Genome-Wide Evolutionary Characterization and Expression Analysis of Major Latex Protein (MLP) Family Genes in Tomato.

Major latex proteins (MLPs) play a key role in plant response to abiotic and biotic stresses. However, little is known about this gene family in tomatoes (Solanum lycopersicum). In this paper, we perform a genome-wide evolutionary characterization and gene expression analysis of the MLP family in tomatoes. We found a total of 34 SlMLP members in the tomato genome, which are heterogeneously distributed on eight chromosomes. The phylogenetic analysis of the SlMLP family unveiled their evolutionary relationships and possible functions. Furthermore, the tissue-specific expression analysis revealed that the tomato MLP members possess distinct biological functions. Crucially, multiple cis-regulatory elements associated with stress, hormone, light, and growth responses were identified in the promoter regions of these SlMLP genes, suggesting that SlMLPs are potentially involved in plant growth, development, and various stress responses. Subcellular localization demonstrated that SlMLP1, SlMLP3, and SlMLP17 are localized in the cytoplasm. In conclusion, these findings lay a foundation for further dissecting the functions of tomato SlMLP genes and exploring the evolutionary relationships of MLP homologs in different plants.

The seahorse genome and the evolution of its specialized morphology.

Seahorses have a specialized morphology that includes a toothless tubular mouth, a body covered with bony plates, a male brood pouch, and the absence of caudal and pelvic fins. Here we report the sequencing and de novo assembly of the genome of the tiger tail seahorse, Hippocampus comes. Comparative genomic analysis identifies higher protein and nucleotide evolutionary rates in H. comes compared with other teleost fish genomes. We identified an astacin metalloprotease gene family that has undergone expansion and is highly expressed in the male brood pouch. We also find that the H. comes genome lacks enamel matrix protein-coding proline/glutamine-rich secretory calcium-binding phosphoprotein genes, which might have led to the loss of mineralized teeth. tbx4, a regulator of hindlimb development, is also not found in H. comes genome. Knockout of tbx4 in zebrafish showed a 'pelvic fin-loss' phenotype similar to that of seahorses.

Evaluation and optimization of a conventional SPCE for FMD post-vaccination monitoring.

BACKGROUND: Foot-and-mouth disease (FMD) can be controlled by either stamping out or vaccination, a choice which depends on both the economic importance of the livestock sector as well as the disease status. In FMD-free countries with vaccination, such as Korea, vaccination programs should guarantee prevention against transmission of FMD. Monitoring of vaccination programs is also essential for ensuring sufficient coverage that will limit the transmission of FMDV. There are several methods to screen FMD virus (FMDV) structural protein (SP) antibodies including SPCE (Solid-phase competitive ELISA), LPBE (Liquid-phase blocking ELISA), and VNT (Virus neutralization test). Among these, SPCE is widely used for serological monitoring since VNT-the gold standard method-has certain practical limitations, such as high costs in terms of time and labor. However, whether SPCE can ensure the vaccination status of individual animals and whole farms is unclear. In this study, SPCE, LPBE and VNT were compared with respect to correlation with each other and sensitivity at commercial pig farms. RESULTS: The positive results obtained by PrioCHECK SPCE differed from those obtained by LPBE and VNT. The sensitivity of SPCE relative to those of the other tests was fairly low. The raw data of SPCE were most highly correlated with those of VNT with XJ strain, while their positivity and negativity were most highly correlated with LPBE. The results of ROC analysis proposed new cut-off for PrioCHECK SPCE higher than the previous 50% inhibition. CONCLUSIONS: The high false positive rate of PrioCHECK SPCE suggested that high seropositivity by SPCE may not guarantee a true vaccination coverage. Adjusting the cut-off percentage (%) inhibition value for SPCE is needed to address this problem, and it is highly recommended that routine FMDV serological monitoring programs using PrioCHECK SPCE should be combined with alternative methods such as LPBE or VNT.

The Potential of Poly[N-(2-hydroxypropyl)methacrylamide] via Reversible Addition-Fragmentation Chain Transfer Polymerization as Safe Nanocarrier.

N-(2-hydroxypropyl)methacrylamide (HPMA) copolymers have been presented as nanoscale drug/gene delivery systems and imaging probes, and the well-defined HPMA copolymers prepared via reversible addition-fragmentation chain transfer (RAFT) polymerization promote their to clinical trials, as the significant enhanced anticancer efficacy. The biosafety is another issue associated with the carriers. In this study, we prepared the linear and branched HPMA copolymers labeled with Cy5.5 via RAFT polymerization and click chemistry, and their potential biosafety was studied. The linear copolymer was prepared via RAFT polymerization mediated by the ends-functionalized peptide chain transfer agent (peptide2CTA), resulting in well-defined and block linear HPMA copolymer with molecular weight (MW) of 98 kDa. Additionally, the branched HPMA copolymer was also prepared via RAFT polymerization. Followed by Cy5.5 labeling, the two copolymers showed negative zeta potential and their accumulation into tumor was studied by in vivo optical fluorescence imaging in the nude mice with breast tumors. The biosafety studies on in vitro cytotoxicity and hemocompatibility studies, including hemolysis tests, plasma coagulation and thromboelastography assay were carried out well, demonstrating that the linear HPMA copolymer-Cy5.5 with MW around 100 kDa and biodegradable moiety in the main chain might be utilized as safe nanoscale carrier.

Improvement of short-term hypothermic preservation of microencapsulated hepatocytes.

OBJECTIVES: To determine the optimal storage solution containing suitable protective agents for the preservation of microencapsulated hepatocytes at 4 °C as well as the optimum incubation time after hypothermic preservation. RESULTS: L15 was the optimum solution for both maintaining microcapsule integrity and cell viability. Furthermore, 5 %(v/v) PEG (20 or 35 kDa) added to Leibovitz-15 medium was optimal for microencapsulated C3A cells, enhancing cell viability and liver-specific functions, including albumin and urea synthesis as well as CYP1A2 and CYP3A4 activities. The transcription levels of several CYP450-related genes were also dramatically increased in cells incubated in the optimal solution. Pre-incubation for 2 h was the optimal time for restoring favorable levels of CYP1A2 and CYP3A4 activities in microencapsulated C3A cells for short term, 2 day storage. CONCLUSIONS: Leibovitz-15 medium supplemented with 5 % (v/v) PEG is a promising cold solution for microencapsulated hepatocytes at 4 °C, with an incubation of 2 h at 37 °C after hypothermic preservation being the best incubation duration for further cell application.

Finite element simulation and optimization of mechanical performance of the magnesium-alloy biliary stent.

A biliary stent is used to expand the bile duct during interventional treatment of biliary stricture. To reduce the complications after stent implantation, it is necessary to understand and improve the comprehensive performance of magnesium alloy biliary stents. In this study, at first, the mechanical performance of magnesium alloy biliary stent in bile ducts with different ellipticitiesis studied, and the influence of the ellipticity of bile duct on the interaction of the stent-bile duct coupling system is better understood. Proposed results show the increment of ellipticity of the bile duct will increase the difficulty of expanding the stent. Second, to improve the mechanical performance and degradation performance of the stent simultaneously, an optimal design method based on a zero-order algorithm is used to reduce the maximum equivalent stress on the stent effectively, to make the stress distribution more uniform, and to expand the bile duct of the lesion more effectively and uniformly. The results of this analysis and optimization are useful to predict the clinical effect of stents and to design and optimize stents for both bile duct and other non-vascular.

Interaction of tetrabromobisphenol A (TBBPA) with microplastics-sediment (MPs-S) complexes: A comparison between binary and simple systems.

The presence of microplastics (MPs) and the associated organic pollutants in the aquatic environment has attracted growing concern in recent years. MPs could compete with chemicals for adsorption sites on the surface of sediment, affecting the sorption processes of pollutants on sediment. However, few studies focused on the binary system of microplastics-sediment (MPs-S), which appear much common in aquatic environment. Herein, we investigated the interactions between a continuously used flame retardant tetrabromobisphenol A (TBBPA) and four MPs-S complexes (PVC-S, PE-S, PP-S and PS-S). The equilibrium adsorption capacities were 17.1, 15.6, 15.4, and 14.0 mg/kg for PVC-S, PS-S, PE-S, and PP-S, respectively. Kinetics suggest that adsorption behavior of TBBPA was fitted by pseudo-second-order model. Co-adsorption of TBBPA in binary systems were much lower than the sum of each simple system, which may be due to the mutually occupied adsorption sites. Higher ionic strength and lower dissolved organic matter strengthened the sorption of TBBPA onto MPs-S complexes. The enhanced sorption capacities for TBBPA were observed with elevated proportion and small particle size of MPs in the MPs-S complexes. This study contributes to the knowledge on the impact of MPs in partitioning of organic pollutants in-between solid and aqueous phases in the aquatic environment.

Liposomal T cell engager and re-director for tumor cell eradication in cancer immunotherapy.

T cells are one of the most important effector cells in cancer immunotherapy. Various T cell-dependent bispecific antibody (TDB) drugs that engage T cells for targeted cancer cell lysis are being developed. Here, we describe supra-molecular T-cell redirecting antibody fragment-anchored liposomes (TRAFsomes) and report their immune modulation and anti-cancer effects. We found that TRAFsomes containing different copies of anti-CD3 fragments displayed different T cell modulation profiles, showing that optimization of surface density is needed to define the therapeutic window for potentiating cancer cell-specific immune reactions while minimizing nonspecific side effects. Moreover, small molecular immunomodulators may also be incorporated by liposomal encapsulation to drive CD8 + T cell biased immune responses. In vivo studies using human peripheral blood mononuclear cell reconstituted mouse models showed that TRAFsomes remained bounded to human T cells and persisted for more than 48 hours after injection. However, only TRAFsomes containing a few anti-CD3 (n = 9) demonstrated significant T cell-mediated anti-cancer activities to reverse tumor growth. Those with more anti-CD3s (n = 70) caused tumor growth and depletion of human T cells at the end of treatments. These data suggested that TRAFsomes can be as potent as traditional TDBs and the liposomal structure offers great potential for immunomodulation and improvement of the therapeutic index.Abbreviation: Chimeric antigen receptor T cells (CAR-T cells), Cytokine release syndrome (CRS) Cytotoxic T cell (CTL) Effector: target ratios (E:T ratios), Heavy chain (HC) Immune-related adverse events (irAE), Large unilamellar vesicle (LUV), Peripheral blood mononuclear cells (PBMCs, Single-chain variable fragment (scFv), T cell-dependent bispecific antibody (TDB), T cell redirecting antibody fragment-anchored liposomes (TRAFsomes), Methoxy poly-(ethylene glycol) (mPEG).

Seadragon genome analysis provides insights into its phenotype and sex determination locus.

The iconic phenotype of seadragons includes leaf-like appendages, a toothless tubular mouth, and male pregnancy involving incubation of fertilized eggs on an open "brood patch." We de novo-sequenced male and female genomes of the common seadragon (Phyllopteryx taeniolatus) and its closely related species, the alligator pipefish (Syngnathoides biaculeatus). Transcription profiles from an evolutionary novelty, the leaf-like appendages, show that a set of genes typically involved in fin development have been co-opted as well as an enrichment of transcripts for potential tissue repair and immune defense genes. The zebrafish mutants for scpp5, which is lost in all syngnathids, were found to lack or have deformed pharyngeal teeth, supporting the hypothesis that the loss of scpp5 has contributed to the loss of teeth in syngnathids. A putative sex-determining locus encoding a male-specific amhr2y gene shared by common seadragon and alligator pipefish was identified.

Early warning of hand, foot, and mouth disease transmission: A modeling study in mainland, China.

BACKGROUND: Hand, foot, and mouth disease (HFMD) is a global infectious disease; particularly, it has a high disease burden in China. This study was aimed to explore the temporal and spatial distribution of the disease by analyzing its epidemiological characteristics, and to calculate the early warning signals of HFMD by using a logistic differential equation (LDE) model. METHODS: This study included datasets of HFMD cases reported in seven regions in Mainland China. The early warning time (week) was calculated using the LDE model with the key parameters estimated by fitting with the data. Two key time points, "epidemic acceleration week (EAW)" and "recommended warning week (RWW)", were calculated to show the early warning time. RESULTS: The mean annual incidence of HFMD cases per 100,000 per year was 218, 360, 223, 124, and 359 in Hunan Province, Shenzhen City, Xiamen City, Chuxiong Prefecture, Yunxiao County across the southern regions, respectively and 60 and 34 in Jilin Province and Longde County across the northern regions, respectively. The LDE model fitted well with the reported data (R2 > 0.65, P < 0.001). Distinct temporal patterns were found across geographical regions: two early warning signals emerged in spring and autumn every year across southern regions while one early warning signals in summer every year across northern regions. CONCLUSIONS: The disease burden of HFMD in China is still high, with more cases occurring in the southern regions. The early warning of HFMD across the seven regions is heterogeneous. In the northern regions, it has a high incidence during summer and peaks in June every year; in the southern regions, it has two waves every year with the first wave during spring spreading faster than the second wave during autumn. Our findings can help predict and prepare for active periods of HFMD.

The degradable performance of bile-duct stent based on a continuum damage model: A finite element analysis.

Biomedical magnesium alloy stents have become a hot bed of research focus in interventional therapy for nonvascular diseases. In this study, a numerical model for a balloon-expandable bile duct stent made of magnesium alloy with laser sculpture is developed to predict the effects of the degradation of the stent on the biomechanical behavior in the stent-bile duct coupling system. Based on a continuum damage model, the degradable model of the stent is built to understand its performance in an idealized bile duct as it is subject to corrosion over time. The degradation model developed in this study addresses the uniform corrosion and pitting corrosion. By means of the secondary development function of commercial numerical software ANSYS, the finite element analysis procedures were written to control the degradation process based on the technology of element "birth and death," and it is shown how the three-dimensional model and approach give the possibility of analyzing for the degradation mechanism of a magnesium alloy stent in the bile duct or other nonvascular cavities.

Comparative genomics reveal shared genomic changes in syngnathid fishes and signatures of genetic convergence with placental mammals.

Syngnathids (seahorses, pipefishes and seadragons) exhibit an array of morphological innovations including loss of pelvic fins, a toothless tubular mouth and male pregnancy. They comprise two subfamilies: Syngnathinae and Nerophinae. Genomes of three Syngnathinae members have been analyzed previously. In this study, we have sequenced the genome of a Nerophinae member, the Manado pipefish (Microphis manadensis), which has a semi-enclosed brood pouch. Comparative genomic analysis revealed that the molecular evolutionary rate of the four syngnathids is higher than that of other teleosts. The loss of all but one P/Q-rich SCPP gene in the syngnathids suggests a role for the lost genes in dentin and enameloid formation in teleosts. Genome-wide comparison identified a set of 118 genes with parallel identical amino acid substitutions in syngnathids and placental mammals. Association of some of these genes with placental and embryonic development in mammals suggests a role for them in syngnathid pregnancy.

Designing a castor oil-based polyurethane as bioadhesive.

Based on the stealth behavior of castor oil and poly(ethylene glycol), we selected a polyurethane system to obtain an ideal surgical adhesive. The polyurethane adhesives with varying concentrations of castor oil were investigated by Fourier transform infrared spectrometer, differential scanning calorimetry, scanning electron microscopy, goniometer, and universal testing machine. Curing results show that a 7-min to 25-min room temperature curing can be achieved, providing one possibility of shortening the surgery time. In vitro biodegradation test demonstrates that a certain proportion of the polyurethane film will be hydrolyzed in a foregone manner after a period of time (7 weeks). The adhesion strengths of these adhesives show a strong bonding between pieces of tissue, which makes them qualified for application in a moist environment.

Ethanol induces apoptosis in hepatocytes by a pathway involving novel protein kinase C isoforms.

UNLABELLED: Ethanol abuse is one of the major etiologies of cirrhosis. Ethanol has been shown to induce apoptosis via activation of oxidative stress, mitogen-activated protein kinases (MAPK), and tyrosine kinases. However, there is a paucity of data that examine the interplay among these molecules. In the present study we have systematically elucidated the role of novel protein kinase C isoforms (nPKC; PKCdelta and PKCepsilon) in ethanol-induced apoptosis in hepatocytes. Ethanol enhanced membrane translocation of PKCdelta and PKCepsilon, which was associated with the phosphorylation of p38MAPK, p42/44MAPK and JNK1/2, and the nuclear translocation of NF-kappaB and AP-1. This resulted in increased apoptosis in primary rat hepatocytes. Inhibition of both PKCdelta and PKCepsilon resulted in a decreased MAPK activation, decreased nuclear translocation of NF-kappaB and AP-1, and inhibition of apoptosis. In addition, ethanol activated the tyrosine phosphorylation of PKCdelta via tyrosine kinase in hepatocytes. The tyrosine phosphorylated PKCdelta was cleaved by caspase-3 and these fragments were translocated to the nucleus. Inhibition of ethanol-induced oxidative stress blocked the membrane translocation of PKCdelta and PKCepsilon, and the tyrosine phosphorylation of PKCdelta in hepatocytes. Inhibition of oxidative stress, tyrosine kinase or caspase-3 activity caused a decreased nuclear translocation of PKCdelta in response to ethanol, and was associated with less apoptosis. CONCLUSION: These results provide a newly-described mechanism by which ethanol induces apoptosis via activation of nPKC isoforms in hepatocytes.

Fabrication of graded TiN coatings on nitinol occluders and effects on in vivo nickel release.

A nano-structured TiN/Ti coating with a total thickness of 0.9 mum was deposited on nitinol cardiac occluders using the filtered multi-arc vacuum ion plating technique at less than 300 degrees C. The coating was composed of laminated TiN/Ti layers with thickness of about 100 nm. The cardiac occluders made of a nitinol mesh with and without a graded nano-structured titanium nitride (TiN) coating were implanted into the hearts of rams. The nickel concentration of the whole blood of the animals were measured one week, one month, three months, and six months after implantation and compared to that before operation. The nickel concentration in the neo-endocardium covered occluders was also measured using graphite furnace atomic absorption spectrophotometry. After one week, the nickel content in the blood increased by a factor of three compared to the level before operation and decreased afterwards returning to the normal level after six months when endothelialization was complete. Statistical analyses showed that the TiN coating could mitigate nickel release into blood (P<0.01). For example, the nickel concentration released from the control increased from about 2.65+/-1.20 microg/kg, the normal concentration, to 7.30+/-1.00 microg/kg but just from 2.56+/-1.16 microg/kg to 4.68+/-1.29 microg/kg from the TiN coated occluder after 7 days. The nickel concentration in the neo-endocardium covered and TiN coated occluders reached 17.0+/-8.05 microg/kg in two months after implantation. In comparison, it was 31.0+/-5.72 microg/kg for the occluder without the TiN coating. While normal concentration of nickel in endocardium is also 2.6+/-1.09 microg/kg. Our results demonstrate that the graded TiN coating can significantly reduce nickel release into the endocardium (P<0.01) under in vivo conditions.

A biocompatible and cathepsin B sensitive nanoscale system of dendritic polyHPMA-gemcitabine prodrug enhances antitumor activity markedly.

In an attempt to improve the therapeutic indices of gemcitabine (GEM), a prodrug was designed by conjugating GEM with a stimuli-responsive dendritic polyHPMA copolymer (dendritic polyHPMA-GEM) and synthesized using the one-pot method of RAFT polymerization. The prodrug with dendritic architectures was able to aggregate and form stable nanoscale systems in the order of 46 nm. The high molecular weight (HMW, 168 kDa) dendritic prodrug could biodegrade into segments of low molecular weight (LMW, 29 kDa) for excretion. The prodrug demonstrates enzyme-responsive drug release features; over 95% GEM was released from the carrier in the presence of cathepsin B within 3 h. Investigation of the cellular mechanism underlying the dendritic prodrug suggests that cytotoxicity is associated with cellular uptake and cell apoptosis. The prodrug shows good hemocompatibility and in vivo biosafety. In particular, the dendritic polymer prodrug displays high accumulation within tumors and markedly improved in vivo antitumor activity in the 4T1 murine breast cancer model compared to free GEM. The in vivo antitumor activities are characterized by a marked suppression in tumor volumes indicating much higher tumor growth inhibition (TGI, 83%) than that in GEM treatment (TGI, 36%). In addition, some tumors were eliminated. The tumor xenograft immunohistochemistry study clearly indicates that tumor apoptosis occurs through antiangiogenic effects. These results suggest that the stimuli-responsive dendritic polymer-gemcitabine has great potential as an efficient anticancer agent.

Stimuli-Sensitive Biodegradable and Amphiphilic Block Copolymer-Gemcitabine Conjugates Self-Assemble into a Nanoscale Vehicle for Cancer Therapy.

The availability and the stability of current anticancer agents, particularly water-insoluble drugs, are still far from satisfactory. A widely used anticancer drug, gemcitabine (GEM), is so poorly stable in circulation that some polymeric drug-delivery systems have been under development for some time to improve its therapeutic index. Herein, we designed, prepared, and characterized a biodegradable amphiphilic block N-(2-hydroxypropyl) methacrylamide (HPMA) copolymer-GEM conjugate-based nanoscale and stimuli-sensitive drug-delivery vehicle. An enzyme-sensitive oligopeptide sequence glycylphenylalanylleucylglycine (GFLG) was introduced to the main chain with hydrophilic and hydrophobic blocks via the reversible addition-fragmentation chain transfer (RAFT) polymerization. Likewise, GEM was conjugated to the copolymer via the enzyme-sensitive peptide GFLG, producing a high molecular weight (MW) product (90 kDa) that can be degraded into smaller MW segments (<50 kDa), and ensuring potential rapid site-specific release and stability in vivo. The amphiphilic copolymer-GEM conjugate can self-assemble into compact nanoparticles. NIR fluorescent images demonstrated that the conjugate-based nanoparticles could accumulate and be retained within tumors, resulting in significant increased antitumor efficacy compared to free GEM. The conjugate was not toxic to organs of the mice as measured by body weight reductions and histological analysis. In summary, this biodegradable amphiphilic block HPMA copolymer-gemcitabine conjugate has the potential to be a stimuli-sensitive and nanoscale drug-delivery vehicle.

Formulation and PEGylation optimization of the therapeutic PEGylated phenylalanine ammonia lyase for the treatment of phenylketonuria.

Phenylketonuria (PKU) is a genetic metabolic disease in which the decrease or loss of phenylalanine hydroxylase (PAH) activity results in elevated, neurotoxic levels of phenylalanine (Phe). Due to many obstacles, PAH enzyme replacement therapy is not currently an option. Treatment of PKU with an alternative enzyme, phenylalanine ammonia lyase (PAL), was first proposed in the 1970s. However, issues regarding immunogenicity, enzyme production and mode of delivery needed to be overcome. Through the evaluation of PAL enzymes from multiple species, three potential PAL enzymes from yeast and cyanobacteria were chosen for evaluation of their therapeutic potential. The addition of polyethylene glycol (PEG, MW = 20,000), at a particular ratio to modify the protein surface, attenuated immunogenicity in an animal model of PKU. All three PEGylated PAL candidates showed efficacy in a mouse model of PKU (BTBR Pahenu2) upon subcutaneous injection. However, only PEGylated Anabaena variabilis (Av) PAL-treated mice demonstrated sustained low Phe levels with weekly injection and was the only PAL evaluated that maintained full enzymatic activity upon PEGylation. A PEGylated recombinant double mutant version of AvPAL (Cys503Ser/Cys565Ser), rAvPAL-PEG, was selected for drug development based on its positive pharmacodynamic profile and favorable expression titers. PEGylation was shown to be critical for rAvPAL-PEG efficacy as under PEGylated rAvPAL had a lower pharmacodynamic effect. rAvPAL and rAvPAL-PEG had poor stability at 4°C. L-Phe and trans-cinnamate were identified as activity stabilizing excipients. rAvPAL-PEG is currently in Phase 3 clinical trials to assess efficacy in PKU patients.

Formation of CdS nanoparticle necklaces with functionalized dendronized polymers.

A one-dimensional assembly of cadmium sulfide (CdS) nanoparticles was prepared by using functionalized polymers with pendant Fréchet-type dendritic wedges of different generation. The novel dendronized polymers acted as both nanoreactors for the formation of CdS nanoparticles and a template to arrange the nanoparticles into a necklace. The polymers of different dendron generation were applied to stabilize CdS particles and the effect of dendron size was observed. CdS nanoparticle necklaces were obtained successfully when polymers bearing second-generation dendrons were applied.