Dynamic evaluation of the protective effect of Dendrobium officinale polysaccharide on acute alcoholic liver injury mice in vitro and in vivo by NIR fluorescence imaging.

Acute alcoholic liver injury (AALI) is a threat to human health. Dendrobium officinale polysaccharide (DOP) has the potential to protect the liver by enhancing the anti-oxidative system to maintain the relative balance of ROS (active oxygen species) and antioxidants in AALI mice. However, the dynamic improvement effect of DOP on AALI is still not clear and accurate medication guidance is not available, which limits the clinical application of DOP. Because of the advantages of high sensitivity, noninvasiveness, and visualization, near-infrared (NIR) fluorescence imaging has been widely studied in biochemistry and biomedicine. As the glutathione (GSH) level in the liver is closely related to the progression of AALI, herein, an NIR fluorescent probe for GSH, HCG was used to dynamically evaluate the effect of DOP on AALI mice. In this study, DOP was proven to maintain the relative balance of GSH content in the liver to protect it from damage. To the best of our knowledge, it is the first time to assess the effect of DOP on AALI mice through a NIR fluorescence imaging technique. This study may also provide a potential NIR imaging agent for the clinical research to improve the management of liver injury-related diseases.

A deep-red emission fluorescent probe with long wavelength absorption for viscosity detection and live cell imaging.

Intracellular viscosity is closely related to a series of biological processes and could be a biomarker for various diseases. Herein, we reported a deep-red emission viscosity probe ACI, which showed a turn-on fluorescence effect with excellent selectivity encountering high viscous medium. To assure the practical biological application, ACI demonstrated not only a long wavelength emission at 634 nm but also a long wavelength excitation at 566 nm, which were crucial to afford deeper penetration depth and higher sensitivity in bioimaging. The photophysical properties and viscosity recognition mechanism of the probe were carefully discussed here. Theoretical calculations furtherly confirmed that high viscous medium could inhibit the twisted intramolecular charge transfer (TICT) process of the probe which quenched the fluorescence in low viscous media, and restore the emission. More importantly, it was successfully applied to visualize the viscosity in living cells. Graphical abstract.

A novel fluorescent probe with red emission and a large Stokes shift for selective imaging of endogenous cysteine in living cells.

Cysteine (Cys) plays crucial roles in physiological and pathological processes, and is related to many diseases. Selective monitoring of Cys over its analogues homocysteine (Hcy) and glutathione (GSH) is of great significance. A probe named ANT with an acrylate group as a recognizing moiety and a red-emission dye with a large Stokes shift (160 nm) as a fluorophore was designed and synthesized. The acrylate group can quench the fluorescence of ANT by a photoinduced electron transfer (PET) process. However, in the presence of Cys, the emission was switched on by the cleavage of the quencher through a concerted reaction including Michael addition and intramolecular cyclization. Furthermore, solution experiments showed that ANT exhibited high sensitivity and selectivity towards Cys with the maximum emission at 640 nm. Besides, ANT was successfully applied to specifically map endogenous Cys in living MCF-7 cells with low toxicity, despite the interference of Hcy and GSH. We hope that such a prepared novel probe will bring advancement in specifically differentiating Cys, Hcy and GSH.

Strategies targeting PD-L1 expression and associated opportunities for cancer combination therapy.

Immunotherapy has achieved great success recently and opened a new avenue for anti-tumor treatment. Programmed cell death 1/programmed cell death ligand 1 (PD-1/PD-L1) are typical immune checkpoints that transmit coinhibitory signals, muting the host immunity. Monoclonal antibodies that block PD-1/PD-L1 axis have benefited many patients with different tumor diseases. However, the objective response rate is still unsatisfactory. In this review, we summarize three strategies targeting PD-L1 based on different forms of PD-L1 and various regulating mechanisms to enhance the therapeutic effect, including blockade of the interaction between PD-L1 and PD-1, downregulation of PD-L1 expression and degradation of mature PD-L1. Thereinto, we describe a variety of materials have been designed to target PD-L1, including antibodies, nanoparticle, peptide, aptamer, RNA, and small molecule. Additionally, we list the drugs with PD-L1 regulation capacity used in clinical and ongoing studies to explore other alternatives for targeting PD-L1 besides anti-PD-L1 monoclonal antibodies. Moreover, we discuss associated opportunities for cancer combination therapy with other modalities such as chemotherapy, radiotherapy, photodynamic therapy (PDT) and photothermal therapy (PTT), as these conventional or emerging modalities are capable of increasing the immune response of tumor cells by altering the tumor microenvironment (TME), and would display synergistic effect. At last, we give a brief summary and outlook regarding the research status and future prospect of immunotherapy.

Exploring the microscopic changes of lipid droplets and mitochondria in alcoholic liver disease via fluorescent probes with high polarity specificity.

Alcoholic liver disease (ALD) has received extensive attention because of the increasing alcohol consumption globally as well as its high morbidity. It is reported that absorbed alcohol can cause lipid metabolism disorder and mitochondria dysfunction, so here in this work, we planned to study the microscopic changes of the two organelles, lipid droplets (LDs) and mitochondria in hepatocyte, under the stimulation of alcohol, hoping to present some meaningful information for the theranostics of ALD by the technique of fluorescence imaging. Guided by theoretical calculation, two fluorescent probes, named CBu and CBuT, were rationally designed. Although constructed by the same chromophore scaffold, they stained different organelles efficiently and emitted distinctively. CBu with high lipophilicity, ascribed to the two butyl groups, can selectively localize in LDs with green fluorescence, while CBuT bearing a triphenylphosphine unit can specifically target mitochondria due to electrostatic interactions with near-infrared (NIR) fluorescence. Both probes displayed remarkable selectivity and sensitivity to polarity, free from the environmental interferences including viscosity, pH and other bio-species. With these two probes, the accumulation of LDs and polarity decrease in mitochondria were clearly monitored at the green and red channels, respectively, in the ALD cell model. CBuT was further applied to image the mice with ALD in vivo. In short, we have confirmed the valuable organelles, LDs and mitochondria, for ALD study and provided two potent molecular tools to visualize their changes through fluorescence imaging, which would be favorable for the further development of theranostics for ALD.

Recent Progress of Glutathione (GSH) Specific Fluorescent Probes: Molecular Design, Photophysical Property, Recognition Mechanism and Bioimaging.

The selective detection of glutathione (GSH) in vitro and in vivo has attracted great attentions, credited to its important role in life activities and association with a series of diseases. Among all kinds of analytical techniques, the fluorescent probe for GSH detection become prevalent recently because of its ease of operation, high temporal-spatial resolution, visualization and noninvasiveness, etc. The special structural features of GSH, such as the nucleophilicity of sulfhydryl group, the concerted reaction ability of amino group, the negative charged nature, the latent hydrogen bonding ability along with its flexible molecular chain, are all potent factors to be employed to design the specific fluorescent probe for GSH and discriminate it from other bio-species including its analogues cysteine (Cys) and homocysteine (Hcy). This paper reviewed the studies in the last 3 years and was organized based on the reaction mechanism of each probe. According to the reactivity of GSH, various recognition mechanisms including Michael addition, nucleophilic aromatic substitution, ordinary nucleophilic substitution, multi-site reaction, and other unique reactions have been utilized to construct the GSH specific fluorescent probes, and the molecular design strategy, photophysical property, recognition mechanism, and bioimaging application of each reported probe were all discussed here systematically. Great progress has been made in this area, and we believe the analyses and summarization of these excellent studies would provide valuable message and inspiration to researchers to advance the research toward clinic applications.

NIR-II imaging-guided diagnosis and evaluation of the therapeutic effect on acute alcoholic liver injury via a nanoprobe.

Acute alcoholic liver injury (AALI) is hard to diagnose on account of no obvious clinical symptoms, and thereby it easily develops into serious liver diseases and threatens people's health. However, traditional methods for detecting AALI are far from satisfactory due to the low sensitivity, invasiveness and non-visualization, and the development of new techniques is in urgent demand. Near-infrared (NIR)-II fluorescence imaging has been widely studied in biochemistry and biomedicine. As the blood flow velocity of the liver is closely related to the progression of AALI, herein, a NIR-II fluorescent nanoprobe, NTPB-NPs, was applied to diagnose AALI by monitoring the fluorescence intensity changes in the liver caused by the variations of the blood flow velocity. More importantly, when medication was applied to alleviate the liver injury of AALI mice, NTPB-NPs could also track the therapeutic effect in situ. In this study, the relationship between hepatic vascular velocity and the progression of AALI was confirmed with NTPB-NPs via NIR-II imaging. To the best of our knowledge, this is the first time that a NIR-II fluorescence imaging technique has been used to diagnose AALI mice and evaluate the therapeutic effect on AALI mice. This study may also provide a potential NIR-II imaging agent for clinical research to improve the management of liver injury related diseases.

Structure modulation on fluorescent probes for biothiols and the reversible imaging of glutathione in living cells.

The detection of small molecular biothiols (cysteine, homocysteine and glutathione) is of great importance, as they involve in a series of physiological and pathological processes and are associated with many diseases. To realize the real-time monitoring of a specific biothiol, a rapid and reversible probe is required. Therefore, three probes, namely, o-MNPy, m-MNPy and p-MNPy, with pyridine substituted α, ß-unsaturated ketone as the recognition site, were reported here, and the reactivity of the recognition site was finely tuned by the connection mode of the pyridine unit. To single out the optimal one, the response performances of three probes toward each biothiol were systemically studied, taking the differences of the intracellular contents of three biothiols into account during the evaluation. Biothiols reacted with the probes through Michael addition, and results showed that the slight structural variations could affect the performances of the probes obviously. p-MNPy with the pyridine unit connected to the recognition site through the para-position of the nitrogen atom, revealed the best sensing ability among the three probes. It demonstrated rapid response, good selectivity and sensitivity, excellent pH adaptability to Cys and GSH, and displayed reversible detection toward GSH. Finally, p-MNPy was successfully applied to track the GSH fluctuations under the oxidative stress stimulated by H2O2 in living cells.

Highly Efficient Near-Infrared Photosensitizers with Aggregation-Induced Emission Characteristics: Rational Molecular Design and Photodynamic Cancer Cell Ablation.

Photosensitizers (PSs) that play a decisive role in effective photodynamic therapy (PDT) have attracted great research interest. PSs with aggregation-induced emission (AIE) characteristics could overcome the deficiencies of traditional PSs that usually suffer from the aggregation-caused fluorescence quenching (ACQ) effect in applications and show enhanced emission and high singlet oxygen (1O2) generation efficiency in aggregates; therefore, they are outstanding candidates for imaging-guided PDT, and the development of AIE PSs with both excellent photophysical properties and 1O2 generation ability is highly desirable. Herein, three AIE fluorogens (AIEgens), BtM, ThM, and NaM, with a donor-π-acceptor (D-π-A) structure were designed and synthesized, and the photosensitizing ability was adjusted by π-linker engineering. All of the three AIEgens showed excellent photostability and high molar absorption coefficients, and their emission edges were extended to the near-infrared (NIR) region, with peaks at 681, 678, and 638 nm, respectively. NaM demonstrated the smallest ΔES1-T1, which was ascribed to its better separation degree of the highest occupied molecular orbital (HOMO) and the lowest unoccupied molecular orbital (LUMO). The AIEgens were fabricated into nanoparticles (NPs) by amphipathic mPEG3000-DSPE encapsulating, and thus the obtained NaM NPs exhibited the best 1O2 generation efficiency under white light irradiation, which was almost 3 times that of the renowned PS rose bengal (RB). Furthermore, under white light irradiation, the cell killing efficiency of NaM NPs was also much better than those of the other two AIE PSs and RB. Therefore, NaM NPs revealed great potential to treat superficial diseases as a PS for PDT.

An ultralow concentration of two-photon fluorescent probe for rapid and selective detection of lysosomal cysteine in living cells.

Herein we reported a two-photon (TP) fluorescence "turn-on" probe MNPO, exhibiting high selectivity and sensitivity towards intracellular cysteine (Cys) with excellent lysosomal localization. The probe displayed fast response towards Cys over homocysteine (Hcy), glutathione (GSH), and other various analytes under physiological conditions. Low cytotoxicity made it successful for TP imaging of Cys in HeLa cells with an ultralow probe concentration of 250 nM, and a rapid response of only 10 min. Simultaneously, colocalization experiments in lysosome demonstrated its ability for specific in situ detection of lysosomal Cys in living cells, which shed light on its potential applications in biomedical applications. Beyond that MNPO was successfully applied for TP imaging of Cys in mice organ tissues such as heart, liver, and spleen, and the penetration depth of mice heart tissue was up to 184 µm, which disclosed the predominant TP characteristic. We believe that this study will provide some useful information toward diagnosis and treatment of pathogenesis associated with Cys or lysosomes in future.

A low background D-A-D type fluorescent probe for imaging of biothiols in living cells.

Two probes, structurally symmetric CBFB and asymmetric CBFM, constructed by a D-A-D (donor-acceptor-donor) type curcuminoid as the fluorophore and the DNBS (2,4-dinitrobenzenesulfonyl) group as the biothiol recognition site were designed and synthesized here. The DNBS group can quench the emission of the fluorophore by the PET (photoinduced electron transfer) process, and in the presence of biothiols, the emission of the probe was switched on as a result of the cleavage of the quencher by a nucleophilic aromatic substitution reaction. Experimental analyses and theoretical calculations revealed that two recognition moieties in the molecule can quench the fluorescence more efficiently, therefore, CBFB showed a much higher SNR (signal to noise ratio) than CBFM in biothiol detection with an emission maximum at 610 nm. This "low background" and "turn-on" fluorescent probe, CBFB, was successfully utilized to map endogenous biothiols in living cells.

Photoisomerization of spiropyran for driving a molecular shuttle.

A novel light-powered molecular shuttle was synthesized which can switch the movement of a macrocycle between two distinct stations-dipeptide and zwitterionic ME-by exploiting the photoisomerization of spiropyran. The macrocycle resides selectively in the dipeptide station in the SP form and moves to the ME station under the irradiation of UV light. This movement process of the macrocycle is accompanied by reversible absorptive output signals which can be detected by the naked eye.

Colorimetric and fluorescent probes for real-time naked eye sensing of copper ion in solution and on paper substrate.

In this paper, BT ((E)-2-(4-(4-(bis(pyridin-2-ylmethyl)amino)styryl)-3-cyano-5,5-dimethylfuran-2(5H)-ylidene)malononitrile) with strong donor-π-acceptor structure was synthesized, which showed both colorimetric and fluorescent sensing ability toward Cu2+ with high selectivity and sensitivity. Job plot and mass spectra measurement revealed a 1 : 1 coordination mode between Cu2+ and probe BT in ethanol/HEPES (1 : 4 v/v) buffer (pH 7.2) solution, and the binding constant was calculated to be 3.6 × 104 M-1. The colour of BT solution (10 µM) immediately turned from purple red to yellow and the red fluorescence was quenched obviously when a certain amount of Cu2+ was added, which enabled a dual-channel detection of Cu2+. A paper strip pre-stained with BT solution was further fabricated and it also showed excellent sensing ability toward Cu2+ with a detection limit as low as 10-6 M with the naked eye, which represents better portability and operation simplicity that is favourable for on-site analysis of Cu2+ in water.

Pyridine-ring containing twisttetraazaacene: Synthesis, physical properties, crystal structure and picric acid sensing.

Novel pyridine-ring containing twisttetraazaacene 9,14-diphenylpyreno[4,5-g]isoquinoline (1) and its full-carbon derivative 9,14-diphenyldibenzo[de,qr]tetracene (2) have been synthesized and fully characterized. Studies showed that compound 1 could identify picric acid (PA) over other common nitro compounds with high selectivity and sensitivity. Upon the addition of PA, the emission peak of compound 1 in CH3CN was red shifted from 447 to 555nm with a fluorescence quenching efficiency as high as 95%, the detection limit was calculated to be 2.42µM, while its full-carbon derivative (2) could not exhibit this kind of performance. The possible mechanism with the enhanced PA detection efficiency in pyridine-ring containing twisttetraazaacene (1) than its full-carbon derivative (2) was also investigated.