Effects of three-dimensional quality assessment nursing intervention on efficacy and disease management of patients undergoing esophageal cancer surgery.

BACKGROUND: Esophageal cancer is one of the most common malignant tumors. The three-dimensional quality structure model is a quality assessment theory that includes three dimensions: Structure, process, and results. AIM: To investigate the effects of nursing interventions with three-dimensional quality assessment on the efficacy and disease management ability of patients undergoing esophageal cancer surgery. METHODS: In this prospective study, the control group received routine nursing, and the intervention group additionally received a three-dimensional quality assessment intervention based on the above routine care. Self-efficacy and patient disease management abilities were evaluated using the General Self-Efficacy Scale (GSES) and Exercise of Self-Care Agency scale, respectively. IBM SPSS Statistics for Windows, version 17.0, was used for the data processing. RESULTS: This study recruited 112 patients who were assigned to the control and experimental groups (n = 56 per group). Before the intervention, there was no significant difference in GSES scores between the two groups (P > 0.05). After the intervention, the GSES scores of both groups increased, with the experimental group showing higher values (P < 0.05). At the time of discharge and three months after discharge, the scores for positive attitudes, self-stress reduction, and total score of health promotion in the experimental group were higher than those in the control group (P < 0.05). CONCLUSION: The implementation of a three-dimensional quality structure model for postoperative patients with esophageal cancer can effectively improve their self-management ability and self-efficacy of postoperative patients.

Oligotrophic state reduces the time dependence of the observed survival fraction for heavy ion beam-irradiated Saccharomyces cerevisiae and provides new insights into DNA repair.

Heavy ion beam (HIB) irradiation is widely utilized in studies of cosmic rays-induced cellular effects and microbial breeding. Establishing an accurate dose-survival relationship is crucial for selecting the optimal irradiation dose. Typically, after irradiating logarithmic-phase cell suspensions with HIB, the survival fraction (SF) is determined by the ratio of clonal-forming units in irradiated versus control groups. However, our findings indicated that SF measurements were time sensitive. For the Saccharomyces cerevisiae model, the observed SF initially declined and subsequently increased in a eutrophic state; conversely, in an oligotrophic state, it remained relatively stable within 120 minutes. This time effect of SF observations in the eutrophic state can be ascribed to HIB-exposed cells experiencing cell cycle arrest, whereas the control proliferated rapidly, resulting in an over-time disproportionate change in viable cell count. Therefore, an alternative involves irradiating oligotrophic cells, determining SF thereafter, and transferring cells to the eutrophic state to facilitate DNA repair-mutation. Transcriptomic comparisons under these two trophic states yield valuable insights into the DNA damage response. Although DNA repair was postponed in an oligotrophic state, cells proactively mobilized specific repair pathways to advance this process. Effective nutritional supplementation should occur within 120 minutes, beyond this window, a decline in SF indicates an irreversible loss of repair capability. Upon transition to the eutrophic state, S. cerevisiae swiftly adapted and completed the repair. This study helps to minimize time-dependent variability in SF observations and to ensure effective damage repair and mutation in microbial breeding using HIB or other mutagens. It also promotes the understanding of microbial responses to complex environments.IMPORTANCEMutation breeding is a vital means of developing excellent microbial resources. Consequently, understanding the mechanisms through which microorganisms respond to complex environments characterized by mutagens and specific physiological-biochemical states holds significant theoretical and practical values. This study utilized Saccharomyces cerevisiae as a microbial model and highly efficient heavy ion beam (HIB) radiation as a mutagen, it revealed the time dependence of observations of survival fractions (SF) in response to HIB radiation and proposed an alternative to avoid the indeterminacy that this variable brings. Meanwhile, by incorporating an oligotrophic state into the alternative, this study constructed a dynamic map of gene expression during the fast-repair and slow-repair stages. It also highlighted the influence of trophic states on DNA repair. The findings apply to the survival-damage repair-mutation effects of single-celled microorganisms in response to various mutagens and contribute to elucidating the biological mechanisms underlying microbial survival in complex environments.

Realization of chiral two-mode Lipkin-Meshkov-Glick models via acoustics.

Thechirality-controlled two-mode Lipkin-Meshkov-Glick (LMG) modelsare mimicked in a potential hybrid quantum system, involving two ensembles of solid-state spins coupled to a pair of interconnected surface-acoustic-wave cavities. With the assistance of dichromatic classical optical drives featuring chiral designs, it can simulate two-mode LMG-type long-range spin-spin interactions with left-right asymmetry. For applications, this unconventional LMG model can not only engineer both ensembles of collective spins into two-mode spin-squeezed states but also simulate novel quantum critical phenomena and time crystal behaviors, among others. Since this acoustic-based system can generate ion-trap-like interactions without requiring any additional trapping techniques, our work is considered a fresh attempt at realizing chiral quantum manipulation of spin-spin interactions using acoustic hybrid systems.

Radiation-Emitting metallic stent for unresectable bismuth type III or IV perihilar cholangiocarcinoma: a multicenter randomized trial.

BACKGROUND AIMS: Self-expandable metallic stents (SEMSs) have been recommended for patients with unresectable malignant biliary obstruction while radiation-emitting metallic stents (REMSs) loaded with 125I seeds have recently been approved to provide longer patency and overall survival in malignant biliary tract obstruction. This trial is to evaluate the efficacy and safety of REMS plus hepatic arterial infusion chemotherapy (REMS-HAIC) versus SEMS plus HAIC (SEMS-HAIC) for unresectable perihilar cholangiocarcinoma (pCCA). METHODS: This multicenter randomized controlled trial recruited patients with unresectable Bismuth type III or IV pCCA between March 2021 and January 2023. Patients were randomly assigned (1:1 ratio) to receive either REMS-HAIC or SEMS-HAIC using permuted block randomization, with a block size of six. The primary endpoint was overall survival (OS). The secondary endpoints were time to symptomatic progression (TTSP), stent patency, relief of jaundice, quality of life, and safety. RESULTS: A total of 126 patients were included in the intent-to-treat population, with 63 in each group. The median OS was 10.2 months versus 6.7 months (P=0.002). The median TTSP was 8.6 months versus 5.4 months (P=0.003). The median stent patency was longer in the REMS-HAIC group than in the SEMS-HAIC group (P=0.001). The REMS-HAIC group showed better improvement in physical functioning scale (P<0.05) and fatigue symptoms (P<0.05) when compared to the SEMS-HAIC group. No significant differences were observed in relief of jaundice (85.7% vs. 84.1%; P=0.803) or the incidence of grade 3 or 4 adverse events (9.8% vs. 11.9%; P=0.721). CONCLUSION: REMS plus HAIC showed better OS, TTSP, and stent patency compared with SEMS plus HAIC in patients with unresectable Bismuth type III or IV pCCA with an acceptable safety profile.

Reciprocal or nonreciprocal bimolecular interface and quantum entanglement.

We study a hybrid system of a plasmonic cavity coupled to a pair of different molecular vibration modes with the strong optomechanical-like interactions. Here, this plasmonic cavity is considered as a quantum data bus and then assist several applications. For instance, it can first establish a bimolecular interface to ensure the reciprocal or non-reciprocal information transmission, and then engineer both molecules into the steady-state quantum entanglement of the continuous variable through the dissipative method. In contrast to the traditional optomechanical system, this hybrid system can provide the stronger optomechanical-like interactions and more convenient controls to the molecular quantum units. This investigation is believed to be able to further expand the practical application range of quantum technology. .

Integrated approach for improving mechanical and high-temperature properties of fast-growing poplar wood using lignin-controlled treatment combined with densification.

Previous studies on the modification of fast-growing wood have extensively examined the effects of density and lignin content on the strength and high-temperature properties of modified wood. However, a comprehensive quantitative analysis of their effects on high-temperature performance remains insufficient. To address this knowledge gap, we applied alkali treatment and compression densification to fast-growing poplar, resulting in modified specimens with varying densities and lignin levels. The quantitative effects of density and lignin content on high-temperature properties were meticulously evaluated. Chemical changes were analyzed using Fourier transform infrared spectroscopy (FT-IR), while the mechanical and high-temperature properties were comprehensively assessed. Delignification was found to be positively correlated with treatment duration, with hemicellulose degradation also detected via FT-IR analysis. Significant enhancements were recorded in flexural strength, tensile strength, and modulus of elasticity, accompanied by improvements in ductility ratio and compressive strength. The modified poplar wood exhibited increased thermal stability at elevated temperatures. Furthermore, density and lignin content were identified as significant factors affecting high-temperature performance, establishing minimum density thresholds for various lignin contents in modified poplar wood to ensure optimal performance. This study enhances to the understanding of the intricate relationships among wood properties, modification techniques, and high-temperature performance.

Comprehensive applications of the artificial intelligence technology in new drug research and development.

Purpose: Target-based strategy is a prevalent means of drug research and development (R&D), since targets provide effector molecules of drug action and offer the foundation of pharmacological investigation. Recently, the artificial intelligence (AI) technology has been utilized in various stages of drug R&D, where AI-assisted experimental methods show higher efficiency than sole experimental ones. It is a critical need to give a comprehensive review of AI applications in drug R &D for biopharmaceutical field. Methods: Relevant literatures about AI-assisted drug R&D were collected from the public databases (Including Google Scholar, Web of Science, PubMed, IEEE Xplore Digital Library, Springer, and ScienceDirect) through a keyword searching strategy with the following terms [("Artificial Intelligence" OR "Knowledge Graph" OR "Machine Learning") AND ("Drug Target Identification" OR "New Drug Development")]. Results: In this review, we first introduced common strategies and novel trends of drug R&D, followed by characteristic description of AI algorithms widely used in drug R&D. Subsequently, we depicted detailed applications of AI algorithms in target identification, lead compound identification and optimization, drug repurposing, and drug analytical platform construction. Finally, we discussed the challenges and prospects of AI-assisted methods for drug discovery. Conclusion: Collectively, this review provides comprehensive overview of AI applications in drug R&D and presents future perspectives for biopharmaceutical field, which may promote the development of drug industry.

Furfural tolerance of mutant Saccharomyces cerevisiae selected via ionizing radiation combined with adaptive laboratory evolution.

BACKGROUND: Lignocellulose is a renewable and sustainable resource used to produce second-generation biofuel ethanol to cope with the resource and energy crisis. Furfural is the most toxic inhibitor of Saccharomyces cerevisiae cells produced during lignocellulose treatment, and can reduce the ability of S. cerevisiae to utilize lignocellulose, resulting in low bioethanol yield. In this study, multiple rounds of progressive ionizing radiation was combined with adaptive laboratory evolution to improve the furfural tolerance of S. cerevisiae and increase the yield of ethanol. RESULTS: In this study, the strategy of multiple rounds of progressive X-ray radiation combined with adaptive laboratory evolution significantly improved the furfural tolerance of brewing yeast. After four rounds of experiments, four mutant strains resistant to high concentrations of furfural were obtained (SCF-R1, SCF-R2, SCF-R3, and SCF-R4), with furfural tolerance concentrations of 4.0, 4.2, 4.4, and 4.5 g/L, respectively. Among them, the mutant strain SCF-R4 obtained in the fourth round of radiation had a cellular malondialdehyde content of 49.11 nmol/mg after 3 h of furfural stress, a weakening trend in mitochondrial membrane potential collapse, a decrease in accumulated reactive oxygen species, and a cell death rate of 12.60%, showing better cell membrane integrity, stable mitochondrial function, and an improved ability to limit reactive oxygen species production compared to the other mutant strains and the wild-type strain. In a fermentation medium containing 3.5 g/L furfural, the growth lag phase of the SCF-R4 mutant strain was shortened, and its growth ability significantly improved. After 96 h of fermentation, the ethanol production of the mutant strain SCF-R4 was 1.86 times that of the wild-type, indicating that with an increase in the number of irradiation rounds, the furfural tolerance of the mutant strain SCF-R4 was effectively enhanced. In addition, through genome-transcriptome analysis, potential sites related to furfural detoxification were identified, including GAL7, MAE1, PDC6, HXT1, AUS1, and TPK3. CONCLUSIONS: These results indicate that multiple rounds of progressive X-ray radiation combined with adaptive laboratory evolution is an effective mutagenic strategy for obtaining furfural-tolerant mutants and that it has the potential to tap genes related to the furfural detoxification mechanism.

Improvement of Saccharomyces cerevisiae strain tolerance to vanillin through heavy ion radiation combined with adaptive laboratory evolution.

Vanillin is an inhibitor of lignocellulose hydrolysate, which can reduce the ability of Saccharomyces cerevisiae to utilize lignocellulose, which is an important factor limiting the development of the ethanol fermentation industry. In this study, mutants of vanillin-tolerant yeast named H6, H7, X3, and X8 were bred by heavy ion irradiation (HIR) combined with adaptive laboratory evolution (ALE). Phenotypic tests revealed that the mutants outperformed the original strain WT in tolerance, growth rate, genetic stability and fermentation ability. At 1.6 g/L vanillin concentration, the average OD600 value obtained for mutant strains was 0.95 and thus about 3.4-fold higher than for the wild-type. When the concentration of vanillin was 2.0 g/L, the glucose utilization rate of the mutant was 86.3 % within 96 h, while that of the original strain was only 70.0 %. At this concentration of vanillin, the mitochondrial membrane potential of the mutant strain recovered faster than that of the original strain, and the ROS scavenging ability was stronger. We analyzed the whole transcriptome sequencing map and the whole genome resequencing of the mutant, and found that DEGs such as FLO9, GRC3, PSP2 and SWF1, which have large differential expression multiples and obvious mutation characteristics, play an important role in cell flocculation, rDNA transcription, inhibition of DNA polymerase mutation and protein palmitoylation. These functions can help cells resist vanillin stress. The results show that combining HIR with ALE is an effective mutagenesis strategy. This approach can efficiently obtain Saccharomyces cerevisiae mutants with improved vanillin tolerance, and provide reference for obtaining robust yeast strains with lignocellulose inhibitor tolerance.

The evolution of cutinase Est1 based on the clustering strategy and its application for commercial PET bottles degradation.

The rapid increase in global plastic consumption, especially the worldwide use of polyethylene terephthalate (PET), has caused serious pollution problems. Due to the low recycling rate of PET, a substantial amount of waste accumulates in the environment, which prompts a growing focus on enzymatic degradation for its efficiency and environmentally friendliness. This study systematically designed and modified a cutinase, Est1 from Thermobifida alba AHK119, known for its potential of plastic-degradation at high temperatures. Additionally, the introduction of clustering algorithms provided the ability to understand and modify biomolecules, to accelerate the process of finding the optimal mutations. K-means was further proceeded based on the positive mutations. After comprehensive screening for thermostability and activity mutation sites, the dominant mutation Est1_5M (Est1 with the mutations of N213M, T215P, S115P, Q93A, and L91W) exhibited satisfying degradation ability for commercial PET bottles. The results showed that Est1_5M achieved a degradation rate of 90.84% in 72 h, 65-fold higher than the wild type. This study offers reliable theoretical and practical support for the development of efficient PET-degrading enzymes, providing a reference for plastic pollution management.

Lysineless HiBiT and NanoLuc Tagging Systems as Alternative Tools for Monitoring Targeted Protein Degradation.

Target protein degradation (TPD) has emerged as a revolutionary approach in drug discovery, leveraging the cell's intrinsic machinery to selectively degrade disease-associated proteins. Nanoluciferase (nLuc) fusion proteins and the NanoBiT technology offer two robust and sensitive screening platforms to monitor the subtle changes in protein abundance induced by TPD molecules. Despite these advantages, concerns have arisen regarding potential degradation artifacts introduced by tagging systems due to the presence of lysine residues on them, prompting the development of alternative tools. In this study, we introduce HiBiT-RR and nLucK0, variants devoid of lysine residues, to mitigate such artifacts. Our findings demonstrate that HiBiT-RR maintains a similar sensitivity and binding affinity with the original HiBiT. Moreover, the comparison between nLucWT and nLucK0 constructs reveals variations in degradation patterns induced by certain TPD molecules, emphasizing the importance of choosing appropriate tagging systems to ensure the reliability of experimental outcomes in studying protein degradation processes.

Anomalous normal-state gap in an electron-doped cuprate.

In the underdoped n-type cuprate Nd2-xCexCuO4, long-range antiferromagnetic order reconstructs the Fermi surface, resulting in a putative antiferromagnetic metal with small Fermi pockets. Using angle-resolved photoemission spectroscopy, we observe an anomalous energy gap, an order of magnitude smaller than the antiferromagnetic gap, in a wide portion of the underdoped regime and smoothly connecting to the superconducting gap at optimal doping. After considering all the known ordering tendencies in tandem with the phase diagram, we hypothesize that the normal-state gap in the underdoped n-type cuprates originates from Cooper pairing. The high temperature scale of the normal-state gap raises the prospect of engineering higher transition temperatures in the n-type cuprates comparable to those of the p-type cuprates.

Diagnosis and Treatment of Hepatic Hematoma After Liver Transplantation in a Timely Manner.

OBJECTIVE: Hematomas of the liver graft, that is, postintervention, subcapsular or intrahepatic are rare yet potentially fatal complications following liver transplantation (LT), necessitating immediate diagnosis and management to avert devastating outcomes. This study was aimed to introduce our approach to manage graft hematoma subsequent to LT. METHODS: Among 131 orthotopic liver transplantations (OLT) conducted at our institution between January 2017 and May 2023, 3 cases of intrahepatic (n = 2) and extrahepatic (n = 1) hematoma were confirmed through computed tomography (CT) within 10 days after LT. The clinical outcomes of various treatment modalities for these three cases were analyzed. RESULTS: Three out of 131 (2.3%) LT recipients developed graft hematoma. Patient 1 developed a spontaneous intrahepatic hematoma, without evident predisposing factors, while patient 2 developed an intrahepatic hematoma following endoscopic retrograde cholangiopancreatography (ERCP). The third case that is extrahepatic hematoma was speculated to be a result of minor hepatic parenchymal injury stemming from compressive and volume-reducing manipulation of a large graft, or secondary to focal ischemic necrosis of the liver. Our management protocol was summarized as follows: (1). Immediate ultrasound and CT, particularly enhanced CT; (2). Puncture and percutaneous drainage (PD) of the hematoma; (3). Arterial embolization if the origin could be identified as a ruptured vessel; (4). Surgical evacuation of the hematoma in the presence of bile leakage, to avoid a compartment respectably secondary infection. All three patients responded favorably to treatment and remained alive to date. CONCLUSION: Prompt diagnosis and sequential individualized management can successfully deal with intra-/extrahepatic graft hematoma after LT. Our results underscored that an individualized management considering potential future complications into account.

Prodrug-inspired adenosine triphosphate-activatable celastrol-Fe(III) chelate for cancer therapy.

Celastrol (CEL), an active compound isolated from the root of Tripterygium wilfordii, exhibits broad anticancer activities. However, its poor stability, narrow therapeutic window and numerous adverse effects limit its applications in vivo. In this study, an adenosine triphosphate (ATP) activatable CEL-Fe(III) chelate was designed, synthesized, and then encapsulated with a reactive oxygen species (ROS)-responsive polymer to obtain CEL-Fe nanoparticles (CEL-Fe NPs). In normal tissues, CEL-Fe NPs maintain structural stability and exhibit reduced systemic toxicity, while at the tumor site, an ATP-ROS-rich tumor microenvironment, drug release is triggered by ROS, and antitumor potency is restored by competitive binding of ATP. This intelligent CEL delivery system improves the biosafety and bioavailability of CEL for cancer therapy. Such a CEL-metal chelate strategy not only mitigates the challenges associated with CEL but also opens avenues for the generation of CEL derivatives, thereby expanding the therapeutic potential of CEL in clinical settings.

A Simple Prognostic Scoring System for Hepatocellular Carcinoma Treated with DEB-TACE.

Objective: To develop a simple and effective prognostic scoring system to predict the efficacy of drug-eluting bead-transcatheter arterial chemoembolization (DEB-TACE) in the treatment of hepatocellular carcinoma (HCC). Methods: Data were retrospectively collected from 230 patients with HCC who received DEB-TACE treatment at six medical centers between January 2019 and December 2022. We developed a predictive score based on independent risk factors for overall survival (OS), validated the model using a validation cohort, and compared its prognostic accuracy with commonly used HCC staging systems. Results: The number of tumors, albumin-bilirubin levels, alpha-fetoprotein levels, and portal vein thrombus grade were identified as independent factors influencing OS. Based on these factors, we established the DEB-TACE treatment of HCC (DTH) scoring system. The DTH score correlated well with OS, which decreased as the DTH score increased. According to the DTH score, patients were categorized into three risk groups: low-risk (DTH-A, 0-4 points), medium-risk (DTH-B, 5-6 points), and high-risk (DTH-A, 7 points). The OS of each risk group was 18.73±0.62 months, 12.73±0.10 months, and 6.93±0.19 months, respectively (p<0.001). The external cohort validation confirmed the accuracy of the DTH score, demonstrating superior predictive performance compared to other commonly used HCC scoring systems. Conclusion: The DTH-HCC scoring system effectively predicts the outcomes of HCC patients undergoing DEB-TACE as initial treatment. This model can aid in the initial planning and decision-making process for DEB-TACE treatment in HCC patients.

5,7,4'-Trimethoxyflavone triggers cancer cell PD-L1 ubiquitin-proteasome degradation and facilitates antitumor immunity by targeting HRD1.

Targeting the programmed cell death 1/programmed cell death ligand 1 (PD-1/PD-L1) pathway has been identified as a successful approach for tumor immunotherapy. Here, we identified that the small molecule 5,7,4'-trimethoxyflavone (TF) from Kaempferia parviflora Wall reduces PD-L1 expression in colorectal cancer cells and enhances the killing of tumor cells by T cells. Mechanistically, TF targets and stabilizes the ubiquitin ligase HMG-CoA reductase degradation protein 1 (HRD1), thereby increasing the ubiquitination of PD-L1 and promoting its degradation through the proteasome pathway. In mouse MC38 xenograft tumors, TF can activate tumor-infiltrating T-cell immunity and reduce the immunosuppressive infiltration of myeloid-derived suppressor cells and regulatory T cells, thus exerting antitumor effects. Moreover, TF synergistically exerts antitumor immunity with CTLA-4 antibody. This study provides new insights into the antitumor mechanism of TF and suggests that it may be a promising small molecule immune checkpoint modulator for cancer therapy.

Integration of food raw materials, food microbiology, and food additives: systematic research and comprehensive insights into sweet sorghum juice, Clostridium tyrobutyricum TGL-A236 and bio-butyric acid.

Introduction: Sweet sorghum juice is a typical production feedstock for natural, eco-friendly sweeteners and beverages. Clostridium tyrobutyricum is one of the widely used microorganisms in the food industry, and its principal product, bio-butyric acid is an important food additive. There are no published reports of Clostridium tyrobutyricum producing butyric acid using SSJ as the sole substrate without adding exogenous substances, which could reach a food-additive grade. This study focuses on tailoring a cost-effective, safe, and sustainable process and strategy for their production and application. Methods: This study modeled the enzymolysis of non-reducing sugars via the first/second-order kinetics and added food-grade diatomite to the hydrolysate. Qualitative and quantitative analysis were performed using high-performance liquid chromatography, gas chromatography-mass spectrometer, full-scale laser diffraction method, ultra-performance liquid chromatography-tandem mass spectrometry, the cell double-staining assay, transmission electron microscopy, and Oxford nanopore technology sequencing. Quantitative real-time polymerase chain reaction, pathway and process enrichment analysis, and homology modeling were conducted for mutant genes. Results: The treated sweet sorghum juice showed promising results, containing 70.60 g/L glucose and 63.09 g/L fructose, with a sucrose hydrolysis rate of 98.29% and a minimal sucrose loss rate of 0.87%. Furthermore, 99.62% of the colloidal particles and 82.13% of the starch particles were removed, and the concentrations of hazardous substances were effectively reduced. A food microorganism Clostridium tyrobutyricum TGL-A236 with deep utilization value was developed, which showed superior performance by converting 30.65% glucose and 37.22% fructose to 24.1364 g/L bio-butyric acid in a treated sweet sorghum juice (1:1 dilution) fermentation broth. This titer was 2.12 times higher than that of the original strain, with a butyric acid selectivity of 86.36%. Finally, the Genome atlas view, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and evolutionary genealogy of genes: Non-supervised Orthologous (eggNOG) functional annotations, three-dimensional structure and protein cavity prediction of five non-synonymous variant genes were obtained. Conclusion: This study not only includes a systematic process flow and in-depth elucidation of relevant mechanisms but also provides a new strategy for green processing of food raw materials, improving food microbial performance, and ensuring the safe production of food additives.

Development of Biochemical and Cellular Probes to Study RIPK1 Target Engagement.

RIPK1 inhibitors have emerged as promising candidates for treating diverse diseases, including inflammatory diseases, autoimmune disorders, Alzheimer's disease, and cancer. However, the previously reported binding assays have limited sensitivity and stability, impeding high-throughput screening and robust characterization of the RIPK1 inhibitors. To address this challenge, we introduced two probes, T2-BDP-FL and T3-BDP-FL, derived from distinct RIPK1 inhibitors with different binding modes to establish time-resolved fluorescence resonance energy transfer (TR-FRET) displacement assays. Employing our TR-FRET displacement assays, we quantified the biochemical binding affinities of a series of RIPK1 inhibitors with diverse structural and binding modes for human RIPK1. Consistent results were obtained with these two probes in the TR-FRET displacement assay. Furthermore, we developed a RIPK1 fluorescent probe, T2-BDP589, for the NanoBRET assay. This assay enabled the characterization of RIPK1 target engagement by various RIPK1 inhibitors for both human and mouse RIPK1 in live cells. Our developed fluorescent probe displacement assays offer a sensitive and high-throughput approach to identify RIPK1 inhibitors based on both biochemical and cellular activities.

Application of LAMP coupled with NALF for precise detection of mycoplasma pneumoniae.

Mycoplasma pneumoniae (MP),as the most commonly infected respiratory pathogen in community-acquired pneumonia in preschool children,has becoming a prominent factor affecting children's respiratory health.Currently, there is a lack of easy, rapid, and accurate laboratory testing program for MP infection, which causes comparatively difficulty for clinical diagnostic.Here,we utilize loop-mediated isothermal amplification (LAMP) to amplify and characterize the P1 gene of MP, combined with nucleic acid lateral flow (NALF) for fast and visuallized detection of MP.Furthermore, we evaluated and analyzed the sensitivity, specificity and methodological consistency of the method.The results showed that the limit of detection(LoD) of MP-LAMP-NALF assay was down to 100 copys per reaction and there was no cross-reactivity with other pathogens infected the respiratory system. The concordance rate between MP-LAMP-NALF assay with quantitative real-time PCR was 94.3 %,which exhibiting excellent testing performance.We make superior the turnaround time of the MP-LAMP-NALF assay, which takes only about 50 min. In addition, there is no need for precision instruments and no restriction on the laboratory site.Collectively, LAMP-NALF assay targeting the P1 gene for Mycoplasma pneumoniae detection was a easy, precise and visual test which could be widely applied in outpatient and emergency departments or primary hospitals.When further optimized, it could be used as "point-of-care testing" of pathogens or multiple testing for pathogens.

Lysineless HiBiT and NanoLuc Tagging Systems as Alternative Tools Monitoring Targeted Protein Degradation.

Target protein degradation (TPD) has emerged as a revolutionary approach in drug discovery, leveraging the cell's intrinsic machinery to selectively degrade disease-associated proteins. Proteolysis-Targeting Chimeras (PROTACs) exemplify this strategy, exploiting heterobifunctional molecules to induce ubiquitination and subsequent degradation of target proteins. The clinical advancement of PROTACs underscores their potential in therapeutic intervention, with numerous projects progressing through clinical stages. However, monitoring subtle changes in protein abundance induced by TPD molecules demands highly sensitive assays. Nano-luciferase (nLuc) fusion proteins, or the NanoBiT technology derived from it, offer a robust screening platform due to their high sensitivity and stability. Despite these advantages, concerns have arisen regarding potential degradation artifacts introduced by tagging systems due to the presence of lysine residues on them, prompting the development of alternative tools. In this study, we introduce HiBiT-RR and nLuc K0 , variants devoid of lysine residues, to mitigate such artifacts. Our findings demonstrate that HiBiT-RR maintains similar sensitivity and binding affinity with the original HiBiT. Moreover, the comparison between nLuc WT and nLuc K0 constructs reveals variations in degradation patterns induced by certain PROTAC molecules, emphasizing the importance of choosing appropriate tagging systems to ensure the reliability of experimental outcomes in studying protein degradation processes.