A Perspective Looking Backward and Forward on the 25th Anniversary of Conjugated Polyelectrolytes.

Conjugated polyelectrolytes are π-conjugated polymers that contain ionic charged groups such as sulfonate (R-SO3-), carboxylate (R-COO-), or ammonium (R-NR3+) combined with a π-conjugated backbone. This perspective provides a summary review of the key developments in the field, starting from the first reports of their synthesis and properties to application-focused developments. The applications include optical sensors for molecular and biomolecular targets, organic electronic applications, and specific biological applications including cellular imaging and photodynamic therapy. This perspective concludes with a discussion of where the field of conjugated polyelectrolytes is expected to lead in the coming years.

Simple, Visual, Point-of-Care SARS-CoV-2 Detection Incorporating Recombinase Polymerase Amplification and Target DNA-Protein Crosslinking Enhanced Chemiluminescence.

The ongoing COVID-19 pandemic, driven by persistent SARS-CoV-2 transmission, threatens human health worldwide, underscoring the urgent need for an efficient, low-cost, rapid SARS-CoV-2 detection method. Herein, we developed a point-of-care SARS-CoV-2 detection method incorporating recombinase polymerase amplification (RPA) and DNA-protein crosslinking chemiluminescence (DPCL) (RPADPCL). RPADPCL involves the crosslinking of biotinylated double-stranded RPA DNA products with horseradish peroxidase (HRP)-labeled streptavidin (SA-HRP). Modified products are captured using SA-labeled magnetic beads, and then analyzed using a chemiluminescence detector and smartphone after the addition of a chemiluminescent substrate. Under optimal conditions, the RPADPCL limit of detection (LOD) was observed to be 6 copies (within the linear detection range of 1-300 copies) for a plasmid containing the SARS-CoV-2 N gene and 15 copies (within the linear range of 10-500 copies) for in vitro transcribed (IVT) SARS-CoV-2 RNA. The proposed method is convenient, specific, visually intuitive, easy to use, and does not require external excitation. The effective RPADPCL detection of SARS-CoV-2 in complex matrix systems was verified by testing simulated clinical samples containing 10% human saliva or a virus transfer medium (VTM) spiked with a plasmid containing a SARS-CoV-2 N gene sequence or SARS-CoV-2 IVT RNA. Consequently, this method has great potential for detecting targets in clinical samples.

Azobenzene-Based Conjugated Polymers: Synthesis, Properties, and Biological Applications.

Conjugated polymers (CPs) have been developed quickly as an emerging functional material with applications in optical and electronic devices, owing to their highly electron-delocalized backbones and versatile side groups for facile processibility, high mechanical strength, and environmental stability. CPs exhibit multistimuli responsive behavior and fluorescence quenching properties by incorporating azobenzene functionality into their molecular structures. Over the past few decades, significant progress has been made in developing functional azobenzene-based conjugated polymers (azo-CPs), utilizing diverse molecular design strategies and synthetic pathways. This article comprehensively reviews the rapidly evolving research field of azo-CPs, focusing on the structural characteristics and synthesis methods of general azo-CPs, as well as the applications of charged azo-CPs, specifically azobenzene-based conjugated polyelectrolytes (azo-CPEs). Based on their molecular structures, azo-CPs can be broadly categorized into three primary types: linear CPs with azobenzene incorporated into the side chain, linear CPs with azobenzene integrated into the main chain, and branched CPs containing azobenzene moieties. These systems are promising for biomedical applications in biosensing, bioimaging, targeted protein degradation, and cellular apoptosis.

mRNA PROTACs: engineering PROTACs for high-efficiency targeted protein degradation.

Proteolysis-targeting chimeras (PROTACs) are essential bifunctional molecules that target proteins of interest (POIs) for degradation by cellular ubiquitination machinery. Despite significant progress made in understanding PROTACs' functions, their therapeutic potential remains largely untapped. As a result of the success of highly flexible, scalable, and low-cost mRNA therapies, as well as the advantages of the first generation of peptide PROTACs (p-PROTACs), we present for the first time an engineering mRNA PROTACs (m-PROTACs) strategy. This design combines von Hippel-Lindau (VHL) recruiting peptide encoding mRNA and POI-binding peptide encoding mRNA to form m-PROTAC and promote cellular POI degradation. We then performed proof-of-concept experiments using two m-PROTACs targeting two cancer-related proteins, estrogen receptor alpha and B-cell lymphoma-extra large protein. Our results demonstrated that m-PROTACs could successfully degrade the POIs in different cell lines and more effectively inhibit cell proliferation than the traditional p-PROTACs. Moreover, the in vivo experiment demonstrated that m-PROTAC led to significant tumor regression in the 4T1 mouse xenograft model. This finding highlights the enormous potential of m-PROTAC as a promising approach for targeted protein degradation therapy.

Universal smartphone-assisted label-free CRISPR/Cas12a-DNAzyme chemiluminescence biosensing platform for on-site detection of nucleic acid and non-nucleic acid targets.

The clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein (Cas) (CRISPR/Cas) system enables sensitive and specific detection of biomolecules, thanks to its programmability, high fidelity, and powerful signal amplification capabilities. Herein, a universal smartphone-assisted label-free G-quadruplex (G4) DNAzyme-based chemiluminescence CRISPR/Cas12a biosensing platform (G4CLCas) is firstly described that achieves on-site, ultrasensitive visual detection of nucleic acid and non-nucleic acid targets. The G4CLCas-based sensing platform relies on Cas12a trans-cleavage activation that triggers the cleavage of the G4 DNAzyme, resulting in chemiluminescence signals off/on compared to that of the control. Chemiluminescence signals are captured as images that are quantitatively analyzed and visualized using a smartphone-assisted imaging cartridge. Under optimal conditions, G4CLCas achieves a low limit of detection (LOD) of 8.6 aM (∼5.2 copies/µL) for monkeypox virus (MPXV) DNA within the linear concentration range of 10-300 aM and can accurately quantify viral DNA in spiked samples. G4CLCas can also detect non-nucleic acid targets, whereby it achieves a low LOD value of 84.3 nM for adenosine triphosphate (ATP) within the linear concentration range of 2-2000 µM. Here, a label-free, portable, on-site CRISPR/Cas12a chemiluminescence biosensing platform based on the G4 DNAzyme substrates is proposed with potential applications in clinical detection and bioanalytical chemistry research.

Recent Advances in Optically Controlled PROTAC.

Proteolysis-targeting chimera (PROTAC) technology is a groundbreaking therapeutic approach with significant clinical potential for degrading disease-inducing proteins within targeted cells. However, challenges related to insufficient target selectivity raise concerns about PROTAC toxicity toward normal cells. To address this issue, researchers are modifying PROTACs using various approaches to enhance their target specificity. This review highlights innovative optically controlled PROTACs as anti-cancer therapies currently used in clinical practice and explores the challenges associated with their efficacy and safety. The development of optically controlled PROTACs holds the potential to significantly expand the clinical applicability of PROTAC-based technology within the realm of drug discovery.

Conjugated Polyelectrolyte/Single Strand DNA Hybrid Polyplexes for Efficient Nucleic Acid Delivery and Targeted Protein Degradation.

Nucleic acid-based therapeutics have gained increasing attention due to their ability to regulate various genetic disorders. However, the safe and effective delivery of nucleic acids to their intended cellular sites remains a challenge, primarily due to poor cell membrane permeation and low in vivo stability. Limitations associated with the commonly used nucleic acid delivering agent viral vectors such as carcinogenesis and immunogenicity have driven scientists to develop various nonviral vectors. In this study, we present a highly efficient nucleic acid delivery system based on cationic conjugated polyelectrolytes and single-strand DNA polyplexes with further application in efficient ubiquitin-regulated targeting protein degradation. These polyplexes, formed by 9TC, an aptamer sequence for estrogen receptor (ERα), and cationic PPET3N2 through electrostatic and hydrophobic interactions, demonstrate improved cellular uptake efficiency as well as enhanced stability against nuclease degradation. Furthermore, by incorporation of 9TC into a proteolysis targeting chimera (PROTAC) molecule (P9TC), PPET3N2/P9TC polyplexes significantly enhance the target protein ERα degradation efficiency. Collectively, our findings suggest that PPET3N2 provides a versatile, low cytotoxicity platform for safe, efficient, and simplified delivery of nucleic acids.

Infrared Spectral Analysis for Prediction of Functional Groups Based on Feature-Aggregated Deep Learning.

Infrared (IR) spectroscopy is a powerful and versatile tool for analyzing functional groups in organic compounds. A complex and time-consuming interpretation of massive unknown spectra usually requires knowledge of chemistry and spectroscopy. This paper presents a new deep learning method for transforming IR spectral features into intuitive imagelike feature maps and prediction of major functional groups. We obtained 8272 gas-phase IR spectra from the NIST Chemistry WebBook. Feature maps are constructed using the intrinsic correlation of spectral data, and prediction models are developed based on convolutional neural networks. Twenty-one major functional groups for each molecule are successfully identified using binary and multilabel models without expert guidance and feature selection. The multilabel classification model can produce all prediction results simultaneously for rapid characterization. Further analysis of the detailed substructures indicates that our model is capable of obtaining abundant structural information from IR spectra for a comprehensive investigation. The interpretation of our model reveals that the peaks of most interest are similar to those often considered by spectroscopists. In addition to demonstrating great potential for spectral identification, our method may contribute to the development of automated analyses in many fields.

Establishment, verification, and application of concentration intervals corresponding to dipstick grades for urinary protein.

BACKGROUND: To address the situation that the accuracy of concentration intervals (CI) corresponding to dipstick grades is not given by the manufacturers or literature, we developed a method that determined reasonable dipstick grades with concentration intervals (GCIs) based on the percent agreement (PA) and discussed the GCI application to comparability among currently dipstick tests. METHODS: By comparing the results of 2 dipstick tests (iChem and KU-500) with the quantitative test (AU5800), the GCIs were verified and established based on the PAs, which were calculated and used as an indicator of GCI's accuracy. The overlap (percent) between the 2 GCIs with the same grade (2 dipstick devices), was calculated and used to evaluate the agreement between their test results. RESULTS: After verification and adjustment, the GCI and PA combinations for iChem Velocity were as follows: - (<0.1 g/l, 85 %), ± (0.1-0.3 g/l, 66 %), 1+ (0.3-1 g/l, 78 %), 2+ (1-3 g/l, 74 %), 3+ (3-6 g/l, 77 %), and 4+ (≥6 g/l, 84 %). The determined GCI and PA combinations for KU-500 were: - (<0.1.2 g/l, 75 %), ± (0.12-0.5 g/l, 63 %), 1+ (0.5-1.2 g/l, 69 %), 2+ (1.2-3.2 g/l, 76 %), and 3+ (≥3.2 g/l, 82 %). The GCI overlaps between the 2 dipstick devices were - (83 %), ± (45 %), 1+ (56 %), 2+ (82 %), and 3+ or ≥3+ (94 %). The overall overlap was 72 %. Since the overlaps ± (45 %) and 1+ (56 %) were within the overlap reject limit for any grade (70 %), and the overall overlap (72 %) was within the overall overlap reject limit (80 %), the test results of the 2 devices were not comparable. CONCLUSIONS: GCIs can be verified and established correctly based on PAs, and industry standards for dipstick tests can be established based on GCIs and PAs. Comparability between dipstick devices, historical data, and literature data can be roughly determined based on the overlap.

Transcriptome and Metabolomics Integrated Analysis Reveals MdMYB94 Associated with Esters Biosynthesis in Apple (Malus × domestica).

Volatile esters are major aromas contributing to the organoleptic quality of apple fruit. However, the molecular mechanisms underlying the regulation of volatile ester biosynthesis in apple remain elusive. This study investigated the volatile profiles and transcriptomes of 'Qinguan' (QG) apple fruit during development and/or postharvest storage. Although the constitution of volatiles varied widely between the peel and flesh, the volatile profiles of the peel and flesh of ripening QG fruit were dominated by volatile esters. WGCNA results suggested that 19 genes belonging to ester biosynthesis pathways and 11 hub transcription factor genes potentially participated in the biosynthesis and regulation of esters. To figure out key regulators of ester biosynthesis, correlation network analysis, dual-luciferase assays, and yeast one-hybrid assay were conducted and suggested that MdMYB94 trans-activated the MdAAT2 promoter and participated in the regulation of ester biosynthesis. This study provides a framework for understanding ester biosynthesis and regulation in apple.

Effects of enrofloxacin's exposure on the gut microbiota of Tilapia fish (Oreochromis niloticus).

Enrofloxacin (ENFX) has a broad-spectrum antibiotic activity, which is widely used in aquaculture. The effect of different ENFX exposure ways on the gut microbiota of tilapia is unclear. This study was conducted to investigate the effects of ENFX exposure on the gut microbiota of tilapia fish (Oreochromis niloticus). Three methods of ENFX exposure were selected: injection (IEG), oral administration (OEG) and soaking (SEG). After 48 h of exposure period, the intestine of tilapia was collected for high-throughput sequencing. PCoA analysis revealed a distinct clustering of control group, and which was located rather far away from ENFX exposure groups. The dominant phyla in the gut microbiota of tilapia fish were Proteobacteria, Actinobacteriota, Fusobacteria and Firmicutes. Compared to the control group, phylum Fusobacteriota was increased in SEG and IEG while decreased in OEG. ENFX treatment led to a decline in Corynebacterium, Clostridium sensu stricto_3 and Bacillus in treated fish compared with control fish, accompanied by an increase in Akkermansia, Ralstonia and Romboutsia. IEG had the least effect on gut microbiota of tilapia because it retained more microbes among treatment groups. Alpha- diversity decreased the most in SEG, but retained more probiotics such as Cetobacterium and Akkermansia. We assessed the effect of enrofloxacin on tilapia by changes in intestinal flora. The result indicated that either exposure method significantly reduced the diversity of tilapia gut microbiota. It may provide basic data for the scientific use of ENFX in aquaculture.

A universal platform for one-pot detection of circulating non-coding RNA combining CRISPR-Cas12a and branched rolling circle amplification.

Multiple circulating non-coding RNAs (ncRNAs) in serum may serve as vital biomarkers for use in diagnosing early-stage colorectal cancer (CRC). Herein, a universal platform for one-pot detection of CRC-related ncRNAs was developed based on branched rolling circle amplification and CRISPR-Cas12a (BRCACas). For the implementation of the method, primers incorporating ncRNA sequences of circulating CRC-associated RNAs (piRNA or miRNA) were designed that could specifically hybridize with circular probes to initiate the BRCA process. Thereafter, the generation of dendritic DNA products triggered Cas12a trans-cleavage activity to elicit a fluorescent signal. The proposed method, combining high BRCA reaction efficiency with powerful Cas12a trans-cleavage activity, provided greatly enhanced detection sensitivity, as reflected by limits of detection (LODs) for model piRNA (piR-54265) and model miRNA (miR21) of 0.76 fM and 0.87 fM, respectively. Notably, the proposed BRCACas platform, assaying two different types of CRC-associated ncRNAs in patient samples, produced consistent results with the conventional reverse transcription-quantitative PCR (RT-qPCR) method. Therefore, the one-pot, isothermal, and specific BRCACas platform provided excellent performance, thus demonstrating its promise as a rapid, adaptable, and practical diagnostic/prognostic cancer screening method.

Overexpression of ameloblastin in secretory ameloblasts results in demarcated, hypomineralized opacities in enamel.

Introduction: Developmental defects of the enamel manifest before tooth eruption and include amelogenesis imperfecta, a rare disease of underlying gene mutations, and molar-incisor hypomineralization (MIH), a prevalent disease in children originating from environmental and epigenetic factors. MIH enamel presents as the abnormal enamel marked by loss of translucency, demarcation between the healthy and affected enamel, and reduced mineral content. The pathophysiology of opaque, demarcated enamel lesions is not understood; however, the retention of enamel proteins in the matrix has been suggested. Ameloblastin (Ambn) is an enamel protein of the secreted calcium-binding phosphoproteins (SCPPs) critical for enamel formation. When the Ambn gene is mutated or deleted, teeth are affected by hypoplastic amelogenesis imperfecta. Methods: In this study, enamel formation in mice was analyzed when transgenic Ambn was overexpressed from the amelogenin promoter encoding full-length Ambn. Ambn was under- and overexpressed at six increasing concentrations in separate mouse lines. Results: Mice overexpressing Ambn displayed opaque enamel at low concentrations and demarcated lesions at high concentrations. The severity of enamel lesions increased starting from the inner enamel close to the dentino-enamel junction (DEJ) to span the entire width of the enamel layer in demarcated areas. Associated with the opaque enamel were 17-kDa Ambn cleavage products, a prolonged secretory stage, and a thin basement membrane in the maturation stage. Ambn accumulations found in the innermost enamel close to the DEJ and the mineralization front correlated with reduced mineral content. Demarcated enamel lesions were associated with Ambn species of 17 kDa and higher, prolonged secretory and transition stages, a thin basement membrane, and shortened maturation stages. Hypomineralized opacities were delineated against the surrounding mineralized enamel and adjacent to ameloblasts detached from the enamel surface. Inefficient Ambn cleavage, loss of contact between ameloblasts, and the altered basement membrane curtailed the endocytic activity; thus, enamel proteins remained unresorbed in the matrix. Ameloblasts have the ability to distinguish between Ambn concentration and Ambn cleavage products through finely tuned feedback mechanisms. The under- or overexpression of Ambn in murine secretory ameloblasts results in either hypoplastic amelogenesis imperfecta or hypomineralization with opaque or sharply demarcated boundaries of lesions, similar to MIH.

Effect of Continuous Nursing Model Based on WeChat Public Health Education on Self-Management Level and Treatment Compliance of Stroke Patients.

Background: To analyze the effect of continuous nursing model based on WeChat public health education on self-management level and treatment compliance of stroke patients. Methods: Overall, 98 stroke patients admitted to the Second Affiliated Hospital of Chongqing Medical University, ChongQing, 400000,China from Feb 2018 to Feb 2019 were enrolled as the research objects. They were randomly and equally divided into experimental group and control group. The control group received routine nursing management model while the experimental group received continuous nursing model based on WeChat public health education to compare the self-management level, treatment compliance, life quality and other indexes between the two groups. Results: The overall self-management level in the experimental group was significantly higher than that in the control group (P<0.05). The life quality scores in both groups after nursing intervention increased, and the scores in the experimental group after nursing intervention were significantly higher than those in the control group (P<0.001). The total treatment compliance rate in the experimental group was higher than that in the control group (P=0.001). After nursing intervention, the readmission rate and complication rate in the experimental group were significantly lower than those in the control group (P<0.05). Conclusion: The implementation of continuous nursing model based on WeChat public health education for stroke patients can significantly enhance the self-management level, improve the cognitive level on stroke related-health knowledge, improve treatment compliance of patients in the nursing process, promote their physical and mental health, improve the life quality and effectively reduce the complication rate and readmission rate.

PROTAC Degrader of Estrogen Receptor α Targeting DNA-Binding Domain in Breast Cancer.

PROteolysis-TArgeting Chimeras (PROTACs) are a powerful class of drugs that selectively degrade the proteins of interest (POIs) through cellular ubiquitination mechanisms. Estrogen receptor α (ERα) plays a vital role in the pathogenesis and treatment of breast cancer. In this work, the DNA-binding domain (DBD) of ERα was selected as the target to avoid drug resistance caused by the ligand-binding domain (LBD) of ERα. The estrogen response element (ERE), a natural DNA sequence binding with DBD of ERα, was chosen as a recognized unit of PROTAC. Therefore, we designed a nucleic acid-conjugated PROTAC, ERE-PROTAC, via a click reaction, in which the ERE sequence recruits ERα and the typical small molecule VH032 recruits the von Hippel-Lindau (VHL) E3 ligase. The proposed ERE-PROTAC showed to efficiently and reversibly degrade ERα in different breast cancer cells by targeting the DBD, indicating its potential to overcome the current resistance caused by LBD mutations.

Lactococcus lactis' Effect on the Intestinal Microbiota of Streptococcus agalactiae-Infected Zebrafish (Danio rerio).

Streptococcus agalactiae is a common pathogen in aquaculture that disrupts the balance of the intestinal microbiota and threatens fish health, causing enormous losses to the aquaculture industry. In this study, we isolated and screened a Lactococcus lactis KUST48 (LLK48) strain with antibacterial effect against S. agalactiae in vitro and used it as a potential probiotic to explore its therapeutic effect on zebrafish (Danio rerio) infected with S. agalactiae. This study divided zebrafish into 3 groups: control group, injected with phosphate-buffered saline; infection group, injected with S. agalactiae; and treatment group, treated with LLK48 after S. agalactiae injection. Then, the 16S rRNA gene sequences of the intestinal microbiota of these 3 groups were sequenced using Illumina high-throughput sequencing technology. The results showed that the relative abundance of intestinal bacteria was significantly decreased in the infection group, and a high relative abundance of S. agalactiae was observed. The relative abundance of the intestinal microbiota was increased in the treatment group, with a decrease in the relative abundance of S. agalactiae compared to that in the control group. In the Cluster of Orthologous Groups of proteins function classification, the relative abundance of each biological function in the infection group was significantly lower than that of the control and treatment groups, showing that LLK48 has a positive therapeutic effect on zebrafish infected with S. agalactiae. This study provides a foundation for exploring the pathogenic mechanism of S. agalactiae on fish and their intestinal symbionts, and also presents a new approach for the treatment of S. agalactiae infections in fish aquaculture systems. IMPORTANCE L. lactis KUST48 (LLK48) with a bacteriostatic effect against S. agalactiae was isolated from tilapia intestinal tracts. S. agalactiae infection significantly reduced the relative abundance of intestinal bacteria and various physiological functions in zebrafish intestines. LLK48 demonstrated infection and subsequent therapeutic effects on the S. agalactiae infection in the zebrafish intestine. Therefore, the potential probiotic LLK48 can be considered as a therapeutic treatment for S. agalactiae infections in aquaculture, which can reduce the use of antibiotics and help maintain fish health.

Molecular Design and Photothermal Application of Thienoisoindigo Dyes with Aggregation-Induced Emission.

Organic fluorescent dyes with aggregation-induced emission (AIE) property have an extensive application range, especially in the fields of imaging, labeling, and adjusting microprocesses in aggregated environments. In particular, the thienoisoindigo skeleton, which exhibits an outstanding electron-withdrawing capacity in optoelectronic materials, has been defined as a promising AIE candidate. For instance, by installing AIE blocks or other rotatable groups at two terminal sites, such as various arylamine groups, thienoisoindigo derivatives can be efficiently turned to be functional AIE structures. In this work, a thienoisoindigo derivative with AIE characteristics, namely, TII-TPE, was developed. This AIE system was expanded by linking typical AIE fragments, namely, tetraphenylethene, with the proposed thienoisoindigo derivative, which exhibited typical AIE fluorescence in the 600-850 nm range and maintained high photostability. Then, employing the reported derivative TII-TPA coating thienoisoindigo and triphenylamine as a contrast, aggregated TII-TPE and TII-TPA nanoparticles were prepared and demonstrated photothermal conversion efficiencies of 36.2 and 35.6%, respectively. Moreover, both nanoparticles were evaluated as photothermal therapeutic (PTT) agents in a tumor mouse model, which showed to significantly inhibit tumor growth after four treatment cycles in vivo. This work not only presents an enriched thienoisoindigo system but also provides a pattern for subsequent construction of functional AIE molecules.