A single mitochondria-targetable fluorescent probe for visualizing cysteine and glutathione in ferroptosis of myocardial ischemia/reperfusion injury.

Ferroptosis plays an important role in the early stage of myocardial ischemia/reperfusion (MI/R) injury, which is closely associated with the antioxidant damage of mitochondrial cysteine (Cys)/glutathione (GSH)/glutathione peroxidase 4 (GPX4) axis. Visualization of Cys and GSH in mitochondria is meaningful to value ferroptosis and further contributes to understanding and preventing MI/R injury. Herein a mitochondria-targetable thiols fluorescent probe (MTTP) was designed and synthesized based on sulfonyl benzoxadiazole (SBD) chromophore with a triphenylphosphine unit as the mitochondria-targeted functional group. Cys and GSH can be differentiated by MTTP with two distinguishable emission bands (583 nm and 520 nm) through the controllable aromatic substitution-rearrangement reaction. Importantly, MTTP is capable of monitoring ferroptosis and its inhibition by measuring mitochondrial Cys and GSH. MTTP was also employed to non-invasively detect ferroptosis during oxygen and glucose deprivation/reoxygenation (OGD/R)-induced MI/R injury in H9C2 cells. In a word, MTTP provides a visual tool that can simultaneously detect Cys and GSH to monitor ferroptosis processes during MI/R injury, which helps for more deeper understanding of the role of ferroptosis in MI/R injury-related diseases.

Engineering a near-infrared LAP fluorescent probe with high sensitivity and selectivity for surgical resection of liver cancer.

Hepatocellular carcinoma (HCC) is a type of cancer associated with a high rate of mortality and morbidity. In order to achieve precise HCC theranostics, it is important to develop excellent fluorescent probes. However, the existing probes are not sensitive or specific enough to accurately identify HCC margins and contours. For diagnosing HCC and identifying tumors during surgery, it is urgent to engineer highly sensitive and selective fluorescent probes. Liver tumor progression is closely associated with leucine aminopeptidase (LAP) overexpression, a biomarker of liver cancer. Herein, we have rationally designed a NIR fluorescent probe, NLAP, which is specially activated by LAP. The probe exhibited high sensitivity (detection limit = 6.8 mU L-1) and superior affinity (Km = 2.98 µM) for LAP. With this probe, we distinguished cancer cells overexpressing LAP from normal cells and applied it intraoperatively to guide liver tumor excisions. Furthermore, NLAP was employed to successfully detect the LAP of intestinal and splenic metastatic tumors in orthotopic liver tumor mice by "in situ spraying" and good performances were demonstrated.

Development of a Golgi-targeted superoxide anion fluorescent probe for elucidating protein GOLPH3 function in myocardial ischemia-reperfusion injury.

Superoxide anion (O2•-) is an important reactive oxygen species (ROS) and participates in various physiological and pathological processes in the organism. The O2•- burst induced by ischemia-reperfusion (I/R) is associated with cardiovascular disease and promotes the cell apoptosis. In this work, a turn-on type Golgi-targeting fluorescent probe Gol-Cou-O2•- was rationally designed for sensitive and selective detection of O2•-. The minimum detection limit concentration for O2•- was about 3.9 × 10-7 M in aqueous solution. Gol-Cou-O2•- showed excellent capacity of detecting exogenous and endogenous O2•- in living cells and zebrafish, and was also used to capture the up-regulated O2•- level during the duration of I/R process in cardiomyocytes. Golgi Phosphoprotein 3 (GOLPH3) is a potential Golgi stress marker protein and plays a key role in cells apoptosis during I/R. The fluorescence imaging and flow cytometry assay results indicated that silencing GOLPH3 through siRNA could give rise to the down-regulated O2•- level and alleviation of apoptosis in I/R myocardial cells. Thus, development of Gol-Cou-O2•- provides a diagnostic tool for myocardial oxidative stress injury and distinct insights on roles of GOLPH3 in myocardial I/R injury.

Development of a Golgi-targeted fluorescent chemosensor for detecting ferrous ions overload under Golgi stress.

Ferrous ion (Fe2+) is a crucial metal ion in the body and participates in the diseases related to oxidation and reduction. Golgi apparatus is the main subcellular organelle of Fe2+ transport in cells, and the stability of its structure is related to the Fe2+ at an appropriate concentration. In this work, a turn-on type Golgi-targeting fluorescent chemosensor Gol-Cou-Fe2+ was rationally designed for sensitive and selective detection of Fe2+. Gol-Cou-Fe2+ showed excellent capacity of detecting exogenous and endogenous Fe2+ in HUVEC and HepG2 cells. It was used to capture the up-regulated Fe2+ level during the hypoxia. Moreover, the fluorescence of sensor was enhanced over time under Golgi stress combining with the reduce of Golgi matrix protein GM130. However, elimination of Fe2+ or addition of nitric oxide (NO) would restore the fluorescence intensity of Gol-Cou-Fe2+ and the expression of GM130 in HUVEC. Thus, development of chemosensor Gol-Cou-Fe2+ provides a new window for tracking Golgi Fe2+ and elucidating Golgi stress-related diseases.

Construction of HClO activated near-infrared fluorescent probe for imaging hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common malignancies in the liver with poor prognosis. In order to improve the prognosis and overall survival of patients with HCC, it is important to identify it at early stage and resect it precisely. Cell microenvironment, active compounds, and enzymes may change during the cancerization of hepatocytes. Hypochlorous acid (HClO), one of the most significant signal molecules in the cellular signaling pathway, plays an important role in many cellular processes. To detect and treat liver cancers, it is imperative to study how HClO levels change in hepatocytes. However, developing fluorescent probes specific to liver cells to detect HClO still challenging. Herein, we designed and synthesized a NIR hepatocyte-specific fluorescent probe (MBH-MT) that displayed excellent optical properties for detecting HClO in biological samples. Cell imaging experiment conducted with the unique probe MBH-MT, showed that the biocompatible sensor is capable of monitoring HClO and distinguishing normal cells from cancer cells (e.g., HepG2, HUVEC, RAW264.7, L02 and HK-2 cells). An organ imaging experiment with the probe MBH-MT demonstrated its effectiveness in diagnosing and imaging hepatocellular carcinoma in vivo. MBH-MT's in situ imaging also demonstrated that it can target and image mouse hepatocellular carcinomas. Furthermore, MBH-MT has also successfully been used to diagnose and guide liver cancer surgery early. In the future, we expect that this powerful tool may be help in the detection and imaging of hepatocellular carcinoma, which may affect a large number of people.

Cysteine-Activatable Near-Infrared Fluorescent Probe for Dual-Channel Tracking Lipid Droplets and Mitochondria in Epilepsy.

Dual-channel fluorescent probes could respond to a specific target and emit different wavelengths of fluorescence before and after the response. Such probes could alleviate the influence caused by the variation of the probe concentration, excitation intensity, and so on. However, for most dual-channel fluorescent probes, the probe and fluorophore faced spectral overlap, which reduced sensitivity and accuracy. Herein, we introduced a cysteine (Cys)-responsive and near-infrared (NIR) emissive AIEgen (named TSQC) with good biocompatibility to dual-channel monitor Cys in mitochondria and lipid droplets (LDs) during cell apoptosis through wash-free fluorescence bio-imaging. TSQC can label mitochondria with bright fluorescence around 750 nm, and after reacting with Cys, the reaction product TSQ could spontaneously target LDs with emissions around 650 nm. Such spatially separated dual-channel fluorescence responses could significantly improve detection sensitivity and accuracy. Furthermore, the Cys-triggered dual-channel fluorescence imaging in LDs and mitochondria during apoptosis induced by UV light exposure, H2O2, or LPS treatment is clearly observed for the first time. Besides, we also report here that TSQC can be used to image subcellular Cys in different cell lines by measuring the fluorescence intensities of different emission channels. In particular, TSQC shows superior utility for the in vivo imaging of apoptosis in acute and chronic epilepsy mice. In brief, the newly designed NIR AIEgen TSQC can respond to Cys and separate two fluorescence signals to mitochondria and LDs, respectively, to study Cys-related apoptosis.

Advances in Multifunctional Chemotherapeutic Prodrugs for Near-infrared Fluorescence Imaging-guided Therapy.

Conventional chemotherapy (CT) is associated with severe side effects and inducible resistance, making it difficult to meet clinical requirements, forcing the development of new multifunctional prodrugs for precision medicine. In recent decades, researchers and clinicians have focused on developing of multifunctional chemotherapeutic prodrugs with tumor-targeting capability, activatable and traceable chemotherapeutic activity, as a powerful tool to improve theranostic outcomes in cancer treatment. The conjugates of near-infrared (NIR) organic fluorophores and chemotherapy reagents create an exciting avenue for real-time monitoring of drug delivery and distribution, as well as the combination of chemotherapy and photodynamic therapy (PDT). Therefore, there are great opportunities for researchers to conceive and exploit multifunctional prodrugs that can visualize chemo-drugs release and tumor treatment in vivo. In this review, the design strategy and the recent progress of multifunctional organic chemotherapeutic prodrugs for activating NIR fluorescence imaging-guided therapy are described and discussed in detail. Finally, the prospects and challenges of multifunctional chemotherapeutic prodrugs for NIR fluorescence imaging-guided therapy are provided.

A mitochondrial-targeted near-infrared fluorescent probe for visualizing the fluctuation of hypochlorite acid in idiopathic pulmonary fibrosis mice.

Idiopathic pulmonary fibrosis (IPF) is a chronic inflammatory disease destroying lungs irreversibly with high mortality rates. There are challenges in diagnosing IPF and treating it at an early stage. Mounting evidence suggests that hypochlorous acid (HClO) can help in diagnosing inflammation and relevant conditions. Pulmonary fibrosis is linked to the mitochondrial oxidative stress where excessive HClO production is a key molecular mechanism. Measuring mitochondrial HClO levels assists in the investigations of how the mitochondrial oxidative stress affects IPF. Herein, NIR-PTZ-HClO was developed and optimized as a probe for detecting fluctuations in HClO concentrations of cells and mice models through near-infrared (NIR) fluorescence. The probe featured large Stokes shift of 150 nm, NIR turn-on signal at 650 nm, high sensitivity (45-fold) and quick HClO detection (2 s). The probe is selective for HClO in the presence of range of other analytes. NIR-PTZ-HClO visualized both endogenous and exogenous HClO in living cells (RAW264.7, H460 and A549). The probe monitored HClO in mice models with IPF and moreover the HClO profile could be tracked during the IPF process. The probe also detected precipitous decrease in HClO levels in IPF mice treated with OFEV. NIR-PTZ-HClO probe has thus the potential for earlier diagnosis of lung fibrosis, thereby improving the treatment efficacy.

Introduction to the Meta-analysis method of functional magnetic resonance imaging research and the exploration of its application in the field of acupuncture. / åŠŸèƒ½ç£å ±æŒ¯ç ”ç©¶å ƒåˆ†æžæ–¹æ³•ä»‹ç»åŠå ¶åœ¨é’ˆåˆºé¢†åŸŸçš„åº”ç”¨æŽ¢æž.

In recent years, the number of functional magnetic resonance imaging (fMRI) research in acupuncture grows increasingly. However, due to the differences in acupoint selection, acupuncture technique and sample size, the problems get more prominent in terms of the diverse results and the lack of common rules of acupuncture among researches. By taking the fMRI research for post-stroke motor dysfunction (PSMD) treated with acupuncture as the example, this paper introduces the fMRI Meta-analysis technology for integrating the relevant research results and extracting the common rules, namely image-based Meta-analysis (IBMA) and coordinate-based Meta-analysis (CBMA). Considering the higher feasibility of CBMA, three available CBMA methods are explained specially, including activation likelihood estimation (ALE), kernel density analysis (KDA), and seed-based d mapping (SDM). Focusing on the precautions and operation procedure of CBMA, the review is conducted systematically on the type of fMRI research, task design, analytical method, and the thinking integrity of fMRI Meta-analysis, and the review findings are collated in charts. It aims to assist readers to understand the abstract and complex theories and practical information of this technology efficiently, conveniently and systematically, and hopes to provide the references for the future learning and the application.

Golgi-Targeting Fluorescent Probe for Monitoring CO-Releasing Molecule-3 In Vitro and In Vivo.

CO-releasing molecule-3 (CORM-3), mainly metal carbonyl compounds, is widely used as experimental tools to deliver CO, a biological "gasotransmitter", in mammalian systems. CORM-3 is also proposed as a potential new antimicrobial agent, which kills bacteria effectively and rapidly in vitro and in animal models. Organelle-targeting therapy, as a highly effective therapeutic strategy with little toxic and side effects, has important research significance and development prospects. Therefore, the development of effective methods for detecting and tracking CORM-3 at the subcellular level has important implications. In this paper, an easily available Golgi-targetable fluorescent probe (Golgi-Nap-CORM-3) was proposed for CORM-3 detection. In the probe Golgi-Nap-CORM-3, the phenyl sulfonamide group was selected as the Golgi-targetable unit, naphthalimide dye was chosen as a fluorophore, and the nitro group was selected as a CORM-3-responsive unit. Golgi-Nap-CORM-3 shows a CORM-3-reponsive increase of fluorescence emission at 520 nm. Using the excellent probe, the change of CORM-3 in HeLa cells, HepG2 cells, and zebrafish is successfully monitored. This study demonstrates very important information for the study of CORM-3 in vivo systems.

A NIR-emissive probe with a remarkable Stokes shift for CO-releasing molecule-3 detection in cells and in vivo.

Carbon monoxide (CO) is regarded as one of the most important gaseous transmitters, playing a vital role in biological systems; meanwhile, abnormal levels of CO can be correlated with conditions such as lung disease, Alzheimer's disease, and cardiovascular disease. CO-releasing molecules (CORMs) are chemical agents used to release CO as an endogenous, biologically active molecule in order to treat diseases. CO-releasing molecule-3 (CORM-3), as a convenient and safe CO donor and therapeutic drug molecule, has been widely used to release exogenous CO in living cells to study the physiological and pathological roles of CO in living systems. Herein, we designed a NIR-emitting probe (NIR-CORM-3) with a large Stokes shift based on a 4-(dimethylamino)cinnamaldehyde lepidine derived fluorophore. A 4-nitrobenzyl group was selected as the CORM-3 recognizing moiety, and the probe is able to selectively and sensitively respond to CORM-3 (within only 15 min). Upon encountering CORM-3, NIR-CORM-3 releases a fluorophore with a response at 670 nm, and it shows a remarkable Stokes shift (up to 250 nm). In addition, NIR-CORM-3 has low cytotoxicity and exhibits outstanding NIR imaging abilities in living cells and mice.

Recent Progresses in NIR-I/II Fluorescence Imaging for Surgical Navigation.

Cancer is still one of the main causes of morbidity and death rate around the world, although diagnostic and therapeutic technologies are used to advance human disease treatment. Currently, surgical resection of solid tumors is the most effective and a prior remedial measure to treat cancer. Although medical treatment, technology, and science have advanced significantly, it is challenging to completely treat this lethal disease. Near-infrared (NIR) fluorescence, including the first near-infrared region (NIR-I, 650-900 nm) and the second near-infrared region (NIR-II, 1,000-1,700 nm), plays an important role in image-guided cancer surgeries due to its inherent advantages, such as great tissue penetration, minimal tissue absorption and emission light scattering, and low autofluorescence. By virtue of its high precision in identifying tumor tissue margins, there are growing number of NIR fluorescence-guided surgeries for various living animal models as well as patients in clinical therapy. Herein, this review introduces the basic construction and operation principles of fluorescence molecular imaging technology, and the representative application of NIR-I/II image-guided surgery in biomedical research studies are summarized. Ultimately, we discuss the present challenges and future perspectives in the field of fluorescence imaging for surgical navigation and also put forward our opinions on how to improve the efficiency of the surgical treatment.

Design of Activatable NIR-II Molecular Probe for In Vivo Elucidation of Disease-Related Viscosity Variations.

A clear elucidation of a disease-related viscosity change in vivo is significant yet highly challenging as well. Fluorescence imaging in the second near-infrared region (NIR-II, 1000-1700 nm) has gained increasing attention for observation in living organisms, but a viscosity-activatable fluorescent probe emitting at this region remains a vacancy. Herein, we report the first panel of a viscosity-activated NIR-II emissive fluorescent probe WD-X. By embedding different substituents into the WD-X platform and screening, we obtained an ideal probe, WD-NO2, which displayed the best combination of properties, including a 31-fold fluorescence enhancement in response to viscosity, insensitivity to environments (pH, polarity), and relatively high quantum yield (1.6% in glycerol). WD-NO2 was successfully applied to track the variation of viscosity in diabetes-induced liver injury in vivo.

In Situ Imaging of Cysteine in the Brains of Mice with Epilepsy by a Near-Infrared Emissive Fluorescent Probe.

Epilepsy is characterized by oxidative stress in the brain. As the crucial reductive biothiol, cysteine (Cys) directly regulates the occurrence of oxidative stress in the brain. Observations suggest that the decreased cysteine in plasma could potentially serve as a redox biomarker in temporal lobe epilepsy. However, due to the complexity of the brain and lack of appropriate in situ detecting tools, the concentration change and regulation of Cys in epileptic brains remain unclear. Here, we report a near-infrared imaging probe (named Mito-CP) for tracking endogenous Cys in brains of pentylenetetrazole (PTZ)-induced epileptic seizures with high sensitivity and selectivity. Using this probe, we achieved an in situ visualization of the increased Cys in PC12 cells under dithiothreitol stimulation. In addition, Mito-CP was able to real-time monitor Cys fluctuation in lipopolysaccharide-mediated oxidative stress in zebrafish. Ultimately, we directly visualized the precipitous reduction of Cys in epileptic brains for the first time. Mito-CP also revealed the fluctuation of Cys in epileptic brains during the treatment by an antiepileptic drug, curcumin. This work provides an effective tool for Cys imaging in brains and will help to expand the understanding of the pathogenesis of epilepsy.

Simultaneous imaging of mitochondrial viscosity and hydrogen peroxide in Alzheimer's disease by a single near-infrared fluorescent probe with a large Stokes shift.

It has been speculated that both the intracellular viscosity and H2O2 level in Alzheimer's disease (AD) brains are higher than that in healthy brains, but direct evidence from living beings is scarce. Herein, we report a NIR emissive fluorescent probe with a large Stokes shift for the associated detection of mitochondrial viscosity and H2O2 in live rat brains with AD for the first time.

Construction of a two-photon fluorescent probe for ratiometric imaging of hypochlorous acid in alcohol-induced liver injury.

Alcohol-induced liver injury has been a terrible threat to human health and life. The relationship between HClO and the process is unclear. Thus, a ratiometric two-photon fluorescent probe for HClO was deliberately constructed and revealed the generation of HClO in the alcohol-induced liver injury process for the first time.

Efficient Two-Photon Fluorescent Probe for Imaging of Nitric Oxide during Endoplasmic Reticulum Stress.

Nitric oxide (NO) is a vital gaseous signal molecule and plays an important role in diverse physiological and pathological processes including regulation of vascular functions. Endoplasmic reticulum (ER) stress is caused by the accumulation of misfolded or unfolded protein in the ER. Besides, ER stress induced by NO can be involved in the pathogenesis of various vascular diseases. Unfortunately, to the best of our knowledge, no ER-targeting probe for NO is reported to study the relationship between ER stress and the level of NO in a biological system. Herein, an ER-targeted fluorescent probe named ER-Nap-NO for imaging of NO is designed and synthesized. ER-Nap-NO consists of three main parts: naphthalimide (two-photon fluorophore), o-phenylenediamino (NO recognition group), and methyl sulfonamide (ER-targetable group). The probe itself is nonfluorescent because a photoinduced electron transfer (PET) process exists. After the addition of NO, the PET process is inhibited and thus strong fluorescence is released. Moreover, the response mechanism is confirmed by 1H NMR and mass spectra and DFT calculation in detail. In addition, from the experimental results, we can conclude that the probe displays several obvious advantages including high sensitivity, selectivity, and ER-targetable ability. Based on these excellent properties, the probe is used for the two-photon imaging of exogenous and endogenous NO in ER of living cells. Most importantly, the ER-targetable probe has potential capability as a tool for investigating the level of NO during tunicamycin-induced ER stress in cells and tissues, which is beneficial for revealing the role of NO in ER-associated vascular diseases.

A Dual-Response Fluorescent Probe for the Detection of Viscosity and H2S and Its Application in Studying Their Cross-Talk Influence in Mitochondria.

Intracellular viscosity is an essential microenvironmental parameter and H2S is a critical gaseous signaling molecule, which are both related to various physiological processes. It is reported that the change of viscosity and an imbalance of H2S production in the mitochondria are both associated with overexpression of amyloid betapeptide (Aß), which is thought to play a central role in the pathogenesis of Alzheimer's disease (AD). However, to our best knowledge, no fluorescent probe is found for dual detection of mitochondrial viscosity and H2S. Herein, a dual-response fluorescent probe (Mito-VS) is designed and synthesized to monitor the level of viscosity and H2S, respectively. Mito-VS itself is nonfluorescent due to a free intramolecular rotation between dimethylaniline and pyridine. After the increase of viscosity, the rotation is prohibited and an intense red fluorescence is released. Upon the addition of H2S, the probe can react with H2S to form compound 3 and a strong green fluorescence can be observed. Moreover, the probe possesses a good mitochondrion-targeting ability and is applied for imaging the change of viscosity on the red channel and visualizing the variation of exogenous and endogenous H2S concentration on the green channel in mitochondria. Most importantly, the probe is capable of studying the cross-talk influence of viscosity and H2S in mitochondria, which is very beneficial for knowing the pathogenesis of AD.

Mitochondria-targeted near-infrared fluorescent probe for the detection of carbon monoxide in vivo.

Carbon monoxide is a critical gasotransmitter in the body and related with mitochondrial respiration. To date, various fluorescent probes for CO have been well proposed, but two main problems remain. One is that most of the probes are not mitochondria-targeting, even if the probes claim to be able to detect CO in living cells. The other is that the probes for CO display excitation and emission within the ultraviolet or visible range, which hinders their applications in vivo. Herein, a hemicyanine-based near-infrared (NIR) fluorescent probe named CyAPC is first synthesized and used to detect mitochondrial CO. The characteristics of probe CyAPC are as follows: (1) The fluorescence emission of the sensing system is at 736 nm belonging to NIR region, which is suitable for bioimaging in vivo. (2) CyAPC, a positively charged molecule, would have a high tendency to localize in mitochondria of cells. (3) The fluorescence change of the probe is attributed to the fact that CO with Pd2+ induced cleavage of the allyl formate group from the probe and CyAPC (fluorescence off) is transformed into CyOH (fluorescence on), which is proved by HPLC, MS and DFT calculation. (4) The NIR fluorescent probe is applied for the detection of exogenous and endogenous CO in various biological samples such as cell, tissue and in vivo with satisfactory results.