Multiplexed Organelles Portrait Barcodes for Subcellular MicroRNA Array Detection in Living Cells

Multiplexed profiling of microRNAs’ subcellular expression and distribution is essential to understand their spatiotemporal function information, but it remains a crucial challenge. Herein, we report an encoding approach that leverages combinational fluorescent dye barcodes, organelle targeting elements, and an independent quantification signal, termed Multiplexed Organelles Portrait Barcodes (MOPB), for high-throughput profiling of miRNAs from organelles. The MOPB barcodes consist of heterochromatic fluorescent dye-loaded shell–core mesoporous silica nanoparticles modified with organelle targeting peptides and molecular beacon detection probes. Using mitochondria and endoplasmic reticulum as models, we encoded four Cy3/AMCA ER-MOPB and four Cy5/AMCA Mito-MOPB by varying the Cy3 and Cy5 intensity for distinguishing eight organelles’ miRNAs. Significantly, the MOPB strategy successfully and accurately profiled eight subcellular organelle miRNAs’ alterations in the drug-induced Ca2+ homeostasis breakdown. The approach should allow more widespread application of subcellular miRNAs and multiplexed subcellular protein biomarkers’ monitoring for drug discovery, cellular metabolism, signaling transduction, and gene expression regulation readout

Aptamer-Conjugated Graphene Quantum Dots/Porphyrin Derivative Theranostic Agent for Intracellular Cancer-Related MicroRNA Detection and Fluorescence-Guided Photothermal/Photodynamic Synergetic Therapy

Multifunctional theranostic platform coupling diagnostic and therapeutic functions holds great promise for personalized nanomedicine. Nevertheless, integrating consistently high performance in one single agent is still challenging. This work synthesized a sort of porphyrin derivatives (P) with high singlet oxygen generation ability and graphene quantum dots (GQDs) possessing good fluorescence properties. The P was conjugated to polyethylene glycol (PEG)­ylated and aptamer-functionalized GQDs to gain a multifunctional theranostic agent (GQD-PEG-P). The resulting GQD-PEG-P displayed good physiological stability, excellent biocompatibility and low cytotoxicity. The intrinsic fluorescence of the GQDs could be used to discriminate cancer cells from somatic cells, whereas the large surface facilitated gene delivery for intracellular cancer-related microRNA (miRNA) detection. Importantly, it displayed a photothermal conversion efficiency of 28.58% and a high quantum yield of singlet oxygen generation up to 1.08, which enabled it to accomplish advanced photothermal therapy (PTT) and efficient photodynamic therapy (PDT) for cancer treatment. The combined PTT/PDT synergic therapy led to an outstanding therapeutic efficiency for cancer cell treatment

Fabricating Pt/Sn–In₂O₃ Nanoflower with Advanced Oxygen Reduction Reaction Performance for High-Sensitivity MicroRNA Electrochemical Detection

Herein, an efficient electrochemical tracer with advanced oxygen reduction reaction (ORR) performance was designed by controllably decorating platinum (Pt) (diameter, 1 nm) on the surface of compositionally tunable tin-doped indium oxide nanoparticle (Sn–In₂O₃) (diameter, 25 nm), and using the Pt/Sn–In₂O₃ as electrochemical tracer and interfacial term hairpin capture probe, a facile and ultrasensitive microRNA (miRNA) detection strategy was developed. The morphology and composition of the generated Pt/Sn–In₂O₃ NPs were comprehensively characterized by spectroscopic and microscopic measurements, indicating numerous Pt uniformly anchored on the surface of Sn–In₂O₃. The interaction between Pt and surface Sn as well as high Pt(111) exposure resulted in the excellent electrochemical catalytic ability and stability of the Pt/Sn–In₂O₃ ORR. As proof-of-principle, using streptavidin (SA) functionalized Pt/Sn–In₂O₃ (SA/Pt/Sn–In₂O₃) as electrochemical tracer to amplify the detectable signal and a interfacial term hairpin probe for target capture probe, a miRNA biosensor with a linear range from 5 pM to 0.5 fM and limit of detection (LOD) down to 1.92 fM was developed. Meanwhile, the inherent selectivity of the term hairpin capture probe endowed the biosensor with good base discrimination ability. The good feasibility for real sample detection was also demonstrated. The work paves a new avenue to fabricate and design high-effective electrocatalytic tracer, which have great promise in new bioanalytical applications

Intelligent MnO₂/Cu_2–<i>x</i>S for Multimode Imaging Diagnostic and Advanced Single-Laser Irradiated Photothermal/Photodynamic Therapy

Lately, photothermal therapy (PTT) and photodynamic therapy (PDT) dual-modal therapy has attracted much attention in cancer therapy as a synergistic therapeutic model. However, the integration of PDT and PTT in a single nanoagent for cancer therapy is still a challenging task. Herein, an intelligent MnO₂/Cu_2–<i>x</i>S-siRNA nanoagent simultaneously overcoming inherent limitations of PDT and PTT with remarkable PTT&PDT therapeutic efficiency enabling a multimode accurate tumor imaging diagnostic is designed. We first develop a general method to decorate Cu_2–<i>x</i>S on the surface of MnO₂ nanosheet (MnO₂/Cu_2–<i>x</i>S); then, it is loaded with heat shock protein (HSP) 70 siRNA to obtain MnO₂/Cu_2–<i>x</i>S-siRNA. The intracellular microRNA (miRNA) imaging can be realized by loading miRNA detection probes. In the tumor acidic microenvironment, the MnO₂ is reduced to Mn²⁺ ion and triggers the decomposition of H₂O₂ into O₂ to relieve tumor hypoxia. The reduced Mn²⁺ ions significantly enhance magnetic resonance imaging (MRI) contrast, and the Cu_2–<i>x</i>S acts as a powerful photoacoustic (PA) and photothermal (PT) imaging agent, leading to trimodal accurate tumor-specific imaging and detection. Under a single NIR laser irradiation, the nanosystem exhibits superiority of PTT&PDT efficiency owing to siRNA-mediated blocked heat-shock response and MnO₂-related relieved tumor hypoxia. This work highlights the great promise of modulating the tumor cellular defense mechanism and microenvironment with intelligent multifunctional nanoagents to achieve a comprehensive fighting cancer effect

Charge-Reversal NaCl/G-Quartets for Aggregation-Induced Mitochondrial MicroRNA Imaging and Ion-Interference Therapy

Mitochondrial therapy is a promising new strategy that offers the potential to achieve precise disease diagnosis or maximum therapeutic response. However, versatile mitochondrial theranostic platforms that integrate biomarker detection and therapy have rarely been exploited. Here, we report a charge-reversal nanomedicine activated by an acidic microenvironment for mitochondrial microRNA (mitomiR) detection and ion-interference therapy. The transporter liposome (DD-DC) was constructed from a pH-responsive polymer and a positively charged phospholipid, encapsulating NaCl nanoparticles with coloading of the aggregation-induced emission (AIE) fluorogens AIEgen-DNA/G-quadruplexes precursor and brequinar (NAB@DD-DC). The negatively charged nanomedicine ensured good blood stability and high tumor accumulation, while the charge-reversal to positive in response to the acidic pH in the tumor microenvironment (TME) and lysosomes enhanced the uptake by tumor cells and lysosome escape, achieving accumulation in mitochondria. The subsequently released Na+ in mitochondria not only contributed to the formation of mitomiR-494 induced G-quadruplexes for AIE imaging diagnosis but also led to an osmolarity surge that was enhanced by brequinar to achieve effective ion-interference therapy

Engineered Exosome-Mediated Near-Infrared-II Region V₂C Quantum Dot Delivery for Nucleus-Target Low-Temperature Photothermal Therapy

The limited penetration depth of photothermal agents (PTAs) active in the NIR-I biowindow and the thermoresistance caused by heat shock protein (HSP) significantly limit the therapeutic efficiency of photothermal therapy (PTT). To address the problem, we introduce a strategy of low-temperature nucleus-targeted PTT in the NIR-II region achieving effective tumor killing by combining the vanadium carbide quantum dots (V2C QDs) PTA and an engineered exosomes (Ex) vector. The small fluorescent V2C QDs with good photothermal effect in the NIR-II region were modified with TAT peptides and packaged into Ex with RGD modification (V2C-TAT@Ex-RGD). The resulting nanoparticles (NPs) exhibited good biocompatibility, long circulation time, and endosomal escape ability, and they could target the cell and enter into the nucleus to realize low-temperature PTT with advanced tumor destruction efficiency. The fluorescent imaging, photoacoustic imaging (PAI), and magnetic resonance imaging (MRI) capability of the NPs were also revealed. The low-temperature nucleus-targeted PTT in the NIR-II region provides more possibilities toward successful clinical application of PTT

Mitochondria MicroRNA Spatial Imaging via pH-Responsive Exonuclease-Assisted AIE Nanoreporter

Mitochondrial microRNAs (mitomiRs) critically orchestrate mitochondrial functions. Spatial imaging of mitomiRs is essential to understand its clinical value in diagnosis and prognosis. However, the direct monitoring of mitomiRs in living cells remains a key challenge. Herein, we report an AIE nanoreporter strategy for mitomiRs imaging in living cells through pH-controlled exonuclease (Exo)-assisted target cycle signal amplification. The AIE-labeled DNA detection probes are conjugated on Exo III encapsulated polymeric nanoparticles (NPs) via consecutive adenines (polyA). The amplified sensing functions are off during the cytoplasm delivery process, and it can be spatially switched from off to on when in the alkaline mitochondria (about pH 8) after triphenylphosphonium (TPP)-mediated mitochondrial targeting. Where the NPs degraded to release Exo III and cancer-specific mitomiRs hybridize with AIE-labeled DNA detection probes to expose the cleavage site of released Exo III, enabling spatially restricted mitomiRs imaging. The mitomiRs expression fluctuation was also realized. This study contributes to a facile strategy that could easily extend to a broad application for the understanding of mitomiRs-related pathological processes

Purely Chemical Approach for Preparation of d‑α-Amino Acids via (<i>S</i>)‑to‑(<i>R</i>)‑Interconversion of Unprotected Tailor-Made α‑Amino Acids

Unnatural (<i>R</i>)-α-amino acids (α-AAs) are in growing demand in the biomedical research and pharmaceutical industries. In this work, we present development of a purely chemical approach for preparation of (<i>R</i>)-α-AAs via (<i>S</i>)-to-(<i>R</i>)-interconversion of natural and tailor-made (<i>S</i>)-α-AAs. The method can be used on free, unprotected α-AAs and features a remarkable structural generality including substrates bearing tertiary alkyl chains and reactive functional groups. These attractive characteristics, combined with simplicity of reaction conditions and virtually complete stereochemical outcome, constitute a true methodological advance in this area, rivaling previously reported chemical and biocatalytic approaches

Self-Propelled Janus Mesoporous Micromotor for Enhanced MicroRNA Capture and Amplified Detection in Complex Biological Samples

The slow mass transport of the target molecule essentially limits the biosensing performance. Here, we report a Janus mesoporous microsphere/Pt-based (meso-MS/Pt) nanostructure with greatly enhanced target transport and accelerated recognition process for microRNA (miRNA) amplified detection in complex biological samples. The mesoporous MS was synthesized via double emulsion interfacial polymerization, and Pt nanoparticles (PtNPs) were deposited on the half-MS surface to construct Janus meso-MS/Pt micromotor. The heterogeneous meso-MS/Pt with a large surface available was attached to an entropy-driven DNA recognition system, termed meso-MS/Pt/DNA, and the tremendous pores network was beneficial to enhanced receptor–target interaction. It enabled moving around complex biological samples to greatly enhance target miRNA mass transport and accelerate recognition procedure due to the self-diffusiophoretic propulsion. Coupling with the entropy-driven signal amplification, extremely sensitive miRNA detection in Dulbecco’s modified Eagle medium (DMEM), and cell lysate without preparatory and washing steps was realized. Given the free preparatory and washing steps, fast mass transport, and amplified capability, the meso-MS/Pt/DNA micromotor provides a promising method for miRNAs analysis in real biological samples