The N1-Methyladenosine Methylome of Petunia mRNA.

N1-methyladenosine is a unique type of base methylation in that it blocks Watson-Crick base pairing and introduces a positive charge. m1A is prevalent in yeast and mammalian mRNA and plays a functional role. However, little is known about the abundance, dynamics, and topology of this modification in plant mRNA. Dot blotting and liquid chromatography tandem mass spectrometry analyses revealed a dynamic pattern of m1A mRNA modification in various tissues and at different developmental stages in petunia (Petunia hybrida), a model system for plant growth and development. We performed transcriptome-wide profiling of m1A in petunia mRNA by m1A mRNA immunoprecipitation followed by a deep-sequencing approach (m1A-seq, using an m1A-specific antibody). m1A-seq analysis identified 4,993 m1A peaks in 3,231 genes expressed in petunia corollas; there were 251 m1A peaks in which A residues were partly replaced by thymine and/or reverse transcription stopped at an adenine site. m1A was enriched in coding sequences, with single peaks located immediately after start codons. Ethylene treatment upregulated 400 m1A peaks in 375 mRNAs and downregulated 603 m1A peaks in 530 mRNAs in petunia corollas; 975 m1A peaks in mRNA were only detected in corollas treated with air and 430 were only detected in corollas treated with ethylene. Silencing of petunia tRNA-specific methyltransferase 61A (PhTRMT61A) reduced the m1A level in mRNA in vivo and in vitro. In addition, PhTRMT61A silencing caused abnormal leaf development, and the PhTRMT61A protein was localized to the nucleus. Thus, m1A in mRNA is an important epitranscriptome marker and plays a role in plant growth and development.

Knee loading repairs osteoporotic osteoarthritis by relieving abnormal remodeling of subchondral bone via Wnt/ß-catenin signaling.

Osteoporotic osteoarthritis (OPOA) is a common bone disease mostly in the elderly, but the relationship between Osteoporotic (OP) and osteoarthritis (OA) is complex. It has been shown that knee loading can mitigate OA symptoms. However, its effects on OPOA remain unclear. In this study, we characterized pathological linkage of OP to OA, and evaluated the effect of knee loading on OPOA. We employed two mouse models (OA and OPOA), and conducted histology, cytology, and molecular analyses. In the OA and OPOA groups, articular cartilage was degenerated and Osteoarthritis Research Society International score was increased. Subchondral bone underwent abnormal remodeling, the differentiation of bone marrow mesenchymal stem cells (BMSCs) to osteoblasts and chondrocytes was reduced, and migration and adhesion of pre-osteoclasts were enhanced. Compared to the OA group, the pathological changes of OA in the OPOA group were considerably aggravated. After knee loading, however, cartilage degradation was effectively prevented, and the abnormal remodeling of subchondral bone was significantly inhibited. The differentiation of BMSCs was also improved, and the expression of Wnt/ß-catenin was elevated. Collectively, this study demonstrates that osteoporosis aggravates OA symptoms. Knee loading restores OPOA by regulating subchondral bone remodeling, and may provide an effective method for repairing OPOA.

Studies on the Detection, Expression, Glycosylation, Dimerization, and Ligand Binding Properties of Mouse Siglec-E.

CD33-related Siglecs are a family of proteins widely expressed on innate immune cells. Binding of sialylated glycans or other ligands triggers signals that inhibit or activate inflammation. Immunomodulation by Siglecs has been extensively studied, but relationships between structure and functions are poorly explored. Here we present new data relating to the structure and function of Siglec-E, the major CD33-related Siglec expressed on mouse neutrophils, monocytes, macrophages, and dendritic cells. We generated nine new rat monoclonal antibodies specific to mouse Siglec-E, with no cross-reactivity to Siglec-F. Although all antibodies detected Siglec-E on transfected human HEK-293T cells, only two reacted with mouse bone marrow neutrophils by flow cytometry and on spleen sections by immunohistochemistry. Moreover, whereas all antibodies recognized Siglec-E-Fc on immunoblots, binding was dependent on intact disulfide bonds and N-glycans, and only two antibodies recognized native Siglec-E within spleen lysates. Thus, we further investigated the impact of Siglec-E homodimerization. Homology-based structural modeling predicted a cysteine residue (Cys-298) in position to form a disulfide bridge between two Siglec-E polypeptides. Mutagenesis of Cys-298 confirmed its role in dimerization. In keeping with the high level of 9-O-acetylation found in mice, sialoglycan array studies indicate that this modification has complex effects on recognition by Siglec-E, in relationship to the underlying structures. However, we found no differences in phosphorylation or SHP-1 recruitment between dimeric and monomeric Siglec-E expressed on HEK293A cells. Phylogenomic analyses predicted that only some human and mouse Siglecs form disulfide-linked dimers. Notably, Siglec-9, the functionally equivalent human paralog of Siglec-E, occurs as a monomer.

In situ synthesis of robust conductive cellulose/polypyrrole composite aerogels and their potential application in nerve regeneration.

Nanostructured conductive polymers can offer analogous environments for extracellular matrix and induce cellular responses by electric stimulation, however, such materials often lack mechanical strength and tend to collapse under small stresses. We prepared electrically conductive nanoporous materials by coating nanoporous cellulose gels (NCG) with polypyrrole (PPy) nanoparticles, which were synthesized in situ from pyrrole monomers supplied as vapor. The resulting NCG/PPy composite hydrogels were converted to aerogels by drying with supercritical CO2, giving a density of 0.41-0.53 g cm(-3), nitrogen adsorption surface areas of 264-303 m(2) g(-1), and high mechanical strength. The NCG/PPy composite hydrogels exhibited an electrical conductivity of up to 0.08 S cm(-1). In vitro studies showed that the incorporation of PPy into an NCG enhances the adhesion and proliferation of PC12 cells. Electrical stimulation demonstrated that PC12 cells attached and extended longer neurites when cultured on NCG/PPy composite gels with DBSA dopant. These materials are promising candidates for applications in nerve regeneration, carbon capture, catalyst supports, and many others.

Study on the quality characteristics of hot-dry noodles by microbial polysaccharides.

The effect of curdlan gum (CG), gellan gum (GG), and xanthan gum (XG) on the quality characteristics of hot-dry noodles (HDN) was investigated. The rheology properties were used to evaluate the quality of the dough, the textural, viscosity, cooking characteristics and water states were investigated to study the quality changes of HDN. Three microbial polysaccharides were found that it could improve the quality of wheat flour and significantly increase the starch viscosity of HDN and delay the water migration rate of HDN. When 0.2% CG, 0.5% GG, and 0.5% XG were added, the HDN showed the best flour swelling power, texture, and tensile properties, and the structure of gluten network was significantly improved. The flourier transform infrared spectroscopy results showed that microbial polysaccharides with appropriate concentrations changed the formation of hydrogen bond in HDN, decreased α-helix and increased ß-turn content. Meanwhile, the relative continuous and complete gluten network was formed, which could be proven by microstructure observation. This study provides a reference for functionality applications of HDN with microbial polysaccharides.

Study on inhibitory activity and mechanism of chitosan oligosaccharides on Aspergillus Flavus and Aspergillus Fumigatus.

Chitosan oligosaccharides (COS) are a derivative of low molecular weight chitosan and are potent natural antimicrobial agents. The antimicrobial activity of COS against Aspergillus flavus and Aspergillus fumigatus was evaluated by minimum inhibitory concentration (MIC) and inhibition of mycelial growth. The MICs of COS against these two fungi were 31.2 and 15.6 mg/mL, respectively. COS treatment rendered fungal mycelia wrinkled and deformed with a fractured appearance. COS also increased cellular permeability leading to a significant leakage of cellular components indicating membrane damage. This compound also dose-dependently reduced chitin production and enhanced chitinase activity while enhancing the accumulation of reactive oxygen species (ROS). These characteristics suggested that COS has inhibitory effects against food spoilage fungi and acts on the cell wall and membrane and alters cellular metabolism. COS shows promise for food industry applications since it is non-toxic to higher organisms.

Mechanical loading mitigates osteoarthritis symptoms by regulating the inflammatory microenvironment in a mouse model.

Osteoarthritis (OA) is one of the most common chronic diseases, in which inflammatory responses in the articular cavity induce chondrocyte apoptosis and cartilage degeneration. While mechanical loading is reported to mitigate synovial inflammation, the mechanism and pathways for the loading-driven improvement of OA symptoms remain unclear. In this study, we evaluated the loading effects on M1/M2 polarization of synovial macrophages via performing histology, cytology, and molecular analyses. In the OA group, the cell layer of the synovial lining was enlarged with an increase in cell density. Also, M1 macrophages were polarized and proinflammatory cytokines were increased. In contrast, in the OA group with mechanical loading, cartilage degradation was reduced and synovial inflammation was alleviated. Notably, the M1 macrophages were diminished by mechanical loading, while M2 macrophages were increased. Furthermore, mechanical loading decreased the levels of proinflammatory cytokines, such as interleukin-1 beta and tumor necrosis factor-α, and suppressed PI3K/AKT/NF-κB signaling. Taken together, this study demonstrates that mechanical loading changes the ratio of M1 and M2 macrophages via regulation of PI3K/AKT/NF-κB signaling and provides cartilage protection in the mouse OA model.

Single Molecular Layer of Chitin Sub-Nanometric Nanoribbons: One-Pot Self-Exfoliation and Crystalline Assembly into Robust, Sustainable, and Moldable Structural Materials.

Sub-nanometric materials (SNMs) represent a series of unprecedented size-/morphology-related properties applicable in theoretical research and diverse cutting-edge applications. However, in-depth investigation and wide utilization of organic SNMs are frequently hindered, owing to the complex synthesis procedures, insufficient colloidal stability, poor processability, and high cost. In this work, a low-cost, energy-efficient, convenient, effective, and scalable method is demonstrated for directly exfoliating chitin SNMs from their natural sources through a one-pot "tandem molecular intercalation" process. The resultant solution-like sample, which exhibits ribbon-like feature and contains more than 85% of the single molecular layer (thickness <0.6 nm), is capable of being solution-processed to different types of materials. Thanks to the sub-nanometric size and rich surface functional groups, chitin SNMs reveal versatile intriguing properties that rarely observe in their nano-counterparts (nanofibrils), e.g., crystallization-like assembly in the colloidal state and alcoplasticity/self-adhesiveness in the bulk aggregate state. The finding in this work not only opens a new avenue for the high value-added utilization of chitin, but also provides a new platform for both the theoretical study and practical applications of organic SNMs.

Effects of chitosan oligosaccharide and hyriopsis cumingii polysaccharide on the quality of wheat flour and extruded flour products.

ABSTRACT: Effects of chitosan oligosaccharide (COS) and hyriopsis cumingii polysaccharide (HCP) on the quality of wheat flour and corresponding extruded flour products were investigated in this work. The results showed that both COS and HCP are conducive to the improvement of dough quality. Moreover, compared to control group samples, the moisture content, expansion ratio and oil absorption rate of the samples were increased and the hardness were decreased with the addition of COS. These phenomena indicate the quality of extruded flour products became better in the presence of COS as well. However, HCP has little or no effect on the quality of extruded flour products may be due to its degradation under high temperature and pressure extrusion. COS with higher stability exhibited better improvement effects on the quality of extruded flour products and showed a promising prospect for application in extruded food industry.

Pseudosolvent Intercalator of Chitin: Self-Exfoliating into Sub-1 nm Thick Nanofibrils for Multifunctional Chitinous Materials.

Traditionally, energy-intensive and time-consuming postmechanical disintegration processes are inevitable in extracting biopolymer nanofibrils from natural materials and thereby hinder their practical applications. Herein, a new, convenient, scalable, and energy-efficient method for exfoliating nanofibrils (ChNFs) from various chitin sources via pseudosolvent-assisted intercalation process is proposed. These self-exfoliated ChNFs possess controllable thickness from 2.2 to 0.8 nm, average diameter of 4-5 nm, high aspect ratio up to 103 and customized surface chemistries. Particularly, compared with elementary nanofibrils, ChNFs with few molecular layers thick exhibit greater potential to construct high-performance structural materials, e.g., ductile nanopapers with large elongation up to 70.1% and toughness as high as 30.2 MJ m-3 , as well as soft hydrogels with typical nonlinear elasticity mimicking that of human-skin. The proposed self-exfoliation concept with unique advantages in the combination of high yield, energy efficiency, scalable productivity, less equipment requirements, and mild conditions opens up a door to extract biopolymer nanofibrils on an industrial scale. Moreover, the present modular ChNFs exfoliation will facilitate researchers to study the effect of thickness on the properties of nanofibrils and provide more insight into the structure-function relationship of biopolymer-based materials.

Effect of hydrocolloids on physical, thermal and microstructure properties of par-baked baguette during frozen storage.

The retrogradation of starch occurs in the process of freezing storage of par-baked baguette, resulting in easy staling and a decrease of consumer acceptance. The objective of this study was to assess whether the staling of par-baked baguette could be improved by the addition of Arabic gum (AG), Sodium alginate (SA), and Sesbania gum (SG). The physical, thermal dynamic, and microstructure properties of par-baked baguette during frozen storage were analyzed. The addition of hydrocolloid increased the moisture of the baguette and delayed the water migration, which was beneficial to improve the dough formation and gas capacity, hinder the growth of ice crystals, and reduce the hardness of the baguette. These properties were more pronounced with increasing freezing storage periods. These hydrocolloids could slow down the rate of recrystallization, which reduced the enthalpy change and crystallinity of par-baked baguette. It was also found that the hydrocolloids incorporated baguette was smooth in the crumb microstructure. In general, these results suggested that the incorporation of hydrocolloids improved the quality and anti-staling mechanism of the par-baked baguette during frozen storage which can be used as potential improvers to increase freezing stability in the formulation of the baguette.

Effects of inulin on protein in frozen dough during frozen storage.

Effects of inulin on protein in frozen dough during frozen storage were investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC) and scanning electron microscopy (SEM). The strength of electrophoretic bands in A1 (32-57 kDa) and A2 (20-25 kDa) regions and the content of freezable water and sulfhydryl in frozen dough with inulin were lower than those of the blank under the same storage time. The gluten structure of frozen dough with 2.5 wt% long-chain inulin was more dense and compact than that of the sample with 5.0 wt% short-chain inulin after 2 weeks. Moreover, 4 weeks later, the change of the α-helix and ß-sheet with 2.5 wt% long-chain inulin was lowest. These characteristics suggested that long-chain inulin exhibited a better protection effect on protein in frozen dough and showed a promising prospect for application in the food industry.

Novel decoy cellular vaccine strategy utilizing transgenic antigen-expressing cells as immune presenter and adjuvant in vaccine prototype against SARS-CoV-2 virus.

A novel approach modifying cells to express viral markers to elicit protective immunity responses (decoy cellular vaccination) in the prevention of COVID-19 disease is currently being explored. Our approach entails utilizing SARS-CoV-2 Spike antigen-expressing, non-replicating cells as carriers and presenters of immunogenic antigens, so called "I-cells". By using irradiated cells as presenting vehicles of SARS-CoV-2 viral antigens(s) in a cellular context, these presented viral proteins can be recognized by the host immune system, thus, an efficient protective immune response might be elicited. Another advantage of this strategy is that the manufacturing process is scalable and yields uniform cell products allowing for "off-the-shelf" frozen supply availability. To prevent engraftment and proliferation of the cells after administration, the cells will be irradiated post-harvesting abolishing in vivo replication potential. Specifically, immunoreactive Spike-1 proteins from SARS-CoV-2 are expressed on the surface of irradiated target I-cells. Utilizing this innovative strategy, these viral antigen-displaying decoy cells will be developed as a vaccine to protect against COVID-19 disease.

Natural rubber bio-nanocomposites reinforced with self-assembled chitin nanofibers from aqueous KOH/urea solution.

Rigid chitin nanofibers (ChNFs) self-assembled from dilute α-chitin/KOH/urea aqueous solution were utilized as 1D filler to reinforce soft natural rubber (NR). The prepared ChNFs suspension has good compatibility with natural rubber latex (NRL) and thus showing favorable dispersibility in NR matrix at nanoscale. The bio-nanocomposites were fabricated by casting and evaporating the pre-mixed NRL/ChNFs suspensions with different ChNFs loadings. Gratifyingly, the NR/ChNFs bio-nanocomposite with only 0.3 wt% ChNFs content presented distinct improvement in both the strength and toughness due to the large aspect ratio of ChNFs and its homogeneous dispersion in NRL matrix. Moreover, the introduction of ChNFs can promote the proliferation of mBMSCs effectively and endow NR/ChNFs bio-nanocomposites with good biocompatibility, enabling expanded applications of NR in biomedical field, such as artificial blood vessel, cosmetology prosthesis and human diaphragm materials.

Ultrafine and carboxylated ß-chitin nanofibers prepared from squid pen and its transparent hydrogels.

TEMPO-mediated oxidation has been successfully used to prepare carboxylated chitin nanofibers (ChNFs) with purified chitins originating from the outer shells of crab and shrimp (α-form) or tubeworm (ß-form). However, the method for obtaining carboxylated ChNFs with squid pen chitin (hydrated ß-form) has not been developed yet. It might be due to the existence of the small amount of partial deacetylation (DD ≈ 9%) in the squid pen ß-chitins. Herein, ultrafine (2-4 nm in width and several micrometers in length) and carboxylated ß-ChNFs were fabricated directly from the squid pen with simultaneously removing of protein, CaCO3 and other non-chitin components by one-step oxidation procedure using ammonium persulfate (APS) and followed ultrasonic disintegration under acid conditions. When 45 wt% APS was used to react with squid pen, the carboxylate content of ChNFs reaches 0.802 mmol/g. Therefore, the ß-ChNFs with anionically charged groups (COO-) can be dispersed stably in aqueous solution under basic conditions. Meanwhile, thus-obtained ß-ChNFs aqueous solution even with very low concentration (0.8%) can be transformed to transparent, robust and moldable hydrogels by gas coagulation of acetic acid.

PhDHS Is Involved in Chloroplast Development in Petunia.

Deoxyhypusine synthase (DHS) is encoded by a nuclear gene and is the key enzyme involved in the post-translational activation of the eukaryotic translation initiation factor eIF5A. DHS plays important roles in plant growth and development. To gain a better understanding of DHS, the petunia (Petunia hybrida) PhDHS gene was isolated, and the role of PhDHS in plant growth was analyzed. PhDHS protein was localized to the nucleus and cytoplasm. Virus-mediated PhDHS silencing caused a sectored chlorotic leaf phenotype. Chlorophyll levels and photosystem II activity were reduced, and chloroplast development was abnormal in PhDHS-silenced leaves. In addition, PhDHS silencing resulted in extended leaf longevity and thick leaves. A proteome assay revealed that 308 proteins are upregulated and 266 proteins are downregulated in PhDHS-silenced plants compared with control, among the latter, 21 proteins of photosystem I and photosystem II and 12 thylakoid (thylakoid lumen and thylakoid membrane) proteins. In addition, the mRNA level of PheIF5A-1 significantly decreased in PhDHS-silenced plants, while that of another three PheIF5As were not significantly affected in PhDHS-silenced plants. Thus, silencing of PhDHS affects photosynthesis presumably as an indirect effect due to reduced expression of PheIF5A-1 in petunia. Significance: PhDHS-silenced plants develop yellow leaves and exhibit a reduced level of photosynthetic pigment in mesophyll cells. In addition, arrested development of chloroplasts is observed in the yellow leaves.

Effect of surface chemistry on the dispersion and pH-responsiveness of chitin nanofibers/ natural rubber latex nanocomposites.

Chitin nanofibers (ChNFs) have emerged as a rising nanomaterial due to their excellent mechanical properties, biocompatibility and biodegradability etc. Herein, carbonylated ChNFs (C-ChNFs) and zwitterionic ChNFs (NC-ChNFs) decorated with both amino (-NH2) and carboxyl (-COOH) groups were used to prepare ChNFs/natural rubber (NR) nanocomposite films by dip molding method, respectively. The results showed that C-ChNFs had better dispersion than that of NC-ChNFs in NRL matrix. Moreover, C-ChNFs/NR nanocomposite films demonstrated obvious pH-responsiveness owing to the association and disassociation of the hydrogen bonding between C-ChNFs under various pH conditions. In contrast, NC-ChNFs/NR nanocomposite films showed pH-stable mechanical properties because the protonation of -NH2 at pH < 7 and the ionization of -COOH at pH > 7 resulted in electrostatic repulsive force between NC-ChNFs. This study demonstrates that surface chemistry of ChNFs plays an important role on tailoring the performance of ChNFs/NRL nanocomposites.

Simultaneous improvement of thermal stability and redispersibility of cellulose nanocrystals by using ionic liquids.

Cellulose nanocrystals (CNCs) are predominantly obtained by the traditional sulfuric acid hydrolysis process. However, as-prepared CNCs powder features low thermal stability and poor redispersibility due to the existence of sulfonate groups and the hydrogen bond interaction among particles. Herein, by mixing the ionic liquid [BMIm][BF4] with freshly prepared CNCs without dialysis through a simple rotary evaporate procedure, the simultaneous improvement of thermal stability and redispersibility of CNCs has been achieved. By combining FTIR, TGA and DLS measurements, the critical role of rotary evaporates process for improving the thermal stability of CNCs has been discussed. Furthermore, the poly(lactic acid) (PLLA)/IL-CNC nanocomposites with enhanced mechanical properties were prepared by the melt-mixing method. This study provides a green and simple strategy for preparing dried CNC powders, which has a great potential in large-scale production of fully bio-based nanocomposites.

One-pot synthesis of graphene/chitin nanofibers hybrids and their remarkable reinforcement on Poly(vinyl alcohol).

Novel hybrid nanomaterials composed of graphene and chitin nanofibers (ChNFs) were successfully prepared by one-pot ball milling. Under strong shear and collision force of ball milling, graphite was exfoliated to mono-layer or few-layer graphene with the assistance of chitin nanofibers. Unexpectedly, the hybridization of exfoliated graphene and ChNFs was realized simultaneously. Morphology analysis observed that the ChNFs were adsorbed tightly on the surface of graphene, providing for reduced graphene hydrophobicity and enhanced stability of the hybrid dispersion. In addition, the concentration of exfoliated graphene reaches up to 1.5 mg ml-1. Strong interaction between graphene and ChNFs may benefit from the large amounts of carboxylate groups on the surface of ChNFs, which was prepared by TEMPO-mediated oxidation of chitin. As prepared graphene/ChNFs hybrids can remarkably enhance both the tensile strength and toughness of Poly(vinyl alcohol). This study provides a green, simple and large-scale synthesis method for preparing water-dispersible graphene/ChNFs hybrid nanobuilding blocks, which shows great promise potential in various applications requiring biocompatibility, hydrophilicity, electrical conductivity and strong mechanical properties.