Rational design and comparison of three curcumin-based fluorescent probes for viscosity detection in living cells and zebrafish.

Viscosity is a crucial indicator of the cellular microenvironment, which can affect the normal level of cellular metabolism. Aberrant levels of viscosity can result in the emergence of a variety of physiological problems including diabetes, Parkinson's disease, inflammation, etc. Therefore, it is crucial to exploit effective assays that can detect viscosity levels in living cells and organisms. Three new nitrogen-containing heterocyclic fluorescent probes, CNO, CNN and CNNB, were designed and prepared by coupling curcumin with isoxazole, pyrazole, and phenylpyrazole rings, respectively. The fluorescence response properties of these probes to the viscosity level were analyzed in parallel. All the probes, CNO, CNN and CNNB, exhibited a significantly enhanced fluorescence response to viscosity in a broad pH range with excellent photostability, sensitivity and anti-interference ability. The sensing mechanisms of these probes for viscosity were verified by DFT calculations. In addition, these probes were successfully employed for detecting viscosity levels in living HeLa cells and zebrafish. This research compares the viscosity-responsive capabilities of curcumin-based fluorescent probes containing different nitrogen-containing heterocyclic structures, and provides a new design strategy and guidance for developing curcumin-based fluorescent probes for viscosity analysis.

Computational design and engineering of self-assembling multivalent microproteins with therapeutic potential against SARS-CoV-2.

Multivalent drugs targeting homo-oligomeric viral surface proteins, such as the SARS-CoV-2 trimeric spike (S) protein, have the potential to elicit more potent and broad-spectrum therapeutic responses than monovalent drugs by synergistically engaging multiple binding sites on viral targets. However, rational design and engineering of nanoscale multivalent protein drugs are still lacking. Here, we developed a computational approach to engineer self-assembling trivalent microproteins that simultaneously bind to the three receptor binding domains (RBDs) of the S protein. This approach involves four steps: structure-guided linker design, molecular simulation evaluation of self-assembly, experimental validation of self-assembly state, and functional testing. Using this approach, we first designed trivalent constructs of the microprotein miniACE2 (MP) with different trimerization scaffolds and linkers, and found that one of the constructs (MP-5ff) showed high trimerization efficiency, good conformational homogeneity, and strong antiviral neutralizing activity. With its trimerization unit (5ff), we then engineered a trivalent nanobody (Tr67) that exhibited potent and broad neutralizing activity against the dominant Omicron variants, including XBB.1 and XBB.1.5. Cryo-EM complex structure confirmed that Tr67 stably binds to all three RBDs of the Omicron S protein in a synergistic form, locking them in the "3-RBD-up" conformation that could block human receptor (ACE2) binding and potentially facilitate immune clearance. Therefore, our approach provides an effective strategy for engineering potent protein drugs against SARS-CoV-2 and other deadly coronaviruses.

A highly effective "naked eye" colorimetric and fluorimetric curcumin-based fluorescent sensor for specific and sensitive detection of H2O2in vivo and in vitro.

Hydrogen peroxide (H2O2) is involved in many important tasks in normal cell metabolism and signaling. However, abnormal levels of H2O2 are associated with the occurrence of several diseases. Therefore, it is important to develop a new method for the detection of H2O2in vivo and in vitro. A turn-off sensor, 2,2-difluoro-4,6-bis(3-methoxy-4-((4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)oxy)styryl)-2H-1,3,2-dioxaborine (DFCB), based on curcumin was developed for the detection of H2O2. The DFCB, an orange-emitting sensor, was constructed by employing 2,2-difluoro-4,6-bis(4-hydroxy-3-methoxystyryl)-2H-1,3,2-dioxaborine (DFC) as the main carrier, and 2-(4-bromomethylphenyl)-4,4,5,5-tetramethyl-1,3,2-doxaborolane as the recognition site. The recognition group on the DFCB sensor could be completely cleaved by H2O2 to generate the intermediate DFC, which would lead to a colorimetric change from bright orange to light blue accompanying by a significantly quenched fluorescence, which could be seen by the naked eye. This sensor exhibited a highly specific fluorescence response to H2O2, in preference to other relevant species, with an excellent anti-interference performance. The sensor DFCB also possessed some advantages including a wide pH response range (6-11), a broad linear range (0-300 µM), and a low detection limit (1.31 µM). The sensing mechanism of the DFCB sensor for H2O2 was verified by HRMS analysis, 1H-NMR titration and DFT calculations. In addition, the use of the DFCB sensor was compatible with the fluorescence imaging of H2O2 in living cells and zebrafish.

Full-Length Model of SaCas9-sgRNA-DNA Complex in Cleavage State.

Staphylococcus aureus Cas9 (SaCas9) is a widely used genome editing tool. Understanding its molecular mechanisms of DNA cleavage could effectively guide the engineering optimization of this system. Here, we determined the first cryo-electron microscopy structure of the SaCas9-sgRNA-DNA ternary complex. This structure reveals that the HNH nuclease domain is tightly bound to the cleavage site of the target DNA strand, and is in close contact with the WED and REC domains. Moreover, it captures the complete structure of the sgRNA, including the previously unresolved stem-loop 2. Based on this structure, we build a full-length model for the ternary complex in cleavage state. This model enables identification of the residues for the interactions between the HNH domain and the WED and REC domains. Moreover, we found that the stem-loop 2 of the sgRNA tightly binds to the PI and RuvC domains and may also regulate the position shift of the RuvC domain. Further mutagenesis and molecular dynamics simulations supported the idea that the interactions of the HNH domain with the WED and REC domains play an important role in the DNA cleavage. Thus, this study provides new mechanistic insights into the DNA cleavage of SaCas9 and is also useful for guiding the future engineering of SaCas9-mediated gene editing systems.

Single-Disulfide Conopeptide Czon1107, an Allosteric Antagonist of the Human α3ß4 Nicotinic Acetylcholine Receptor.

Conopeptides are peptides in the venom of marine cone snails that are used for capturing prey or as a defense against predators. A new cysteine-poor conopeptide, Czon1107, has exhibited non-competitive inhibition with an undefined allosteric mechanism in the human (h) α3ß4 nicotinic acetylcholine receptors (nAChRs). In this study, the binding mode of Czon1107 to hα3ß4 nAChR was investigated using molecular dynamics simulations coupled with mutagenesis studies of the peptide and electrophysiology studies on heterologous hα3ß4 nAChRs. Overall, this study clarifies the structure-activity relationship of Czon1107 and hα3ß4 nAChR and provides an important experimental and theoretical basis for the development of new peptide drugs.

Truxene-Centered Electron Acceptors for Non-Fullerene Solar Cells: Alkyl Chain and Branched Arm Engineering.

A series of symmetrical truxene-centered and 3-ethylrhodanine end-capped electron acceptors with high absorption coefficient, namely Tr(Hex)6-3RD, Tr(Dec)6-3RD, and Tr(Hex)6-6RD, were prepared and constructed for non-fullerene solar cells. To satisfy solution-processability, multiple energy levels, and suitable morphology, these three acceptors were comparatively studied through alkyl chain (hexyl/decyl) and branched-arm engineering (three/six branched arms). The six-bladed propeller acceptor of Tr(Hex)6-6RD recorded the power conversion efficiency (PCE) of 1.1% blending with PTB7-Th without additional additives and post-processing. This work highly broadens the potential applications of star-shaped truxene building blocks in the fields of organic electronics.

Mechanical behavior in the interior and boundary of magnesium aluminate spinel (MgAl2O4) grain under nanoindentation.

The understanding of mechanical behavior in magnesium aluminate spinel (MgAl2O4) at the nanoscale lays a foundation for its material removal mechanism in ultraprecision machining. Nanoindentation tests are carried out in the interior and boundary of spinel grain with different loads. An obvious indentation size effect exhibits in both of these areas. First, the nano-hardness and elastic modulus decrease, followed by stabilization due to an increase of pressure. The measured elastic modulus, hardness, and fracture toughness of the grain interior are 277.7±8.4GPa, 19.79±0.83GPa, and 1.12±0.02MPa⋅m1/2, respectively. Deformation of spinel transits from elastic to plastic at approximately 0.8 mN load, which corresponds to the discontinuous steps of load-displacement curves. By comparing the fracture toughness and the residual indent morphology, the grain boundary exhibits lower brittleness than the grain interior. Radial cracks form on the grain surface as indentation load exceeds 29 mN, whose propagation is influenced by the loading conditions and the grain boundary effect.

Synthesis and biological evaluation of 2-phenyl-4-aminoquinolines as potential antifungal agents.

A series of 2-phenyl-4-aminoquinolines were designed, synthesized and evaluated for their antifungal activities against three phytopathogenic fungi in vitro. All of the target compounds were fully elucidated by 1H NMR, 13C NMR and HRMS spectra. The results indicated that most of the target compounds demonstrated significant activities against the tested fungi. Among them, compound 6e exhibited more promising inhibitory activities against C. lunata (EC50 = 13.3 µg/mL), P. grisea (EC50 = 14.4 µg/mL) and A. alternate (EC50 = 15.6 µg/mL), superior to azoxystrobin, a commercial agricultural fungicide. The structure-activity relationship (SAR) revealed that the aniline moiety at position 4 of the quinoline scaffold played a key role in the potency of a compound. And the substitution positions of the aniline moiety significantly influenced the activities. These encouraging results yielded a variety of 2-phenylquinolines bearing an aniline moiety acting as promising antifungal agents.

Research on the Recognition Performance of Bionic Sensors Based on Active Electrolocation for Different Materials.

Underwater object identification by optical sensors is usually difficult in turbid or dark environments. The objective of this paper was to identify different underwater materials using active electrolocation technology. We proposed a bionic sensor inspired by the weakly electric fish. The material identification was completed by analyzing electric signal images, since the electric signal changes when different materials are identified. Firstly, the effective lift-off distance for identification was researched. The materials used in this paper can be effectively identified by the sensor at a lift-off distance of 10 mm. Furthermore, the performance of the sensor for identifying and locating was studied in the presence of multiple materials. The results indicated that the sensor can effectively identify and locate the objects when the distance between objects is greater than 30 mm, while the location error is less than 5% in most cases. Our research proves that the bionic sensor we made can effectively recognize different materials underwater in short-range, which is about 10 mm. Therefore, we expect that the bionic sensor we made can be utilized as a useful tool for underwater object identification.

The effects of ethoxyquin and Angelica sinensis extracts on lipid oxidation in fish feeds and growth, digestive and absorptive capacities and antioxidant status in juvenile red carp (Cyprinus carpio var. xingguonensis): a comparative study.

Firstly, a linoleic and linolenic acid emulsion and fish feeds were incubated with graded levels of ethoxyquin (EQ) and petroleum ether extract, ethyl acetate extract (EAE), ethanol extract and aqueous extract of Angelica sinensis. The results showed that EQ and extracts of Angelica sinensis (EAs) inhibited lipid oxidation in material above. Of all of the examined EAs, EAE showed the strongest protective effects against the lipid oxidation. Moreover, EAE at high concentrations showed a stronger inhibitory effect on lipid oxidation than that of EQ. Next, 7 experimental diets that respectively supplemented 0.0, 0.2, 0.8 and 3.2 g kg-1 of EQ and EAE were fed to 280 juvenile red carp (Cyprinus carpio var. xingguonensis) with seven treatment groups for 30 days. The results indicated that dietary EAE improved growth performance in carp. Moreover, dietary EAE increased the activities of trypsin, lipase, alpha-amylase, alkaline phosphatase, glutamate-oxaloacetate transaminase and glutamate-pyruvate transaminase (GPT) and decreased plasma ammonia content in carp. Meanwhile, dietary EAE reduced the levels of malondialdehyde and raised the activities of anti-superoxide anion, anti-hydroxyl radical, superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase and the content of reduced glutathione in the hepatopancreas and intestine of carp. However, with the exception of GPT, dietary EQ got the opposite results to dietary EAE in carp. These results revealed that dietary EAE improved the digestive, absorptive and antioxidant capacities in fish. However, dietary EQ inhibited the digestive, absorptive and antioxidant capacities in fish. So, EAE could be used as a natural antioxidant for replacing EQ in fish feeds.

A Novel Method of Improving the Mechanical Properties of Propellant Using Energetic Thermoplastic Elastomers with Bonding Groups.

The relatively poor mechanical properties of extruded modified double base (EMDB) propellants limit their range of applications. To overcome these drawbacks, a novel method was proposed to introduce glycidyl azide polymer-based energetic thermoplastic elastomers (GAP-ETPE) with bonding groups into the propellant adhesive. The influence of the molecular structure of three kinds of elastomers on the mechanical properties of the resultant propellant was analyzed. It was found that the mechanical properties of the propellant with 3% CBA-ETPE (a type of GAP-ETPE that features chain extensions using N-(2-Cyanoethyl) diethanolamine and 1,4-butanediol) were improved at both 50 °C and -40 °C compared to a control propellant without GAP-ETPE. The elongation and impact strength of the propellant at -40 °C were 7.49% and 6.58 MPa, respectively, while the impact strength and maximum tensile strength of the propellant at 50 °C reached 21.1 MPa and 1.19 MPa, respectively. In addition, all three types of GAP-ETPE improved the safety of EMDB propellants. The friction sensitivity of the propellant with 3% CBA-ETPE was found to be 0%, and its characteristic drop height H50 was found to be 39.0 cm; 126% higher than the traditional EMDB propellant. These results provide guidance for studies aiming to optimize the performance of EMDB propellants.

Textbook outcome of laparoscopic hepatectomy in the context of precision surgery: A single center experience.

OBJECTIVE: Laparoscopic hepatectomy (LH) has become a common surgery for the treatment of liver tumor. To evaluate the surgical quality of laparoscopic hepatectomy under the context of precision surgery with Textbook outcome (TO), a comprehensive and holistic assessment approach. METHODS: A total of 1056 patients who underwent laparoscopic hepatectomy from May 2016 and December 2022 were enrolled in the study. All the patients were performed hepatectomy. The rate of TO and factors associated with achieving TO were examined. RESULTS: Among the 1056 patients, 75 % patients achieved TO. The main reason limited patients achieving textbook outcomes was prolonged length of hospital stay (LOS). The univariate analysis indicated that age>65, ASA classification ≥3, liver cirrhosis, tumor size > 3 cm, tumor number ≥2, type of primary cancer, and IWATE DSS were significantly associated with non-achievement of TO. The multivariate analysis indicated that the ASA classification ≥3 and advanced difficulty level in IWATE DSS independent factors associated with achieving TO. Reaching TO can significantly prolong the postoperative recurrence time and overall survival time of hepatocellular carcinoma patients. CONCLUSION: In the context of precision surgery, 75 % patients undergoing laparoscopic hepatectomy achieved a TO. Patients who achieved TO had significantly improved survival.

Discovery of a PROTAC degrader for METTL3-METTL14 complex.

N6-methyladenosine (m6A) methylation is the most abundant type of RNA modification that is mainly catalyzed by the METTL3-METTL14 methyltransferase complex. This complex has been linked to multiple cancers and is considered a promising therapeutic target for acute myeloid leukemia (AML). However, only a few METTL3 inhibitors targeting the catalytic activity were developed recently. Here, we present the discovery of WD6305 as the potent and selective proteolysis-targeting chimera (PROTAC) degrader of METTL3-METTL14 complex. WD6305 suppresses m6A modification and the proliferation of AML cells, and promotes apoptosis much more effectively than its parent inhibitor. WD6305 also affects a variety of signaling pathways related to the development and proliferation of AML. Collectively, our study reveals PROTAC degradation of METTL3-METTL14 complex as a potential anti-leukemic strategy and provides desirable chemical tool for further understanding METTL3-METTL14 protein functions.

Preparation and Properties of Hydrophobic Polyurethane Based on Silane Modification.

Waterborne coatings have obtained more and more attention from researchers with increasing concerns in environmental protection, and have the advantages of being green, environmentally friendly and safe. However, the introduction of hydrophilic groups leads to lower hydrophobicity and it is difficult to meet the requirements of complex application environments. Herein, we proposed an optimization approach of waterborne polyurethane (WPU) with vinyl tris(ß-methoxyethoxy) silane (A172), and it was found that the surface roughness, mechanical properties, thermal stability and water resistance of WPU will be increased to a certain extent with the addition of A172. Moreover, the hydrophobicity of the coating film is best when the silicon content is 10% of the acrylic monomer mass and the water contact angle reaches 100°, which could exceed two-thirds of the research results in the last decade. Therefore, our study can provide some theoretical basis for the research of hydrophobic polyurethane coatings.

Scoparia dulcis L. Extract Relieved High Stocking Density-Induced Stress in Crucian Carp (Carassius auratus).

The objective of this study was to investigate the effects of Scoparia dulcis extract (SDE) on stress induced by high stocking density and Cu and trichlorfon exposure in crucian carp (Carassius auratus). The results showed that these stressors exerted detrimental effects in fish, such as inhibition of growth performance, reduced feed intake, and interruption of fish locomotion. Under high stocking density, dietary SDE supplementation increased the content of reduced glutathione (GSH) and the activities of amylase, catalase (CAT), and glutathione reductase (GR) and decreased the content of malonaldehyde (MDA) in the intestine of crucian carp. A similar trend was presented in the hepatopancreas under Cu exposure. Dietary SDE supplementation enhanced the activities of CAT, superoxide dismutase (SOD), glutathione peroxidase (GPx), lactate dehydrogenase, glutamate-oxaloacetate transaminase, and glutamate-pyruvate transaminase in the muscle of crucian carp under trichlorfon exposure. The optimum dietary SDE supplementation levels were 4.07, 4.33, and 3.95 g kg-1 diet based on the recovery rate of weight gain (RWG), feed intake (FI), and inhibitory rate of rollover (IR) for crucian carp under high stocking density and Cu and trichlorfon exposure, respectively. Overall, dietary supplementation with SDE may be a useful nutritional strategy for relieving these stresses in aquatic animals.

Sodium alginate crosslinker engineered UCST hydrogel towards superior mechanical properties and controllable dye removal.

Smart hydrogels that can adsorb both anionic and cationic dyes and have excellent mechanical properties have not been reported yet, despite bright prospect in dye adsorption. The emergence of polyampholyte made it possible for the preparation of novel hydrogels. PMAD (copolymer of AM, AA and DMC) is a novel thermo-responsive hydrogel based on polyampholyte, whereas poor mechanical properties limit its application. Herein, we designed a novel polysaccharide crosslinker, which integrates chemical-crosslinking network and physical-entanglements to fabricate PMAD hydrogels. Chemical-crosslinking network maintains the shape of hydrogel as backbone, whereas physical-entanglements enhance the energy-dissipation capacity by disentangle upon deformation. Synergistic effect of the above structure improves the fracture strength and elongation of PMAD by nearly 4700% and 900%, respectively. Moreover, the hydrogel can be used as controllable adsorption material towards both cationic and anionic dyes, which could greatly increase adsorption capacity after reaching the critical temperature due to its unique temperature-sensitive characteristics.

Strong and Flexible High-Performance Anion Exchange Membranes with Long-Distance Interconnected Ion Transport Channels for Alkaline Fuel Cells.

Anion exchange membrane fuel cells (AEMFCs), which operate on a variety of green fuels, can achieve high power without emitting greenhouse gases. However, the lack of high ionic conductivity and long-term durability of anion-exchange membranes (AEMs) as their key components is a major obstacle hindering the commercial application of AEMFCs. Here, a series of homogeneous semi-interpenetrating network (semi-IPN) AEMs formed by cross-linking a copolymer of styrene (St) and 4-vinylbenzyl chloride (VBC) with branched polyethylenimine (BPEI) were designed. The pure carbon copolymer skeleton without sulfone/ether bonds accompanied by the semi-IPN endows the AEMs with excellent chemical stability. Moreover, the cross-linking effect of flexible BPEI chains is supposed to promote the "strong-flexible" mechanical properties, while the presence of multiquaternary ammonium groups can boost the formation of microphase separation, thereby enhancing the ionic conductivity of these AEMs. Consequently, the optimized (S1V1)3Q AEM exhibits an excellent hydroxide conductivity of 106 mS cm-1 at 80 °C, as well as more than 81% residual conductivity after soaking in 1 M NaOH at 60 °C for 720 h. Furthermore, the H2/O2 fuel cell assembled with (S1V1)3Q AEM delivers a peak power density of 150.2 mW cm-2 at 60 °C and 40% relative humidity. All results indicate that the approach of combining a pure carbon backbone polymer with a semi-IPN structure may be a viable strategy for fabricating AEMs that can be used in AEMFCs for long-term applications.

Nucleolus localization of SpyCas9 affects its stability and interferes with host protein translation in mammalian cells.

The CRISPR/Cas9 system, originally derived from the prokaryotic adaptive immune system, has been developed as efficient genome editing tools. It enables precise gene manipulation on chromosomal DNA through the specific binding of programmable sgRNA to target DNA, and the Cas9 protein, which has endonuclease activity, will cut a double strand break at specific locus. However, Cas9 is a foreign protein in mammalian cells, and the potential risks associated with its introduction into mammalian cells are not fully understood. In this study, we performed pull-down and mass spectrometry (MS) analysis of Streptococcus pyogenes Cas9 (SpyCas9) interacting proteins in HEK293T cells and showed that the majority of Cas9-associated proteins identified by MS were localized in the nucleolus. Interestingly, we further discovered that the Cas9 protein contains a sequence encoding a nucleolus detention signal (NoDS). Compared with wild-type (WT) Cas9, NoDS-mutated variants of Cas9 (mCas9) are less stable, although their gene editing activity is minimally affected. Overexpression of WT Cas9, but not mCas9, causes general effects on transcription and protein translation in the host cell. Overall, identification of NoDS in Cas9 will improve the understanding of Cas9's biological function in vivo, and the removal of NoDS in Cas9 may enhance its safety for future clinical use.

Discovery of a potent and selective proteolysis targeting chimera (PROTAC) degrader of NSD3 histone methyltransferase.

Nuclear receptor binding SET domain protein 3 (NSD3) is an attractive potential target in the therapy for human cancers. Herein, we report the discovery of a series of small-molecule NSD3 degraders based on the proteolysis targeting chimera (PROTAC) strategy. The represented compound 8 induces NSD3 degradation with DC50 values of 1.43 and 0.94 µM in NCI-H1703 and A549 lung cancer cells, respectively, and shows selectivity over two other NSD proteins. 8 reduces histone H3 lysine 36 methylation and induces apoptosis and cell cycle arrest in lung cancer cells. Moreover, the RNA sequencing and immunohistochemistry assays showed that 8 downregulates NSD3-associated gene expression. Significantly, 8, but not 1 (a reported NSD3-PWWP antagonist) could inhibit the cell growth of NCI-H1703 and A549 cells. A single administration of 8 effectively decreases the NSD3 protein level in lung cancer xenograft models. Therefore, this study demonstrated that inducing NSD3 degradation is a more effective approach inhibiting the function of NSD3 than blocking the NSD3-PWWP domain, which may provide a potential therapeutic approach for lung cancer.

Metabolomic and transcriptomic responses of mouse testis to the dextran sulfate sodium induced colitis.

Inflammatory bowel diseases (IBDs), including Crohn's disease (CD) and ulcerative colitis, are widespread in developed countries and gradually increasing in developing countries. Evidences showed that man with CD has a decrease of serum testosterone, but how IBD take effects on testicular testosterone synthesis is not well elucidated. To investigate the effects of IBD on testis, we analyzed testicular metabolome and transcriptome data of the dextran sulfate sodium (DSS) induced IBD mice. As a result, metabolomic data showed that DSS indeed induced androgen decrease in mouse testis. Correspondingly, androgen synthesis associated genes, especially Lhcgr, were down-regulated in DSS testis. From the metabolomic data, we found vitamin intake associated metabolites vitamin B2 and pyridoxamine were significantly decreased, whereas fatty acid metabolism associated molecules N-lauroylglycine and N-decanoylglycine were increased in DSS testis. In addition, we found 8-hydroxy-deoxyguanosine, a DNA oxidative damage marker, and 8-oxoguanine, a molecule responsible for DNA damage repair, were also changed in DSS testis. Simultaneously, our data also showed that DSS up-regulated the expression of meiosis initiation associated gene Stra8 and oxygen transport associated genes in testis. In summary, these results depicted the complex effects of colitis on testis. These metabolites and transcripts changed in DSS testis could be used as potential targets for IBD treatment or symptom relieve.