Integrated analysis of mRNAs and lncRNAs reveals candidate marker genes and potential hub lncRNAs associated with growth regulation of the Pacific Oyster, Crassostrea gigas.

BACKGROUND: The Pacific oyster, Crassostrea gigas, is an economically important shellfish around the world. Great efforts have been made to improve its growth rate through genetic breeding. However, the candidate marker genes, pathways, and potential lncRNAs involved in oyster growth regulation remain largely unknown. To identify genes, lncRNAs, and pathways involved in growth regulation, C. gigas spat was cultured at a low temperature (15 ℃) to yield a growth-inhibited model, which was used to conduct comparative transcriptome analysis with spat cultured at normal temperature (25 ℃). RESULTS: In total, 8627 differentially expressed genes (DEGs) and 1072 differentially expressed lncRNAs (DELs) were identified between the normal-growth oysters (cultured at 25 ℃, hereinafter referred to as NG) and slow-growth oysters (cultured at 15 ℃, hereinafter referred to as SG). Functional enrichment analysis showed that these DEGs were mostly enriched in the AMPK signaling pathway, MAPK signaling pathway, insulin signaling pathway, autophagy, apoptosis, calcium signaling pathway, and endocytosis process. LncRNAs analysis identified 265 cis-acting pairs and 618 trans-acting pairs that might participate in oyster growth regulation. The expression levels of LNC_001270, LNC_003322, LNC_011563, LNC_006260, and LNC_012905 were inducible to the culture temperature and food abundance. These lncRNAs were located at the antisense, upstream, or downstream of the SREBP1/p62, CDC42, CaM, FAS, and PIK3CA genes, respectively. Furthermore, the expression of the trans-acting lncRNAs, including XR_9000022.2, LNC_008019, LNC_015817, LNC_000838, LNC_00839, LNC_011859, LNC_007294, LNC_006429, XR_002198885.1, and XR_902224.2 was also significantly associated with the expression of genes enriched in AMPK signaling pathway, insulin signaling pathway, autophagy, apoptosis, calcium signaling pathway, and endocytosis process. CONCLUSIONS: In this study, we identified the critical growth-related genes and lncRNAs that could be utilized as candidate markers to illustrate the molecular mechanisms underlying the growth regulation of Pacific oysters.

MSCs-engineered biomimetic PMAA nanomedicines for multiple bioimaging-guided and photothermal-enhanced radiotherapy of NSCLC.

BACKGROUND: The recently developed biomimetic strategy is one of the mostly effective strategies for improving the theranostic efficacy of diverse nanomedicines, because nanoparticles coated with cell membranes can disguise as "self", evade the surveillance of the immune system, and accumulate to the tumor sites actively. RESULTS: Herein, we utilized mesenchymal stem cell memabranes (MSCs) to coat polymethacrylic acid (PMAA) nanoparticles loaded with Fe(III) and cypate-an derivative of indocyanine green to fabricate Cyp-PMAA-Fe@MSCs, which featured high stability, desirable tumor-accumulation and intriguing photothermal conversion efficiency both in vitro and in vivo for the treatment of lung cancer. After intravenous administration of Cyp-PMAA-Fe@MSCs and Cyp-PMAA-Fe@RBCs (RBCs, red blood cell membranes) separately into tumor-bearing mice, the fluorescence signal in the MSCs group was 21% stronger than that in the RBCs group at the tumor sites in an in vivo fluorescence imaging system. Correspondingly, the T1-weighted magnetic resonance imaging (MRI) signal at the tumor site decreased 30% after intravenous injection of Cyp-PMAA-Fe@MSCs. Importantly, the constructed Cyp-PMAA-Fe@MSCs exhibited strong photothermal hyperthermia effect both in vitro and in vivo when exposed to 808 nm laser irradiation, thus it could be used for photothermal therapy. Furthermore, tumors on mice treated with phototermal therapy and radiotherapy shrank 32% more than those treated with only radiotherapy. CONCLUSIONS: These results proved that Cyp-PMAA-Fe@MSCs could realize fluorescence/MRI bimodal imaging, while be used in phototermal-therapy-enhanced radiotherapy, providing desirable nanoplatforms for tumor diagnosis and precise treatment of non-small cell lung cancer.

Crosstalk between dopamine and insulin signaling in growth control of the oyster.

Neuroendocrine hormones such as dopamine and insulin/insulin-like peptides play indispensable roles in growth regulation of animals, while the interplay between dopamine and insulin signaling pathways remains largely unknown in invertebrates. In the present study, we showed that tyrosine hydroxylase (TH), the rate-limiting enzyme of dopamine synthesis, was highly expressed in all tissues of the fast-growing oysters, and gradually increased with the development, which indicated the potential role of dopamine in growth regulation. Incubated with dopamine hydrochloride and insulin-like peptide recombinant proteins in vitro induced the expression of TH, suggesting a mutual regulatory relationship between insulin and dopamine signaling. Fasting and re-feeding experiments confirmed the role of TH in food intake regulation, also provide a clue about the potential regulatory relationship between the FoxO and TH. Further luciferase assay experiment confirmed that FoxO was involved in transcriptional regulation of TH gene through binding to its specific promoter region. This work provided insights into the crosstalk between dopamine and insulin signaling in growth control of mollusks.

Transient Receptor Potential (TRP) Channels in the Pacific Oyster (Crassostrea gigas): Genome-Wide Identification and Expression Profiling after Heat Stress between C. gigas and C. angulata.

Transmembrane proteins are involved in an array of stress responses, particularly in thermo-sensation and thermo-regulation. In this study, we performed a genome-wide identification and characterization of the Transient Receptor Potential (TRP) genes in the Pacific oyster (Crassostrea gigas) and investigated their expression profiles after heat stress to identify critical TRPs potentially associated with thermal regulation. A total of 66 TRP genes were identified in the C. gigas, which showed significant gene expansion and tandem duplication. Meta-analysis of the available RNA-Seq data generated from samples after acute heat stress revealed a set of heat-inducible TRPs. Further examination of their expression profiles under chronic heat stress, and comparison between C. gigas and C. angulata, two oyster species with different tolerance levels to heat stress, led to the identification of TRPC3.6, TRPC3.7, and TRPV4.7 as important TRPs involved in thermal regulation in oysters. This work provided valuable information for future studies on the molecular mechanism of TRP mediated thermal tolerance, and identification of diagnostic biomarker for thermal stress in the oysters.

Insulin-Like Peptide Receptor-Mediated Signaling Pathways Orchestrate Regulation of Growth in the Pacific Oyster (Crassostrea gigas), as Revealed by Gene Expression Profiles.

The involvement of insulin/insulin-like growth factor signaling (IIS) pathways in the growth regulation of marine invertebrates remains largely unexplored. In this study, we used a fast-growing Pacific oyster (Crassostrea gigas) variety "Haida No.1" as the material with which to unravel the role of IIS systems in growth regulation in oysters. Systematic bioinformatics analyses allowed us to identify major components of the IIS signaling pathway and insulin-like peptide receptor (ILPR)-mediated signaling pathways, including PI3K-AKT, RAS-MAPK, and TOR, in C. gigas. The expression levels of the major genes in IIS and its downstream signaling pathways were significantly higher in "Haida No.1" than in wild oysters, suggesting their involvement in the growth regulation of C. gigas. The expression profiles of IIS and its downstream signaling pathway genes were significantly altered by nutrient abundance and culture temperature. These results suggest that the IIS signaling pathway coupled with the ILPR-mediated signaling pathways orchestrate the regulation of energy metabolism to control growth in Pacific oysters.

The roles of two myostatins and immune effects after inhibition in Qi river crucian carp (Carassius auratus).

Myostatin, through type I receptor (kinase 4, 5, ALK4/5), functions to participate in the immune system and negatively regulate muscle growth in mammals. However, the role of myostatin (mstn) in the immune system of teleosts is largely unknown. In a previous study, we cloned the mstn1 cDNA encoding myostatin in Qi river crucian carp (Carassius auratus). In the present study, we have cloned mstn2 cDNA, which was characterized and analyzed together with mstn1. Tissue distribution analysis showed that both mstn genes are expressed in numerous tissues, with mstn1 dominantly expressed in the muscle and brain, whereas mstn2 is mainly expressed in the brain. During embryogenesis, mstn1 and mstn2 exhibit different expression patterns. Both mstn1 and mstn2 expression increased stepwise in the brain at different developmental stages. Furthermore, both genes are differentially regulated during different periods of fasting/re-feeding. Following the exposure of C. auratus to polyI:C, lipopolysaccharide (LPS), and Aeromonas hydrophila, both genes were upregulated in different tissues, which indicated that they might be involved in the immune response against pathogenic invasion. Blocking the Mstn signal pathway with SB-431542 (a chemical inhibitor of ALK4/5) resulted in significantly increased body length and weight. However, the mortality of SB-431542-treated fish was higher after A. hydrophila challenge. Moreover, decreased expression of lysozymes (lyz), complement component 3 (c3), ß-defensin 3 (defb3), and interferon γ (ifnγ) were exhibited in treated fish, compared with the controls. Furthermore, the expression of nf-κb1, three pro-inflammatory cytokines (il1ß, il6, and tnfα), and inflammatory cytokines (il8 and il10) were significantly increased in both the SB-431542-treated group and the control after A. hydrophila infection, suggesting that the NF-κB pathway was not suppressed in the SB-431542-treated fish. Taken together, our data suggest that both mstn1 and mstn2 play important roles in early body development, muscle growth, and the immune system by acting downstream of the NF-κB signal pathway.

Cloning and characterization of wnt4a gene in a natural triploid teleost, Qi river crucian carp (Carassius auratus).

WNT4 (wingless-type MMTV integration site family, member 4) plays a key role in the ovarian differentiation and development in mammals. However, the possible roles of Wnt4 during gonadal differentiation and development need further clarification in teleosts. In this study, we cloned and characterized the full-length cDNA of Qi river crucian carp (Carassius auratus) wnt4a gene (CA-wnt4a). The cDNA of CA-wnt4a is 2337 bp, including the ORF of 1059 bp, encoding a putative protein with a transmembrane domain and a WNT family domain. Sequence and phylogenetic analyses revealed that the CA-Wnt4a identified is a genuine Wnt4a. Tissue distribution analysis showed that CA-wnt4a is expressed in all the tissues examined, including ovary. CA-wnt4a undergoes a stepwise increase in the embryonic stages, suggesting that CA-wnt4a might be involved in the early developmental stage. Ontogenic analysis demonstrated that CA-wnt4a expression is upregulated in the ovaries at 30-50 days after hatching (dah), the critical period of sex determination/differentiation in Qi river crucian carp. From 90 dah, the expression of CA-wnt4a was gradually downregulated in the developing ovaries. Immunohistochemistry demonstrated that CA-Wnt4a was expressed in the somatic and germ cells of the ovary by 30 dah, thereafter, positive signals of Wnt4a were detected in the somatic cells, oogonia and primary growth oocytes from 60 dah. In the sex-reversed testis induced by letrozole treatment, the expression level of CA-wnt4a was significantly downregulated. When CA-wnt4a expression was inhibited by injection of FH535 (an inhibitor of canonical Wnt/ß-catenin signal pathway) in the ovaries, levels of cyp19a1a, foxl2 mRNA were significantly downregulated, while sox9b and cyp11c1 were upregulated, which suggested that together with Foxl2-leading estrogen pathway, CA-wnt4a signaling pathway might be involved in ovarian differentiation and repression of the male pathway gene expression in Qi river crucian carp.

[Resources industrialization channels and strategies of active polysaccharide from Lycii Fructus based on analysis of domestic and foreign intellectual properties].

Lycii Fructus is a traditional medicinal and edible herb, with the function of liver and kidney nourishing, blood and eyesight replenishing. As the most important active substance in the fruits of Lycium barbarum, Lycium barbarum polysaccharides (LBP) have been demonstrated to play multiple pharmacological activities, with broad prospects for development and utilization. Based on the comprehensive deep analysis of global LBP patent output, the current patent LBP features were explored from the perspective of development trend, technology field distribution, time dimension, technology life curve and patent applicant. The development trend of Chinese LBP industry was also revealed. At present, the research and development of LBP is in the "development period", with a good development track in which the main research institutions remain domestic ones. At the same time, problem still presents in the lack of industrialization, which means that the advantage of natural resources has not been transformed into industrial advantages of LBP. The format of potential product group and prospect of LBP were also analyzed to provide scientific information for the effective development,comprehensive utilization and collaborative innovation mechanism of Chinese Lycium barbarum resources and LBP.

Integrative proteome and metabolome analyses reveal molecular basis underlying growth and nutrient composition in the Pacific oyster, Crassostrea gigas.

In order to comprehend the molecular basis of growth, nutrient composition, and color pigmentation in oysters, comparative proteome and metabolome analyses of two selectively bred oyster strains with contrasting growth rate and shell color were used in this study. A total of 289 proteins and 224 metabolites were identified differentially expressed between the two strains. We identified a series of specifically enriched functional clusters implicated in protein biosynthesis (RPL4, MRPS7, and CARS), fatty acid metabolism (ACSL5, PEX3, ACOXI, CPTIA, FABP6, and HSD17B12), energy metabolism (FH, PPP1R7, CLAM2, and RGN), cell proliferation (MYB, NFYC, DOHH, TOP2a, SMARCA5, and SMARCC2), material transport (ABCB1, ABCB8, VPS16, and VPS33a), and pigmentation (RDH7, RDH13, Retsat, COX15, and Cyp3a9). Integrated proteome and metabolome analyses indicate that fast-growing strain utilize energy-efficient mechanisms of ATP generation while promoting protein and polyunsaturated fatty acid synthesis, activating the cell cycle to increase cell proliferation and thus promoting their biomass increase. These results uncovered molecular mechanisms underlying growth regulation, nutrition quality, and pigmentation and provided candidate biomarkers for molecular breeding in oysters. SIGNIFICANCE: Rapid growth has always been the primary breeding objective to increase the production profits of Pacific oyster (Crassostrea gigas), while favorable nutritional quality and beautiful color add commercial value. In recent years, proteomic and metabolomic techniques have been widely used in marine organisms, although these techniques are seldom utilized to study oyster growth and development. In this study, two C. gigas strains with contrasted phenotypes in growth and shell color provided an ideal model for unraveling the molecular basis of growth and nutrient composition through a comparison of the proteome and metabolome. Since proteins and metabolites are the critical undertakers and the end products of cellular regulatory processes, identifying the differentially expressed proteins and metabolites would allow for discovering biomarkers and pathways that were implicated in cell growth, proliferation, and other critical functions. This work provides valuable resources in assistance with molecular breeding of oyster strains with superior production traits of fast-growth and high-quality nutrient value.

Cognitive control training with domain-general response inhibition does not change children's brains or behavior.

Cognitive control is required to organize thoughts and actions and is critical for the pursuit of long-term goals. Childhood cognitive control relates to other domains of cognitive functioning and predicts later-life success and well-being. In this study, we used a randomized controlled trial to test whether cognitive control can be improved through a pre-registered 8-week intervention in 235 children aged 6-13 years targeting response inhibition and whether this leads to changes in multiple behavioral and neural outcomes compared to a response speed training. We show long-lasting improvements of closely related measures of cognitive control at the 1-year follow-up; however, training had no impact on any behavioral outcomes (decision-making, academic achievement, mental health, fluid reasoning and creativity) or neural outcomes (task-dependent and intrinsic brain function and gray and white matter structure). Bayesian analyses provide strong evidence of absent training effects. We conclude that targeted training of response inhibition does little to change children's brains or their behavior.

Mechanical Behavior of Al-Si10-Mg P-TPMS Structure Fabricated by Selective Laser Melting and a Unified Mathematical Model with Geometrical Parameter.

Compared with the traditional lattice structure, the triply periodic minimal surface (TPMS) structure can avoid stress concentration effectively. Here, it is promising in the fields of lightweight and energy absorption. However, the number of structural parameters and mechanical properties of the TPMS structure is plentiful, and the relationship between them is unclassified. In this paper, for the first time, a unified mathematical model was proposed to establish the relationship between TPMS structural design parameters and mechanical properties. Fifteen primitive models were designed by changing the structural parameters (level-set value C and thickness T) and manufacturing by selective laser melting. The geometric defects and surface quality of the structures were explored by optical microscope and scanning electron microscopy (SEM). The mechanical properties were investigated by quasi-static compression test and finite element simulation. The influence of building direction on structural mechanical behavior (failure mode, stress-strain curve) was studied. The real mechanical properties (Young's modulus and plateau stress) of the structure could be predicted according to different C and T combinations. Finally, the energy absorption characteristics were explored. The results showed that when the C value is 0.6 in the range of 0-0.6, the energy absorption performance of the structure is at the maximum level.

Structural and expression analysis of the dopamine receptors reveals their crucial roles in regulating the insulin signaling pathway in oysters.

Dopamine performs its critical role upon binding to receptors. Since dopamine receptors are numerous and versatile, understanding their protein structures and evolution status, and identifying the key receptors involved in the modulation of insulin signaling will provide essential clues to investigate the molecular mechanism of neuroendocrine regulating the growth in invertebrates. In this study, seven dopamine receptors were identified in the Pacific oysters (Crassostrea gigas) and were classified into four subtypes according to their protein secondary and tertiary structures, and ligand-binding activities. Of which, DR2 (dopamine receptor 2) and D(2)RA-like (D(2) dopamine receptor A-like) were considered the invertebrate-specific type 1 and type 2 dopamine receptors, respectively. Expression analysis indicated that the DR2 and D(2)RA-like were highly expressed in the fast-growing oyster "Haida No.1". After in vitro incubation of ganglia and adductor muscle with exogenous dopamine and dopamine receptor antagonists, the expression of these two dopamine receptors and ILPs (insulin-like peptides) was also significantly affected. Dual-fluorescence in situ hybridization results showed that D(2)RA-like and DR2 were co-localized with MIRP3 (molluscan insulin-related peptide 3) and MIRP3-like (molluscan insulin-related peptide 3-like) in the visceral ganglia, and were co-localized with ILP (insulin-like peptide) in the adductor muscle. Furthermore, the downstream components of dopamine signaling, including PKA, ERK, CREB, CaMKK1, AKT, and GSK3ß were also significantly affected by the exogenous dopamine and dopamine receptor antagonists. These findings confirmed that dopamine might affect the secretion of ILPs through the invertebrate-specific dopamine receptors D(2)RA-like and DR2, and thus played crucial roles in the growth regulation of the Pacific oysters. Our study establishes the potential regulatory relationship between the dopaminergic system and insulin-like signaling pathway in marine invertebrates.

Strength and Microscopic Mechanism of Cement-Fly Ash-Slag-Desulfurization Gypsum Solidified Mica Schist Weathered Soil.

Mica schist weathered soil possesses a number of poor engineering characteristics, which make it difficult to use as a subgrade material for resource utilization. Therefore, in this study, a new type of curing agent, CFSD (cement-fly ash-slag-desulfurized gypsum), is proposed for this soil. The effects of different curing agent dosages, age of preservation, and confining pressure on the stress-strain curves were analyzed via the uniaxial compression test and triaxial compression test, while the micromorphological characteristics of cured soil were analyzed via X-ray diffraction analysis and the SEM test combined with Image J software. In this paper, we also establish a microscopic mechanism model to determine how curing agents increase the strength of mica schists. The results reveal that the compressive strength of solidified soil increases rapidly within 28 days; the CFSD dosage of 4% at 7 d increased by 103.23% by 28 d. After 28 d, the trend of compressive strength growth was flat. The CFSD dosage of 4% at 7 d increased by 128.34% by 90 d; with the increase in the dosage, the curve transformed from flat to steep. These results suggest that the CFSD dosage is positively correlated with the damage strain and damage bias stress of solidified soil. The curves for the strain softening type with a 4% dosage as the initial effective confining pressure increased from 50 kPa to 300 kPa; the failure stress and failure strain increased by 202.09% and 90.85%, respectively. With the increase in curing agent dosage and maintenance age, the pore size of 2~5 µm, >5 µm interval decreased from 56.46% to 27.92%, the porosity decreased from 12.51% to 4.6%, and the hydrate produced by the curing agent cemented and filled up the pore space between the loose particles of the soil body. Thus, the large pore space became microporous, and the pore structure densification was greatly improved.

Contribution of HIF-1α to Heat Shock Response by Transcriptional Regulation of HSF1/HSP70 Signaling Pathway in Pacific Oyster, Crassostrea gigas.

Ocean temperature rising drastically threatens the adaptation and survival of marine organisms, causing serious ecological impacts and economic losses. It is crucial to understand the adaptive mechanisms of marine organisms in response to high temperature. In this study, a novel regulatory mechanism that is mediated by hypoxia-inducible factor-1α (HIF-1α) was revealed in Pacific oyster (Crassostrea gigas) in response to heat stress. We identified a total of six HIF-1α genes in the C. gigas genome, of which HIF-1α and HIF-1α-like5 were highly induced under heat stress. We found that the HIF-1α and HIF-1α-like5 genes played critical roles in the heat shock response (HSR) through upregulating the expression of heat shock protein (HSP). Knocking down of HIF-1α via RNA interference (RNAi) inhibited the expression of heat shock factor 1 (HSF1) and HSP70 genes in C. gigas under heat stress. Both HIF-1α and HIF-1α-like5 promoted the transcriptional activity of HSF1 by binding to hypoxia response elements (HREs) within the promoter region. Furthermore, the survival of C. gigas under heat stress was significantly decreased after knocking down of HIF-1α. This work for the first time revealed the involvement of HIF-1α/HSF1/HSP70 pathway in response to heat stress in the oyster and provided an insight into adaptive mechanism of bivalves in the face of ocean warming.

Damage Model of Steel Fiber-Reinforced Coal Gangue Concrete under Freeze-Thaw Cycles Based on Weibull Distribution.

In order to improve the utilization rate of coal gangue and expand the application range of coal gangue concrete (CGC), a certain proportion of steel fiber was added to the concrete, and the freeze-thaw cycles (FTCs) and flexural tests were used to explore the effects of different mass replacement rates of coal gangue (0%, 25%, 50%, 75%, and 100%) and different proportions of the volumetric blending of the steel fiber (0%, 0.8%, 1.0%, and 1.2%) on the frost resistance of steel fiber-reinforced CGC (SCGC). The governing laws of mass loss rate, relative dynamic elastic modulus and load-midspan deflection curve were obtained on the base of the analysis of testing results. The damage mechanisms of the SCGC under the FTCs were analyzed using the results of scanning electron microscopy (SEM). Based on the Lemaitre's strain equivalence principle and Krajcinovic's vector damage theory, a damage evolution model of the SCGC under the FTCs was established by introducing the damage variable of the SCGC satisfying Weibull distribution. The results show an increasing mass loss rate of the SCGC and a decreasing relative dynamic elastic modulus with an increasing mass replacement rate of coal gangue. The proper content of the steel fiber can reduce the mass loss rate of concrete by 10~40% and the relative loss rate of dynamic elastic modulus of concrete by 2~8%, thus significantly improving the ductility and toughness of the concrete. The established damage evolution model is well validated by the experimental results, which further help to improve the modelling accuracy. This study provides key experimental data and a theoretical basis for a wider range of proper utilization of coal gangue in cold regions.

Rapid preparation of N-CNTs/P(AA-co-AM) composite hydrogel via frontal polymerization and its mechanical and conductive properties.

Deep eutectic solvent (DES) was prepared by using acrylic acid (AA) and acrylamide (AM) as hydrogen bonding donors (HBD) and choline chloride (ChCl) as hydrogen bonding receptors (HBA). Nitrogen-doped carbon nanotubes (N-CNTs) were dispersed in DES as fillers, and N-CNTs/P(AA-co-AM) composite hydrogels were prepared by FP. The interaction mode between the hydrogel and N-CNTs was characterized by Fourier infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). The mechanical properties, pH response and electrical conductivity of the composite hydrogels were studied. The results showed that the mechanical properties of the hydrogel were significantly enhanced with the increase of N-CNT content. The tensile strength and compressive strength of the FP4 composite hydrogel reached 5.42 MPa and 4.29 MPa, respectively. Due to the dissociation of carboxyl groups in AA in an alkaline environment the composite hydrogel showed excellent pH response performance. The conductivity of the hydrogel was also found to be improved with the content of N-CNTs. When the content of N-CNTs is 1.0 wt%, the conductivity of the hydrogel was 4.2 times higher than that of the hydrogel without N-CNTs, and connecting it to a circuit can make an LED lamp emit bright light. In this study, a simple and green method was proposed to prepare composite hydrogels with excellent mechanical properties and electrical conductivity by FP of DES in less than 5 min.

Comparative Methylome Analysis Reveals Epigenetic Signatures Associated with Growth and Shell Color in the Pacific Oyster, Crassostrea gigas.

Fast growth is one of the most important breeding goals for all economic species such as the Pacific oyster (Crassostrea gigas), an aquaculture mollusk with top global production. Although the genetic basis and molecular mechanisms of growth-related traits have been widely investigated in the oyster, the role of DNA methylation involved in growth regulation remains largely unexplored. In this study, we performed a comparative DNA methylome analysis of two selectively bred C. gigas strains with contrasted phenotypes in growth and shell color based on whole-genome bisulfite sequencing (WGBS). Genome-wide profiling of DNA methylation at the single-base resolution revealed that DNA methylations were widely spread across the genome with obvious hotspots, coinciding with the distribution of genes and repetitive elements. Higher methylation levels were observed within genic regions compared with intergenic and promoter regions. Comparative analysis of DNA methylation allowed the identification of 339,604 differentially methylated CpG sites (DMCs) clustering in 27,600 differentially methylated regions (DMRs). Functional annotation analysis identified 11,033 genes from DMRs which were enriched in biological processes including cytoskeleton system, cell cycle, signal transduction, and protein biosynthesis. Integrative analysis of methylome and transcriptome profiles revealed a positive correlation between gene expression and DNA methylation within gene-body regions. Protein-protein interaction (PPI) analysis of differentially expressed and methylated genes allowed for the identification of integrin beta-6 (homolog of human ITGB3) as a hub modulator of the PI3K/Akt signaling pathway that was involved in various growth-related processes. This work provided insights into epigenetic regulation of growth in oysters and will be valuable resources for studying DNA methylation in invertebrates.

Entropy-Driven Quick Loading of Functional Proteins in Nanohydrogels for Highly Efficient Tumor Targeting Therapy.

With the gradual deep understanding of the tumorigenesis and development process, nanodrug are thought to have great prospects for individualized treatment of tumors. To deliver adequate concentration of active ingredients to targeted tissues, proteins are usually used as carriers to avoid clearance by the immune system. Herein, a new strategy is developed for preparation of the protein-functionalized targeting nanodrugs; different kinds of proteins (albumin, horseradish, transferrin, and ricin) can be quickly loaded in polyacrylic acid nanohydrogels (PAA-NGs) without discrimination within 1 min under the strong driving force of entropy; and the loading efficiency can reach 99% with about 50% loading content. Meanwhile, the activity of the released protein can be well retained. After oriented binding of the targeting agent on the surface of the nanocarriers by a unique and facile technique, the protein-loaded nanodrug exhibits excellent tumor cell uptake and targeting effect. The excellent targeting ability from the oriented binding is further proved by comparing with the non-oriented targeting system. With quick loading of the anti-tumor protein of ricin and oriented binding of transferrin protein (Tf), the targeting nanodrug (PAA-BB@Ricin/Tf) shows a remarkable anti-tumor effect. This study proves a new universal delivery and targeting strategy for improving the nanodelivery system, which has great potentials for clinical application.

Combined detection of preoperative neutrophil to lymphocyte ratio and interleukin-6 as an independent prognostic factor for patients with non-metastatic colorectal cancer.

BACKGROUND: This study sought to explore the value of the neutrophil to lymphocyte ratio (NLR) and interleukin-6 (IL-6) in predicting the prognosis of patients with non-metastatic colorectal cancer (CRC). METHODS: The data of 88 surgical CRC patients were retrospectively analyzed. A receiver operating characteristic (ROC) curve analysis was conducted to determine the patients' thresholds for the NLR and IL-6. Kaplan-Meier curve and Cox regression models were used to assess the prognostic values. RESULTS: A ROC analysis was conducted to calculate the NLR cut-off value. The area under the curve (AUC) of the NLR was 0.739 [95% confidence interval (CI): 0.634 to 0.844] for overall survival (OS), and 0.799 (95% CI: 0.705 to 0.892) for disease-free survival (DFS). The AUC of IL-6 was 0.773 (95% CI: 0.670 to 0.876) for OS, and 0.817 (95% CI: 0.728 to 0.906) for DFS. The AUC of NLR + IL-6 was 0.805 (95% CI: 0.710 to 0.899) for OS and 0.853 (95% CI: 0.774 to 0.933) for DFS, which were higher than the NLR or IL-6 alone AUCs for OS and DFS. In addition, a high NLR and IL-6 value was significantly correlated with tumor differentiation and tumor-node-metastasis staging. The NLR was positively correlated with IL-6 level (r=0.481). The results of the Kaplan-Meier analysis showed that a high NLR + IL-6 value was correlated with worse OS and DFS. CONCLUSIONS: A high NLR and IL-6 value is a better independent prognostic biomarker of CRC than the NLR or IL-6 level alone, and may be applied in clinical practice to identify high-risk patients.

[Bioinformatics Analysis and Verification of Acute B-Lymphocytic Leukemia in Children with Isolated Bone Marrow Relapse].

OBJECTIVE: To analyze the hub genes affecting the solely bone marrow relapse of childish acute B-cell lymphoblastic leukemia (B-ALL). METHODS: The high-throughput RNA sequencing data were downloaded from TCGA database, the differentially expressed genes were screened by DESeq2 package of R, and the differentially expressed genes were grouped by GO function enrichment analysis and KEGG pathway enrichment analysis. Further, the data of STRING database and Cytoscape software were used to construct protein interaction network, screen hub genes and highly interaction protein sub network, perform GO and KEGG analysis of the hub genes and protein sub network respectively. JASPAR database was used to screen the upstream transcription factor of the hub gene promoter. Survival analysis based on the expression of hub genes was performed with clinical information attached to TCGA database. The bone marrow samples and clinical data of the patients were collected, the analysis results of hub genes were verified through clinical samples. RESULTS: 847 differentially expressed genes were collected, including 813 up-regulated genes, 34 down-regulated genes, 11 hub genes were screened out. The results of survival analysis showed that RPS5、RPS15、RPL23、RPL35、RPS8、RPS27A、RPS3、RPL9、RPS21、RPS7 and RPL38 showed significant effect on the survival of the children, and ZNF460 might be involved in their regulation. The high expressions of RPS3, RPS15, RPS8, RPS27A, and RPS21 had been verified in clinical samples of solely bone marrow relapsed patients. CONCLUSION: RPS3, RPS15, RPS8, RPS27A, RPS21 can be used as biomarkers to indicate the malignant event of solely bone marrow relapse, which may be regulated by ZNF460.