Elucidation of CKAP4-remodeled cell mechanics in driving metastasis of bladder cancer through aptamer-based target discovery.

Metastasis contributes to the dismal prognosis of bladder cancer (BLCA). The mechanical status of the cell membrane is expected to mirror the ability of cell migration to promote cancer metastasis. However, the mechanical characteristics and underlying molecular profile associated with BLCA metastasis remain obscure. To study the unique cellular architecture and traits associated with cell migration, using a process called cell-based systematic evolution of ligands by exponential enrichment (cell-SELEX) we generated an aptamer-based molecular probe, termed spl3c, which identified cytoskeleton-associated protein 4 (CKAP4). CKAP4 was associated with tumor metastasis in BLCA, but we also found it to be a mechanical regulator of BLCA cells through the maintenance of a central-to-peripheral gradient of stiffness on the cell membrane. Notably, such mechanical traits were transportable through exosome-mediated intercellular CKAP4 trafficking, leading to significant enhancement of migration in recipient cells and, consequently, aggravating metastatic potential in vivo. Taken together, our study shows the robustness of this aptamer-based molecular tool for biomarker discovery, revealing the dominance of a CKAP4-induced central-to-peripheral gradient of membrane stiffness that benefits cell migration and delineating the role of exosomes in mediating mechanical signaling in BLCA metastasis.

Label-Free Multiplex Profiling of Exosomal Proteins with a Deep Learning-Driven 3D Surround-Enhancing SERS Platform for Early Cancer Diagnosis.

Identification of protein profiling on plasma exosomes by SERS can be a promising strategy for early cancer diagnosis. However, it is still challenging to detect multiple exosomal proteins simultaneously by SERS since the Raman signals of exosomes detected by conventional colloidal nanocrystals or two-dimensional SERS substrates are incomplete and complex. Herein, we develop a novel three-dimensional (3D) surround-enhancing SERS platform, named 3D se-SERS, for the multiplex detection of exosomal proteins. In this 3D se-SERS, proteins and exosomes are covered with "hotspots" generated by the gold nanoparticles, which surround the analytes densely and three-dimensionally, providing sensitive and comprehensive SERS signals. Combining this 3D se-SERS with a deep learning model, we successfully quantitatively profiled seven proteins including CD63, CD81, CD9, CD151, CD171, TSPAN8, and PD-L1 on the surface of plasma exosomes from patients, which can predict the occurrence and advancement of lung cancer. This 3D se-SERS integrating deep learning technique benefits from high sensitivity and significant multiplexing ability for comprehensive analysis of proteins and exosomes, demonstrating the potential of deep learning-driven 3D se-SERS technology for plasma exosome-based early cancer diagnosis.

Tomographic FLEET with a wedge array for multi-point three-component velocimetry.

Femtosecond laser electronic excitation tagging (FLEET) velocimetry is an important diagnostic technique for seedless velocimetry measurements particularly in supersonic and hypersonic flows. Typical FLEET measurements feature a single laser line and camera system to achieve one-component velocimetry along a line, although some multiple-spot and multiple-component configurations have been demonstrated. In this work, tomographic imaging is used to track the three-dimensional location of many FLEET spots. A quadscope is used to combine four unique views onto a single high-speed image intensifier and camera. Tomographic reconstructions of the FLEET emission are analyzed for three-component velocimetry from multiple FLEET spots. Glass wedges are used to create many (nine) closely spaced FLEET spots with less than 10% transmission losses. These developments lead to a significant improvement in the dimensionality and spatial coverage of a FLEET instrument with some increases in experimental complexity and data processing. Multiple-point three-component FLEET velocimetry is demonstrated in an underexpanded jet.

Predicting the unpredictable: a robust nomogram for predicting recurrence in patients with ampullary carcinoma.

OBJECTIVE: To screen the risk factors affecting the recurrence risk of patients with ampullary carcinoma (AC)after radical resection, and then to construct a model for risk prediction based on Lasso-Cox regression and visualize it. METHODS: Clinical data were collected from 162 patients that received pancreaticoduodenectomy treatment in Hebei Provincial Cancer Hospital from January 2011 to January 2022. Lasso regression was used in the training group to screen the risk factors for recurrence. The Lasso-Cox regression and Random Survival Forest (RSF) models were compared using Delong test to determine the optimum model based on the risk factors. Finally, the selected model was validated using clinical data from the validation group. RESULTS: The patients were split into two groups, with a 7:3 ratio for training and validation. The variables screened by Lasso regression, such as CA19-9/GGT, AJCC 8th edition TNM staging, Lymph node invasion, Differentiation, Tumor size, CA19-9, Gender, GPR, PLR, Drinking history, and Complications, were used in modeling with the Lasso-Cox regression model (C-index = 0.845) and RSF model (C-index = 0.719) in the training group. According to the Delong test we chose the Lasso-Cox regression model (P = 0.019) and validated its performance with time-dependent receiver operating characteristics curves(tdROC), calibration curves, and decision curve analysis (DCA). The areas under the tdROC curves for 1, 3, and 5 years were 0.855, 0.888, and 0.924 in the training group and 0.841, 0.871, and 0.901 in the validation group, respectively. The calibration curves performed well, as well as the DCA showed higher net returns and a broader range of threshold probabilities using the predictive model. A nomogram visualization is used to display the results of the selected model. CONCLUSION: The study established a nomogram based on the Lasso-Cox regression model for predicting recurrence in AC patients. Compared to a nomogram built via other methods, this one is more robust and accurate.

A Novel Near-Infrared Fluorescent Probe for the Detection of Endogenous Peroxynitrite (ONOO-) in Living Cells.

In this study, we present a novel near-infrared (NIR) fluorescent probe Nile-ONO designed for the selective and sensitive detection of ONOO-. The probe Nile-ONO employed Nile red as the fluorophore, with diphenylphosphinate serving as the reaction site. In the presence of ONOO-, the probe Nile-ONO exhibits remarkable fluorescence enhancement at 659 nm, with a response time of less than 20 min and a low detection limit of 0.32 µM. Importantly, MTT assays demonstrate low cytotoxicity in living cells. Furthermore, Nile-ONO has excellent imaging capabilities for endogenous ONOO-. Overall, this work introduces a valuable new method for the rapid detection of ONOO- in biological systems.

Lack of causal association between epilepsy and dementia: A Mendelian randomization analysis.

OBJECTIVE: Epidemiological studies have reported an association between epilepsy and dementia. However, the causal relationship between epilepsy and the risk of dementia is not clear. We aimed to inspect the causal effect of epilepsy on memory loss and dementia. METHODS: We analyzed summary data of epilepsy, memory loss, and dementia from the genome-wide association study (GWAS) using the two-sample Mendelian randomization (MR) method. We used the estimated odds ratio of memory loss and dementia associated with each of the genetically defined traits to infer evidence for a causal relationship with the following exposures: all epilepsy, focal epilepsy (including focal epilepsy with hippocampal sclerosis, lesion-negative focal epilepsy, and focal epilepsy with other lesions), and genetic generalized epilepsy (including childhood absence epilepsy, generalized tonic-clonic seizures alone, Juvenile absence epilepsy, and Juvenile myoclonic epilepsy). RESULTS: According to the result of MR using the inverse variance weighted method (IVW), we found that genetically predicted epilepsy did not causally increase the risk of memory loss and dementia (p > 0.05). Results of the MR-Egger and weighted median method were consistent with the IVW method. CONCLUSIONS: No evidence has been found to support the notion that epilepsy can result in memory loss and dementia. The associations observed in epidemiological studies could be attributed, in part, to confounding or nongenetic determinants.

Evaluation of a biomarker (NO) dynamics in inflammatory zebrafish and periodontitis saliva samples via a fast-response and sensitive fluorescent probe.

Many pathological processes include nitric oxide (NO), a signaling transduction molecule. Tumors, cardiovascular, cerebrovascular, neurodegenerative, and other illnesses are linked to abnormal NO levels. Thus, evaluating NO levels in vitro and in vivo is crucial for studying chemical biology process of associated disorders. This work devised and manufactured a coumarin-based fluorescent probe ZPS-NO to detect nitric oxide (NO). The reaction between ZPS-NO and NO produced a highly selective and sensitive optical response that caused a powerful fluorescence "turn-on" effect with a ultra-low NO detection limit of 14.5 nM. Furthermore, the probe was applied to sense and image NO in living cells and inflammatory model of zebrafish, as well as to detect NO in periodontitis patients' saliva samples. We anticipate that probe ZPS-NO will serve as a practical and effective tool for assessing the interactions and evaluation of periodontitis development.

Factors Influencing Noise Following Primary Ceramic-on-Ceramic Total Hip Arthroplasty.

BACKGROUND: The noise associated with ceramic-on-ceramic (CoC) total hip arthroplasty (THA) has been a concerning issue, while its underlying causes remain unclear. METHODS: We conducted a retrospective analysis of 119 patients (174 primary CoC THAs) who had a mean follow-up of 28 months (range, 12 to 106). A questionnaire was designed to collect information on nature, frequency, onset, duration, and impact of the noise. Postoperative x-rays were evaluated. Clinical evaluations, including Harris and Oxford hip scores, were documented at follow-up time points (6 weeks, 3 months, 6 months, and 1 year). RESULTS: Of the 174 hips, 31.6% reported noise, including 26 popping (14.9%), 24 clicking (12.1%), and 5 grinding (2.9%). No patients reported squeaking. Noisy hips had lower age (P = .009) and body mass index (P = .019). Among patients with developmental dysplasia of the hip, 17 of 55 hips reported noise associated with smaller cup anteversion angle (P = .004), greater body height (P = .022), and larger acetabular cup size (P = .049). Noise typically began at a mean of 193 days (range, 1 to 2,598) after surgery and disappeared spontaneously in 50.9% of hips before final follow-up, with an average disappearance time of 211 days (range, 60 to 730). Noise did not affect daily life in 74.5% of patients, while 26.9% of patients who had popping reported painful sensations. One patient experienced joint dislocation, and another experienced a ceramic liner fracture during follow-up. No statistical difference was observed in outcome scores between noise and silent groups at 4 follow-up time points. CONCLUSIONS: Incidence of noise after primary CoC THA is relatively high. Smaller cup anteversion and larger acetabular cup size were associated with noise production in patients who had developmental dysplasia of the hip.

Who Are the Anatomic Outliers Undergoing Total Knee Arthroplasty? A Computed Tomography-Based Analysis of the Hip-Knee-Ankle Axis Across 1,352 Preoperative Computed Tomographies Using a Deep Learning and Computer Vision-Based Pipeline.

BACKGROUND: Dissatisfaction after total knee arthroplasty (TKA) ranges from 15 to 30%. While patient selection may be partially responsible, morphological and reconstructive challenges may be determinants. Preoperative computed tomography (CT) scans for TKA planning allow us to evaluate the hip-knee-ankle axis and establish a baseline phenotypic distribution across anatomic parameters. The purpose of this cross-sectional analysis was to establish the distributions of 27 parameters in a pre-TKA cohort and perform threshold analysis to identify anatomic outliers. METHODS: There were 1,352 pre-TKA CTs that were processed. A 2-step deep learning pipeline of classification and segmentation models identified landmark images and then generated contour representations. We used an open-source computer vision library to compute measurements for 27 anatomic metrics along the hip-knee axis. Normative distribution plots were established, and thresholds for the 15th percentile at both extremes were calculated. Metrics falling outside the central 70th percentile were considered outlier indices. A threshold analysis of outlier indices against the proportion of the cohort was performed. RESULTS: Significant variation exists in pre-TKA anatomy across 27 normally distributed metrics. Threshold analysis revealed a sigmoid function with a critical point at 9 outlier indices, representing 31.2% of subjects as anatomic outliers. Metrics with the greatest variation related to deformity (tibiofemoral angle, medial proximal tibial angle, lateral distal femoral angle), bony size (tibial width, anteroposterior femoral size, femoral head size, medial femoral condyle size), intraoperative landmarks (posterior tibial slope, transepicondylar and posterior condylar axes), and neglected rotational considerations (acetabular and femoral version, femoral torsion). CONCLUSIONS: In the largest non-industry database of pre-TKA CTs using a fully automated 3-stage deep learning and computer vision-based pipeline, marked anatomic variation exists. In the pursuit of understanding the dissatisfaction rate after TKA, acknowledging that 31% of patients represent anatomic outliers may help us better achieve anatomically personalized TKA, with or without adjunctive technology.

Foldback-crRNA-Enhanced CRISPR/Cas13a System (FCECas13a) Enables Direct Detection of Ultrashort sncRNA.

The CRISPR/Cas13a system has promising applications in clinical small noncoding RNA (sncRNA) detection because it is free from the interference of genomic DNA. However, detecting ultrashort sncRNAs (less than 20 nucleotides) has been challenging because the Cas13a nuclease requires longer crRNA-target RNA hybrids to be activated. Here, we report the development of a foldback-crRNA-enhanced CRISPR/Cas13a (FCECas13a) system that overcomes the limitations of the current CRISPR/Cas13a system in detecting ultrashort sncRNAs. The FCECas13a system employs a 3'-terminal foldback crRNA that hybridizes with the target ultrashort sncRNA, forming a double strand that "tricks" the Cas13a nuclease into activating the HEPN structural domain and generating trans-cleavage activity. The FCECas13a system can accurately detect miRNA720 (a sncRNA currently known as tRNA-derived small RNA), which is only 17 nucleotides long and has a concentration as low as 15 fM within 20 min. This FCECas13a system opens new avenues for ultrashort sncRNA detection with significant implications for basic biological research, disease prognosis, and molecular diagnosis.

Development of Uveal Melanoma-Specific Aptamer for Potential Biomarker Discovery and Targeted Drug Delivery.

Uveal melanoma (UM) is the most common primary intraocular malignancy in adults. However, challenges in early diagnosis, high risk of liver metastasis, and lack of effective targeted therapy lead to poor prognosis and high mortality of UM. Therefore, generating an effective molecular tool for UM diagnosis and targeted treatment is of great significance. In this study, a UM-specific DNA aptamer, PZ-1, was successfully developed, which could specifically distinguish molecular differences between UM cells and noncancerous cells with nanomolar-range affinity and presented excellent recognition ability for UM in vivo and clinical UM tissues. Subsequently, the binding target of PZ-1 on UM cells was identified as JUP (junction plakoglobin) protein, which held great potential as a biomarker and therapeutic target for UM. Meanwhile, the strong stability and internalization capacity of PZ-1 were also determined, and a UM-specific aptamer-guided "nanoship" was engineered to load and selectively release doxorubicin (Dox) to targeted UM cells, with lower toxicity to nontumor cells. Taken together, the UM-specific aptamer PZ-1 could serve as a molecular tool to discover the potential biomarker for UM and to achieve the targeted therapy of UM.

Generation of DNA Aptamers with Functional Activity in Mammalian Cells by Mimicking Retroviruses.

DNA aptamers are single-stranded DNA oligonucleotide sequences that bind to specific targets with high affinity. Currently, DNA aptamers can be produced only by in vitro synthesis. It is difficult for DNA aptamers to have a sustained impact on intracellular protein activity, which limits their clinical application. In this study, we developed a DNA aptamer expression system to generate DNA aptamers with functional activity in mammalian cells by mimicking retroviruses. Using this system, DNA aptamers targeting intracellular Ras (Ra1) and membrane-bound CD71 (XQ2) were successfully generated in cells. In particular, the expressed Ra1 not only specifically bound to the intracellular Ras protein but also inhibited the phosphorylation of downstream ERK1/2 and AKT. Furthermore, by inserting the DNA aptamer expression system for Ra1 into a lentivirus vector, the system can be delivered into cells and stably produce Ra1 over time, resulting in the inhibition of lung cancer cell proliferation. Therefore, our study provides a novel strategy for the intracellular generation of DNA aptamers with functional activity and opens a new avenue for the clinical application of intracellular DNA aptamers in disease treatment.

Aptamer-Mediated Enrichment of Rare Circulating Fetal Nucleated Red Blood Cells for Noninvasive Prenatal Diagnosis.

Isolation of circulating fetal nucleated red blood cells (cfNRBCs) from maternal peripheral blood provides a superior strategy for noninvasive prenatal genetic diagnosis. Recent technical advances in single-cell isolation and genetic analyses have promoted the clinical application of circulating fetal cell-based noninvasive prenatal diagnosis. However, the lack of highly specific ligands for rare circulating fetal cell enrichment from massive maternal cells significantly impedes the clinical transformation progress. In this work, aptamers specific to NRBCs were developed through clinical sample-based cell-SELEX. Herein, the complex clinical system provides natural selection stringency through binding competition between target and background cells, and it empowers aptamers with high specificity. An aptamer-based strategy was also established to isolate cfNRBCs from maternal peripheral blood. Results show the remarkable selectivity and affinity of developed aptamers, enabling efficient enrichment of cfNRBCs from abundant maternal cells. Moreover, screening for fetal sex and trisomy syndrome achieved high accuracy through chromosome analysis of enriched cfNRBCs. To the best of our knowledge, this is the first report to develop aptamer ligands for cfNRBC enrichment, providing an efficient strategy to screen cfNRBC-specific ligands and demonstrating broad application potential for cfNRBC-based noninvasive prenatal diagnosis.

Identification of QTL for reducing loss of grain yield under salt stress conditions in bi-parental populations derived from wheat landrace Hongmangmai.

KEY MESSAGE: A novel QTL (QSt.nftec-2BL) was mapped to a 0.7 cM interval on chromosome 2B. Plants carrying QSt.nftec-2BL produced higher grain yields by up to 21.4% than otherwise in salinized fields. Wheat yield has been limited by soil salinity in many wheat-growing areas globally. The wheat landrace Hongmangmai (HMM) possesses salt tolerance as it produced higher grain yields than other tested wheat varieties including Early Premium (EP) under salt stresses. To detect QTL underlying this tolerance, wheat cross EP × HMM was chosen to serve as mapping population that was homozygous at Ppd (photoperiod response gene), Rht (reduced plant height gene) and Vrn (vernalization gene); thus, interference with QTL detection by these loci could be minimized. QTL mapping was conducted firstly using 102 recombinant inbred lines (RILs) that were selected from the EP × HMM population (827 RILs) for similarity in grain yield under non-saline condition. Under salt stresses, however, the 102 RILs varied significantly in grain yield. These RILs were genotyped using a 90 K SNP (single nucleotide polymorphism) array; consequently, a QTL (QSt.nftec-2BL) was detected on chromosome 2B. Then, using 827 RILs and new simple sequence repeat (SSR) markers developed according to the reference sequence IWGSC RefSeq v1.0, location of QSt.nftec-2BL was refined to a 0.7 cM (6.9 Mb) interval flanked by SSR markers 2B-557.23 and 2B-564.09. Selection for QSt.nftec-2BL was performed based on the flanking markers using two bi-parental wheat populations. Trials for validating effectiveness of the selection were conducted in salinized fields in two geographical areas and two crop seasons, demonstrating that wheat plants with the salt-tolerant allele in homozygous status at QSt.nftec-2BL produced higher grain yields by up to 21.4% than otherwise.

Enantioselective conjugate addition of malonates to α,ß-unsaturated aldehydes catalysed by 4-oxalocrotonate tautomerase.

The enantioselective conjugate addition of malonates to α,ß-unsaturated aldehydes catalysed by 4-oxalocrotonate tautomerase is described. High conversions, high enantioselectivities, and good isolation yields were achieved for a range of substrates. We further completed a four-step synthesis of the antidepressant (+)-femoxetine by utilizing this reaction and an enzymatic reductive amination reaction.

Near-infrared fluorescent probe based on rhodamine derivative for detection of NADH in live cells.

A near-infrared fluorescent probe was prepared for selective detection of reduced nicotinamide adenine dinucleotide (NADH) in live cells. The probe turns off the fluorescence with a closed spironolactone switch. However, reduction of the probe by NADH turns on fluorescence at 740 nm. Theoretical calculations suggest a more planar arrangement between the rhodamine and quinoline moieties with increased π-delocalization resulting from reduction.

Nucleic acid aptamer controls mycoplasma infection for inhibiting the malignancy of esophageal squamous cell carcinoma.

Esophageal cancer is one of the most frequent malignant tumors of the digestive tract, among which esophageal squamous cell carcinoma (ESCC) is the main pathological type worldwide. Previous studies have shown microbial infections in the upper digestive tract to be a potential risk factor in ESCC etiology. In this study, we identified that Mycoplasma hyorhinis infection promoted the malignancy of ESCC. In response, we generated a single-stranded DNA aptamer, ZY3A, against M. hyorhinis using the cell-SELEX strategy. The underlying recognition mechanism of ZY3A on M. hyorhinis involves its binding to M. hyorhinis-specific p37 protein. This tool allowed us to provide the first proof-of-concept evidence using a nucleic acid aptamer to control mycoplasma infection. More specifically, we found that ZY3A could neutralize M. hyorhinis infection on ESCC cells by blocking the interaction between p37 protein and its receptor TLR4 on the ESCC cell membrane. As a result, ZY3A inhibited the migration and invasion of M. hyorhinis-infected ESCC cells in vitro and metastasis in vivo. Taken together, these findings indicate that aptamer ZY3A is a potential candidate for development into a novel molecular tool for treatment of M. hyorhinis infection and a safe first-in-class M. hyorhinis-targeting antitumor agent.

Visualization of photothermal therapy by semiconducting polymer dots mediated photoacoustic detection in NIR II.

Visualization of photothermal therapy mediated by photothermal transduction agents (PTAs) is important to promote individual treatment of patients with low side effects. Photoacoustic detection has emerged as a promising noninvasive method for the visualization of PTAs distribution but still has limitations in temperature measurement, including poor measurement accuracy and low tissue penetration depth. In this study, we developed biocompatible semiconducting polymer dots (SPD) for in situ coupling of photothermal and photoacoustic detection in the near-infrared II window. SPD has dual photostability under pulsed laser and continuous-wave laser irradiation with a photothermal conversion efficiency of 42.77%. Meanwhile, a strong correlation between the photoacoustic signal and the actual temperature of SPD can be observed. The standard deviation of SPD-mediated photoacoustic thermometry can reach 0.13 °C when the penetration depth of gelatin phantom is 9.49 mm. Preliminary experimental results in vivo show that SPD-mediated photoacoustic signal has a high signal-to-noise ratio, as well as good performance in temperature response and tumor enrichment. Such a study not only offers a new nanomaterial for the visualization of photothermal therapy but will also promote the theranostic platform for clinical applications.

Machine-learning screening of luminogens with aggregation-induced emission characteristics for fluorescence imaging.

Due to the excellent biocompatible physicochemical performance, luminogens with aggregation-induced emission (AIEgens) characteristics have played a significant role in biomedical fluorescence imaging recently. However, screening AIEgens for special applications takes a lot of time and efforts by using conventional chemical synthesis route. Fortunately, artificial intelligence techniques that could predict the properties of AIEgen molecules would be helpful and valuable for novel AIEgens design and synthesis. In this work, we applied machine learning (ML) techniques to screen AIEgens with expected excitation and emission wavelength for biomedical deep fluorescence imaging. First, a database of various AIEgens collected from the literature was established. Then, by extracting key features using molecular descriptors and training various state-of-the-art ML models, a multi-modal molecular descriptors strategy has been proposed to extract the structure-property relationships of AIEgens and predict molecular absorption and emission wavelength peaks. Compared to the first principles calculations, the proposed strategy provided greater accuracy at a lower computational cost. Finally, three newly predicted AIEgens with desired absorption and emission wavelength peaks were synthesized successfully and applied for cellular fluorescence imaging and deep penetration imaging. All the results were consistent successfully with our expectations, which demonstrated the above ML has a great potential for screening AIEgens with suitable wavelengths, which could boost the design and development of novel organic fluorescent materials.

Correction: lncRNA PSMB8-AS1 promotes colorectal cancer progression through sponging miR-1299 to upregulate ADAMTS5.

This corrects the article DOI: 10.4149/neo_2022_220111N42.