Unraveling the immune functions of large yellow croaker Tmem208 in response to Pseudomonas plecoglossicida: Insights from cloning, expression profiling, and transcriptome analysis.

Pseudomonas plecoglossicida, the causative agent of Visceral White Spot Disease, poses substantial risks to large yellow croaker (Larimichthys crocea) aquaculture. Previous genome-wide association studies (GWAS), directed towards elucidating the resistance mechanisms of large yellow croaker against this affliction, suggested that the transmembrane protein 208 (named Lctmem208) may confer a potential advantage. TMEM proteins, particularly TMEM208 located in the endoplasmic reticulum, plays significant roles in autophagy, ER stress, and dynamics of cancer cell. However, research on TMEM's function in teleost fish immunity remains sparse, highlighting a need for further study. This study embarks on a comprehensive examination of LcTmem208, encompassing cloning, molecular characterization, and its dynamics in immune function in response to Pseudomonas plecoglossicida infection. Our findings reveal that LcTmem208 is highly conserved across teleost species, exhibiting pronounced expression in immune-relevant tissues, which escalates significantly upon pathogenic challenge. Transcriptome analysis subsequent to LcTmem208 overexpression in kidney cells unveiled its pivotal role in modulating immune-responsive processes, notably the p53 signaling pathway and cytokine-mediated interactions. Enhanced phagocytic activity in macrophages overexpressing LcTmem208 underscores its importance in innate immunity. Taken together, this is the first time reported the critical involvement of LcTmem208 in regulating innate immune responses of defensing P. plecoglossicida, thereby offering valuable insights into teleost fish immunity and potential strategies for the selective breeding of disease-resistant strains of large yellow croaker in aquaculture practices.

Catalytic role of the enol ether intermediate in the intramolecular Stetter reaction: a computational perspective.

The intramolecular Stetter reaction catalyzed by a carbene is investigated by density functional theory (DFT) calculations and kinetic simulations. Catalyst 1 first reacts with aldehyde 2 to give the primary adduct (PA). The PA undergoes the intramolecular oxa-Michael reaction to irreversibly generate enol ether intermediate 9. The conversion of the enol ether to the Breslow intermediate (BI) requires the assistance of a base such as the PA. The next step involves formation of a carbon-carbon bond through the Michael addition, and expulsion of the catalyst generates the Stetter product 7. Calculations show that the catalytic cycle is composed of two irreversible processes: the first one involves the exergonic formation of the enol ether intermediate, while the second one is the conversion of the enol ether to the final product. Kinetic simulations using initial concentrations of [1]0 = 0.005 M and [2]0 = 0.025 M demonstrate that under a steady-state condition, 35% of the catalyst rests on the state of the enol ether (0.0018 M). The catalyst resting state therefore consists of the unbound form (the free catalyst) and its bound form (the enol ether species). According to variable time normalization analysis, the reaction exhibits a second-order dependence (first order in catalyst and first order in substrate), which agrees with experiments. The oxa-Michael reaction to form the enol ether is identified to be turnover limiting in the intramolecular Stetter reaction, which rationalizes the observed electronic effect of the Michael acceptor on the reactivity, as well as the measured isotope effect with respect to the aldehydic proton/deuteron. The base that participates in the BI formation has a significant effect on the build-up of the resting state 9 and the active catalyst concentration. In addition, the thermodynamic stability of the enol ether is found to depend on the tether length between the aromatic aldehyde and the Michael acceptor, as well as the chemical nature of the carbene catalyst. The favorability for the oxa-Michael reaction is therefore suggested to govern the reactivity of the intramolecular Stetter transformation.

Facile fabrication of microfibrillated cellulose-based aerogels incorporated with nisin/ß-cyclodextrin microcapsule for channel catfish preservation.

In this study, a hydrophobic and antibacterial pad was prepared to preserve Channel Catfish (Ictalurus punctatus). The pad composite the microfibrillated cellulose and ß-cyclodextrin/nisin microcapsules. The hydrophobic pad ensures a dry surface in contact with the fish, reducing microbial contamination. The pad has a low density and high porosity, making it lightweight and suitable for packaging applications, while also providing a large surface area for antibacterial activity. Results demonstrated that this antibacterial pad exhibits an ultralow density of 9.0 mg/cm3 and an ultrahigh porosity of 99.10%. It can extend the shelf life of Channel Catfish fillets to 9 days at 4 °C, with a total volatile base nitrogen below 20 mg/100 g. The study proposes a novel solution for preserving aquatic products by combining antibacterial substances with the natural base material aerogel. This approach also extends the utilization of aerogel and nisin in food packaging.

Formation of Breslow Intermediates under Aprotic Conditions: A Computational Study.

The mechanism of formation of the Breslow intermediate (BI) under aprotic conditions is investigated with density functional theory (DFT) calculations. The zwitterionic adduct (ZA) is formed by the first addition of an imidazolinylidene to benzaldehyde. The forward reaction is found to proceed through the second addition of the ZA to another benzaldehyde, and subsequent proton migration gives a hemiacetal. The bimolecular reaction enables the conversion of the ZA to a more reactive hemiacetal, which is further decomposed to the BI with the assistance of the ZA. During the ZA-assisted process, the hemiacetal and the BI act as hydrogen bond donors to stabilize the ZA. The hydrogen bond interactions between the ZA and the BI or hemiacetal are analyzed. The DFT computations demonstrate that along the proposed route, the proton migration leading to the hemiacetal intermediate is the rate-determining step (ΔG⧧ = 21.2 kcal mol-1). The bimolecular mechanism provides an alternative pathway to explain BI formation under aprotic conditions.

Idebenone reduces sepsis-induced oxidative stress and apoptosis in hepatocytes via RAGE/p38 signaling.

Background: Sepsis-induced liver dysfunction is believed to be an independent risk factor for multiple organ dysfunction and death. Idebenone (IDE), a synthetic analog of coenzyme Q10 (CoQ10), possesses an antioxidizing property. The present study aimed to investigate the efficacy of IDE on sepsis-induced liver injury and discuss its reaction mechanism in vivo and in vitro. Methods: To establish an in vivo model of sepsis-induced liver injury, rats were treated with high-grade cecal ligation and puncture (CLP). Hematoxylin-eosin staining was applied to observe the liver pathological changes, and liver function was examined using alanine aminotransferase (ALT) and aspartate aminotransferase (AST) assay kits. Enzyme-linked immunosorbent assay (ELISA) kits were employed to assess the levels of inflammatory cytokines in serum and tissues. The activities of malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) were analyzed using MDA, SOD, and GSH-Px assay kits, respectively. The apoptosis of liver tissues was measured by terminal deoxynucleotidyl transferase-mediated nick-end labeling (TUNEL) staining, and western blot was employed to estimate apoptosis-related proteins. In vitro, 0.5 µg/mL lipopolysaccharide (LPS) was adopted to administrate primary hepatocytes. The expressions of receptor for advanced glycation end products (RAGE)/p38-related proteins were evaluated by western blot. Cell counting kit-8 (CCK-8) and dichloro-dihydro-fluorescein diacetate (DCFH-DA) assays were utilized to estimate cell viability and reactive oxygen species (ROS) content. Moreover, the transfection efficacy of overexpression (Ov)-RAGE in primary hepatocytes was tested by real-time quantitative polymerase chain reaction (RT-qPCR) and western blot. Results: IDE could improve liver function and reduce sepsis-induced pathological damage. The levels of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and interleukin-6 (IL-6) in the serum and liver tissue of sepsis rats were suppressed by IDE. Additionally, IDE repressed the oxidative stress and apoptosis of liver tissues in sepsis-induced rats. IDE also inhibited RAGE/p38 signaling. Furthermore, IDE revived the decreased viability in LPS-induced hepatocytes concentration-dependently. After overexpressing RAGE, RAGE expression in hepatocytes was significantly elevated. Further functional experiments revealed that IDE attenuated cell viability injury, apoptosis, oxidative stress, and inflammatory damage in LPS-induced hepatocytes via RAGE/p38 signaling. Conclusions: IDE helped to protect against sepsis-induced liver injury via the regulation of RAGE/p38 signaling.

Multimodal imaging and electroretinography highlights the role of VEGF in the laser-induced subretinal fibrosis of monkey.

Age-related macular degeneration (AMD) is a leading cause of blindness. Laser-induced nonhuman primate choroidal neovascularization (CNV) is a widely used animal model of neovascular AMD. Subretinal fibrosis (SFb) is the major limiting factor of effective anti-VEGF therapy for neovascular AMD, yet SFb has never been systematically analyzed in the primate CNV model and if VEGF directly affect SFb is unknown. We recruited a large cohort of rhesus macaques to study the occurrence, multimodal imaging and electroretinography (ERG) features, and related cytokines of SFb. Here we show that among 33 rhesus macaques, 88% CNV eyes developed SFb. Spectral domain optical coherence tomography (SD-OCT) identified four types of subretinal hyper-reflective material (SHRM) of SFb in primate. Multimodal imaging is reliable for monitoring SFb and matches the histological results well. Reduced amplitude of oscillatory potentials correlates with the thinning of inner retina layers and is a possible SFb indicator. Iba1+ microglia/macrophage cells infiltrated in the fibrotic lesions, and aqueous cytokine analysis identified four fibrosis-related factors (GM-CSF, IL-10, TGFß2 and VEGF). Unexpectedly, we found sustained expression of VEGF may be an important inducer of SFb, and anti-VEGF therapy actually partially suppresses SFb. Taken together, our data suggest the laser-induced primate SFb model, coupled with multimodal imaging and ERG recording, is a useful system to dissect the pathogenesis and explore the rationale of treatment for SFb; and combined therapy with anti-VEGF and anti-fibrosis agents is necessary for AMD treatment.

Effects of simulated saliva-gastrointestinal digestion on the physicochemical properties and bioactivities of okra polysaccharides.

This study was to investigate the effects of in vitro simulated saliva-gastrointestinal digestion on the physicochemical properties and bioactivities of okra polysaccharides (OPS). Results showed that the digestibilities of OPS were about 5.1%, 37.5%, and 41.3% after saliva digestion (SD), saliva-gastric digestion (SGD), and saliva-gastrointestinal digestion (SGID), respectively. The SGID significantly changed the physicochemical properties of OPS, such as total uronic acids, total flavonoids, monosaccharide composition, rheological properties, and molecular weights (Mw). Especially, Mw changes resulted in the breakdown of glycosidic bonds during SGD, and the degradation of OPS during SGID was mainly caused by disrupting aggregates. Furthermore, the bioactivities of OPS were also affected by SGID. After SGID, OPS still possessed strong antioxidant activities, binding capacities, and prebiotic activities, but the α-glucosidase inhibitory effect was obviously decreased. Overall, results can provide valuable and scientific support on the oral administration of OPS as functional foods and medicines in the future.

Analyses on Molecular Properties of the Diamidinate CrI-CrI Complex by Multireference and DFT Approaches.

The model system of the diamidinate CrI-CrI complex is investigated by wave function theory (WFT) and Kohn-Sham density functional theory (KS-DFT). The multireference perturbation theory (RASPT2) estimates a stabilization energy of ca. 20 kcal mol-1 for the δ bonding. The multiconfiguration pair-density functional theory (MC-PDFT) with the ftPBE functional well predicts the singlet energy curve comparable to the RASPT2 level. For the KS-DFT scheme based on a single determinant, seven functionals including BP86, BLYP, PBE, B3LYP, M06-L, M06, and ωB97X-D are assessed: two types of functionals are classified according to the nature of the restricted and broken symmetry potential energy curves. The broken symmetry scheme with the type I functionals can give good results for the energy curve in agreement with the multireference calculations. In regard to the metal-metal bonding, the restricted KS-DFT calculations performed by all of the seven functionals yield inferior description due to the lack of significant multiconfigurational character. The Mayer bond order, the electron localization function, and electron density predicted by the broken symmetry formalism with the type II functionals are consistent with those obtained with the multireference theory.

Can the Radical Channel Contribute to the Catalytic Cycle of N-Heterocyclic Carbene in Benzoin Condensation?

NHC can catalyze benzoin condensation via the key Breslow intermediate. EPR spectroscopy recently confirmed the existence of the radical species, but its catalytic role is still unclear. Herein, we use density functional approaches to study the radical-associated pathway in comparison with the nonradical mechanism reported previously. Theoretical investigations show that the nonradical path (Δ G⧧ = 18.7 kcal/mol) is more kinetically favorable than the radical route (Δ G⧧ = 27.6 kcal/mol), which is initialized by the hydrogen abstraction from the Breslow intermediate by benzaldehyde, leading to a radical pair. The product formation is thus dominated by the nonradical pathway. In addition, the Breslow intermediate is less stable than its keto form, which blocks the benzoin condensation, and the radical species could play an important role in assisting the tautomerization and promoting the catalytic reaction.

Reversible Cleavage/Formation of the Chromium-Chromium Quintuple Bond in the Highly Regioselective Alkyne Cyclotrimerization.

Herein we report the employment of the quintuply bonded dichromium amidinates [Cr{κ2 -HC(N-2,6-i Pr2 C6 H3 )(N-2,6-R2 C6 H3 )}]2 (R=iPr (1), Me (7)) as catalysts to mediate the [2+2+2] cyclotrimerization of terminal alkynes giving 1,3,5-trisubstituted benzenes. During the catalysis, the ultrashort Cr-Cr quintuple bond underwent reversible cleavage/formation, corroborated by the characterization of two inverted arene sandwich dichromium complexes (µ-η6 :η6 -1,3,5-(Me3 Si)3 C6 H3 )[Cr{κ2 -HC(N-2,6-i Pr2 C6 H3 )(N-2,6-R2 C6 H3 )}]2 (R=i Pr (5), Me (8)). In the presence of σ donors, such as THF and 2,4,6-Me3 C6 H2 CN, the bridging arene 1,3,5-(Me3 Si)3 C6 H3 in 5 and 8 was extruded and 1 and 7 were regenerated. Theoretical calculations were employed to disclose the reaction pathways of these highly regioselective [2+2+2] cylcotrimerization reactions of terminal alkynes.

Enantiodivergent Steglich rearrangement of O-carboxylazlactones catalyzed by a chirality switchable helicene containing a 4-aminopyridine unit.

A pseudo-enantiomeric pair of optically switchable helicenes containing a catalytic 4-N-methylaminopyridine (MAP) bottom unit and a C2-symmetric, (10R,11R)-dimethoxymethyl-dibenzosuberane top template was synthesized. They underwent complementary photoswitching at 290 nm (P/M', <1/>99) and 340 nm (P/M', 91/9) and unidirectional thermo-rotation at 130 °C (P/M', >99/<1). They were utilized to catalyze enantiodivergent Steglich rearrangement of O- to C-carboxylazlactones, with formation of either enantiomer with up to 91% ee (R) and 94% ee (S), respectively.

Enzyme Catalysis that Paves the Way for S-Sulfhydration via Sulfur Atom Transfer.

S-sulfhydration is generally anticipated to proceed through the transfer of the SH group (Nu-SH···(-)S-R → Nu(-)···HS-S-R). The other route involves the sulfur atom (S(0)) transfer between two sulfhydryl anions (Nu-S(-)···(-)S-R → Nu(-)···(-)S-S-R) and is considered electrostatically unfavorable. Mercaptopyruvate sulfurtransferase (MST, PDB code: 4JGT ) catalyzes sulfur transfer from mercaptopyruvate to sulfur acceptors, and the first step of the reaction is the formation of cysteine (Cys248) persulfide via S-sulfhydration. Mechanistic studies on S-sulfhydration in MST using QM/MM methods show that the sulfur atom transfer initialized by the deprotonation of the Ser250/His74/Asp63 triad is kinetically preferred to the SH-promoted sulfur transfer. The calculated barrier of approximately 16 kcal mol(-1) for the S(0) transfer agrees well with experimental results. The electrostatic repulsion during the S(0) transfer can be sophisticatedly reduced by the aid of the Cys248-Gly249-Ser250-Gly251-Val252-Thr253 (CGSGVT) loop. Electrostatic potentials and frontier orbitals are also analyzed for the persulfide anion surrounded by the loop. The sulfur atom transfer which is seldom regarded possible is therefore facilitated with the assistance of the triad and the loop in the enzyme.

A computational study of the activation of allenoates by Lewis bases and the reactivity of intermediate adducts.

Several chemical properties of Lewis base-allenoate adducts (LB·allenoate), such as solvent effect, basicity, nucleophilicity and cycloaddition, are studied to provide a detailed foundation for the analysis of LB-catalyzed reactions of allenoates. The zwitterionic LB·allenoates formed between methyl allenoate and Lewis bases, such as N-heterocyclic carbenes (NHCs), phosphines, amines and aza-heterocycles, are studied at the M06-2X/6-31+G* level. The addition of the LBs to the allenoate can yield Z- or E-type adducts. The formation of the Z-type adducts is more favorable in the gas phase due to electrostatic interactions. The yield of the E-type adducts increases with the permittivity of the solvent. The lowest barriers for the addition and the most stable adducts are observed with NHCs as catalysts. It is also shown that the α-carbon atom of the allenic moiety in LB·allenoate is more nucleophilic than the γ-carbon atom. Aza-arenes, phosphines and NHCs stabilize the [3 + 2]-ylides formed by the cycloaddition of LB·allenoate to ethylene; therefore, these LBs thermodynamically support the [3 + 2] cycloadditions. The detailed analysis of [3 + 2]-, [2 + 4]-, [2 + 2]- and [2 + 2 + 2]-cycloadditions with enones/ketones shows that the amine-catalyzed reactions follow the kinetically preferred path, and that the exergonic formation of the P-ylide favors the [3 + 2] cycloaddition in the phosphine-catalyzed reaction. The thermodynamically preferred pathway is followed with NHCs whereas the high stability of NHC·allenoate adducts reduces the overall catalytic efficiency of NHCs.

A computational study: reactivity difference between phosphine- and amine-catalyzed cycloadditions of allenoates and enones.

Allenoates and enones form cyclopentenes via a phosphine-catalyzed [3 + 2] cycloaddition while the amine-catalyzed [2 + 4] cycloaddition yields dihydropyrans or pyrans. The difference between these catalysts is studied with M06-2X/6-31+G* calculations. The addition of the catalyst to the allenoate is the first step in both pathways followed by the reaction with the enone. The formation of the [3 + 2] phosphorus-ylide is exergonic, and hence, the [3 + 2] cycloaddition is kinetically favored over the [2 + 4] addition. Amines do not stabilize [3 + 2] ammonium-ylides. However, electron-withdrawing groups on the enone enable [2 + 4] cycloadditions. The strength of the electron-withdrawing group further controls the α/γ regioselectivity of the [2 + 4] cycloaddition, and the analysis of the HOMO-LUMO interactions explains why only E-dihydropyrans from the direct γ-[2 + 4] cycloaddition have been observed in experiments. The quantum calculations further reveal a new path to the α-[2 + 4] product starting with an intermediate Rauhut-Currier reaction. This new path is kinetically favored over the direct amine-catalyzed α-[2 + 4] cycloaddition.

[Update of research on hyperthermia in relief of bone metastasis pain].

Hyperthermia is an efficient type of cancer treatment in which body tissue is exposed to high temperatures to damage and kill cancer cells. Previous studies have focused on the treatment of tumor, however,it can not substitute for traditional methols. In recent years,new research in shows hyperthermia plays an important role in bone metastasis pain control because of the advantages of width rang,rapid onset and noninvasive, and it is therefore well used in. It is also becoming one of classical methods for bone metastasis from cancer. This article reviews recent research and progress of mechanisms of hyperthermia in relief of bone metastasis pain.