Live-cell super-resolved PAINT imaging of piconewton cellular traction forces.

Despite the vital role of mechanical forces in biology, it still remains a challenge to image cellular force with sub-100-nm resolution. Here, we present tension points accumulation for imaging in nanoscale topography (tPAINT), integrating molecular tension probes with the DNA points accumulation for imaging in nanoscale topography (DNA-PAINT) technique to map piconewton mechanical events with ~25-nm resolution. To perform live-cell dynamic tension imaging, we engineered reversible probes with a cryptic docking site revealed only when the probe experiences forces exceeding a defined mechanical threshold (~7-21 pN). Additionally, we report a second type of irreversible tPAINT probe that exposes its cryptic docking site permanently and thus integrates force history over time, offering improved spatial resolution in exchange for temporal dynamics. We applied both types of tPAINT probes to map integrin receptor forces in live human platelets and mouse embryonic fibroblasts. Importantly, tPAINT revealed a link between platelet forces at the leading edge of cells and the dynamic actin-rich ring nucleated by the Arp2/3 complex.

DNA Nanotechnology for Investigating Mechanical Signaling in the Immune System.

Immune recognition occurs at specialized cell-cell junctions when immune cells and target cells physically touch. In this junction, groups of receptor-ligand complexes assemble and experience molecular forces that are ultimately generated by the cellular cytoskeleton. These forces are in the range of piconewton (pN) but play crucial roles in immune cell activation and subsequent effector responses. In this minireview, we will review the development of DNA based molecular tension sensors and their applications in mapping and quantifying mechanical forces experienced by immunoreceptors including T-cell receptor (TCR), Lymphocyte function-associated antigen (LFA-1), and the B-cell receptor (BCR) among others. In addition, we will highlight the use of DNA as a mechanical gate to manipulate mechanotransduction and decipher how mechanical forces regulate antigen discrimination and receptor signaling.

Diazabicyclic Electroactive Ionenes for Efficient and Stable Organic Solar Cells.

Electroactive ionenes combining caged-shaped diazabicyclic cations and aromatic diimides were developed as interlayers in organic solar cells (OSCs). These ionenes reduce the work-function of air-stable metal electrodes (e.g., Ag, Cu and Au) by generating strong interfacial dipoles, and their optoelectronic and morphological characters can be modulated by aromatic diimides, leading to high conductivity and good compatibility with active layers. The optimal ionene exhibits superior charge-transport, desirable crystallinity, and weak visible-absorption, boosting the efficiency of benchmark PM6 : Y6-based OSCs up to 17.44 %. The corresponding normal devices show excellent stability at maximum power point test under one sun illumination for 1000 h. Replacing Y6 with L8-BO promotes the efficiency to 18.43 %, one of the highest in binary OSCs. Notably, high efficiencies >16 % are maintained as the interlayer thickness increasing to 105 nm, the best result with interlayer-thickness over 100 nm.

The factors influencing the psychological distress of transgender women in Shandong, China: a cross-sectional study.

BACKGROUND: As a group at high-risk for acquired immune deficiency syndrome (AIDS) infection, the psychological distress of transgender women cannot be ignored while preventing and controlling AIDS risks. Transgender women are a vulnerable group, and their psychological distress deserves attention. The purpose of this study was to evaluate the psychological distress of transgender women and further determine the influences of factors on the psychological distress of transgender women. METHODS: From March 2021 to August 2021, a cross-sectional survey was conducted in Shandong province, China. Data were collected by a questionnaire designed for transgender women, and the GHQ-12 scale was used to measure their psychological distress. The questionnaire combined sociodemographic characteristics, HIV/AIDS cognition, related behaviors, substance abuse, social support, gender identity and other factors. Univariate logistic regression and multivariate logistic regression models were used to explore the psychological factors of transgender women. RESULTS: In this study, the rate of transgender women with psychological distress was 20.08%. Earned monthly income between 10,000-15,000 yuan (OR:0.16, 95% CI:0.06-0.45) and a monthly income greater than 15,000 yuan (OR:0.07, 95% CI:0.01-0.43) were protective factors in the psychology of transgender women. Transgender women who never disclosed sexual orientation and identity (OR: 0.19, 95% CI: 0.06-0.58), who only disclosed their sexual orientation and identity to families or friends (OR: 0.41, 95% CI: 0.18-0.93) were also less likely to have psychological distress. Additionally, transgender women who did not desire to be identified with their sexual orientation and identity (OR: 3.31, 95%CI: 1.08-10.16) and who reported that the Internet did not play an essential role in helping determine sexual orientation (OR: 5.96, 95% CI: 2.91-12.20) were more likely to have psychological distress. CONCLUSION: Transgender women were at risk of psychological distress. Earning more money can help transgender women's psychological health. When formulating measures for transgender women, we should pay attention to enhance social inclusion and social acceptance of their gender identity and sexual orientation. Strengthening the role of the internet in transgender women's confirmation of sexual orientation and improving the social acceptance of transgender women will have a positive impact on the psychological status of transgender women.

Massively Parallelized Molecular Force Manipulation with On-Demand Thermal and Optical Control.

In single-molecule force spectroscopy (SMFS), a tethered molecule is stretched using a specialized instrument to study how macromolecules extend under force. One problem in SMFS is the serial and slow nature of the measurements, performed one molecule at a time. To address this long-standing challenge, we report on the origami polymer force clamp (OPFC) which enables parallelized manipulation of the mechanical forces experienced by molecules without the need for dedicated SMFS instruments or surface tethering. The OPFC positions target molecules between a rigid nanoscale DNA origami beam and a responsive polymer particle that shrinks on demand. As a proof-of-concept, we record the steady state and time-resolved mechanical unfolding dynamics of DNA hairpins using the fluorescence signal from ensembles of molecules and confirm our conclusion using modeling.

Prevalence and associated factors of psychological distress in tuberculosis patients in Northeast China: a cross-sectional study.

BACKGROUND: Psychological distress, a major comorbidities of tuberculosis (TB) patients, has posed a serious threat to the progress being made in global TB programs by affecting treatment adherence and health outcomes. However, the magnitude and associated factors of psychological distress have not been fully studied in China. The aim of the current study was to assess the prevalence of psychological distress in TB patients and to further determine the effects of socio-demographic characteristics, health-related variables, substance use status, social support, and experienced stigma on psychological distress. METHODS: A cross-sectional survey was conducted among TB patients attending three medical institutions in Dalian, Liaoning Province, Northeast China from November 2020 to March 2021. A structured questionnaire was developed to collect data on patients' socio-demographic characteristics, health-related information, substance use status, psychological distress, family function, doctor-patient relationship, policy support, experienced stigma and so on. The binary logistics regression model was used to determine the associated factors of psychological distress. RESULTS: A total of 473 TB patients were enrolled in this study, and the prevalence of psychological distress was 64.1%. Binary logistic regression analysis revealed that patients with a middle school education level or above (OR: 0.521, 95%CI: 0.279-0.974), no adverse drug reactions (OR: 0.476, 95%CI: 0.268-0.846), and regular physical exercise (OR: 0.528, 95%CI: 0.281-0.993) were more likely to stay away from psychological distress. However, patients who had a high economic burden (OR: 1.697, 95%CI: 1.014-2.840), diabetes (OR: 2.165, 95%CI: 1.025-4.573), self-rated illness severe (OR: 3.169, 95%CI: 1.081-9.285), perceived poor resistance (OR: 2.065, 95%CI: 1.118-3.815), severe family dysfunction (OR: 4.001, 95%CI: 1.158-13.823), perceived need for strengthen psychological counseling (OR: 4.837, 95%CI: 2.833-8.258), and a high experienced stigma (OR: 3.253, 95%CI: 1.966-5.384) tended to have a psychological distress. CONCLUSIONS: The study found that the proportion of psychological distress among TB patients was high in Northeast China, and it was influenced by a variety of factors. Effective interventions to reduce psychological distress in TB patients urgently need to be developed, and greater attention should be given to patients with risk factors.

Mechanically Triggered Hybridization Chain Reaction.

Cells transmit piconewton forces to receptors to mediate processes such as migration and immune recognition. A major challenge in quantifying such forces is the sparsity of cell mechanical events. Accordingly, molecular tension is typically quantified with high resolution fluorescence microscopy, which hinders widespread adoption and application. Here, we report a mechanically triggered hybridization chain reaction (mechano-HCR) that allows chemical amplification of mechanical events. The amplification is triggered when a cell receptor mechanically denatures a duplex revealing a cryptic initiator to activate the HCR reaction in situ. Importantly, mechano-HCR enables direct readout of pN forces using a plate reader. We leverage this capability and measured mechano-IC50 for aspirin, Y-27632, and eptifibatide. Given that cell mechanical phenotypes are of clinical importance, mechano-HCR may offer a convenient route for drug discovery, personalized medicine, and disease diagnosis.

DNA-Based Microparticle Tension Sensors (µTS) for Measuring Cell Mechanics in Non-planar Geometries and for High-Throughput Quantification.

Mechanotransduction, the interplay between physical and chemical signaling, plays vital roles in many biological processes. The state-of-the-art techniques to quantify cell forces employ deformable polymer films or molecular probes tethered to glass substrates. However, the applications of these assays in fundamental and clinical research are restricted by the planar geometry and low throughput of microscopy readout. Herein, we develop a DNA-based microparticle tension sensor, which features a spherical surface and thus allows for investigation of mechanotransduction at curved interfaces. The micron-scale of µTS enables flow cytometry readout, which is rapid and high throughput. We applied the method to map and measure T-cell receptor forces and platelet integrin forces at 12 and 56 pN thresholds. Furthermore, we quantified the inhibition efficiency of two anti-platelet drugs providing a proof-of-concept demonstration of µTS to screen drugs that modulate cellular mechanics.

Tunable DNA Origami Motors Translocate Ballistically Over µm Distances at nm/s Speeds.

Inspired by biological motor proteins, that efficiently convert chemical fuel to unidirectional motion, there has been considerable interest in developing synthetic analogues. Among the synthetic motors created thus far, DNA motors that undertake discrete steps on RNA tracks have shown the greatest promise. Nonetheless, DNA nanomotors lack intrinsic directionality, are low speed and take a limited number of steps prior to stalling or dissociation. Herein, we report the first example of a highly tunable DNA origami motor that moves linearly over micron distances at an average speed of 40 nm/min. Importantly, nanomotors move unidirectionally without intervention through an external force field or a patterned track. Because DNA origami enables precise testing of nanoscale structure-function relationships, we were able to experimentally study the role of motor shape, chassis flexibility, leg distribution, and total number of legs in tuning performance. An anisotropic rigid chassis coupled with a high density of legs maximizes nanomotor speed and endurance.

Heteromultivalency enables enhanced detection of nucleic acid mutations.

Detecting genetic mutations such as single nucleotide polymorphisms (SNPs) is necessary to prescribe effective cancer therapies, perform genetic analyses and distinguish similar viral strains. Traditionally, SNP sensing uses short oligonucleotide probes that differentially bind the SNP and wild-type targets. However, DNA hybridization-based techniques require precise tuning of the probe's binding affinity to manage the inherent trade-off between specificity and sensitivity. As conventional hybridization offers limited control over binding affinity, here we generate heteromultivalent DNA-functionalized particles and demonstrate optimized hybridization specificity for targets containing one or two mutations. By investigating the role of oligo lengths, spacer lengths and binding orientation, we reveal that heteromultivalent hybridization enables fine-tuned specificity for a single SNP and dramatic enhancements in specificity for two non-proximal SNPs empowered by highly cooperative binding. Capitalizing on these abilities, we demonstrate straightforward discrimination between heterozygous cis and trans mutations and between different strains of the SARS-CoV-2 virus. Our findings indicate that heteromultivalent hybridization offers substantial improvements over conventional monovalent hybridization-based methods.

Quantifying T cell receptor mechanics at membrane junctions using DNA origami tension sensors.

The T cell receptor (TCR) is thought to be a mechanosensor, meaning that it transmits mechanical force to its antigen and leverages the force to amplify the specificity and magnitude of TCR signalling. Although a variety of molecular probes have been proposed to quantify TCR mechanics, these probes are immobilized on hard substrates, and thus fail to reveal fluid TCR-antigen interactions in the physiological context of cell membranes. Here we developed DNA origami tension sensors (DOTS) which bear force sensors on a DNA origami breadboard and allow mapping of TCR mechanotransduction at dynamic intermembrane junctions. We quantified the mechanical forces at fluid TCR-antigen bonds and observed their dependence on cell state, antigen mobility, antigen potency, antigen height and F-actin activity. The programmability of DOTS allows us to tether these to microparticles to mechanically screen antigens in high throughput using flow cytometry. Additionally, DOTS were anchored onto live B cells, allowing quantification of TCR mechanics at immune cell-cell junctions.

MA-MIL: Sampling point-level abnormal ECG location method via weakly supervised learning.

BACKGROUND AND OBJECTIVE: Current automatic electrocardiogram (ECG) diagnostic systems could provide classification outcomes but often lack explanations for these results. This limitation hampers their application in clinical diagnoses. Previous supervised learning could not highlight abnormal segmentation output accurately enough for clinical application without manual labeling of large ECG datasets. METHOD: In this study, we present a multi-instance learning framework called MA-MIL, which has designed a multi-layer and multi-instance structure that is aggregated step by step at different scales. We evaluated our method using the public MIT-BIH dataset and our private dataset. RESULTS: The results show that our model performed well in both ECG classification output and heartbeat level, sub-heartbeat level abnormal segment detection, with accuracy and F1 scores of 0.987 and 0.986 for ECG classification and 0.968 and 0.949 for heartbeat level abnormal detection, respectively. Compared to visualization methods, the IoU values of MA-MIL improved by at least 17 % and at most 31 % across all categories. CONCLUSIONS: MA-MIL could accurately locate the abnormal ECG segment, offering more trustworthy results for clinical application.

Nomograms incorporating hsa_circ_0029325 highly expressed in exosomes of hepatocellular carcinoma predict the postoperative outcomes.

BACKGROUND: Liquid biopsies, for example, exosomal circular RNA (circRNA) can be used to assess potential predictive markers for hepatocellular carcinoma (HCC) in patients after curative resection. This study aimed to search for effective prognostic biomarkers for HCC in patients after surgical resection based on exosomal circRNA expression profiles. We developed two nomograms incorporating circRNAs to predict the postoperative recurrence-free survival (RFS) and overall survival (OS) of HCC patients. METHOD: Plasma exosomes isolated from HCC patients and healthy individuals were used for circRNA microarray analysis to explore differentially expressed circRNAs. Pearson correlation analysis was used to evaluate the correlation between circRNAs and clinicopathological features. Cox regression analysis was used to explore the correlation between circRNA and postoperative survival time as well as recurrence time. A nomogram based on circRNA and clinicopathological characteristics was established and further evaluated to predict prognosis and recurrence. RESULT: Among 60 significantly upregulated circRNAs and 25 downregulated circRNAs, hsa_circ_0029325 was selected to verify its power for predicting HCC outcomes. The high expression level of exosomal hsa_circ_0029325 was significantly correlated with OS (P = 0.001, HR = 2.04, 95% CI 1.41-3.32) and RFS (P = 0.009, HR = 1.62, 95% CI 1.14-2.30). Among 273 HCC patients, multivariate regression analysis showed that hsa_circ_0029325 (HR = 1.96, 95% CI 1.21-3.18), tumor size (HR = 2.11, 95% CI 1.33-3.32), clinical staging (HR = 2.31, 95% CI 1.54-3.48), and tumor thrombus (HR = 1.74, 95% CI 1.12-2.7) were independent risk factors for poor prognosis in HCC patients after radical resection. These independent predictors of prognosis were incorporated into the two nomograms. The AUCs under the 1-year, 3-year, and 5-year survival and recurrence curves of the OS and RFS nomograms were 0.755, 0.749, and 0.742 and 0.702, 0.685, and 0.642, respectively. The C-index, calibration curves, and clinical decision curves showed that the two prediction models had good predictive performance. These results were verified in the validation cohort with 90 HCC patients. CONCLUSION: Our study established two reliable nomograms for predicting recurrence and prognosis in HCC patients. We also show that it is feasible to screen potential predictive markers for HCC after curative resection through exosomal circRNA expression profile analysis.

The immune response of hepatocellular carcinoma after locoregional and systemic therapies: The available combination option for immunotherapy.

Hepatocellular carcinoma (HCC) is associated with a highly heterogeneous immune environment that produces an immune response to various locoregional treatments (LRTs), which in turn affects the effectiveness of immunotherapy. Although LRTs still dominate HCC therapies, 50-60% of patients will ultimately be treated with systemic therapies and might receive those treatments for the rest of their life. TACE, SIRT, and thermal ablation can dramatically increase the immunosuppressive state of HCC, a condition that can be addressed by combination with immunotherapy to restore the activity of lymphocytes and the secretion of cellular immune factors. Immune treatment with locoregional and systemic treatments has dramatically changed the management of HCC. In this review, we examine the research on the changes in the immune microenvironment after locoregional or systemic treatment. We also summarize the regulation of various immune cells and immune factors in the tumor microenvironment and discuss the different infiltration degrees of immune cells and factors on the prognosis of HCC to better compare the efficacy between different treatment methods from the perspective of the tumor microenvironment. This information can be used to help develop treatment options for the upcoming new era of HCC treatment in the future.

Design of a high-performance NIR-II nanoprobe by steric regulation for in vivo vasculature and tumor imaging.

A novel NIR-II nanoprobe was developed through regulating the steric effect of an A-DA'D-A dye. The probe features the properties of strong fluorescence, high stability, and a large Stokes shift, thereby serving as a remarkable contrast agent for the fluorescence imaging of hindlimb vasculature and tumors in live mice.

Immune Regulatory Networks and Therapy of γδ T Cells in Liver Cancer: Recent Trends and Advancements.

The roles of γδ T cells in liver cancer, especially in the potential function of immunotherapy due to their direct cytotoxic effects on tumor cells and secretion of important cytokines and chemokines, have aroused research interest. This review briefly describes the basic characteristics of γδ T cells, focusing on their diverse effects on liver cancer. In particular, different subtypes of γδ T cells have diverse or even opposite effects on liver cancer. We provide a detailed description of the immune regulatory network of γδ T cells in liver cancer from two aspects: immune components and nonimmune components. The interactions between various components in this immune regulatory network are dynamic and pluralistic, ultimately determining the biological effects of γδ T cells in liver cancer. We also integrate the current knowledge of γδ T-cell immunotherapy for liver cancer treatment, emphasizing the potential of these cells in liver cancer immunotherapy.

Prevalence of intestinal colonization and nosocomial infection with carbapenem-resistant Enterobacteriales in children: a retrospective study.

Objective: We investigated the epidemiological surveillance of the intestinal colonization and nosocomial infection of carbapenem-resistant Enterobacteriales (CRE) isolates from inpatients, which can provide the basis for developing effective prevention. Methods: A total of 96 CRE strains were collected from 1,487 fecal samples of hospitalized children between January 2016 and June 2017, which were defined as the "CRE colonization" group. In total, 70 CRE clinical isolates were also randomly selected for the comparison analysis and defined as the "CRE infection" group. The antimicrobial susceptibility of all strains was determined by the microdilution broth method. Polymerase chain reaction (PCR) was used to analyze carbapenemase genes, plasmid typing, and integrons. Multilocus sequence typing was further used to determine clonal relatedness. Results: In the "CRE colonization" group, Klebsiella pneumoniae was mostly detected with a rate of 42.7% (41/96), followed by Escherichia coli (34.4%, 33/96) and Enterobacter cloacae (15.6%, 15/96). The ST11 KPC-2 producer, ST8 NDM-5 producer, and ST45 NDM-1 producer were commonly present in carbapenem-resistant K. pneumoniae (CRKPN), carbapenem-resistant E. coli (CRECO), and carbapenem-resistant E. cloacae (CRECL) isolates, respectively. In the "CRE infection" group, 70% (49/70) of strains were K. pneumoniae, with 21.4% E. cloacae (15/70) and 5.7% E. coli (4/70). The ST15 OXA-232 producer and ST48 NDM-5 producer were frequently observed in CRKPN isolates, while the majority of NDM-1-producing CRECL isolates were assigned as ST45. Phylogenetic analysis showed that partial CRE isolates from intestinal colonization and nosocomial infection were closely related, especially for ST11 KPC-2-producing CRKPN and ST45 NDM-1-producing CRECL. Furthermore, plasmid typing demonstrated that IncF and IncFIB were the most prevalent plasmids in KPC-2 producers, while IncX3/IncX2 and ColE were widely spread in NDM producer and OXA-232 producer, respectively. Then, class 1 integron intergrase intI1 was positive in 74.0% (71/96) of the "CRE colonization" group and 52.9% (37/70) of the "CRE infection" group. Conclusion: This study revealed that CRE strains from intestinal colonization and nosocomial infection showed a partial correlation in the prevalence of CRE, especially for ST11 KPC-2-producing CRKPN and ST45 NDM-1-producing CRECL. Therefore, before admission, long-term active screening of rectal colonization of CRE isolates should be emphasized.

DNA Origami Tension Sensors (DOTS) to study T cell receptor mechanics at membrane junctions.

The T cell receptor (TCR) is thought to be a mechanosensor, meaning that it transmits mechanical force to its antigen and leverages the force to amplify the specificity and magnitude of TCR signaling. The past decade has witnessed the development of molecular probes which have revealed many aspects of receptor mechanotransduction. However, most force probes are immobilized on hard substrates, thus failing to reveal mechanics in the physiological context of cell membranes. In this report, we developed DNA origami tension sensors (DOTS) which bear force sensors on a DNA origami breadboard and allow mapping of TCR mechanotransduction at dynamic intermembrane junctions. We demonstrate that TCR-antigen bonds experience 5-10 pN forces, and the mechanical events are dependent on cell state, antigen mobility, antigen potency, antigen height and F-actin activity. We tethered DOTS onto a microparticle to mechanically screen antigen in high throughput using flow cytometry. Finally, DOTS were anchored onto live B cell membranes thus producing the first quantification of TCR mechanics at authentic immune cell-cell junctions.

Detection of cellular traction forces via the force-triggered Cas12a-mediated catalytic cleavage of a fluorogenic reporter strand.

Molecular forces generated by cell receptors are infrequent and transient, and hence difficult to detect. Here we report an assay that leverages the CRISPR-associated protein 12a (Cas12a) to amplify the detection of cellular traction forces generated by as few as 50 adherent cells. The assay involves the immobilization of a DNA duplex modified with a ligand specific for a cell receptor. Traction forces of tens of piconewtons trigger the dehybridization of the duplex, exposing a cryptic Cas12-activating strand that sets off the indiscriminate Cas12-mediated cleavage of a fluorogenic reporter strand. We used the assay to perform hundreds of force measurements using human platelets from a single blood draw to extract individualized dose-response curves and half-maximal inhibitory concentrations for a panel of antiplatelet drugs. For seven patients who had undergone cardiopulmonary bypass, platelet dysfunction strongly correlated with the need for platelet transfusion to limit bleeding. The Cas12a-mediated detection of cellular traction forces may be used to assess cell state, and to screen for genes, cell-adhesion ligands, drugs or metabolites that modulate cell mechanics.

Unraveling the Emerging Niche Role of Hepatic Stellate Cell-derived Exosomes in Liver Diseases.

Hepatic stellate cells (HSCs) play an essential role in various liver diseases, and exosomes are critical mediators of intercellular communication in local and distant microenvironments. Cellular crosstalk between HSCs and surrounding multiple tissue-resident cells promotes or inhibits the activation of HSCs. Substantial evidence has revealed that HSC-derived exosomes are involved in the occurrence and development of liver diseases through the regulation of retinoid metabolism, lipid metabolism, glucose metabolism, protein metabolism, and mitochondrial metabolism. HSC-derived exosomes are underpinned by vehicle molecules, such as mRNAs and microRNAs, that function in, and significantly affect, the processes of various liver diseases, such as acute liver injury, alcoholic liver disease, nonalcoholic fatty liver disease, viral hepatitis, fibrosis, and cancer. As such, numerous exosomes derived from HSCs or HSC-associated exosomes have attracted attention because of their biological roles and translational applications as potential targets for therapeutic targets. Herein, we review the pathophysiological and metabolic processes associated with HSC-derived exosomes, their roles in various liver diseases and their potential clinical application.