Depolymerization of the polyester-polyurethane by amidase GatA250 and enhancing the production of 4,4'-methylenedianiline with cutinase LCC.

Polyurethane (PU) is a complex polymer synthesized from polyols and isocyanates. It contains urethane bonds that resist hydrolysis, which decreases the efficiency of biodegradation. In this study, we first expressed the amidase GatA250, and then, assessed the enzymatic characterization of GatA250 and its efficiency in degrading the polyester-PU. GatA250 degraded self-synthesized thermoplastic PU film and postconsumption foam with degradation efficiency of 8.17% and 4.29%, respectively. During the degradation, the film released 14.8 µm 4,4'-methylenedianiline (MDA), but 1,4-butanediol (BDO) and adipic acid (AA) were not released. Our findings indicated that GatA250 only cleaved urethane bonds in PU, and the degradation efficiency was extremely low. Hence, we introduced the cutinase LCC, which possesses hydrolytic activity on the ester bonds in PU, and then used both enzymes simultaneously to degrade the polyester-PU. The combined system (LCC-GatA250) had higher degradation efficiency for the degradation of PU film (42.2%) and foam (13.94%). The combined system also showed a 1.80 time increase in the production of the monomer MDA, and a 1.23 and 3.62 times increase in the production of AA and BDO, respectively, compared to their production recorded after treatment with only GatA250 or LCC. This study provides valuable insights into PU pollution control and also proposes applicable solutions to manage PU wastes through bio-recycling.

Identification and characterization of a fungal cutinase-like enzyme CpCut1 from Cladosporium sp. P7 for polyurethane degradation.

Plastic degradation by biological systems emerges as a prospective avenue for addressing the pressing global concern of plastic waste accumulation. The intricate chemical compositions and diverse structural facets inherent to polyurethanes (PU) substantially increase the complexity associated with PU waste management. Despite the extensive research endeavors spanning over decades, most known enzymes exhibit a propensity for hydrolyzing waterborne PU dispersion (i.e., the commercial Impranil DLN-SD), with only a limited capacity for the degradation of bulky PU materials. Here, we report a novel cutinase (CpCut1) derived from Cladosporium sp. P7, which demonstrates remarkable efficiency in the degrading of various polyester-PU materials. After 12-h incubation at 55°C, CpCut1 was capable of degrading 40.5% and 20.6% of thermoplastic PU film and post-consumer foam, respectively, while achieving complete depolymerization of Impranil DLN-SD. Further analysis of the degradation intermediates suggested that the activity of CpCut1 primarily targeted the ester bonds within the PU soft segments. The versatile performance of CpCut1 against a spectrum of polyester-PU materials positions it as a promising candidate for the bio-recycling of waste plastics.IMPORTANCEPolyurethane (PU) has a complex chemical composition that frequently incorporates a variety of additives, which poses significant obstacles to biodegradability and recyclability. Recent advances have unveiled microbial degradation and enzymatic depolymerization as promising waste PU disposal strategies. In this study, we identified a gene encoding a cutinase from the PU-degrading fungus Cladosporium sp. P7, which allowed the expression, purification, and characterization of the recombinant enzyme CpCut1. Furthermore, this study identified the products derived from the CpCut1 catalyzed PU degradation and proposed its underlying mechanism. These findings highlight the potential of this newly discovered fungal cutinase as a remarkably efficient tool in the degradation of PU materials.

METACASPASE8 (MC8) Is a Crucial Protein in the LSD1-Dependent Cell Death Pathway in Response to Ultraviolet Stress.

LESION-SIMULATING DISEASE1 (LSD1) is one of the well-known cell death regulatory proteins in Arabidopsis thaliana. The lsd1 mutant exhibits runaway cell death (RCD) in response to various biotic and abiotic stresses. The phenotype of the lsd1 mutant strongly depends on two other proteins, ENHANCED DISEASE SUSCEPTIBILITY 1 (EDS1) and PHYTOALEXIN-DEFICIENT 4 (PAD4) as well as on the synthesis/metabolism/signaling of salicylic acid (SA) and reactive oxygen species (ROS). However, the most interesting aspect of the lsd1 mutant is its conditional-dependent RCD phenotype, and thus, the defined role and function of LSD1 in the suppression of EDS1 and PAD4 in controlled laboratory conditions is different in comparison to a multivariable field environment. Analysis of the lsd1 mutant transcriptome in ambient laboratory and field conditions indicated that there were some candidate genes and proteins that might be involved in the regulation of the lsd1 conditional-dependent RCD phenotype. One of them is METACASPASE 8 (AT1G16420). This type II metacaspase was described as a cell death-positive regulator induced by UV-C irradiation and ROS accumulation. In the double mc8/lsd1 mutant, we discovered reversion of the lsd1 RCD phenotype in response to UV radiation applied in controlled laboratory conditions. This cell death deregulation observed in the lsd1 mutant was reverted like in double mutants of lsd1/eds1 and lsd1/pad4. To summarize, in this work, we demonstrated that MC8 is positively involved in EDS1 and PAD4 conditional-dependent regulation of cell death when LSD1 function is suppressed in Arabidopsis thaliana. Thus, we identified a new protein compound of the conditional LSD1-EDS1-PAD4 regulatory hub. We proposed a working model of MC8 involvement in the regulation of cell death and we postulated that MC8 is a crucial protein in this regulatory pathway.

Biodegradable microplastics: Uptake by and effects on the rockpool shrimp Palaemon elegans (Crustacea: Decapoda).

Ingestion of microplastics can lead to deleterious consequences for organisms, as documented by numerous laboratory studies. The current knowledge is based on a multitude of effect studies, conducted with conventional fossil-based and non-degradable plastics. However, there is a lack of information about the acceptance and the effects of novel bio-based and biodegradable plastics. Biodegradable plastics are considered an alternative to conventional plastics and are showing rapidly growing production rates. Biodegradable plastics can disperse into the environment in the same way as conventional plastics do, becoming available to marine organisms. This study aims to provide new insights into the uptake and effects of biodegradable microplastics on marine invertebrates. Rockpool shrimp, Palaemon elegans, were fed with algal flakes coated with polylactic acid (PLA), polyhydroxybutyrate-co-valerate (PHBV) and conventional low-density polyethylene (LDPE) microparticles. Live observations showed that all of the different types of microplastics were ingested. After dissection of the shrimp, less LDPE particles were found in the stomachs than PLA and PHBV particles. This indicates a longer retention time of biodegradable microplastics compared to conventional microplastics. Presumably, less LDPE particles were ingested or evacuated from the stomach, probably by regurgitation. The ingestion of microparticles of all types of plastics induced enzymatic activity of short-chain carboxylesterases in the midgut glands of the shrimp. However, only PLA induced enzymatic activity of medium-chain carboxylesterases. Palaemon elegans showed no oxidative stress response after ingestion of microparticles, irrespective of polymer type. From our results we conclude that biodegradable plastics might have different effects than conventional plastics. The longer retention times of biodegradable plastics might enhance exposure to leaching additives and other harmful substances. Our study provides new insights into how biodegradable plastics might affect aquatic fauna and indicate that the use of biodegradable plastics needs to be reconsidered to some extent.

The secreted feruloyl esterase of Verticillium dahliae modulates host immunity via degradation of GhDFR.

Feruloyl esterase (ferulic acid esterase, FAE) is an essential component of many biological processes in both eukaryotes and prokaryotes. This research aimed to investigate the role of FAE and its regulation mechanism in plant immunity. We identified a secreted feruloyl esterase VdFAE from the hemibiotrophic plant pathogen Verticillium dahliae. VdFAE acted as an important virulence factor during V. dahliae infection, and triggered plant defence responses, including cell death in Nicotiana benthamiana. Deletion of VdFAE led to a decrease in the degradation of ethyl ferulate. VdFAE interacted with Gossypium hirsutum protein dihydroflavanol 4-reductase (GhDFR), a positive regulator in plant innate immunity, and promoted the degradation of GhDFR. Furthermore, silencing of GhDFR led to reduced resistance of cotton plants against V. dahliae. The results suggested a fungal virulence strategy in which a fungal pathogen secretes FAE to interact with host DFR and interfere with plant immunity, thereby promoting infection.

Peptidyl-tRNA hydrolase as a key player in the liberation of truncated nascent chains from the ribosomal subunit.

Svetlov et al. identify the enzyme peptidyl-tRNA hydrolase as a ribosome-associated quality-control factor that promotes hydrolysis of the dislodged peptidyl-tRNA, which helps to recycle ribosomal subunits blocked by truncated nascent chains in bacteria.

Co-Reactive Ligand In Situ Engineered Gold Nanoclusters with Ultra-Bright Near-Infrared Electrochemiluminescence for Ultrasensitive and Label-Free Detection of Carboxylesterase Activity.

Ultrasensitive and accurate monitoring of carboxylesterase (CE) activity is extremely crucial for the early diagnosis of hepatocellular carcinoma (HCC), which is still a considerable challenge. Herein, using a co-reactive ligand engineering strategy, ultra-bright near-infrared (λmax = 830 nm) and self-enhanced electrochemiluminescence (ECL) Au nanoclusters (NCs) were in situ prepared with 2-(diethylamino) ethanethiol (DEAET) as a co-reactive ligand. Remarkably, the co-reactive ligand not only acts as a stabilizer like traditional ligands but also plays a crucial role as a co-reactant to ensure a confinement effect to shorten the charge transfer distance and increase the local concentration, significantly improving the collision efficiency between the electrogenerated free radicals. Consequently, the DEAET Au NCs exhibited a record and stable anodal ECL without the addition of an exogenous co-reactant, dramatically superior to classical Au NCs and Ru(bpy)32+ with a certain amount of the co-reactant. As a proof of concept, a convenient and label-free CE biosensor was innovatively constructed using 1-naphthyl acetate as a selective substrate, achieving ultrasensitive detection for CE activity with a low limit of detection of 9.1 × 10-7 U/L. Therefore, this work not only paves a co-reactive ligand engineering strategy for in situ preparation of high-efficiency metal NCs but also provides an ultrasensitive and convenient platform for the early diagnosis of HCC.

A novel thermostable YtnP lactonase from Stenotrophomonas maltophilia inhibits Pseudomonas aeruginosa virulence in vitro and in vivo.

Infections caused by multidrug-resistant pathogens are one of the biggest challenges facing the healthcare system today. Quorum quenching (QQ) enzymes have the potential to be used as innovative enzyme-based antivirulence therapeutics to combat infections caused by multidrug-resistant pathogens. The main objective of this research was to describe the novel YtnP lactonase derived from the clinical isolate Stenotrophomonas maltophilia and to investigate its antivirulence potential against multidrug-resistant Pseudomonas aeruginosa MMA83. YtnP lactonase, the QQ enzyme, belongs to the family of metallo-ß-lactamases. The recombinant enzyme has several advantageous biotechnological properties, such as high thermostability, activity in a wide pH range, and no cytotoxic effect. High-performance liquid chromatography analysis revealed the activity of recombinant YtnP lactonase toward a wide range of N-acyl-homoserine lactones (AHLs), quorum sensing signaling molecules, with a higher preference for long-chain AHLs. Recombinant YtnP lactonase was shown to inhibit P. aeruginosa MMA83 biofilm formation, induce biofilm decomposition, and reduce extracellular virulence factors production. Moreover, the lifespan of MMA83-infected Caenorhabditis elegans was prolonged with YtnP lactonase treatment. YtnP lactonase showed synergistic inhibitory activity in combination with gentamicin and acted additively with meropenem against MMA83. The described properties make YtnP lactonase a promising therapeutic candidate for the development of next-generation antivirulence agents.

Inactivation mechanisms on pectin methylesterase by high pressure processing combined with its recombinant inhibitor.

High pressure processing (HPP) juice often experiences cloud loss during storage, caused by the activity of pectin methylesterase (PME). The combination of HPP with natural pectin methylesterase inhibitor (PMEI) could improve juice stability. However, extracting natural PMEI is challenging. Gene recombination technology offers a solution by efficiently expressing recombinant PMEI from Escherichia coli and Pichia pastoris. Experimental and molecular dynamics simulation were conducted to investigate changes in activity, structure, and interaction of PME and recombinant PMEI during HPP. The results showed PME retained high residual activity, while PMEI demonstrated superior pressure resistance. Under HPP, PMEI's structure remained stable, while the N-terminus of PME's α-helix became unstable. Additionally, the helix at the junction with the PME/PMEI complex changed, thereby affecting its binding. Furthermore, PMEI competed with pectin for active sites on PME, elucidating. The potential mechanism of PME inactivation through the synergistic effects of HPP and PMEI.

Bedside Leukocyte Esterase Testing to aid in Diagnosing Bacterial Conjunctivitis in Children.

BACKGROUND: Conjunctivitis is a frequent symptom in pediatric emergency departments; however, the etiology of conjunctivitis is difficult to clinically differentiate. OBJECTIVE: Our study objective was to evaluate the test performance characteristics of leukocyte esterase (LE) test strips in diagnosing bacterial conjunctivitis. METHODS: Patients aged from 3 months through 21 years presenting to an emergency department with symptoms of conjunctivitis were prospectively enrolled from September 2018 to March 2020. A swab of the affected eye was applied to the LE test strip and another swab was sent for culture processing. The primary outcome was the association between LE test results and eye culture results. RESULTS: We enrolled 189 patients. Overall, 117 eye cultures (62%) were positive. The sensitivity and specificity of LE testing was 96% (95% CI 90-98%) and 14% (95% CI 7-25%), respectively. Positive predictive value was 64% (95% CI 57-71%) and negative predictive value was 67% (95% CI 39-87%). CONCLUSIONS: The LE test strip had limited ability to differentiate bacterial conjunctivitis from other etiologies.

Drug-Drug Interaction Between Cannabidiol, Cyclosporine, and Mycophenolate Mofetil: A Case Report.

Kidney transplantation remains the optimal therapy for many patients with end-stage kidney disease (ESKD). Chronic pain is one of the most common and distressing symptoms among patients with ESKD, and its treatment is a complex and challenging task to accomplish. The benefits of cannabidiol (CBD) in chronic pain treatment have been reported recently. Cannabidiol is metabolized by cytochrome P450, mainly CYP3A4 and CYP2C19, and can also undergo direct conjugation via UDP-glucuronosyltransferase enzymes, with a growing body of evidence suggesting it is also a potent inhibitor or inducer of these pathways. Cannabidiol was also found to be a potent inhibitor of carboxylesterases in vitro. Because cytochrome P450 enzymes and carboxylesterases are also responsible for the clearance and activation of immunosuppressants, respectively, drug-drug interactions are likely to occur. Here, we report a pharmacokinetic drug interaction between CBD and cyclosporine and mycophenolate mofetil in a patient with ESKD with a kidney transplantation. It is thus crucial to take into account these interactions and monitor drug levels to avoid drug toxicity or a lack of efficacy. This study is in accordance with the guidelines of the Declaration of Helsinki and the Declaration of Istanbul.

Peptidyl-tRNA hydrolase is the nascent chain release factor in bacterial ribosome-associated quality control.

Rescuing stalled ribosomes often involves their splitting into subunits. In many bacteria, the resultant large subunits bearing peptidyl-tRNAs are processed by the ribosome-associated quality control (RQC) apparatus that extends the C termini of the incomplete nascent polypeptides with polyalanine tails to facilitate their degradation. Although the tailing mechanism is well established, it is unclear how the nascent polypeptides are cleaved off the tRNAs. We show that peptidyl-tRNA hydrolase (Pth), the known role of which has been to hydrolyze ribosome-free peptidyl-tRNA, acts in concert with RQC factors to release nascent polypeptides from large ribosomal subunits. Dislodging from the ribosomal catalytic center is required for peptidyl-tRNA hydrolysis by Pth. Nascent protein folding may prevent peptidyl-tRNA retraction and interfere with the peptide release. However, oligoalanine tailing makes the peptidyl-tRNA ester bond accessible for Pth-catalyzed hydrolysis. Therefore, the oligoalanine tail serves not only as a degron but also as a facilitator of Pth-catalyzed peptidyl-tRNA hydrolysis.

Mutation of AtPME2, a pH-Dependent Pectin Methylesterase, Affects Cell Wall Structure and Hypocotyl Elongation.

Pectin methylesterases (PMEs) modify homogalacturonan's chemistry and play a key role in regulating primary cell wall mechanical properties. Here, we report on Arabidopsis AtPME2, which we found to be highly expressed during lateral root emergence and dark-grown hypocotyl elongation. We showed that dark-grown hypocotyl elongation was reduced in knock-out mutant lines as compared to the control. The latter was related to the decreased total PME activity as well as increased stiffness of the cell wall in the apical part of the hypocotyl. To relate phenotypic analyses to the biochemical specificity of the enzyme, we produced the mature active enzyme using heterologous expression in Pichia pastoris and characterized it through the use of a generic plant PME antiserum. AtPME2 is more active at neutral compared to acidic pH, on pectins with a degree of 55-70% methylesterification. We further showed that the mode of action of AtPME2 can vary according to pH, from high processivity (at pH8) to low processivity (at pH5), and relate these observations to the differences in electrostatic potential of the protein. Our study brings insights into how the pH-dependent regulation by PME activity could affect the pectin structure and associated cell wall mechanical properties.

Use of tannase-producing bacteria isolated from the rumen to improve the nutritional value of pomegranate peel for fattening lambs.

BACKGROUND: The use of plants and by-products, which are containing a high amount of secondary and anti-nutritional compounds such as tannins, in animal feed is limited. The methods that can reduce these compounds make facilitate their use in animal feed. OBJECTIVES: The aim of this study was to reduce the adverse effects of pomegranate peel (PP) tannin for fattening lambs using the tannase-producing bacteria. METHODS: Twenty-one Arabi male lambs (averagely 35 ± 3.8 kg weight and 8 ± 1.0 months age) were used in a completely randomized design with three treatments and seven replications in the present experiment. The experimental treatments included 1 - control diet (CNT, no PP), 2 - diet containing untreated PP (raw PP, UTPP) and 3 - diet containing PP treated with tannase-producing bacteria (bacteria treating PP, BTPP). RESULTS: Using UTPP decreased nutrient intake compared to the control and treatment with tannase-producing bacteria again significantly increased nutrient intake compared to the UTPP (p < 0.05). The digestibilities of organic matter, neutral detergent fibre and acid detergent fibre in the control treatment were significantly higher than UTPP and BTPP and in the BTPP were significantly higher than the UTPP (p < 0.05). The use of UTPP in the diet significantly decreased the pH, ammonia nitrogen concentration and the total protozoa population of the rumen compared to the control (p < 0.05), and treatment with bacteria increased them again. The lowest total protozoa population was observed in UTPP treatments (p < 0.05). The highest concentration of blood glucose was observed in UTPP; however, the highest concentrations of blood urea nitrogen, cholesterol, triglyceride, high-density lipoprotein (non-significant) and low-density lipoprotein were in the control treatment. The effect of experimental treatments on the dry matter consumption of the whole period was significant; however, there was no significant effect on average daily gain, feed conversion ratio, feed efficiency and longissimus muscle colorimetric systems. CONCLUSIONS: Therefore, considering the positive effects of treatment PP with tannin-degrading bacteria relative to raw PP, using these bacteria is a proper way to reduce tannin, thus improving the nutritional value of PP for ruminants.

Long-lasting blocking of interoceptive effects of cocaine by a highly efficient cocaine hydrolase in rats.

Cocaine dependence is a serious world-wide public health problem without an FDA-approved pharmacotherapy. We recently designed and discovered a highly efficient long-acting cocaine hydrolase CocH5-Fc(M6). The present study examined the effectiveness and duration of CocH5-Fc(M6) in blocking interoceptive effects of cocaine by performing cocaine discrimination tests in rats, demonstrating that the duration of CocH5-Fc(M6) in blocking cocaine discrimination was dependent on cocaine dose and CocH5-Fc(M6) plasma concentration. Particularly, a dose of 3 mg/kg CocH5-Fc(M6) effectively attenuated discriminative stimulus effects of 10 mg/kg cocaine, cumulative doses of 10 and 32 mg/kg cocaine, and cumulative doses of 10, 32 and 56 mg/kg cocaine by ≥ 20% for 41, 19, and 10 days, and completely blocked the discriminative stimulus effects for 30, 13, and 5 days with corresponding threshold plasma CocH5-Fc(M6) concentrations of 15.9, 72.2, and 221 nM, respectively, under which blood cocaine concentration was negligible. Additionally, based on the data obtained, cocaine discrimination model is more sensitive than the locomotor activity to reveal cocaine effects and that CocH5-Fc(M6) itself has no long-term toxicity regarding behavioral activities such as lever pressing and food consumption in rats, further demonstrating that CocH5-Fc(M6) has the desired properties as a promising therapeutic candidate for prevenance of cocaine dependence.

Carboxylesterase-activated near-infrared fluorescence probe for highly sensitive imaging of liver tumors.

Carboxylesterases (CESs) are critical for metabolizing ester-containing biomolecules and are specifically important in liver metabolic disorders. The modulation of CESs is also an important issue in pharmacology and clinical applications. Herein, we present a near-infrared (NIR) CES fluorescent probe (NCES) based on the protection-deprotection of the hydroxyl group for monitoring CES levels in living systems. The NCES probe has good selectivity and sensitivity for CESs with a limit of detection (LOD) of 5.24 mU mL-1, which allows for tracing the fluctuation of cellular CES after treatment with anticancer drugs and under inflammation and apoptosis states. Furthermore, NCES can be successfully applied for guiding liver cancer surgery with high-contrast in vivo imaging and detecting clinical serum samples from liver cancer patients. This work showed that the NCES probe has great potential in drug development, imaging applications for medical diagnosis, and early-stage detection for clinical liver diseases.

Evaluation of antibacterial, antifungal and antibiofilm activities of A. baumannii-derived tannase and gallic acid against uropathogenic microorganisms.

One of the most prevalent infectious diseases and a key driver of antibiotic prescriptions in pediatrics is urinary tract infection (UTI). Due to the emergence of more resistant uropathogenic bacterial and fungal strains, current treatments are no longer effective, necessitating the urgent development of novel antibacterial and antifungal drugs. In this study, the antifungal, antibacterial, and anti-biofilm capabilities of compounds, such as tannase (TN) and gallic acid (GA), which were produced from a novel natural source, Acinetobacter baumannii (AB11) bacteria, were assessed for the inactivation of uropathogenic microorganisms (UMs). Ammonium sulphate precipitation, ion exchange, high-performance liquid chromatography, and gel filtration were used to purify TN and GA that were isolated from A. baumannii. A 43.08 % pure TN with 1221.2 U/mg specific activity and 10.51 mg/mL GA was obtained. The antibacterial, antifungal and anti-biofilm activities of TN and GA were evaluated against UMs and compared to those of commercially available antibiotics including sulfamethoxazole (SXT), levofloxacin (LEV), ciprofloxacin (CIP), amikacin (Ak), and nitrofurantoin (F). The results showed that TN and GA were superior to commercial antibiotics in their ability to inactivate UMs and considerably reduced biofilms formation. Additionally, the GA emerges as the top substitute for currently available medications, demonstrating superior antibacterial and antibiofilm properties against all UMs evaluated in this study. The results of this investigation showed that A. baumannii-derived TN and GA could be utilized as an alternative medication to treat UTIs.

Target Cell Activation of a Structurally Novel NOD-Like Receptor Pyrin Domain-Containing Protein 3 Inhibitor NT-0796 Enhances Potency.

The NOD-like receptor pyrin domain-containing protein 3 (NLRP3) inflammasome is a central regulator of innate immunity, essential for processing and release of interleukin-1ß and pyroptotic cell death. As endogenous NLRP3 activating triggers are hallmarks of many human chronic inflammatory diseases, inhibition of NLRP3 has emerged as a therapeutic target. Here we identify NDT-19795 as a novel carboxylic acid-containing NLRP3 activation inhibitor in both human and mouse monocytes and macrophages. Remarkably, conversion of the carboxylate to an isopropyl-ester (NT-0796) greatly enhances NLRP3 inhibitory potency in human monocytes. This increase is attributed to the ester-containing pharmacophore being more cell-penetrant than the acid species and, once internalized, the ester being metabolized to NDT-19795 by carboxylesterase-1 (CES-1). Mouse macrophages do not express CES-1, and NT-0796 is ineffective in these cells. Mice also contain plasma esterase (Ces1c) activity which is absent in humans. To create a more human-like model, we generated a mouse line in which the genome was modified, removing Ces1c and replacing this segment of DNA with the human CES-1 gene driven by a mononuclear phagocyte-specific promoter. We show human CES-1 presence in monocytes/macrophages increases the ability of NT-0796 to inhibit NLRP3 activation both in vitro and in vivo. As NLRP3 is widely expressed by monocytes/macrophages, the co-existence of CES-1 in these same cells affords a unique opportunity to direct ester-containing NLRP3 inhibitors precisely to target cells of interest. Profiling NT-0796 in mice humanized with respect to CES-1 biology enables critical modeling of the pharmacokinetics and pharmacodynamics of this novel therapeutic candidate. SIGNIFICANCE STATEMENT: Inhibition of NLRP3 represents a desirable therapeutic strategy for the treatment of multiple human disorders. In this study pharmacological properties of a structurally-novel, ester-containing NLRP3 inhibitor NT-0796 are characterized. To study pharmacodynamics of NT-0796 in vivo, a mouse line was engineered possessing more human-like traits with respect to carboxylesterase biology. In the context of these hCES-1 mice, NT-0796 serves as a more effective inhibitor of NLRP3 activation than the corresponding acid, highlighting the full translational potential of the ester strategy.

An ultrasensitive and selective near-infrared fluorescent probe for tracking carboxylesterases with large Stokes shift in living cells and mice.

Carboxylesterases (CEs) play great role in CEs-related diseases and drug metabolism. Selectively monitoring its activity is important to explore its role in CEs-related diseases and drug combination. Herein, a new "turn-on" near-infrared (NIR) fluorescent probe (CHY-1) was reported with large Stokes shift (145 nm) for CEs detection. Dicyanoisophorone-based derivative was chosen as NIR fluorophore and 4-bromobutyrate was the identifying group. What's more, CHY-1 exhibited ultra-sensitivity (LOD âˆ¼ 9.2 × 10-5 U/mL), high selectivity against Acetylcholinesterase (AChE), Butyrylcholinesterase (BChE) and Chymotrypsin for CEs fluorescence detection under physiological pH and temperature. Furthermore, CHY-1 showed little effect on cell viability at high concentration and featured good optical imaging character for the slight change of CEs activity induced by 5-Fu (5-Fluorouridine, anti-tumor drug) and CEs inhibitor in living cells. Moreover, CHY-1 was also used to detect the activity and distribution of CEs in mice. Taken together, CHY-1 had widely applicable value in the diagnosis of CEs-related diseases and drug combination.