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A highly sensitive living probe derived from nanoparticle-remodeled neutrophils for precision tumor imaging diagnosis
摘要： Timely and precise diagnosis of malignant tumors is of great value to patients’ treatment and prognosis. Although nanotechnology-based molecular imaging represents a major advancement in the field of tumor imaging diagnosis, it is restricted by the rapid blood clearance of nanoparticles and the diverse physiological barriers in vivo; hence, its further application is greatly hindered. Cell carriers, ascribed to their natural biological properties, are thus gaining increasing attention for addressing such issues. Here, taking full advantage of the inflammation-homing capability of neutrophils, we constructed a highly sensitive cell probe in which reduced bovine-serum albumin (BSA) nanoparticles, loaded with imaging agents (gadolinium (Gd) and BODIPY), were covalently fixed onto the cellular surface by 5-thio-2-nitrobenzoate (TNB)-mediated fast and efficient disulfide–thiol exchange. Impressively, the remodeling process exerted a negligible effect on the neutrophils’ biological profiles with regard to cell viability, morphology, and cell-surface protein markers. Compared with nanoparticle-based imaging agents in a lung-cancer xenograft model, the living neutrophil probe demonstrated faster targeting and stronger accumulation in the tumor site, as revealed by fluorescence and magnetic-resonance (MR) imaging. These results indicate the great potential of neutrophil-based living probe for precision tumor-diagnosis applications.
关键词： Biomaterials,Neutrophils,Nanoparticles,Molecular imaging,Tumor imaging
更新于2025-09-16 10:30:52
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Automated high-throughput light-sheet fluorescence microscopy of larval zebrafish
摘要： Light sheet fluorescence microscopy enables fast, minimally phototoxic, three-dimensional imaging of live specimens, but is currently limited by low throughput and tedious sample preparation. Here, we describe an automated high-throughput light sheet fluorescence microscope in which specimens are positioned by and imaged within a fluidic system integrated with the sheet excitation and detection optics. We demonstrate the ability of the instrument to rapidly examine live specimens with minimal manual intervention by imaging fluorescent neutrophils over a nearly 0.3 mm3 volume in dozens of larval zebrafish. In addition to revealing considerable inter-individual variability in neutrophil number, known previously from labor-intensive methods, three-dimensional imaging allows assessment of the correlation between the bulk measure of total cellular fluorescence and the spatially resolved measure of actual neutrophil number per animal. We suggest that our simple experimental design should considerably expand the scope and impact of light sheet imaging in the life sciences.
关键词： neutrophils,light sheet fluorescence microscopy,automated imaging,high-throughput imaging,larval zebrafish
更新于2025-09-10 09:29:36
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Transforming Weakness into Strength: Photothermal‐Therapy‐Induced Inflammation Enhanced Cytopharmaceutical Chemotherapy as a Combination Anticancer Treatment
摘要： A new synergistic treatment that combines photothermal therapy (PTT) and inflammation-mediated active targeting (IMAT) chemotherapy based on cytopharmaceuticals is developed. During PTT, the photothermal tumor ablation is accompanied by an inflammatory effect and upregulation of inflammatory factors at the tumor site, which may accelerate tumor regeneration. Moreover, PTT-induced inflammation can also recruit neutrophils (NEs) to the tumor site. To convert the disadvantages of PTT-induced inflammation into strengths, NEs are investigated as cytopharmaceuticals for IMAT chemotherapy to further inhibit the tumor recurrence after PTT due to the chemotaxis of NEs to the inflammatory sites. In this study, PEGylated gold nanorods (PEG-GNRs) are explored as the photothermal agent and paclitaxel-loaded cytopharmaceuticals of NEs as the IMAT chemotherapeutic agent. PTT is conducted at 72 h postinjection of PEG-GNRs, followed by cytopharmaceuticals for IMAT chemotherapy. It is demonstrated that the cytopharmaceuticals effectively accumulate in the tumor sites after PTT, which leads to a significant enhancement of antitumor efficacy and a reduction in systemic toxicity. These studies suggest that PTT-induced inflammation further enhances the chemotherapy of cytopharmaceuticals, and the combination of PTT and IMAT chemotherapy may be a promising synergistic strategy for targeted cancer therapy.
关键词： combination cancer therapy,inflammation,photothermal therapy,cytopharmaceuticals,neutrophils
更新于2025-09-04 15:30:14
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A Flow Cytometry‐Based Assay for High‐Throughput Detection and Quantification of Neutrophil Extracellular Traps in Mixed Cell Populations
摘要： Neutrophil extracellular traps (NETs) are web-like structures composed of decondensed chromatin and antimicrobial proteins that are released into the extracellular space during microbial infections. This active cell death program is known as NETosis. To date, fluorescence microscopy is the widely accepted method for visualization and quantification of NETs. However, this method is subjective, time consuming and yields low numbers of analyzed polymorphonuclear cells (PMNs) per sample. Increasing interest has emerged on the identification of NETs using flow cytometry techniques. However, flow cytometry analysis of NETs requires particular precautions for sample preparation to obtain reproducible data. Herein, we describe a flow cytometry-based assay for high-throughput detection and quantification of NETosis in mixed cell populations. We used fluorescent-labeled antibodies against cell markers on PMNs together with a combination of nucleic acid stains to measure NETosis in whole blood (WB) and purified PMNs. Using plasma membrane-impermeable DNA-binding dye, SYTOX Orange (SO), we found that cell-appendant DNA of NETting PMNs were positive for SO and DAPI. The combination of optimally diluted antibody and nucleic acid dyes required no washing and yielded low background fluorescence. Significant correlations were found for NETosis from WB and purified PMNs. We then validated the assay by comparing with time-lapse live cell fluorescence microscopy and determined very good intraassay and interassay variances. The assay was then applied to a disease associated with NETosis, systemic lupus erythematosus (SLE). We examined PMA-induced NETosis in peripheral PMNs from SLE patients and controls and in bone marrow PMNs from multiple murine models. In summary, this assay is observer-independent and allows for rapid assessment of a large number of PMNs per sample. Use of this assay does not require sophisticated microscopic equipment like imaging flow cytometers and may be a starting point to analyze extracellular trap formation from immune cells other than PMNs.
关键词： neutrophils,flow cytometry,extracellular traps
更新于2025-09-04 15:30:14