait Growth in Rhinoceros Beetles.

Juvenile hormone (JH) is a key insect growth regulator frequently involved in modulating phenotypically plastic traits such as caste determination in eusocial species, wing polymorphisms in aphids, and mandible size in stag beetles. The jaw morphology of stag beetles is sexually-dimorphic and condition-dependent; males have larger jaws than females and those developing under optimum conditions are larger in overall body size and have disproportionately larger jaws than males raised under poor conditions. We have previously shown that large males have higher JH titers than small males during development, and ectopic application of fenoxycarb (JH analog) to small males can induce mandibular growth similar to that of larger males. What remains unknown is whether JH regulates condition-dependent trait growth in other insects with extreme sexually selected structures. In this study, we tested the hypothesis that JH mediates the condition-dependent expression of the elaborate horns of the Asian rhinoceros beetle, Trypoxylus dichotomus. The sexually dimorphic head horn of this beetle is sensitive to nutritional state during larval development. Like stag beetles, male rhinoceros beetles receiving copious food produce disproportionately large horns for their body size compared with males under restricted diets. We show that JH titers are correlated with body size during the late feeding and early prepupal periods, but this correlation disappears by the late prepupal period, the period of maximum horn growth. While ectopic application of fenoxycarb during the third larval instar significantly delayed pupation, it had no effect on adult horn size relative to body size. Fenoxycarb application to late prepupae also had at most a marginal effect on relative horn size. We discuss our results in context of other endocrine signals of condition-dependent trait exaggeration and suggest that different beetle lineages may have co-opted different physiological signaling mechanisms to achieve heightened nutrient-sensitive weapon growth.

Knockout of a transgene by transcription activator-like effector nucleases (TALENs) in the sawfly, Athalia rosae (Hymenoptera) and the ladybird beetle, Harmonia axyridis (Coleoptera).

Transcription activator-like effector nucleases (TALENs) are efficient tools for targeted genome editing and have been utilized in a number of insects. Here, we demonstrate the gene disruption (knockout) caused by TALENs targeting a transgene, 3xP3-driven enhanced green fluorescence protein (EGFP), that is integrated in the genome of two species, the sawfly Athalia rosae (Hymenoptera) and the ladybird beetle Harmonia axyridis (Coleoptera). Messenger RNAs of TALENs targeting the sequences adjacent to the chromophore region were microinjected into the eggs/embryos of each species. In At. rosae, when microinjection was performed at the posterior end of eggs, 15% of G(0) individuals showed a somatic mosaic phenotype for eye EGFP fluorescence. Three-quarters of the somatic mosaics produced EGFP-negative G(1) progeny. When eggs were injected at the anterior end, 63% of the G(0) individuals showed somatic mosaicism, and 17% of them produced EGFP-negative G(1) progeny. In H. axyridis, 25% of posterior-injected and 8% of anterior-injected G(0) individuals produced EGFP-negative G(1) progeny. In both species, the EGFP-negative progeny retained the EGFP gene, and various deletions were detected in the target sequences, indicating that gene disruption was successfully induced. Finally, for both species, 18-21% of G(0) founders produced gene knockout progeny sufficient for establishing knockout strains.

Vestigial and scalloped in the ladybird beetle: a conserved function in wing development and a novel function in pupal ecdysis.

In Drosophila melanogaster, Vestigial (Vg) and Scalloped (Sd) form a transcription factor complex and play a crucial role in wing development. To extend our knowledge of insect wing formation, we isolated vg and sd homologues from two ladybird beetle species, Henosepilachna vigintioctopunctata and Harmonia axyridis. Although the ladybird beetle vg homologues had only low homology with D. melanogaster vg, ectopic expression of H. vigintioctopunctata vg induced wing-like tissues in antennae and legs of D. melanogaster. Subsequent larval RNA interference (RNAi) analysis in H. vigintioctopunctata demonstrated conserved functions of vg and sd in wing development, and an unexpected novel function of sd in pupal ecdysis. Furthermore, our results can be applied to the production of a flightless ladybird beetle for biological control purposes using larval RNAi.

[Intrathoracic chest wall type lipoma with crescent-shaped mass on computed tomography; report of a case].

An asymptomatic 53-year-old woman was referred to our hospital with a slow-growing intrathoracic tumor after 3 years observation. Chest computed tomography (CT) and magnetic resonance imaging (MRI) showed an intrathoracic crescent-shaped mass measuring 13 x 2.5 cm along the left chest wall. Fatty contents were suggested. Surgical resection was successfully performed by video-assisted thracoscopic surgery. At operation, a yellowish, thinly encapsulated, 10 cm tumor was attached with its pedicle 2.5 cm in diameter to the pleura of the 7th intercostal space. The lesion was a soft mass, pedunculated and numerously lobulated, that caused the crescent form of the mass on chest CT and MRI. Histopathological examination revealed mature lipoma.

[Cystic thymoma; report of a case].

We present a case of a 33-year-old man with a noninvasive thymoma undergoing extensive cystic degeneration. The mediastinal tumor was asymptomatic and first noted on a routine chest radiograph. Chest computed tomography (CT) and magnetic resonance imaging (MRI) showed a 11 cm cystic mass with some solid portions in the anterior mediastinum. A cystic thymoma was suggested. The mass and remnants of the thymus were removed by video-assisted thoracoscopic surgery. On cut section, the tumor was predominantly cystic, with several solid nodules randomly attached to the cyst wall. The cystic space was filled with turbid brown fluid. Histopathological examination revealed a World Health Organization (WHO) type B3 thymoma with foci of hemorrhage, necrosis and cystic degeneration, and absence of an epithelial lining of the cyst.

Fractal-feature distance analysis of contrast-detail phantom image and meaning of pseudo fractal dimension and complexity.

The purposes of our studies are to examine whether or not fractal-feature distance deduced from virtual volume method can simulate observer performance indices and to investigate the physical meaning of pseudo fractal dimension and complexity. Contrast-detail (C-D) phantom radiographs were obtained at various mAs values (0.5 - 4.0 mAs) and 140 kVp with a computed radiography system, and the reference image was acquired at 13 mAs. For all C-D images, fractal analysis was conducted using the virtual volume method that was devised with a fractional Brownian motion model. The fractal-feature distances between the considered and reference images were calculated using pseudo fractal dimension and complexity. Further, we have performed the C-D analysis in which ten radiologists participated, and compared the fractal-feature distances with the image quality figures (IQF). To clarify the physical meaning of the pseudo fractal dimension and complexity, contrast-to-noise ratio (CNR) and standard deviation (SD) of images noise were calculated for each mAs and compared with the pseudo fractal dimension and complexity, respectively. A strong linear correlation was found between the fractal-feature distance and IQF. The pseudo fractal dimensions became large as CNR increased. Further, a linear correlation was found between the exponential complexity and image noise SD.

Molecular cloning and characterization of cDNAs encoding dopamine receptor-1 and -2 from brain-suboesophageal ganglion of the silkworm, Bombyx mori.

In order to better understand the relationship between dopamine and the release of diapause hormone into the blood, we cloned and characterized cDNAs encoding Bombyx mori dopamine receptor-1 and -2 (BmDopR1 and 2) from the pupal brain-suboesophageal ganglion. BmDopR1 and 2 had high similarities to group 1 (Drosophila melanogaster DOP1 and Apis mellifera DOP1) and group 2 (D. melanogaster DopR99B, A. mellifera DOP2 and Papilio xuthus DOP1), respectively. When BmDopR1 and 2 were expressed in human embryonic kidney (HEK) cells, they responded to dopamine by increasing intracellular cAMP levels, thus indicating the presence of D1-like receptors. There were no clear differences in BmDopR1 and 2 mRNA levels between brain-suboesophageal ganglion complexes of diapause and nondiapause egg producers during pupal-adult development. BmDopR1 and 2 mRNAs were concentrated in the mushroom body calyx rather than in the suboesophageal ganglion. Taking into account the results of earlier experiments on excised regions corresponding to mushroom bodies, BmDopR1 and 2 in the mushroom body apparently play a role in the release of diapause hormone.

Analysis of streak artefacts on CT images using statistics of extremes.

We have analysed the statistical characteristics of streak artefacts on CT images using the statistics of extremes, and have devised a new method of evaluating streak artefacts on CT images. The CT images of four polymer tubes placed on the chest wall of a commercially available chest phantom were used as the target objects for our analysis. 40 parallel line segments with a length of 20 pixels were placed perpendicular to numerous streak artefacts on the polymer tube image, and the largest difference between adjacent CT values in each of the 40 CT value profiles of these line-segments was employed as a feature variable of a streak artefact; these feature variables have been analysed by extreme value theory. Using the mean rank method, a Gumbel distribution was shown to be the most suitable extreme value distribution for the largest difference between adjacent CT values in each CT value profile. This enabled us to demonstrate that the streak artefacts on CT images can be statistically modelled by a Gumbel distribution. Both the location parameter and the scale parameter of the estimated Gumbel probability density distribution were large on the CT slices in which the shoulder bone or liver was included.

Germ-line transformation and RNAi of the ladybird beetle, Harmonia axyridis.

To elucidate the molecular mechanisms underlying the tremendous diversity of insect wing colour patterns, it is imperative to identify and functionally characterize the genes involved in this developmental process. Here we report the first successful germ-line transformation using the transposable element vector piggyBac in the ladybird beetle Harmonia axyridis, which demonstrates typical genetic polymorphism in its wing colour patterns. The transformation efficiency by piggyBac was 3.7% per fertile G(0). We investigated the effectiveness of RNAi in Harmonia by injecting EGFP (enhanced green fluorescent protein) dsRNA into early transgenic EGFP-expressing embryos and observed substantial reduction of EGFP fluorescence in 87.2% of hatched larvae. Application of these new genetic tools to non-model insects such as Harmonia will facilitate the broad understanding of developmental mechanisms and evolutionary processes that are inaccessible using established model systems.

Extended aortic bypass for atherosclerotic occlusive disease of high abdominal aorta.

We describe a case of acute renal failure caused by extension of an atherosclerotic occlusive process to the suprarenal aorta. A 68-year-old man underwent bypass grafting from the ascending aorta to the femoral arteries using straight and Y-grafts. Additional saphenous vein grafts were placed from the intra-abdominal portion of the graft to the left and right renal arteries and the splenic artery. As a consequence, relief of limb ischemia was achieved and dialysis could be discontinued.

Membrane-penetrating trehalase from silkworm Bombyx mori. Molecular cloning and localization in larval midgut.

The main blood sugar in insects, trehalose, differs from glucose in mammals. To incorporate trehalose into cells and utilize it, tissue cells possess the enzyme trehalase (EC3.2.1.28), which catalyses trehalose into glucose, in the organellar membrane or in the cytoplasm. Soluble and membrane-bound trehalase proteins have been isolated from insects. To date, however, only genes encoding the soluble trehalase have been reported in insects. Soluble trehalase is therefore believed to become localized on the cell surface via modification. In contrast, cDNAs encoding trehalase localized on the apical cell surface via the glycosylphosphatidylinositol-anchor have been isolated from mammalian small intestines. The amino acid sequence contains a specific hydrophobic region and an upstream omega site, which is cleaved for glycosylphosphatidylinositol-attachment, at the C-terminus. Here, we describe a cDNA from the silkworm Bombyx mori that encodes a novel trehalase (type-2) with one transmembrane domain and lacking the omega site. Immunoblotting and immunohistochemical analyses demonstrated that in the midgut tissue of Bombyx larvae, soluble trehalase-1 is present mainly in goblet cell cavities, but membrane-bound trehalase-2 is predominantly seen on the visceral muscle surrounding the midgut. To our knowledge, this is the first report of a cDNA encoding trehalase that penetrates the cell membrane in insects and its cellular localization.

[Aortic dissection induced by symmetry aortic connector system].

A 74-year-old male with severe triple vessel disease underwent off-pump coronary artery bypass grafting (OPCAB). Preoperative computed tomography (CT) showed severely calcified ascending aorta. We revasculize the left coronary arteries with in situ internal thoracic artery (ITA) graft and the right coronary artery with a saphenous vein graft, which was attached to the disease-free portion of the aortic root, using Symmetry aortic connector system (ACS). Although the operation was uncomplicated, and postoperative course was uneventful until the 5th postoperative day when acute type A aortic dissection occurred. The patient died of aortic rupture on the 7th postoperative day. Necropsy disclosed that the entry located just on the proximal anastomotic site of the vein graft. It is possible placement of ACS device would trigger the dissecting process. With regard to the use of these one-shot devices for diseased aorta, its safety needs further investigation, even though it might be placed on an apparently intact portion.

Cloning, expression, and chromosomal localization of the mouse gene (Scgb3a1, alias Ugrp2) that encodes a member of the novel uteroglobin-related protein gene family.

The mouse UGRP gene family consists of two genes, Ugrp1 and Ugrp2. In this study, the genomic structure and expression patterns of Ugrp2 and its alternative spliced form were characterized. The authentic Ugrp2 gene has three exons and two introns, similar to the Ugrp1 gene, which produces a secreted protein. The Ugrp2 variant uses a sequence located between authentic exons 1 and 2, resulting in a cytoplasmic form due to a termination codon within the inserted sequence. Both mouse and human UGRP2 mRNAs are expressed in lung. In the case of human, the mRNA is expressed at the highest level in trachea, followed by salivary gland at a level similar to lung. Weak expression was also found in fetal lung and mammary gland. Ugrp2 was mapped by fluorescence in situ hybridization to mouse chromosome 11A5-B1 and human chromosome 5q35. These regions are known to be homologous. Interspecific mouse backcross mapping was also performed to obtain further detailed localization of mouse Ugrp1 and Ugrp2.

Transforming growth factor beta1 gene polymorphism in rheumatoid arthritis.

OBJECTIVE: Rheumatoid arthritis (RA) is a chronic inflammatory disease and synovial cells, antigen presenting cells, lymphocytes, and their cytokines might be associated with the disease. Transforming growth factor beta1 (TGFbeta1) has been reported to have important roles in unresolved inflammation, immune suppression, fibrosing processes, and angiogenesis. TGFbeta1 is highly expressed in joints in RA and is considered to be a regulator of anti-inflammation in RA. Polymorphisms of TGFbeta1 have been reported to be associated with the production of TGFbeta1 protein, and to increase the risk of acquiring several diseases. It was speculated that these polymorphisms might also be involved in RA, and therefore the TGFbeta1 codon 10 T869C polymorphism in a series of patients and controls was investigated. METHOD: A total of 155 patients with RA and 110 healthy subjects were studied. DNA was extracted from peripheral leucocytes and TGFbeta1 codon 10 T869C polymorphism was determined by polymerase chain reaction restriction fragment polymorphism. RESULTS: A significantly higher proportion of patients with RA with the T allele (CT type or TT type) was found compared with the CC type (p=0.039). CONCLUSION: The T allele, previously reported to be linked with production of TGFbeta1, may be associated with an increased risk of RA.

[Surgical treatment of traumatic rupture of the bicuspid aortic valve; report of a case].

A 27-year-old man was injured during a motocross game. He was suffered from dyspnea, orthopnea, and hemoptysis. The to-and-fro murmur was noticed 3 days after the accident and then the patient was admitted to our hospital. Echocardiography revealed severe aortic regurgitation. Computerized tomography also showed severe pulmonary contusions. Seventeen days after the accident the aortic valve replacement was performed. The aortic valve was anterior-posterior type bicuspid valve (fusion of right coronary cusp and light coronary cusp) and the tear was detected in the anterior cusp. The postoperative course was uneventful. Rupture of the aortic valve due to a blunt chest trauma is rare and reported in 20 cases previously in Japan. This case is the second report of traumatic rupture of the bicuspid aortic valve.

Transforming growth factor-beta gene polymorphism in sarcoidosis and tuberculosis patients.

SETTING: Transforming growth factor-beta (TGF-beta) plays an important role in many diseases, influencing as it does such processes as immune responses, fibrosing processes, and angiogenesis. Recently, polymorphisms have been described for TGF-beta that are associated with the risk of several diseases. In this study, we investigated whether TGF-beta 1 polymorphism has an effect on sarcoidosis and tuberculosis. OBJECTIVE: TGF-beta 1 Codon 10 T869C polymorphism was investigated in 110 healthy control subjects, 104 sarcoidosis patients, and 101 tuberculosis patients. DESIGN: The TGF-beta genotype was determined using polymerase chain reaction restriction fragment length polymorphism. RESULTS: We found no significant differences in TGF-beta genotypes between sarcoidosis patients and healthy controls or tuberculosis patients and controls. The long axis of the tuberculin skin test was larger in the CC type compared with the CT type. However, there was no association between the TGF-beta genotype and the roentgenographic stage, the disappearance of shadows, or organ involvement in sarcoidosis, nor any association between genotype, the extent or type of roentgenographic shadow, or detected volume of tubercle bacilli in tuberculosis. CONCLUSION: From the results, we believe that TGF-beta polymorphisms on the whole do not have a strong influence on disease onset or clinical progression in sarcoidosis and tuberculosis, although this polymorphism might have an effect on the immune response in a tuberculosis host.

Design and synthesis of non-peptidic inhibitors for the Syk C-terminal SH2 domain based on structure-based in-silico screening.

Structure-based in-silico screening was carried out for the Syk C-terminal SH2 domain. Fragments that could interact with the pY or pY+1 pockets were selected by our in-silico screening. After tethering two fragments bound to these pockets, we have designed and synthesized new compounds that show favorable interaction with the pY+3 pocket. One such compound, having a cyclohexylmalonic acid moiety identified as a novel potent phosphotyrosyl mimetic, exhibited an affinity comparable to that of the monophosphorylated ligand peptide.

claudin-18, a novel downstream target gene for the T/EBP/NKX2.1 homeodomain transcription factor, encodes lung- and stomach-specific isoforms through alternative splicing.

T/EBP/NKX2.1, a member of the NKX family of homeodomain-containing transcription factors, regulates the expression of a number of genes in lung and thyroid. Here we describe the isolation and characterization of a novel target gene, termed claudin-18, that is down-regulated in the lungs of T/ebp/Nkx2.1-null mouse embryos. The gene product exhibits an amino acid sequence similar to those of the claudin multigene family of proteins that constitute tight junction strands in epithelial cells. The gene was localized by fluorescence in situ hybridization to mouse chromosome 9 at region 9E3-F1 and to human chromosome 3 at region 3q21-23. The claudin-18 gene has two promoters, each with its own unique exon 1 that is spliced to common exons 2 through 5. Alternative usage of these promoters leads to production of lung and stomach-specific transcripts. The downstream lung-specific promoter contains two T/EBP/NKX2.1 binding sites responsible for trans activation of the gene by T/EBP/NKX2.1 in lung cells. Only claudin-18 was down-regulated in T/ebp/Nkx2.1-null embryo lungs among 11 claudin transcripts examined. Furthermore, the claudin-18 transcript has an alternative 12-bp insertion derived from the 5' end of intron 4, which produces a C-terminally truncated isoform in lung and stomach. Immunohistochemistry demonstrated complete membrane localization of claudin-18 with small focal dots in the lung and stomach epithelial cells. Immunogold electron microscopy analysis revealed that claudin-18 is concentrated at the cell-cell borders of epithelial cells. These unique features suggest a potentially important role for claudin-18 in the structure and function of tight junctions in lung and stomach.