The interaction between 20-hydroxyecdysone and AMPK through PI3K activation in Chinese mitten crab, Eriocheir sinensis.

In crustaceans, the steroid hormone 20-hydroxyecdysone (20E) initiates molting, and the molting process is also regulated by energy metabolism. AMPK is an energy sensor and plays a critical role in systemic energy balance. Here, the regulatory mechanism in the interaction between 20E and AMPK was investigated in Chinese mitten crab, Eriocheir sinensis. The results showed that the 20E concentration and the mRNA expression levels of 20E receptors in hepatopancreas were down-regulated post AMPK activator (AICAR) treatment, and were up-regulated after AMPK inhibitor (Compound C) injection in crabs. Besides, the molt-inhibiting hormone (MIH) gene expression in eyestalk showed the opposite patterns in response to the AICAR and Compound C treatment, respectively. Further investigation found that there was a significant reduction in 20E concentration post PI3K inhibitor (LY294002) treatment, and the phosphorylation level of PI3K was increased in hepatopancreas after AMPK inhibitor injection. On the other hand, the positive regulation of PI3K-mediated activation of AMPK was also observed, the phosphorylation levels of AMPKα, AMPKß and PI3K in hepatopancreas were significantly increased post 20E injection. In addition, the phosphorylation levels of AMPKα and AMPKß induced by 20E were decreased after the injection of PI3K inhibitor. Taken together, these results suggest that the regulatory cross-talk between 20E and AMPK is likely to act through PI3K pathway in E. sinensis, which appeared to be helpful for a better understanding in molting regulation.

Effect of three unsaturated fatty acids on the protein oxidation and structure of myofibrillar proteins from rainbow trout (Oncorhynchus mykiss).

In this study, the impact of three unsaturated fatty acids (Oleic acid: OA, Eicosapentaenoic acid: EPA, Docosahexaenoic acid: DHA) on the oxidation and structure of rainbow trout myofibrillar protein (MP) was explored. The findings revealed a notable increase in carbonyl content (P < 0.05) and a significant decrease in total sulfhydryl content (P < 0.05) of MP with the concentration increase of the three unsaturated fatty acids. Endogenous fluorescence spectroscopy and surface hydrophobicity analyses showed that unsaturated fatty acids can cause unfolding and exposure of hydrophobic groups in MP. In addition, SDS-PAGE showed that disulfide bonds were associated with MP cross-linking and aggregate size induced by unsaturated fatty acids. Overall, three unsaturated fatty acid treatments facilitated the oxidation of myofibrillar proteins, and the extent of protein oxidation was closely associated with the concentration of unsaturated fatty acids.

On-site measurement and environmental impact of vibration caused by construction of double-shield TBM tunnel in urban subway.

The vibration generated during the construction of subway tunnels with double-shield tunnel boring machine (TBM) has a significant impact on the environment, which has caused multiple complaints from residents. Taking a double-shield TBM tunnel project as the background, vibration measurements were conducted by installing vibration sensors on-site. By combining theoretical methods-such as normalization, polynomial fitting prediction, and gray correlation analysis-the vibration characteristics, impact range on the environment, and factors affecting the vibration of TBM construction were studied. The key research results included: (1) The amplified zone of X and Y vibration acceleration occurred on the left-hand side of the tunnel from 3.15 to 13.85 m, but rapidly decayed away from the amplification zone. (2) The impact range of TBM vibrations on residential areas at night and during the day was studied according to the official "Urban Regional Environmental Vibration Standard" and it was found to be larger at night than during the day. (3)The main factors affecting the TBM vibration level was studied-including the cutter-head torque, TBM thrust, cutter-head speed, penetration, field penetration index (FPI) and so on. In summary, when the double-shield TBM construction tunnel is adjacent to residential areas, the vibration generated exceeds the national standard limit. In order to reduce the impact of TBM vibration on residential areas, excavation parameters such as cutter head torque, TBM thrust, cutter head speed, and penetration should be appropriately reduced.

Identification of quorum sensing-regulated Vibrio fortis as potential pathogenic bacteria for coral bleaching and the effects on the microbial shift.

Coastal pollution, global warming, ocean acidification, and other reasons lead to the imbalance of the coral reef ecosystem, resulting in the increasingly serious problem of coral degradation. Coral bleaching is often accompanied by structural abnormalities of coral symbiotic microbiota, among which Vibrio is highly concerned. In this study, Vibrio fortis S10-1 (MCCC 1H00104), isolated from sea cucumber, was used for the bacterial infection on coral Seriatopora guttatus and Pocillopora damicornis. The infection of S10-1 led to coral bleaching and a significant reduction of photosynthetic function in coral holobiont, and the pathogenicity of V. fortis was regulated by quorum sensing. Meanwhile, Vibrio infection also caused a shift of coral symbiotic microbial community, with significantly increased abundant Proteobacteria and Actinobacteria and significantly reduced abundant Firmicutes; on genus level, the abundance of Bacillus decreased significantly and the abundance of Rhodococcus, Ralstonia, and Burkholderia-Caballeronia-Paraburkholderia increased significantly; S10-1 infection also significantly impacted the water quality in the micro-ecosystem. In contrast, S10-1 infection showed less effect on the microbial community of the live stone, which reflected that the microbes in the epiphytic environment of the live stone might have a stronger ability of self-regulation; the algal symbionts mainly consisted of Cladocopium sp. and showed no significant effect by the Vibrio infection. This study verified that V. fortis is the primary pathogenic bacterium causing coral bleaching, revealed changes in the microbial community caused by its infection, provided strong evidence for the "bacterial bleaching" hypothesis, and provided an experimental experience for the exploration of the interaction mechanism among microbial communities, especially coral-associated Vibrio in the coral ecosystem, and potential probiotic strategy or QS regulation on further coral disease control.

The involvement of hypoxia inducible factor-1α on the proportion of three types of haemocytes in Chinese mitten crab under hypoxia stress.

Hypoxia triggers diverse cell physiological processes, and the hypoxia inducible factors (HIFs) are a family of heterodimeric transcription factors that function as master regulators to respond to hypoxia in different cells. However, the knowledge about the hypoxic responses especially cell alteration mediated by HIFs under hypoxia stress is still limited in crustaceans. In the present study, a hypoxia-inducible factor-1α (HIF-1α) gene was identified (designed as EsHIF-1α). The relative mRNA expression level of EsHIF-1α was highest in hyalinocytes and lowest in granulocytes among three types of haemocytes in crabs. Hypoxia could significantly increase the EsHIF-1α protein expression level in haemocytes. Meanwhile, the proportion of hyalinocytes began to increase from 3 h post hypoxia treatment, and reached the highest level at 24 h. However, the opposite variation in proportion of granulocytes was observed under hypoxia stress. Further investigation showed that the inhibition of EsHIF-1α induced by KC7F2 (HIF-1α inhibitor) could lead to the significant decrease in the proportion of hyalinocytes under hypoxia stress, and also resulted in an increase of granulocytes proportion. While, after EsHIF-1α was activated by IOX4 (HIF-1α activator), the proportion of hyalinocytes was significantly up-regulated and the proportion of granulocytes was significantly down-regulated under post hypoxia treatment. These results collectively suggested that EsHIF-1α was involved in the regulation of proportion of three types of haemocytes induced by hypoxia stress, which provided vital insight into the understanding of the crosstalk between hypoxia and cell development in invertebrates.

ERK is involved in the regulation of CpG ODN 2395 on the expression levels of anti-lipopolysaccharide factors in Chinese mitten crab, Eriocheir sinensis.

CpG oligodeoxynucleotides (ODN), as an effective adjuvant or immunopotentiator, activate the immune system and induce various immune responses. Recently, it has also been reported that high dose of CpG ODN can lead to immunosuppression. However, the underlying mechanism of CpG ODN-mediated immune response remains largely unknown in invertebrates. In the present study, the role of ERK in regulating expression levels of anti-lipopolysaccharide factors (ALFs) induced by different doses of CpG ODN 2395 was analyzed in Chinese mitten crab, Eriocheir sinensis. The mRNA expression levels of EsALFs (EsALF1, EsALF2 and EsALF3) and EsERK in haemocytes were observed to increase from 6 h to 48 h post low doses of CpG ODN 2395 (0.5 µg and 2.5 µg) stimulation, while they were suppressed after high dose of CpG ODN 2395 (12.5 µg) injection. Meanwhile, the phosphorylation levels of ERK in haemocytes were significantly promoted after low doses of CpG ODN 2395 injection, and a reduce level of ERK phosphorylation was observed after high dose of CpG ODN 2395 injection. Further investigation showed that the expression levels of EsALFs induced by CpG ODN 2395 were markedly down-regulated after knocking down the expression of EsERK. Similarly, the EsALFs mRNA expression were also inhibited post different doses of CpG ODN 2395 stimulation in PD98059 (ERK inhibitor) injection crabs. These results collectively suggest that ERK is involved in regulating the expression level of EsALFs induced by different dose of CpG ODN 2395 in Chinese mitten crab, which contribute to the understanding of the regulation of CpG ODN involving in immune response in crustaceans.

Incorporation of Al2O3, GO, and Al2O3@GO nanoparticles into water-borne epoxy coatings: abrasion and corrosion resistance.

Nano-Al2O3 particles and graphene oxide (GO) nanosheets were modified by 3-aminopropyltriethoxysilane (KH550), and then dispersed in epoxy resin, and finally modified-Al2O3/epoxy, modified-GO/epoxy and modified-Al2O3@GO/epoxy composite coatings were prepared on steel sheets by the scraping stick method. The microstructure, phase identification, surface bonding and composition of the nanoparticles were characterized by SEM, XRD, FT-IR, and Raman spectroscopy, respectively. The hardness of the coating was assessed by the pencil hardness method. The abrasion resistance of the coating was tested by a sand washing machine. The corrosion resistance of the coating was assessed using salt spray, a long-period immersion test, potentiodynamic polarization curves and electrochemical impedance spectra. With the addition of a small amount of nanoparticles, the dispersion of nanoparticles in the epoxy resin was good. When the content of nano-Al2O3 particles was equal to 1.5 wt%, the particles in the epoxy exhibited the best dispersion and stability. However, the GO and Al2O3@GO nanofillers in the epoxy resin exhibited poor dispersion and stability. The hardness, abrasion and corrosion resistance of the composite coatings were improved with the addition of a small amount of nanoparticles, but the performance began to decline after exceeding a certain content range of the nanoparticles. A relatively good abrasion resistance for the coatings was obtained when the content of Al2O3, GO and Al2O3@GO after modification was 1.5 wt%, 0.2 wt% and 0.4 wt%, respectively. The corrosion resistance of the coatings doped with nano-Al2O3 particles was better than that of the coatings incorporating GO nanosheets and Al2O3@GO hybrids. The corrosion mechanism of the composite coatings in 3.5 wt% NaCl solution was addressed and studied.

An integrated AI model to improve diagnostic accuracy of ultrasound and output known risk features in suspicious thyroid nodules.

OBJECTIVES: From the viewpoint of ultrasound (US) physicians, an ideal thyroid US computer-assisted diagnostic (CAD) system for thyroid cancer should perform well in suspicious thyroid nodules with atypical risk features and be able to output explainable results. This study aims to develop an explainable US CAD model for suspicious thyroid nodules. METHODS: A total of 2992 solid or almost-solid thyroid nodules were analyzed retrospectively. All nodules had pathological results (1070 malignancies and 1992 benignities) confirmed by ultrasound-guided fine-needle aspiration cytology and histopathology after thyroidectomy. A deep learning model (ResNet50) and a multiple risk features learning ensemble model (XGBoost) were used to train the US images of 2794 thyroid nodules. Then, an integrated AI model was generated by combining both models. The diagnostic accuracies of the three AI models (ResNet50, XGBoost, and the integrated model) were predicted in a testing set including 198 thyroid nodules and compared to the diagnostic efficacy of five ultrasonographers. RESULTS: The accuracy of the integrated model was 76.77%, while the mean accuracy of the ultrasonographers was 68.38%. Of the risk features, microcalcifications showed the highest contribution to the diagnosis of malignant nodules. CONCLUSIONS: The integrated AI model in our study can improve the diagnostic accuracy of suspicious thyroid nodules and output the known risk features simultaneously, thus aiding in training young ultrasonographers by linking the explainable results to their clinical experience and advancing the acceptance of AI diagnosis for thyroid cancer in clinical practice. KEY POINTS: • We developed an artificial intelligence (AI) diagnosis model based on both deep learning and multiple risk feature ensemble learning methods. • The AI diagnosis model showed higher diagnostic accuracy for suspicious thyroid nodules than ultrasonographers. • The AI diagnosis model showed partial explainability by outputting the known risk features, thus aiding young ultrasonic doctors in increasing the diagnostic level for thyroid cancer.

Magnetoactive acoustic metamaterials based on nanoparticle-enhanced diaphragm.

Magnetoactive membrane-type acoustic metamaterials are fabricated by coating a layer of magnetic nanoparticles on the polyethylene (PE) membranes and their vibration characters are investigated experimentally. From our experiments, we discovered that, under different magnetic fields by varying the distance between a magnet and the membranes, such membranes exhibit tunable vibration eigenfrequencies (the shift towards lower frequencies), which is caused by the variation of the effective mass density and effective tension coefficient resulted from the second derivative of the magnetic field. The strong magnetic force between the layer of magnetic nanoparticles and the magnet enhances the eigenfrequency shift. A spring oscillator model is proposed and it agrees well with the experimental results. We also experimentally observed that the vibration radius, effective mass density, and effective tension coefficient of the membranes can enormously affect the eigenfrequencies of the membranes. We believe that this type of metamaterials may open up some potential applications for acoustic devices with turntable vibration properties.

Identification of distinct clinical phenotypes of acute respiratory distress syndrome with differential responses to treatment.

BACKGROUND: Acute respiratory distress syndrome (ARDS) is a heterogeneous syndrome, and the identification of homogeneous subgroups and phenotypes is the first step toward precision critical care. We aimed to explore whether ARDS phenotypes can be identified using clinical data, are reproducible and are associated with clinical outcomes and treatment response. METHODS: This study is based on a retrospective analysis of data from the telehealth intensive care unit (eICU) collaborative research database and three ARDS randomized controlled trials (RCTs) (ALVEOLI, FACTT and SAILS trials). We derived phenotypes in the eICU by cluster analysis based on clinical data and compared the clinical characteristics and outcomes of each phenotype. The reproducibility of the derived phenotypes was tested using the data from three RCTs, and treatment effects were evaluated. RESULTS: Three clinical phenotypes were identified in the training cohort of 3875 ARDS patients. Of the three phenotypes identified, phenotype I (n = 1565; 40%) was associated with fewer laboratory abnormalities, less organ dysfunction and the lowest in-hospital mortality rate (8%). Phenotype II (n = 1232; 32%) was correlated with more inflammation and shock and had a higher mortality rate (18%). Phenotype III (n = 1078; 28%) was strongly correlated with renal dysfunction and acidosis and had the highest mortality rate (22%). These results were validated using the data from the validation cohort (n = 3670) and three RCTs (n = 2289) and had reproducibility. Patients with these ARDS phenotypes had different treatment responses to randomized interventions. Specifically, in the ALVEOLI cohort, the effects of ventilation strategy (high PEEP vs low PEEP) on ventilator-free days differed by phenotype (p = 0.001); in the FACTT cohort, there was a significant interaction between phenotype and fluid-management strategy for 60-day mortality (p = 0.01). The fluid-conservative strategy was associated with improved mortality in phenotype II but had the opposite effect in phenotype III. CONCLUSION: Three clinical phenotypes of ARDS were identified and had different clinical characteristics and outcomes. The analysis shows evidence of a phenotype-specific treatment benefit in the ALVEOLI and FACTT trials. These findings may improve the identification of distinct subsets of ARDS patients for exploration in future RCTs.

Artificial Intelligence-Aided Diagnosis Software to Identify Highly Suspicious Pulmonary Nodules.

INTRODUCTION: To evaluate the value of artificial intelligence (AI)-assisted software in the diagnosis of lung nodules using a combination of low-dose computed tomography (LDCT) and high-resolution computed tomography (HRCT). METHOD: A total of 113 patients with pulmonary nodules were screened using LDCT. For nodules with the largest diameters, an HRCT local-target scanning program (combined scanning scheme) and a conventional-dose CT scanning scheme were also performed. Lung nodules were subjectively assessed for image signs and compared by size and malignancy rate measured by AI-assisted software. The nodules were divided into improved visibility and identical visibility groups based on differences in the number of signs identified through the two schemes. RESULTS: The nodule volume and malignancy probability for subsolid nodules significantly differed between the improved and identical visibility groups. For the combined scanning protocol, we observed significant between-group differences in subsolid nodule malignancy rates. CONCLUSION: Under the operation and decision of AI, the combined scanning scheme may be beneficial for screening high-risk populations.

Identification of genes involved in gonadal sex differentiation and the dimorphic expression pattern in Takifugu rubripes gonad at the early stage of sex differentiation.

Quantifying the expression of mRNAs in the gonads at the critical stage of molecular sex differentiation stage might help to clarify the regulatory network during early sex differentiation and provide new information on the role of sex-related genes in gonadal function. In this study, transcriptomic analysis of sex-related genes expression profiles in fugu gonads at 60 and 90 days after hatching (dah) was conducted firstly, and a total of 112,504,991 clean reads, encompassing 28.35 Gb of sequences were retrieved. Twenty-three thousand eight hundred ten genes were found to be expressed in juvenile fugu gonads, and we mainly focused on the differentially expressed genes that have the potential to be involved in the gonadal sex differentiation. For 60-dah juveniles, we identified 1014 genes that were upregulated in the ovary and 1570 that were upregulated in the testis. For 90-dah juveniles, we identified 1287 genes that were upregulated in the ovary and 1500 that were upregulated in the testis. The dimorphic expression patterns of 15 genes in gonads at 30 and 40 dah were further investigate using qPCR. Cyp11b and star were expressed at higher levels in XY than in XX, while cyp11a1 and cyp19a1a were expressed at higher levels in XX than in XY at 30 dah. At 40 dah, the levels of gsdf, dmrt1, dmrt3, cyp11c1, star, and hsd3b expression were higher in XY, while the levels of foxl2, cyp19a1a, wnt9b, and foxD4 expression were higher in XX. Sox9, cyp11a1, cyp17a1, cyp17a2, and nr5a2 were expressed at similar levels in XX and XY at 40 dah. This is the first report of gonadal transcriptome of fugu at early sex differentiation stage, and our results provide an archive for further study on molecular mechanism underlying sex differentiation in this species.

Regioisomeric Tandem Triblock Shape Amphiphiles Based on Polyhedral Oligomeric Silsesquioxanes.

We report the design, synthesis and self-assembly of a series of tandem triblock shape amphiphile regioisomers composed of one rod-like fluorinated polyhedral oligomeric silsesquioxane (FPOSS) cage tethered with two spherical tert-butyl-POSS (tBPOSS) cages in para-, meta-, and ortho-configurations. Precision syntheses are achieved through sequential "click" reactions and the compounds are thoroughly characterized by combined techniques of NMR, FT-IR and MALDI-TOF MS spectroscopy. Only the ortho-isomer forms an ordered lamellae structure induced by the crystallization of perfluoroalkyl-chains and the other two remain amorphous due to frustrated packing by unfavorable regio-configuration. The distinct assembly behaviors of these three regioisomers reflect the delicate influence of a minute structural difference on precision macromolecular assembly.

On-chip DNA preconcentration in different media conductivities by electrodeless dielectrophoresis.

Electrodeless dielectrophoresis is the best choice to achieve preconcentration of nanoparticles and biomolecules due to its simple, robust, and easy implementation. We designed a simple chip with microchannels and nano-slits in between and then studied the trapping of DNA in high conductive medium and low conductive medium, corresponding to positive and negative dielectrophoresis (DEP), respectively. It is very important to investigate the trapping in media with different conductivities since one always has to deal with the sample solutions with different conductivities. The trapping process was analyzed by the fluorescent intensity changes. The results showed that DNA could be trapped at the nano-slit in both high and low conductive media in a lower electric field strength (10 V/cm) compared to the existing methods. This is a significant improvement to suppress the Joule heating effect in DEP related experiments. Our work may give insight to researchers for DNA trapping by a simple and low cost device in the Lab-on-a-Chip system.

Applications of micro/nanoparticles in microfluidic sensors: a review.

This paper reviews the applications of micro/nanoparticles in microfluidics device fabrication and analytical processing. In general, researchers have focused on two properties of particles--electric behavior and magnetic behavior. The applications of micro/nanoparticles could be summarized on the chip fabrication level and on the processing level. In the fabrication of microfluidic chips (chip fabrication level), particles are good additives in polydimethylsiloxane (PDMS) to prepare conductive or magnetic composites which have wide applications in sensors, valves and actuators. On the other hand, particles could be manipulated according to their electric and magnetic properties under external electric and magnetic fields when they are travelling in microchannels (processing level). Researchers have made a great progress in preparing modified PDMS and investigating the behaviors of particles in microchannels. This article attempts to present a discussion on the basis of particles applications in microfluidics.

Identification of an L-phenylalanine binding site enhancing the cooperative responses of the calcium-sensing receptor to calcium.

Functional positive cooperative activation of the extracellular calcium ([Ca(2+)]o)-sensing receptor (CaSR), a member of the family C G protein-coupled receptors, by [Ca(2+)]o or amino acids elicits intracellular Ca(2+) ([Ca(2+)]i) oscillations. Here, we report the central role of predicted Ca(2+)-binding site 1 within the hinge region of the extracellular domain (ECD) of CaSR and its interaction with other Ca(2+)-binding sites within the ECD in tuning functional positive homotropic cooperativity caused by changes in [Ca(2+)]o. Next, we identify an adjacent L-Phe-binding pocket that is responsible for positive heterotropic cooperativity between [Ca(2+)]o and L-Phe in eliciting CaSR-mediated [Ca(2+)]i oscillations. The heterocommunication between Ca(2+) and an amino acid globally enhances functional positive homotropic cooperative activation of CaSR in response to [Ca(2+)]o signaling by positively impacting multiple [Ca(2+)]o-binding sites within the ECD. Elucidation of the underlying mechanism provides important insights into the longstanding question of how the receptor transduces signals initiated by [Ca(2+)]o and amino acids into intracellular signaling events.

Elucidation of a novel extracellular calcium-binding site on metabotropic glutamate receptor 1{alpha} (mGluR1{alpha}) that controls receptor activation.

Metabotropic glutamate receptor 1α (mGluR1α) exerts important effects on numerous neurological processes. Although mGluR1α is known to respond to extracellular Ca(2+) ([Ca(2+)](o)) and the crystal structures of the extracellular domains (ECDs) of several mGluRs have been determined, the calcium-binding site(s) and structural determinants of Ca(2+)-modulated signaling in the Glu receptor family remain elusive. Here, we identify a novel Ca(2+)-binding site in the mGluR1α ECD using a recently developed computational algorithm. This predicted site (comprising Asp-318, Glu-325, and Asp-322 and the carboxylate side chain of the receptor agonist, Glu) is situated in the hinge region in the ECD of mGluR1α adjacent to the reported Glu-binding site, with Asp-318 involved in both Glu and calcium binding. Mutagenesis studies indicated that binding of Glu and Ca(2+) to their distinct but partially overlapping binding sites synergistically modulated mGluR1α activation of intracellular Ca(2+) ([Ca(2+)](i)) signaling. Mutating the Glu-binding site completely abolished Glu signaling while leaving its Ca(2+)-sensing capability largely intact. Mutating the predicted Ca(2+)-binding residues abolished or significantly reduced the sensitivity of mGluR1α not only to [Ca(2+)](o) and [Gd(3+)](o) but also, in some cases, to Glu. The dual activation of mGluR1α by [Ca(2+)](o) and Glu has important implications for the activation of other mGluR subtypes and related receptors. It also opens up new avenues for developing allosteric modulators of mGluR function that target specific human diseases.

Characterization of a Rel\NF-kappaB homologue in a gastropod abalone, Haliotis diversicolor supertexta.

Rel\NF-kappaB signal transduction pathway is evolutionarily conserved and involved in numerous biological processes. We report here, for the first time that a homologue of Rel\NF-kappaB transcription factor, Ab-Rel, was identified and functionally characterized in a gastropod abalone, H. diversicolor supertexta. The full-length Ab-Rel cDNA consists of 1943 bp with an ORF encoding a 584 amino acids protein. Amino acid sequence analysis revealed that Ab-Rel shares conserved signature motifs with other Rel proteins, including the Rel homology domain (RHD), nuclear localization signal (NLS) and phosphorylation site, RRPS. Northern hybridization and real-time PCR indicated that Ab-Rel was ubiquitously and constitutively expressed in abalone. The recombinant Ab-Rel RHD protein was confirmed to specifically bind the consensus NF-kappaB binding site. Furthermore, EMSA showed that NF-kappaB activity was induced in abalone hemocytes by stimulation with LPS. These results strongly suggest that Ab-Rel is a Rel homologue, which plays a conserved role in the immune response of the ancient invertebrate, abalone, allowing us to study the Rel\NF-kappaB signaling pathway in an evolutionary context.

Magnetocapture of abalone transcription factor NF-kappaB: a new strategy for isolation and detection of NF-kappaB both in vitro and in vivo.

Rel\NF-kappaB plays an important role in the immune response. Recently, a homologue of Rel\NF-kappaB transcription factor was identified from a gastropod abalone, Haliotis diversicolor supertexta, named Ab-Rel. We present here a new strategy, so called magnetocapture, based on magnetic separation that was developed to isolate and detect this abalone transcriptional factor. With this method, Ab-Rel was successfully isolated from abalone haemocytes and the recombinant Ab-Rel protein from an insect cell line. Western blot analysis was performed to verify the effectiveness of this magnetocapture assay. Since only the activated NF-kappaB can be captured and separated from the whole cell extracts, this method was simultaneously used to detect the NF-kappaB activity in vivo, as well as the biological activity of the recombinant protein in vitro. In addition, several proteins which might interact with the abalone NF-kappaB were together isolated, suggesting that the magnetocapture method would be useful for identifying interaction molecules. These results illustrated that the magnetocapture strategy is rapid, reliable and versatile, providing a promising tool for studying Rel\NF-kappaB protein.

A novel RUNX2 missense mutation predicted to disrupt DNA binding causes cleidocranial dysplasia in a large Chinese family with hyperplastic nails.

BACKGROUND: Cleidocranial dysplasia (CCD) is a dominantly inherited disease characterized by hypoplastic or absent clavicles, large fontanels, dental dysplasia, and delayed skeletal development. The purpose of this study is to investigate the genetic basis of Chinese family with CCD. METHODS: Here, a large Chinese family with CCD and hyperplastic nails was recruited. The clinical features displayed a significant intrafamilial variation. We sequenced the coding region of the RUNX2 gene for the mutation and phenotype analysis. RESULTS: The family carries a c.T407C (p.L136P) mutation in the DNA- and CBFbeta-binding Runt domain of RUNX2. Based on the crystal structure, we predict this novel missense mutation is likely to disrupt DNA binding by RUNX2, and at least locally affect the Runt domain structure. CONCLUSION: A novel missense mutation was identified in a large Chinese family with CCD with hyperplastic nails. This report further extends the mutation spectrum and clinical features of CCD. The identification of this mutation will facilitate prenatal diagnosis and preimplantation genetic diagnosis.