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As a general mechanism for governing the bioactivity of membrane receptors, allosteric modulation is critical in cell signaling and cell communications but remains difficult to be measured in situ....

As a general mechanism, ligand-induced receptor clustering on cell membrane plays determinative roles in pattern recognition and transmembrane signaling. Nevertheless, probing the dynamic characteristics for the complicated interactions between receptor clusters remains difficult because of the lack of strategy for receptor cluster labeling and long-term monitoring in live cells. Herein, we proposed a data-mining-integrated plasmon coupling microscopy to study the dynamic cluster-cluster interactions on cell surface. The receptor clusters were activated and labeled with multivalent plasmonic nanoprobes, which enables the real-time monitoring of individual receptor clusters and the measurement of cluster-cluster interactions from the analysis of plasmonic coupling for the nanoprobe pairs beyond the diffraction limit. Using this method, we found that the protease-activated receptor 1 (PAR1) clusters would experience an initial contact and then form a weakly bound cluster-cluster complex, followed by cluster fusion to generate large-sized signaling complexes. The underlying state transitions for the cluster-cluster fusion process were uncovered using a data-mining technique named the K-means-based hidden Markov model with the scattering intensity of coupled nanoprobe pairs as observations. All of the findings from single-particle analysis and bulk measurements suggested that the allosteric inhibitors could suppress the dynamic transitions from the weakly bound cluster-cluster complexes to fused signaling complexes, leading to the subsequent downregulation of intracellular calcium signaling pathways. We believe that this strategy is promising for imaging and monitoring receptor clustering as well as protein phase separation on the cell surface in various biological and physiological processes.

Phagosome maturation in macrophage is essential to the clearance of pathogenic materials in host defence but the dynamic features remain difficult to be measured in real time. Herein, we reported the multilayered Au@MnOx @SiO2 nanoparticle as a robust pH-sensitive plasmonic nanosensor for monitoring the dynamic acidification features over the phagosome maturation process in macrophage under darkfield microscopy. For this multilayered nanosensor, the gold nanoparticle core plays a role of signal reporter, the MnOx shell and the outmost SiO2 act as the sensing layer and the protecting layer, respectively. After subject to the acidic buffer solution, the MnOx layer in the multilayered nanoprobe could be decomposed rapidly, resulting in a remarkable spectral shift and color change under darkfield microscopy. We demonstrated this nanosensor for the investigation of single phagosome acidification dynamics by monitoring the color changes of nanoprobes after phagocytosis over time. The nanoprobes after phagocytosized in macrophage displayed a slight color change within the first hour and then cost several minutes to change from red to green in the next stage, indicating the phagosome undergoes a slow first and then fast acidification feature as well as a slow-to-fast acidification translation over the phagosome maturation process. Moreover, we validated that the slow-to-fast acidification translation was dependent on the activation of V-ATPase from the ATP depletion assay. We believed that this nanosensor is promising for studying the dynamic acidification features as well as disorders in phagosome maturation in phagocytic cells, which might provide valuable information for understanding the disease pathogenesis related to phagosome dysfunctions.

Mechanical signal transduction is fundamental for maintaining and regulating cellular processes and functions. Here, we proposed a novel near-infrared (NIR) light-responsive optomechanical actuator for the directional regulation of collective cell adhesion and migration. This optomechanical actuator that is made up of a thermal-responsive copolymer hydrogel and gold nanorods (AuNRs), enables non-invasive activation by NIR light stimulation. The activation of the optomechanical actuator leads to hydrogel contraction and an increase in Young's modulus, which could be used for applying contraction force to cells cultured on the surface of the hydrogel actuator. By grafting cell adhesive peptide ligands, the cells could attach onto the surface of the actuator and displayed a NIR light illumination intensity dependent migration rate along a random orientation. To achieve the controllable modulation of cell behaviors, we employed a microcontact printing strategy for patterned presentation of adhesive ligands on this actuator and achieved directional cell alignment and cell migration through optomechanical actuation. These demonstrations suggest that this robust optomechanical actuator is promising for the optical modulation of cellular events and cell functions in various bioapplications.

Acute pneumonia can greatly increase the vulnerable risk of atherosclerotic plaque and contribute to the mortality of cardiovascular disease. To accurately assess the rupture risk caused by acute pneumonia, we developed a novel kind of ratiometric semiconducting polymer nanoparticle (RSPN) for photoacoustic imaging of vulnerable plaque in apolipoprotein E-deficient mice complicated with pneumonia. Specifically, RSPN can react with O2•- and exhibit the enhanced photoacoustic signals at about 690 nm, while 800 nm is regarded as an internal photoacoustic reference. As a result, RSPN can provide reliable determination of O2•- within aortic atherosclerosis by analyzing the ratios of photoacoustic signals, which can successfully reflect the oxidative stress level in vulnerable plaque. Therefore, RSPN enable to specifically distinguish plaque-bearing mice and plaque-bearing mice complicated with pneumonia from healthy mice, which provides a promising tool to predict the vulnerability of plaque for reducing the mortality of atherosclerotic-induced cardiovascular disease.

Phagosome maturation in macrophage is essential to the clearance of pathogenic materials in host defence but the dynamic features remain difficult to be measured in real time. Herein, we reported the multilayered Au@MnO

Here, we introduced a single-particle mobility analysis-based ratiometric strategy for quantitative detection of disease-related biomarkers using antibody-conjugated gold nanoparticles (AuNPs) as probes under darkfield tracking microscopy (DFTM). On the basis of the capability of discriminating nanoparticles with different hydrodynamic sizes and detecting the changes in hydrodynamic effect, single-particle mobility analysis enables us to determine the amount of aggregated and monodispersed nanoprobes for the sandwich-like immunoassay strategy, making it possible to quantify the biotargets by analyzing the relative changes in the aggregate-to-monomer ratio of nanoprobes. By using capture antibody and detection antibody conjugated AuNPs as nanoprobes, we demonstrated ratiometric detection of carcinoembryonic antigen (CEA) over a linear dynamic range from 50 to 750 pM, which is acceptable for clinical diagnostic analysis of CEA in tumor patients. This ratiometric detection technique exhibited excellent anti-interference ability in the presence of nonspecific proteins or complicated protein mixtures. It can be anticipated that this robust technique is promising for the accurate detection of disease biomarkers and other biomolecules for biochemical and diagnostic applications.