Far-Red Aggregation-Induced Emission Hydrogel-Reinforced Tissue Clearing for 3D Vasculature Imaging of Whole Lung and Whole Tumor.

Understanding the vascular formation and distribution in metastatic lung tumors is a significant challenge due to autofluorescence, antibody/dye diffusion in dense tumor, and fluorophore stability when exposed to solvent-based clearing agents. Here, an approach is presented that redefines 3D vasculature imaging within metastatic tumor, peritumoral lung tissue, and normal lung. Specifically, a far-red aggregation-induced emission nanoparticle with surface amino groups (termed as TSCN nanoparticle, TSCNNP) is designed for in situ formation of hydrogel (TSCNNP@Gel) inside vasculatures to provide structural support and enhance the fluorescence in solvent-based tissue clearing method. Using this TSCNNP@Gel-reinforced tissue clearing imaging approach, the critical challenges are successfully overcome and comprehensive visualization of the whole pulmonary vasculature up to 2 µm resolution is enabled, including its detailed examination in metastatic tumors. Importantly, features of tumor-associated vasculature in 3D panoramic views are unveiled, providing the potential to determine tumor stages, predict tumor progression, and facilitate the histopathological diagnosis of various tumor types.

Dual-targeted fluorescent probe for tracking polarity and phase transition processes during lipophagy.

Eukaryotic cells regulate various cellular processes through membrane-bound and membrane-less organelles, enabling active signal communication and material exchange. Lysosomes and lipid droplets are representative organelles, contributing to cell lipophagy when their interaction and metabolism are disrupted. Our limited understanding of the interacting behaviours and physicochemical properties of different organelles during lipophagy hinders accurate diagnosis and treatment of related diseases. In this contribution, we report a fluorescent probe, PTZ, engineered for dual-targeting of lipid droplets and lysosomes. PTZ can track liquid-liquid phase separation and respond to polarity shifts through ratiometric fluorescence emission, elucidating the lipophagy process from the perspective of organelle behavior and physicochemical properties. Leveraging on the multifunctionality of PTZ, we have successfully tracked the polarity and dynamic changes of lysosomes and lipid droplets during lipophagy. Furthermore, an unknown homogeneous transition of lipid droplets and lysosomes was discovered, which provided a new perspective for understanding lipophagy processes. And this work is expected to serve as a reference for diagnosis and treatment of lipophagy-related diseases.

LIMPID: a versatile method for visualization of brain vascular networks.

Visualization of cerebrovascular networks is crucial for understanding the pathogenesis of many neurological diseases. Recently developed optical clearing techniques offer opportunities in deep tissue imaging, and have been successfully applied in many research studies. The development of nanotechnology enables the labeling of brain vessels with functionalized micro/nanoparticles embedded with fluorescent dyes. We herein report an efficient method, named LIMPID (Labeled and Interlinked Micro/nanoparticles for Imaging and Delipidation), specific for the precise fluorescence imaging of vascular networks in clearing-treated tissues. This robust vessel labeling technique replaces conventional fluorescence dyes with functionalized polymer micro/nanoparticles that are able to cross-link with polyacrylamide to form dense hydrogels in vessels. LIMPID shows high-robustness during the clearing process without sacrificing fluorescence signals and clearing performance. LIMPID enables three dimension (3D) visualization of elaborate vascular networks in mouse brains and is compatible with other fluorescence-labeling techniques. We have successfully applied this method to acquire cortical vasculature images simultaneously with the neurons or microglia, as well as to evaluate vascular damage in a mouse model of stroke. The LIMPID method provides a novel tool for the precise analysis of vascular dysfunction and vascular diseases.

Nonadditive Transport in Multi-Channel Single-Molecule Circuits.

As stated in the classic Kirchhoff's circuit laws, the total conductance of two parallel channels in an electronic circuit is the sum of the individual conductance. However, in molecular circuits, the quantum interference (QI) between the individual channels may lead to apparent invalidity of Kirchhoff's laws. Such an effect can be very significant in single-molecule circuits consisting of partially overlapped multiple transport channels. Herein, an investigation on how the molecular circuit conductance correlates to the individual channels is conducted in the presence of QI. It is found that the conductance of multi-channel circuit consisting of both constructive and destructive QI is significantly smaller than the addition of individual ones due to the interference between channels. In contrast, the circuit consisting of destructive QI channels exhibits an additive transport. These investigations provide a new cognition of transport mechanism and manipulation of transport in multi-channel molecular circuits.

NIR-emitting semiconducting polymer nanoparticles for in vivo two-photon vascular imaging.

Two-photon fluorescence (TPF) imaging holds great promise for real-time monitoring of cerebral ischemia-reperfusion injury, which is important for the clinical diagnosis of stroke. However, biocompatible and photostable NIR-emitting probes for TPF imaging of ischemic stroke are lacking. Herein, we report the first NIR-emitting TPF probe (named NESPN) prepared using semiconducting polymers for TPF imaging of cerebral ischemia. By virtue of its excellent biocompatibility with the nervous system and bright fluorescence NIR emission, NESPN enables the real-time imaging of mouse brain vasculature with micrometer-scale spatial resolution, realizing clear visualization of ultrafine capillaries (∼3.16 µm). Moreover, NESPN can be utilized in the dynamic monitoring of cerebral blood flow velocity. Microangiography using NESPN was successfully used to indicate the openness of the penumbra area in the mouse brain stroke model. More importantly, this technique allows us to continuously monitor the whole process of ischemic stroke and subsequent reperfusion. This work provides a new and versatile tool for vascular research and diagnosis of vascular diseases.