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Two large abalone species prevalent in the Japanese archipelago, Haliotis discus discus and H. madaka, are considered to have different morphological characteristics, and previous population genomics analyses have suggested that they have distinct genomic features. However, integrated analyses comparing the genetic and morphological data from these species are lacking; thus, it remains unclear whether these two species are distinct biological species. In this study, we performed integrated genetic and morphological analyses on individuals of the two species captured in Tokushima Prefecture, Japan. Genetic analysis based on 118 species-diagnostic single-nucleotide polymorphism (SNP) loci showed that H. discus discus and H. madaka were clearly assigned to distinct genetic clusters corresponding to the two species, with the exception of one H. madaka individual showing evidence of genomic introgression from H. discus discus. Multiple statistical analyses based on quantitative characteristics of shell shape and foot muscle color suggested that this H. madaka individual with genomic introgression from H. discus discus had H. discus discus-like quantitative characteristics consistently. In summary, this study suggests that H. discus discus and H. madaka are distinct biological species reproductively isolated in natural environments, but genomic introgression possibly blurs the species boundary between H. discus discus and H. madaka. [ABSTRACT FROM AUTHOR]

Remote ischemic perconditioning (RIPerC) is a novel neuroprotective method against cerebral infarction that has shown efficacy in animal studies but has not been consistently neuroprotective in clinical trials. We focused on the temporal regulation of ischemia–reperfusion by RIPerC to establish an optimal method for RIPerC. Rats were assigned to four groups: 10 min ischemia, 5 min reperfusion; 10 min ischemia, 10 min reperfusion; 5 min ischemia, 10 min reperfusion; and no RIPerC. RIPerC interventions were performed during ischemic stroke, which was induced by a 60-min left middle cerebral artery occlusion. Infarct volume, sensorimotor function, neurological deficits, and cellular expressions of brain-derived neurotrophic factor (BDNF), B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), and caspase 3 were evaluated 48 h after the induction of ischemia. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling (TUNEL) was also performed. RIPerC of 10 min ischemia/10 min reperfusion, and 5 min ischemia/10 min reperfusion decreased infarct volume, improved sensorimotor function, decreased Bax, caspase 3, and TUNEL-positive cells, and increased BDNF and Bcl-2 expressions. Our findings suggest RIPerC with a reperfusion time of approximately 10 min exerts its neuroprotective effects via an anti-apoptotic mechanism. This study provides important preliminary data to establish more effective RIPerC interventions. [ABSTRACT FROM AUTHOR]

Subarachnoid hemorrhage (SAH) is a catastrophic form of stroke responsible for significant morbidity and mortality. Oxidative stress, inflammation, and neuronal apoptosis are important in the pathogenesis of early brain injury (EBI) following SAH. Preconditioning exercise confers neuroprotective effects, mitigating EBI; however, the basis for such protection is unknown. We investigated the effects of preconditioning exercise on brain damage and sensorimotor function after SAH. Male rats were assigned to either a sham-operated (Sham) group, exercise (Ex) group, or no-exercise (No-Ex) group. After a 3-week exercise program, they underwent SAH by endovascular perforation. Consciousness level, neurological score, and sensorimotor function were studied. The expression of nuclear factor erythroid 2 p45-related factor 2 (Nrf2), heme oxygenase 1 (HO-1), 4-hydroxynonenal (4HNE), nitrotyrosine (NT), ionized calcium-binding adaptor molecule 1 (Iba1), tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), interleukin 1β (IL-1β), 14–3-3γ, p-β-catenin Ser37, Bax, and caspase-3 were evaluated by immunohistochemistry or western blotting. The terminal deoxynucleotidyl transferase-mediated biotinylated dUTP nick end labeling (TUNEL) assay was also performed. After SAH, the Ex group had significantly reduced neurological deficits, sensorimotor dysfunction, and consciousness disorder compared with the No-Ex group. Nrf2, HO-1, and 14–3-3γ were significantly higher in the Ex group, while 4HNE, NT, Iba1, TNF-α, IL-6, IL-1β, Bax, caspase-3, and TUNEL-positive cells were significantly lower. Our findings suggest that preconditioning exercise ameliorates EBI after SAH. The expression of 4HNE and NT was reduced by Nrf2/HO-1 pathway activation; additionally, both oxidative stress and inflammation were reduced. Furthermore, preconditioning exercise reduced apoptosis, likely via the 14–3-3γ/p-β-catenin Ser37/Bax/caspase-3 pathway. [ABSTRACT FROM AUTHOR]

Aquaculture production is expected to increase with the help of genomic selection (GS). The possibility of performing GS using only a small number of SNPs has been examined in order to reduce genotyping costs; however, the practicality of this approach is still unclear. Here, we tested whether the effects of reducing the number of SNPs impaired the prediction accuracy of GS for standard length, body weight, and testes weight in the tiger pufferfish (Takifugu rubripes). High values for predictive ability (0.563–0.606) were obtained with 4000 SNPs for all traits under a genomic best linear unbiased predictor (GBLUP) model. These values were still within an acceptable range with 1200 SNPs (0.554–0.588). However, predictive abilities and prediction accuracies deteriorated using less than 1200 SNPs largely due to the reduced power in accurately estimating the genetic relationship among individuals; family structure could still be resolved with as few as 400 SNPs. This suggests that the SNPs informative for estimation of genetic relatedness among individuals differ from those for inference of family structure, and that non-random SNP selection based on the effects on family structure (e.g., site-FST, principal components, or random forest) is unlikely to increase the prediction accuracy for these traits. [ABSTRACT FROM AUTHOR]

The novel non-targeted PCR-based genotyping system, namely Genotyping by Random Amplicon Sequencing, Direct (GRAS-Di), is characterized by the simplicity in library construction and robustness against DNA degradation and is expected to facilitate advancements in genetics, in both basic and applied sciences. In this study, we tested the utility of GRAS-Di for genetic analysis in a cultured population of the tiger pufferfish Takifugu rubripes. The genetic analyses included family structure analysis, genetic map construction, and quantitative trait locus (QTL) analysis for the male precocious phenotype using a population consisting of four full-sib families derived from a genetically precocious line. An average of 4.7 million raw reads were obtained from 198 fish. Trimmed reads were mapped onto a Fugu reference genome for genotyping, and 21,938 putative single-nucleotide polymorphisms (SNPs) were obtained. These 22 K SNPs accurately resolved the sibship and parent–offspring pairs. A fine-scale linkage map (total size: 1,949 cM; average interval: 1.75 cM) was constructed from 1,423 effective SNPs, for which the allele inheritance patterns were known. QTL analysis detected a significant locus for testes weight on Chr_14 and three suggestive loci on Chr_1, Chr_8, and Chr_19. The significant QTL was shared by body length and body weight. The effect of each QTL was small (phenotypic variation explained, PVE: 3.1–5.9%), suggesting that the precociousness seen in the cultured pufferfish is polygenic. Taken together, these results indicate that GRAS-Di is a practical genotyping tool for aquaculture species and applicable for molecular breeding programs, such as marker-assisted selection and genomic selection. [ABSTRACT FROM AUTHOR]

Paleoenvironmental changes in marginal seas on the coast of the northwestern Pacific during the Quaternary have caused genetic differentiation and subsequent secondary contacts between populations of coastal organisms in this region. However, little is known about how geographical isolation and the subsequent connection events associated with these multiple marginal seas have shaped the population structure of coastal organisms in the boundary area. The coastal area of Kyushu Island of the Japanese Archipelago, where the East China Sea (ECS), the Sea of Japan (SJ), and the Pacific Ocean (PO) adjoin within a narrow region, would be a suitable field for elucidating the effect of multiple vicariances. Herein, we conducted phylogeographical analyses of the intertidal goby Chaenogobius annularis, focusing on populations around Kyushu Island, using partial mitochondrial (mt) DNA sequences and analysis of 8 microsatellite (ms) DNA loci. We discovered a new lineage distributed along the coast of the ECS distinct from the already-known SJ and PO lineages, which we termed the ECS lineage. This ECS lineage showed remarkable mito-nuclear discordance: mtDNA analyses indicated that the ECS lineage is a highly diverged lineage branched from the PO lineage, whereas msDNA analyses suggested that the ECS lineage is closely related to the SJ lineage. This discordance could have been due to ancient hybridization. In addition, we detected SJ-PO and ECS-SJ hybrid zones on the western Seto Inland Sea and on the northwestern coast of Kyushu, respectively. Such a complex population structure suggests that the secondary contacts after multiple isolation events among marginal seas around Kyushu have a great impact on coastal biodiversity in the area and emphasize the peculiarities of the region. [ABSTRACT FROM AUTHOR]

Testes of the tiger pufferfish Takifugu rubripes are a delicacy in Japan, and selective breeding for a male precocious phenotype, i.e., with early initiation of testes development, is desirable. However, it is unknown if precocious gonad development in this species is under genetic control. Here, we investigated genetic involvement in precociousness by using progeny tests with sires from two cultured populations, including a family line anecdotally known for its precociousness, and a wild population. Progeny derived from the "precocious" line consistently had greater testes weight than that from the other lines, even after accounting for effects of body weight, which indicates that precociousness is truly heritable. We also compared chronological changes in plasma steroid hormones between progenies sired by males from the precocious line and a wild population, and found that the precocious family line had higher levels of plasma estradiol-17β (E2) prior to the initiation of testicular development. Our findings suggest that selective breeding for testes precociousness in the tiger pufferfish is feasible, and that plasma E2 may be an indicator of this phenotype, which would allow for phenotype evaluation without the need to sacrifice specimens. [ABSTRACT FROM AUTHOR]

Population genetic studies of marine species sometimes find no clear patterns of genetic differentiation, but signals of geographic gene flow typified by isolation by distance (IBD) due to high dispersal ability. A previous study of mitochondrial DNA of the yellowfin goby Acanthogobius flavimanus indicated that this species also does not have clearly differentiated population groups around the Japanese archipelago, in contrast with other coastal goby species that have highly differentiated groups between the Sea of Japan and Pacific Ocean populations. Here, to clearly reveal phylogeographic structures of the yellowfin goby around the Japanese archipelago, we conducted large-scale RAD-sequencing analysis of their nuclear genomic SNP loci. As a result, IBD patterns were confirmed across the yellowfin goby populations, and the larval dispersal distance of this species was revealed to be approximately 19 km on average. In addition, different larval dispersal distances were estimated for its Sea of Japan and Pacific Ocean populations, suggesting that different coastal environments and/or ecological features may affect its dispersal rates. These results would provide foundations for ecological and conservation studies of the yellowfin goby and coastal fishes having similar dispersal strategy to this species. [ABSTRACT FROM AUTHOR]

The tiger pufferfish Takifugu rubripes is one of the most valuable aquaculture species in Japan. In this study, we investigated the genetic variation in resistance to heterobothriosis caused by a host-specific monogenean parasite, Heterobothrium okamotoi, in the tiger pufferfish, and examined the possibility of selective breeding for enhancing resistance traits. A total of 299 tiger pufferfish produced from ten males and one female caught from the wild were experimentally exposed to oncomiracidia of H. okamotoi. The number of parasites on the gills at 3 weeks post-infection ranged from 0 to 68, with great variation observed both within and between families. The estimated heritability for parasite density was small, but significantly different from zero (H2 = 0.098; 95% confidence interval: 0.025–0.201). Although we may have sampled a limited fraction of the genetic variance as the number of families was small, the results suggest that there are genetic components that affect heterobothriosis resistance in the species. A positive phenotypic correlation was observed between parasite count and standard length (r = 0.66). Further investigation of the genetic correlation between the two traits using larger families is needed to assess the impacts of selective breeding on heterobothriosis resistance in this species. [ABSTRACT FROM AUTHOR]

14-3-3γ is an important early ischemia-inducible protective factor against ischemic cell death in cerebral cortical neurons. We investigated the anti-apoptosis mechanism of enhanced 14-3-3γ mediated by preconditioning exercise-induced brain ischemic tolerance after stroke. Rats were assigned to four groups: exercise and ischemia (Ex group), ischemia and no exercise (No-Ex group), exercise and no ischemia (Ex-only group), and no exercise and ischemia (control group). Rats were trained on a treadmill for 5 days a week for 3 weeks (running speed, 25 m/min; running duration, 30 min/day). After the exercise program, stroke was induced by left middle cerebral artery occlusion. The infarct volume, neurological deficits, and motor function, as well as expression levels of hypoxia-induced factor-1α (HIF-1α), 14-3-3γ, P2X7 receptors, p-β-catenin Ser37, Bax, and caspase 3 were evaluated by immunohistochemistry and western blotting. The expression of HIF-1α and 14-3-3γ significantly increased in neurons and astrocytes in the Ex-only group. HIF-1α was co-expressed with P2X7 receptor- and GFAP-positive astrocytes. After stroke, the Ex group had significantly reduced brain infarction. HIF-1α and 14-3-3γ significantly increased in the Ex group compared to the No-Ex group. In addition, p-β-catenin Ser37 significantly increased following elevated 14-3-3γ; in contrast, Bax and caspase 3 were significantly reduced in the Ex group. Our findings suggest that preconditioning exercise prior to ischemia induces neuron- and astrocyte-mediated brain ischemic tolerance through increased expression of HIF-1α and 14-3-3γ, which are intrinsic protective factors; the upregulated 14-3-3γ induced by preconditioning exercise reduces ischemic neuronal cell death through the 14-3-3γ/p-β-catenin Ser37/Bax/caspase 3 anti-apoptotic pathway. [ABSTRACT FROM AUTHOR]

The tiger puffer Takifugu rubripes is one of the most popular aquacultural fish; however, there are two major obstacles to selective breeding. First, they have a long generation time of 2 or 3 years until maturation. Second, the parental tiger puffer has a body size (2-5 kg) much larger than average market size (0.6-1.0 kg). The grass puffer Takifugu niphobles is closely related to the tiger puffer and matures in half the time. Furthermore, grass puffer can be reared in small areas since their maturation weight is about 1/150 that of mature tiger puffer. Therefore, to overcome the obstacles of maturation size and generation time of tiger puffer, we generated surrogate grass puffer that can produce tiger puffer gametes through germ cell transplantation. Approximately 5000 tiger puffer testicular cells were transplanted into the peritoneal cavity of triploid grass puffer larvae at 1 day post hatching. When the recipient fish matured, both males and females produced donor-derived gametes. Through their insemination, we successfully produced donor-derived tiger puffer offspring presenting the same body surface dot pattern, number of dorsal fin rays, and DNA fingerprint as those of the donor tiger puffer, suggesting that the recipient grass puffer produced functional eggs and sperm derived from the donor tiger puffer. Although fine tunings are still needed to improve efficiencies, surrogate grass puffer are expected to accelerate the breeding process of tiger puffer because of their short generation time and small body size. [ABSTRACT FROM AUTHOR]

Correlated suits of behaviors (behavioral syndrome) are commonly observed in both inter- and intraspecific studies. In order to understand the genetic basis of such a correlation between species, we compared ten behaviors classified into five categories (acclimation, feeding, normal swimming, reaction to a novel object and activity in a novel environment) between two pufferfish species, Takifugu rubripes and T. niphobles. The two species showed consistent differences in nine behaviors with a significant correlation among behaviors. Quantitative trait locus (QTL) analysis using second generation hybrids revealed that different sets of small effect QTL are associated with the observed interspecific behavioral disparity. This indicates that correlations in temperament traits between them are governed by many genes with small effects, and each behavior has been selected to form particular combination patterns. One of the QTL showing small pleiotropic effect includes the Drd4 gene known for its association with behavioral traits in some animal taxa including mammals. [ABSTRACT FROM AUTHOR]

Acute ischemic stroke (AIS) is a major cause of mortality and disability and remains a serious and significant global health problem. The development of neurovascular protectants to treat AIS successfully has been beset by disappointments and setbacks. Many promising candidates have lacked significant pleiotropic protective activity for brain tissue and cerebral blood vessels in clinical trials, while those with protective activity have had poor bioavailability or high toxicity. Moreover, the majority of agents did not confer significant neurovascular protection or clinical efficacy, as measured by standard behavioral endpoints in clinical trials of heterogeneous populations of patients with AIS. The recombinant tissue plasminogen activator alteplase is approved in many countries for the treatment of AIS in the first 3 h after symptom onset. Many drug candidates have been subject to clinical trials, including those with anti-excitotoxic, anti-inflammatory, antioxidant, antiapoptotic/regenerative, calcium/adrenergic-modulating/antihypertensive, thrombolytic, nootropic/stimulant, fluid regulatory, or oxygen-delivering mechanisms of action. Some agents, such as tenecteplase, edaravone and minocycline, may be approved for global use in the future. This review evaluates almost all neurovascular protectants subject to clinical trial evaluation for the treatment of AIS, and includes 241 studies conducted between 1978 and 2014. The development of agents that reduce brain injury after AIS will require new and different approaches based on a deeper understanding of the pathophysiology of AIS. Moreover, the future treatment for AIS is likely to lie in combination therapy rather than monotherapy. Additional approaches to the testing and use of neurovascular protectants should be considered. [ABSTRACT FROM AUTHOR]

The tiger pufferfish (fugu) is one of the most important food fishes in East Asia. Since its testes are regarded as a delicacy, sex determination is economically relevant. Previous studies have identified a missense single-nucleotide polymorphism (SNP) in the Amhr2 ( anti-Müllerian hormone receptor type II) gene as a strong candidate for a master sex-determining polymorphism. To distinguish genotypic sex efficiently, we developed a high-resolution melting (HRM) assay for this SNP site. By screening 396 fish from two independent crosses reared under controlled conditions, we observed perfect concordance between the SNP genotype and phenotypic sex. Thus, this method holds great potential for use in high-throughput sexing. When analyzing 293 progeny from a third cross reared under unknown conditions, we unexpectedly found that 25 % of phenotypic males exhibited female genotype. These results suggest that environmental factors such as rearing conditions could influence the sex-determination pathway in pufferfish. Alternatively, genetic modifiers might override the signals from Amhr2. This finding raises a concern regarding enhanced stock management of this species, because sex-reversed fish could compromise the sex ratio in subsequent generations. The HRM assay will also be useful for monitoring the degree of sex reversal before release. [ABSTRACT FROM AUTHOR]

We investigated the influence of paternal effects on progeny survival of the fugu Takifugu rubripes when multiple families were reared in communal tanks. Four males were crossed with four females in a factorial cross to give 16 families, and the half-siblings sharing the same mother were reared together. Male parentage was determined using microsatellite markers at 35 days. Although the maternal and tank effects were large, it was possible to detect paternal effects in relative survival rate and standard length (standardized in each tank), which were found to be significantly correlated. Paternal effects on growth rate suggest the existence of family-based differences in competition for food. Furthermore, genetic differences affecting larval growth appeared to be partly responsible for the disparity in survival rate observed among families. [ABSTRACT FROM AUTHOR]

Artificial cultivation of the Japanese eel has not been feasible due to difficulties in seed production and larval rearing, leading to poor seed and larval survival. We have focused on the importance of the thymus and T-cell function in eel larval stages because only the thymus, of all the immune system organs, develops well in eel leptocephalus. In this study, we identified lymphocyte-specific protein tyrosine kinase ( lck) as the T-cell marker gene and analyzed lck expression in eel by reverse transcription (RT)-PCR and in situ hybridization. Eel lck gene encodes a predicted protein made up of 506 amino acids, including an essential motif that binds to the cytoplasmic domain of both CD4 and CD8 in T cells. RT-PCR analysis revealed that eel lck was expressed in major lymphoid organs (thymus, kidney, and spleen), mucosal organs (intestine and skin), and T-cell populations. In situ hybridization demonstrated the presence of lck-positive cells in the thymus. Eel lck was also expressed in the leptocephalus and even preleptocephalus at 3 and 7 days post-fertilization (artificial fertilization). Our results show that eel lck is an appropriate molecular marker for T-cell identification and for the study of thymus development, and that it offers a better understanding of the eel immune system. [ABSTRACT FROM AUTHOR]

Costello syndrome (CS) is a congenital disease that is characterized by a distinctive facial appearance, failure to thrive, mental retardation and cardiomyopathy. In 2005, we discovered that heterozygous germline mutations in HRAS caused CS. Several studies have shown that CS-associated HRAS mutations are clustered in codons 12 and 13, and mutations in other codons have also been identified. However, a comprehensive comparison of the substitutions identified in patients with CS has not been conducted. In the current study, we identified four mutations (p.G12S, p.G12A, p.G12C and p.G12D) in 21 patients and analyzed the associated clinical manifestations of CS in these individuals. To examine functional differences among the identified mutations, we characterized a total of nine HRAS mutants, including seven distinct substitutions in codons 12 and 13, p.K117R and p.A146T. The p.A146T mutant demonstrated the weakest Raf-binding activity, and the p.K117R and p.A146T mutants had weaker effects on downstream c-Jun N-terminal kinase signaling than did codon 12 or 13 mutants. We demonstrated that these mutant HRAS proteins induced senescence when overexpressed in human fibroblasts. Oncogene-induced senescence is a cellular reaction that controls cell proliferation in response to oncogenic mutation and it has been considered one of the tumor suppression mechanisms in vivo. Our findings suggest that the HRAS mutations identified in CS are sufficient to cause oncogene-induced senescence and that cellular senescence might therefore contribute to the pathogenesis of CS. [ABSTRACT FROM AUTHOR]

Japanese spotted fever (JSF) is severe and can progress to disseminated intravascular coagulation (DIC) with a poor prognosis. We considered whether patient factors are related to serious complications. Between August 1999 and March 2009, all patients with JSF and retrievable clinical data (age, gender, length of hospital stay, medication, comorbidities), vital signs (blood pressure, heart rate, temperature), and laboratory test results [blood cell count, liver function, renal function, electrolytes, blood sugar, C-reactive protein (CRP), CRP normalization period, and aspartate aminotransferase normalization period] from the Integrated Intelligent Management System (IIMS) database, were retrospectively analyzed by logistic regression. There were 51 JSF patients (24 men, 27 women) with a mean age of 63.0 years. Six patients (11.8%) had DIC, but there were no in-hospital deaths. The time between fever onset and initiation of medication was approximately 5 days, but this delay was not associated with disease severity. We identified values correlating with disease severity ( p < 0.1) by univariate analysis and then applied logistic regression. We found renal dysfunction [serum creatinine (Cr) ≥1.5 mg/dl] at the time of initial presentation to be predictive of DIC. Cr was also predictive of a prolonged disease course. In patients with JSF, renal function must be carefully monitored when determining clinical management. [ABSTRACT FROM AUTHOR]

The effects of steroid hormones on the apoptosis and IgM production of LPS-stimulated lymphocytes from peripheral blood, spleen and head kidney of the fugu were investigated. High dose of cortisol induced apoptosis in lymphocytes from all of those origins, while testosterone, 11-ketotestosterone and estradiol-17β did not. This result indicates that the elevation of cortisol due to stress causes immunosuppression in fugu. The rate of apoptosis in the lymphocytes from spleen and head kidney was higher than from peripheral blood lymphocytes, indicating that lymphocytes from different origin differ in their sensitivities to glucocorticoids. In addition, cortisol suppressed the expression of immunoglobulin M mRNA (IgM mRNA) in cultured lymphocytes within a short period. However, lipopolysaccharide (LPS) increased the expression of IgM mRNA in the spleen and head kidney, suggesting that stimulation of lymphocytes due to antigenic challenges might increase the transcription rate of IgM mRNA. However, when LPS-treated cells were cultured in the presence of cortisol, the transcription of IgM mRNA was again suppressed. This result indicates that cortisol is an important regulator of IgM mRNA transcription as well as IgM secretion. [ABSTRACT FROM AUTHOR]

We report herein the case of 64-year-old man with gastrointestinal stromal tumor (GIST), who was treated by partial resection of the duodenum after preoperative transarterial embolization. He presented to our hospital with a history of tarry stools, dizziness, and severe anemia (hemoglobin, 7.5 g/dl). Gastroduodenal endoscopy revealed the presence of a submucosal tumor in the second portion of the duodenum. The presence of the tumor was subsequently confirmed by double-contrast gastrointestinal radiography and abdominal computed tomography. Super-selective angiography showed tumor staining fed from the anterior and posterior superior pancreaticoduodenal arteries, and the inferior pancreaticoduodenal artery. Two weeks after transarterial embolization through these vessels, the tumor size was found to have shrunk to 40% of its original size. Partial resection of the duodenum was performed and absence of tumor cells at the surgical margin was confirmed by intraoperative frozen-section examination. Histopathological examination revealed that the duodenal submucosal tumor consisted of spindle cells, and immunohistochemical analysis revealed positive tumor staining for c-kit protein, CD34 and α-smooth muscle actin (SMA), and negative staining for desmin and S-100; the positivity rate for MIB-1 staining was 2.2%. Based on these findings, the tumor was diagnosed as a GIST of low-grade malignancy, classified as the muscular type. It is considered that preoperative treatment of duodenal GISTs, such as transarterial embolization, may be useful for reducing the extent of resection, from pancreaticoduodenenctomy to a partial resection. [ABSTRACT FROM AUTHOR]

CD3 is an essential component of the CD3-TCR complex. In this report, we describe the cloning, characterization, and expression analysis of the CD3? and CD3?/d chain genes from fugu,Takifugu rubripes. Two distinct CD3? homologue cDNAs, designated asCD3?-1andCD3?-2, and a CD3?/d homologue cDNA were isolated from the fugu thymus. The deduced amino acid sequences of these cDNAs exhibit conserved essential CD3 chain motifs and overall structures. RT-PCR analysis demonstrated that theCD3?andCD3?/dgenes were expressed in lymphoid organs (e.g. thymus, head kidney, trunk kidney and spleen), mucosal tissues (gill, skin, and intestine), and peripheral blood leucocytes (PBL). TheCD3andTCRagenes were expressed only in the surface IgM- population, which were separated from PBL using an anti-fugu IgM monoclonal antibody. In addition, in situ hybridization confirmed thatCD3-expressing cells were distributed randomly in the head kidney, trunk kidney, and spleen, but in the thymus were restricted to the lymphoid outer zone and epithelioid inner zone only. Collectively, these results suggest that CD3 molecules are useful markers for the identification of T cells in teleost fish. The present study thus provides a critical step in identifying T cells in this model organism. [ABSTRACT FROM AUTHOR]

We have isolated and characterized the cDNA that encodes IgD of fugu (Takifugu rubripes). Though the splicing of µ1 with the d1 domain was similar to those reported for teleost IgDs, highly unusual and unprecedented domain duplications were found in the constant region of the fugu IgD. The structure of the fugu IgD is like VDJ-µ1-(d1-d2-d3-d4-d5-d6)2-d7-dm1-dm2. Genomic sequence analysis of the fuguIgDgene supported the results of cDNA sequencing that the first six d domains in the constant region are duplicated. Such a novel duplication pattern has not been reported in any other vertebrates. However, IgD secretory domains could not be identified in this study. The deduced amino acid sequence of the fugu IgD constant region showed high identity (35-55%) to the sequences of previously reported teleost IgDs. Gene expression analyses based on RT-PCR demonstrated that theIgDgene is preferentially expressed in presumptive lymphoid tissues; moreover, in situ hybridization showed that IgD-positive cells are distributed throughout the spleen and head kidney. The expression pattern is similar to that of IgM, corroborating the hypothesis that IgD plays an important role in the humoral immune system of this species. [ABSTRACT FROM AUTHOR]

Presents a study that examined several restriction landmark genomic scanning spots that are commonly altered in hepatocellular carcinoma (HCC) profiles. Background on HCC; Multivariate analysis of the spots; Factors contributing to HCC.

SUMMARY: Common carp (Cyprinus carpio L.) were acclimated to 30°C from 20°C over 24 h and maintained at that temperature for 35 days. Changes in the expression of mRNA transcripts for fast myosin heavy chain (MyHC) isoforms, E12 of the E protein, and myogenic regulatory factors belonging to the MyoD (MyoD, myogenin, myf-5) and myocyte enhancer factor-2 (MEF2A, MEF2C) gene families were investigated in the fast myotomal muscle using northern blot analysis. Myofibrillar Mg2+-ATPase activity gradually declined over 21 days and was significantly different from the starting level after only 3–5 days. Over the same time period there was a dramatic decrease in the mRNA transcripts of the MyHC isoform predominantly expressed in cold-acclimated carp (10°C-type MyHC) and a significant increase in the transcripts of the MyHC isoform predominantly expressed in warm-acclimated carp (30°C-type MyHC). The levels of myogenin mRNA transcripts almost doubled over 3–7 days before declining to 30% of the starting levels for the remainder of the experiment. While the levels of E12 mRNA gradually decreased, MEF2A and MEF2C mRNA transcripts showed a significant decrease over 24 h and then stabilized at the lower levels. In contrast, the levels of MyoD mRNA were not affected by temperature acclimation. The possible role of myogenic regulatory factors in the expression of temperature-specific isoforms of MyHC in common carp is briefly discussed. [ABSTRACT FROM AUTHOR]

Molecular genetic analyses have clarified that accumulation of genomic changes provides important steps in carcinogenesis and have identified a number of valuable genetic markers for certain cancers. To date, however, no prognostic markers have been identified for hepatocellular carcinoma (HCC). In this study, we used restriction landmark genomic scanning (RLGS), a new high-speed screening method for multiple genomic changes, to detect unknown genetic alterations in HCC. Thirty-one HCC samples and their normal counterparts were examined by RLGS. Eight spot changes were common in several cases, and all were seen only on the HCC profile. Five of these spots were detected in more than 12 of 31 cases (38.7%). Viral infection had no influence on changes in the RLGS spots. The disease-free survival rate for patients with 16 changed RLGS spots was significantly lower than that for patients with fewer changed RLGS spots (15 spots) (P<0.001). In multivariate analysis, the number of changed spots was proven to retain an independent prognostic value (relative risk 1.095: P=0.0031). These results suggest that the number of changed RLGS spots may be a useful biological marker for recurrence of HCC. Oncogene (2000) 19, 1676–1683 [ABSTRACT FROM AUTHOR]

SUMMARY:A complementary DNA (cDNA) encoding salmon type gonadotropin-releasing hormone (sGnRH) has already been isolated and characterized in the goldfish Carassius auratus. In the present study, we isolated and characterized a second sGnRH cDNA which was different from the reported sGnRH cDNA from the brain of goldfish by rapid amplification of cDNA ends. This cDNA included an open reading frame of 282 bp encoding the 94 amino acid residues of sGnRH precursor. Amino acids identity between the second sGnRH precursor and the previously reported sGnRH precursor was 86.2%. Genomic Southern blot analysis showed that the goldfish harbors these two distinct sGnRH genes in its genome. Expressions of the two sGnRH genes were observed in the brain, pituitary, and gonad of goldfish by reverse transcription–polymerase chain reaction analysis. Moreover, the levels of two sGnRH mRNA in these tissues were different depending on sex and maturity of the fish. The differential expressions of two sGnRH genes suggest the existence of the differential regulatory mechanism of the two sGnRH genes and the multiple physiological functions of sGnRH. [ABSTRACT FROM AUTHOR]

Mammalian pancreatic ribonuclease (RNase) was conjugated chemically via a disulfide bond to human or murine epidermal growth factor (EGF). The conjugation between EGF and RNase was ascertained by SDS-PAGE using reduced and nonreduced conjugates. The EGF-RNase conjugate retained potent RNase activity and competed with 125I-EGF for binding to EGFR to the same extent as unconjugated EGF. Both the human and murine EGF-RNase conjugates showed dose-dependent cytotoxicity against EGFR-overexpressing A431 human squamous carcinoma cells with IC50 values of 3 x 10(-7) M and 6 x 10(-7) M, respectively, whereas free RNase had an IC50 of 10(-4) M. Against the EGFR-deficient small-cell lung cancer cell line H69, the EGF-RNase conjugate had no cytotoxic effect. The Human EGF-RNase conjugate showed dose-dependent cytotoxicity against other squamous carcinoma cell lines (TE-5, TE-1) and breast cancer cell lines (BT-20, SK-BR-3, MCF-7) and the cytotoxicity of the conjugate correlated positively with the level of expression of EGFR by each cell line. An unconjugated mixture of EGF and RNase had no greater effect than RNase alone on any cell line. Excess free EGF blocked EGF-RNase conjugate cytotoxicity against A431 cells. These results suggest that the EGF-RNase conjugate may be a more effective anticancer agent with less immunogenicity than coventional chimeric toxins. [ABSTRACT FROM AUTHOR]

In the original publication of the article, both the Fig. 2e and Fig. 7e have been published incorrectly and the correct figures are given below. [ABSTRACT FROM AUTHOR]

A correction to this paper has been published: https://doi.org/10.1007/s12562-021-01505-w [ABSTRACT FROM AUTHOR]

Parasite resistance traits in aquaculture species often have moderate heritability, indicating the potential for genetic improvements by selective breeding. However, parasite resistance is often synonymous with an undesirable negative correlation with body size. In this study, we first tested the feasibility of genomic selection (GS) on resistance to heterobothriosis, caused by the monogenean parasite Heterobothrium okamotoi, which leads to huge economic losses in aquaculture of the tiger pufferfish Takifugu rubripes. Then, using a simulation study, we tested the possibility of simultaneous improvement of parasite resistance, assessed by parasite counts on host fish (HC), and standard length (SL). Each trait showed moderate heritability (square-root transformed HC: h2 = 0.308 ± 0.123, S.E.; SL: h2 = 0.405 ± 0.131). The predictive abilities of genomic prediction among 12 models, including genomic Best Linear Unbiased Predictor (GBLUP), Bayesian regressions, and machine learning procedures, were also moderate for both transformed HC (0.248‒0.344) and SL (0.340‒0.481). These results confirmed the feasibility of GS for this trait. Although an undesirable genetic correlation was suggested between transformed HC and SL (rg = 0.228), the simulation study suggested the desired gains index can help achieve simultaneous genetic improvements in both traits. [ABSTRACT FROM AUTHOR]

We report the case of a 2-year-old boy who developed a small bowel intussusception during treatment failure of his first episode of nephrotic syndrome. Despite the absence of typical symptoms other than abdominal pain, the intussusception was diagnosed by ultrasonography and computed tomography and successfully reduced by air enema. No pathological lead point was discovered, and no symptoms of Henoch-Schönlein purpura developed later. Intussusception should be considered in the differential diagnosis of abdominal pain in patients with nephrotic syndrome, especially in patients exhibiting prolonged edema. Ultrasonography or computed tomography should be performed, even in the absence of other typical symptoms suggestive of intussusception. We should also bear in mind that the intussusception associated with nephrotic syndrome might occur at regions other than the typical ileocolic region, such as within the small intestine. [ABSTRACT FROM AUTHOR]

A case of cystadenocarcinoma of the appendix with a large cystic lesion is reported. A 49-year-old man undergoing a routine ultrasonic scan was incidentally found to have an abdominal mass measuring some 30 cm in size. The clinical presentation was asymptomatic, and the patient underwent a laparotomy without ascertaining a diagnosis preoperatively. The lesion, which derived from the appendix, was removed and was found to be cystic and contained huge amounts of mucin. The histological findings revealed a well-differentiated cystadenocarcinoma of the appendix, and immunohistochemical staining of the epithelium and mucinous implants in the mass demonstrated a positive reaction for carcinogenic antigens, including carcinoembryonic antigen and carbohydrate antigen. [ABSTRACT FROM AUTHOR]

Renal proximal tubular function was assessed in neonates by measuring urinary β-microglobulin (βM) concentrations on days 1, 4, 7, 14 and 28. Values were elevated in stable preterm low-birthweight (LBW) neonates but not in stable term LBW neonates, suggesting that proximal tubular maturation is related to gestational age rather than birthweight. The urinary βM was significantly increased on day 1 in neonates with the meconium aspiration syndrome but was not significantly different from normal subsequently. This indicated that although the proximal tubular cells may be susceptible to perinatal hypoxia, they maintain a remarkable capacity to recover in a relatively short period. Neonates with transient tachypnoea of the newborn had normal urinary levels of βM indicating their renal tubular function was not impaired. [ABSTRACT FROM AUTHOR]

Serum and tissue miR-21 expression in patients with breast cancer (BC) is a useful biomarker for cancer diagnosis, progression, and treatment. Matrix metalloproteinase-1 (MMP-1) is also important in breast cancer carcinogenesis. However, miR-21 and MMP-1/CD63 in urine exosomes in these patients have not been examined. Urine samples were collected from patients with BC and 26 healthy females. Urinary exosomes were isolated and confirmed by western blotting with anti-CD63 antibody and electron microscopy observation. MiR-21 and MMP-1/CD63 expression was examined by quantitative RT-PCR and western blotting, respectively. Patients with very early stage breast cancer were evaluated. MiR-21 expression in the patients was 0.26 [95% CI: 0.20–0.78], which was significant lower than in the 26 controls (1.00 [95% CI: 1.01–3.37], p = 0.0947). MMP-1/CD63 expression in patients was significantly higher than in controls (1.74 [95% CI: 0.86–5.08] vs 0.535 [95% CI: −0.01–2.81], p = 0.0001). Sensitivity and specificity were 0.708 and 0.783 for miR-21 and 0.792 and 0.840 for MMP-1/CD63, respectively. Sensitivity and specificity of combined expression were 95% and 79%, respectively. The sensitivity of MMP-1/CD63 expression in urinary exosomes was better than that of miR-21 expression. Thus, miR-21 and MMP/CD63 may be useful markers for BC screening. [ABSTRACT FROM AUTHOR]

Studies using genome-wide single nucleotide polymorphisms (SNPs) have become commonplace in genetics and genomics, due to advances in high-throughput sequencing technologies. Since the numbers of required SNPs and samples vary depending on each research goal, genotyping technologies with high flexibility in the number of SNPs/samples and high repeatability have been intensively investigated. For example, the ultrahigh-multiplexed amplicon sequencing, Ion AmpliSeq, has been used as a high-throughput genotyping method mainly for diagnostic purposes. Here, we designed a custom panel targeting 3,187 genome-wide SNPs of fugu, Takifugu rubripes, and applied it for genotyping farmed fugu to test its feasibility in aquaculture studies. We sequenced two libraries consisting of different pools of individuals (n = 326 each) on the Illumina MiSeq sequencer. Consequently, over 99% target regions (3,178 SNPs) were amplified and 2,655 SNPs were available after filtering steps. Strong correlation was observed in the mean depth of coverage of each SNP between duplicate runs (r = 0.993). Genetic analysis using these genotype data successfully detected the known population structure and the sex determining locus of fugu. These results show the method is superior in repeatability and flexibility, and suits genetic studies including molecular breeding, such as marker assisted and genomic selection. [ABSTRACT FROM AUTHOR]

Uric acid (UA) therapy may prevent early ischemic worsening after acute stroke in thrombolysis patients. The aim of this study was to examine the influence of UA on the thrombolytic efficacy of alteplase in human blood samples by measuring thrombolysis under flow conditions using a newly developed microchip-based flow-chamber assay. Human blood samples from healthy volunteers were exposed to UA, alteplase, or a combination of UA and alteplase. Whole blood and platelet-rich plasma were perfused over a collagen- and thromboplastin-coated microchip, and capillary occlusion was monitored with a video microscope and flow-pressure sensor. The area under the curve (extent of thrombogenesis or thrombolysis) at 30 minutes was 92% lower in the UA-alteplase-treated group compared with the alteplase-treated group. D-dimers were measured to evaluate these effects in human platelet-poor plasma samples. Although hydrogen peroxide significantly decreased the elevation of D-dimers by alteplase, UA significantly inhibited the effect of hydrogen peroxide. Meanwhile, rat models of thromboembolic cerebral ischemia were treated with either alteplase or UA-alteplase combination therapy. Compared with alteplase alone, the combination therapy reduced the infarct volume and inhibited haemorrhagic transformation. UA enhances alteplase-mediated thrombolysis, potentially by preventing oxidative stress, which inhibits fibrinolysis by alteplase in thrombi. [ABSTRACT FROM AUTHOR]

We report on an 11-year-old boy who developed rhabdomyolysis and acute renal failure following Salmonella enteritidis gastroenteritis. Rhabdomyolysis should be considered as a potentially fatal complication in patients with Salmonella gastroenteritis. [ABSTRACT FROM AUTHOR]