cGAS-STING pathway agonists are promising vaccine adjuvants.

Adjuvants are of critical value in vaccine development as they act on enhancing immunogenicity of antigen and inducing long-lasting immunity. However, there are only a few adjuvants that have been approved for clinical use, which highlights the need for exploring and developing new adjuvants to meet the growing demand for vaccination. Recently, emerging evidence demonstrates that the cGAS-STING pathway orchestrates innate and adaptive immunity by generating type I interferon responses. Many cGAS-STING pathway agonists have been developed and tested in preclinical research for the treatment of cancer or infectious diseases with promising results. As adjuvants, cGAS-STING agonists have demonstrated their potential to activate robust defense immunity in various diseases, including COVID-19 infection. This review summarized the current developments in the field of cGAS-STING agonists with a special focus on the latest applications of cGAS-STING agonists as adjuvants in vaccination. Potential challenges were also discussed in the hope of sparking future research interests to further the development of cGAS-STING as vaccine adjuvants.

Visualizing intracellular membrane interactions and cell type-specific differentiation in ferroptosis and apoptosis with Boranil-Carbazole derivative.

Intracellular lipid systems play essential roles in various physiological functions and cell growth processes. However, our understanding of the intricate interactions within this system, especially between mitochondria and lipid droplets, is limited, particularly in the context of cancer cells' altered lipid metabolism. To address this, our study introduces an N-B-O BODIPY-hexylcarbazole derivative, named Cz-Boranil, that sets a new benchmark in visualizing these critical interactions. Cz-Boranil's unique capability lies in its ability to display distinct intracellular distribution patterns in both normal and cancer cells, offering nuanced cell type-specific differentiation. More impressively, this probe tracks the coordinated interactions of lipid droplets and mitochondria during the critical processes of ferroptosis and apoptosis. We believe that the innovative capabilities of Cz-Boranil will revolutionize our understanding of intracellular lipid interactions and prove pivotal in identifying and studying cancerous cells.

Nanoscale myelinogenesis image in developing brain via super-resolution nanoscopy by near-infrared emissive curcumin-BODIPY derivatives.

Understanding the intricate nanoscale architecture of neuronal myelin during central nervous system development is of utmost importance. However, current visualization methods heavily rely on electron microscopy or indirect fluorescent method, lacking direct and real-time imaging capabilities. Here, we introduce a breakthrough near-infrared emissive curcumin-BODIPY derivative (MyL-1) that enables direct visualization of myelin structure in brain tissues. The remarkable compatibility of MyL-1 with stimulated emission depletion nanoscopy allows for unprecedented super-resolution imaging of myelin ultrastructure. Through this innovative approach, we comprehensively characterize the nanoscale myelinogenesis in three dimensions over the course of brain development, spanning from infancy to adulthood in mouse models. Moreover, we investigate the correlation between myelin substances and Myelin Basic Protein (MBP), shedding light on the essential role of MBP in facilitating myelinogenesis during vertebral development. This novel material, MyL-1, opens up new avenues for studying and understanding the intricate process of myelinogenesis in a direct and non-invasive manner, paving the way for further advancements in the field of nanoscale neuroimaging.

Three-Photon AIE Pt(II) Complexes as Cysteine-Targeting Theranostic Agents for Tumor Imaging and Chemotherapy.

Herein, we have synthesized a series of three-photon fluorescent Pt(II) complexes targeting a tumor-associated biothiol, cysteine (Cys), which allows it to be detected without any interference from other intracellular proteins. We focused on how to significantly improve the fluorescence response of Cys via regulating the recognition units in probes. The reaction of K2PtCl4 with L-CH3 or L-COOEt in DMSO solution gave Lyso-Pt-CH3 and Lyso-Pt-COOEt, respectively, which present four-coordinated square-planar geometries in mononuclear structures. Lyso-Pt-CH3 consists of a Cys aptamer labeled with typical aggregation-induced emission (AIE) characteristics, which shows strong three-photon absorption cross section (3PA) only in the presence of Cys. It was found that Lyso-Pt-CH3 displayed a perfect signal-to-noise ratio for imaging lysosomes and for rapid detection of Cys. Using Lyso-Pt-CH3, Cys-related cellular mechanisms were proposed. We confirm that cystine (Cyss) could be absorbed in cells through cystine/glutamate antiporters (system xc-) and is then converted to Cys under the effect of enzymes. All of these suggest that Lyso-Pt-CH3 might be a potential candidate as a simple and straightforward biomarker of lysosome-related Cys in vitro. Lyso-Pt-CH3 can effectively identify tumor tissues with excessive levels of Cys. Lyso-Pt-CH3 also showed excellent antitumor activity than cisplatin. This work provides a novel strategy for the rational design of controllably activated and Cys-targeted Pt(II) anticancer prodrugs for clinical diagnosis and treatment.

Fine Tuning of Multiphoton AIE Emission Behavior, Organelle Targeting, and Fluorescence Lifetime Imaging of Terpyridine Derivatives by Alkyl Chain Engineering.

In this work, a series of multiphoton terpyridine agents (ZA, ZA-Mex, and ZA-Hex) for fluorescence lifetime imaging microscopy (FLIM) are designed and synthesized. The results from photophysical property research reveal that ZA-Hex, as an N-hexylated terpyridine salt, has stronger three-photon aggregation-induced emission (AIE) properties compared to ZA-Mex due to enhanced intramolecular charge transfer (ICT) performance. All three terpyridine derivatives possess suitable fluorescence intensities and stable fluorescence lifetimes under different pH conditions (pH = 4.0-8.0), thereby performing multiphoton fluorescence lifetime imaging. For biological imaging applications, it is found that ZA shows good lipid droplet (LD) turn-on fluorescence performance, and ZA-Hex could easily accumulate in mitochondria with high specificity. This is the first report of terpyridine salts as three-photon AIE probes used for multiphoton FLIM imaging.

Coordination-Regulated Terpyridine-Mn(II) Complexes for Photodynamic Therapy Guided by Multiphoton Fluorescence/Magnetic Resonance Imaging.

The synergy of multiphoton fluorescence imaging (MP-FI) and magnetic resonance imaging (MRI) provides an imaging platform with high resolution and unlimited penetration depth for early disease detection. Herein, two kinds of terpyridine-Mn(II) complexes (FD-Mn-O2NO and FD-Mn-FD) possessing seven and six coordination modes, respectively, were designed rationally for photodynamic therapy (PDT) guided by MP-FI/MRI. The complexes obtain different multiphoton fluorescence/magnetic resonance properties by adjusting the number of terpyridine ligands. Among them, FD-Mn-FD exhibits the following superiorities: (1) The optimal three-photon excitation wavelength of FD-Mn-FD falls at 1450 nm (NIR-II), which brings high sensitivity and deep tissue penetration in MP-FI. (2) FD-Mn-FD has effective longitudinal relaxation efficiency (r1 = 2.6 m M-1 s-1), which can be used for T1-weighted MRI, overcoming the problems of limited tissue penetration depth and low spatial resolution. (3) FD-Mn-FD generates endogenous 1O2 under irradiation by 808 nm light, thereby enhancing the PDT effect in vitro and in vivo. To the best of our knowledge, the complex FD-Mn-FD is the first complex to guide PDT through MP-FI/MRI, providing a blueprint for accurate and effective early detection and timely treatment of the complex in the early stages of cancer.

Curcumin- and resveratrol-co-loaded nanoparticles in synergistic treatment of hepatocellular carcinoma.

BACKGROUND: Currently, systemic therapies for patients with advanced-stage hepatocellular carcinoma (HCC) rely mainly on systemic drugs. However, traditional systemic drugs have a high rate of serious adverse events, and the curative effects of some potential anticancer drugs, such as curcumin (CUR) and resveratrol (RSV), are less apparent due to their poor bioavailability. Therefore, it is urgent to develop a highly effective therapy to improve patient prognosis. Herein, an injectable HCC-targeted nanoparticle (NP) was designed to deliver CUR and RSV to hepatoma cells. RESULTS: The molecular self-assembled NPs showed higher tumour retention through the enhanced permeability and retention (EPR) effect of the NPs and surface modification with the HCC-specific peptide moiety SP94 to effectively treat HCC. These HCC-targeted NPs led to a significant reduction in the drug dosage, delayed the rate of drug release and improved the bioavailability of the encapsulated drugs. The drug concentrations in the vicinity of the tumour increased, and a good therapeutic effect was observed without obvious side effects. CONCLUSIONS: These SP94-mediated NPs allowed large amounts of antitumor drugs to accumulate in tumours, providing a novel strategy for innovative HCC therapy. This nanoplatform also offers an idea for exploring other potential chemotherapeutics.

Revealing Sulfur Dioxide Regulation to Nucleophagy in Embryo Development by an Adaptive Coloration Probe.

Understanding signaling molecules in regulating organelles dynamics and programmed cell death is critical for embryo development but is also challenging because current imaging probes are incapable of simultaneously imaging the signaling molecules and the intracellular organelles they interact with. Here, we report a chemically and environmentally dual-responsive imaging probe that can react with gasotransmitters and label cell nuclei in distinctive fluorescent colors, similar to the adaptive coloration of chameleons. Using this intracellular chameleon-like probe in three-dimensional (3D) super-resolution dynamic imaging of live cells, we discovered SO2 as a critical upstream signaling molecule that activates nucleophagy in programmed cell death. An elevated level of SO2 prompts kiss fusion between the lysosomal and nuclear membranes and nucleus shrinkage and rupture. Significantly, we revealed that the gasotransmitter SO2 is majorly generated in the yolk, induces autophagy there at the initial stage of embryo development, and is highly related to the development of the auditory nervous system.

Halogen-modified carbazole derivatives for lipid droplet-specific bioimaging and two-photon photodynamic therapy.

Lipid droplets (LDs) are dynamic multifunctional organelles that participate in the regulation of many metabolic processes, visualization of which is necessary for biological research. In this work, a series of two-photon responsive fluorescent probes (C-H, C-Br, and C-I) based on carbazole units were designed and synthesized. Thereinto, an iodine-modified carbazole derivative C-I exhibited an exciting lipid droplet targeting ability due to its excellent lipophilicity. Meanwhile, benefiting from its larger Stokes shift and two-photon absorption cross-section, C-I was employed for two-photon confocal laser scanning microscopy (CLSM) and stimulated emission depletion (STED) microscopy imaging to observe LDs more accurately. In addition, given the heavy atom effect, C-I can effectively generate reactive oxygen species (ROS) leading to cancer cell apoptosis under near-infrared light irradiation. Notably, we explained the process of cell apoptosis through in vitro simulation experiments. This study provides a promising platform for visualization of lipid droplets.

A multi-photon fluorescent probe based on quinoline groups for the highly selective and sensitive detection of lipid droplets.

Compared to general fluorescent probes, multi-photon fluorescent probes exhibit deeper tissue penetration, lower auto-fluorescence and lower photo-toxicity in the bio-imaging field. Herein, we synthesized a series multi-photon fluorescent probe (L1-L3) based on quinolone groups. Of notably, the three-photon fluorescence of L3 significantly enhanced when L3 interacted with liposome; moreover, L3 exhibited high selectivity towards lipid droplets in living cells. Due to its large Stokes shift, high selectivity and photon-stability, L3 was successfully used in lipid droplet imaging via multi-photon fluorescence bio-imaging.

Functional Platinum(II) Complexes with Four-Photon Absorption Activity, Lysosome Specificity, and Precise Cancer Therapy.

Multiphoton materials are in special demand in the field of photodynamic therapy and multiphoton fluorescence imaging. However, rational design methodology for these brands of materials is still nascent. This is despite transition-metal complexes favoring optimized nonlinear-optical (NLO) activity and heavy-atom-effected phosphorescent emission. Here, three four-photon absorption (4PA) platinum(II) complexes (Pt1-Pt3) are achieved by the incorporation of varied functionalized C^N^C ligands with high yields. Pt1-Pt3 exhibit triplet metal-to-ligand charge-transfer transitions at ∼460 nm, which are verified multiple times by transient absorption spectra, time-dependent density functional theory calculations, and low-temperature emission spectra. Further, Pt1-Pt3 undergo 4PA. Notably, one of the complexes, Pt2, has maximum 4PA cross-sectional values of up to 15.2 × 10-82 cm8 s3 photon-3 under excitation of a 1600 nm femtosecond laser (near-IR II window). The 4PA cross sections vary when Pt2 is binding to lecithin and when it displays its lysosome-specific targeting behavior. On the basis of the excellent 4PA property of Pt2, we believe that those 4PA platinum(II) complexes have great potential applications in cancer theranostics.

A nitroxides-based macromolecular MRI contrast agent with an extraordinary longitudinal relaxivity for tumor imaging via clinical T1WI SE sequence.

BACKGROUND: Macromoleculization of nitroxides has been an effective strategy to improve low relaxivities and poor in vivo stability, however, nitroxides-based metal-free magnetic resonance imaging (MRI) macromolecular contrast agents (mCAs) are still under-performed. These mCAs do not possess a high nitroxides content sufficient for a cumulative effect. Amphiphilic nanostructures in these mCAs are not stable enough for highly efficient protection of nitroxides and do not have adequate molecular flexibility for full contact of the paramagnetic center with the peripheral water molecules. In addition, these mCAs still raise the concerns over biocompatibility and biodegradability due to the presence of macromolecules in these mCAs. RESULTS: Herein, a water-soluble biodegradable nitroxides-based mCA (Linear pDHPMA-mPEG-Ppa-PROXYL) was prepared via covalent conjugation of a nitroxides (2,2,5,5-tetramethyl-1-pyrrolidinyl-N-oxyl, PROXYL) onto an enzyme-sensitive linear di-block poly[N-(1, 3-dihydroxypropyl) methacrylamide] (pDHPMA). A high content of PROXYL up to 0.111 mmol/g in Linear pDHPMA-mPEG-Ppa-PROXYL was achieved and a stable nano-sized self-assembled aggregate in an aqueous environment (ca. 23 nm) was formed. Its longitudinal relaxivity (r1 = 0.93 mM- 1 s- 1) was the highest compared to reported nitroxides-based mCAs. The blood retention time of PROXYL from the prepared mCA in vivo was up to ca. 8 h and great accumulation of the mCA was realized in the tumor site due to its passive targeting ability to tumors. Thus, Linear pDHPMA-mPEG-Ppa-PROXYL could provide a clearly detectable MRI enhancement at the tumor site of mice via the T1WI SE sequence conventionally used in clinical Gd3+-based contrast agents, although it cannot be compared with DTPA-Gd in the longitudinal relaxivity and the continuous enhancement time at the tumor site of mice. Additionally, it was demonstrated to have great biosafety, hemocompatibility and biocompatibility. CONCLUSIONS: Therefore, Linear pDHPMA-mPEG-Ppa-PROXYL could be a potential candidate as a substitute of metal-based MRI CAs for clinical application.

Light-Up Lipid Droplets Dynamic Behaviors Using a Red-Emitting Fluorogenic Probe.

Intracellular lipid metabolism occurs in lipid droplets (LDs), which is critical to the survival of cells. Imaging LDs is an intuitive way to understand their physiology in live cells. However, this is limited by the availability of specific probes that can properly visualize LDs in vivo. Here, an LDs-specific red-emitting probe is proposed to address this need, which is not merely with an ultrahigh signal-to-noise (S/N) ratio and a large Stokes shift (up to 214 nm) but also with superior resistance to photobleaching. The probe has been successfully applied to real-time tracking of intracellular LDs behaviors, including fusion, migration, and lipophagy processes. We deem that the proposed probe here offers a new possibility for deeper understanding of LDs-associated behaviors, elucidation of their roles and mechanisms in cellular metabolism, and determination of the transition between adaptive lipid storage and lipotoxicity as well.

Activated Type I and Type II Process for Two-Photon Promoted ROS Generation: The Coordinated Zn Matters.

The construction of novel classes of photosensitizers for efficient reactive oxygen species (ROS) generation is of great interest, yet it is a challenge. In this work, a bis(terpyridine)zinc(II) complex (namely, ZnL1) with two-photon absorption activity as an efficient ROS photogenerator was synthesized. Benefiting from the coordinated Zn, the decreased singlet-triplet energy gap favors the intersystem crossing process facilitating the singlet oxygen (1O2) generation via energy transfer. In addition, it makes the superoxide radical (O2·-) generation easier. This is an extremely rare study on two-photon excited ROS generation by activating type I and type II processes based on a cheaper and bioaccessible Zn complex.

A Multi-responsive Fluorescent Probe Reveals Mitochondrial Nucleoprotein Dynamics with Reactive Oxygen Species Regulation through Super-resolution Imaging.

Understanding the biomolecular interactions in a specific organelle has been a long-standing challenge because it requires super-resolution imaging to resolve the spatial locations and dynamic interactions of multiple biomacromolecules. Two key difficulties are the scarcity of suitable probes for super-resolution nanoscopy and the complications that arise from the use of multiple probes. Herein, we report a quinolinium derivative probe that is selectively enriched in mitochondria and switches on in three different fluorescence modes in response to hydrogen peroxide (H2 O2 ), proteins, and nucleic acids, enabling the visualization of mitochondrial nucleoprotein dynamics. STED nanoscopy reveals that the proteins localize at mitochondrial cristae and largely fuse with nucleic acids to form nucleoproteins, whereas increasing H2 O2 level leads to disassociation of nucleic acid-protein complexes.

Coumarin-Based Fluorescent Probes for Super-resolution and Dynamic Tracking of Lipid Droplets.

Visualizing and dynamic tracking lipid droplets (LDs) are of great importance to biological research. Herein, two-photon absorption fluorescent small bioprobes based on lipophilic coumarin were developed, which exhibited high selectivity toward LDs in HeLa cells. Because of good biocompatibility and excellent photostability, the probes were applied to realize specific super-resolution visualization of the intracellular LDs in HeLa cells, offering us the quantitative results of the amount and diameters of LDs as well. Furthermore, the bioprobes were capable of monitoring the movements of the LDs in real time. We believe that bioprobes would provide new avenues to designing bioimaging and biological diagnosis.

Gasotransmitter Regulation of Phosphatase Activity in Live Cells Studied by Three-Channel Imaging Correlation.

Enzyme activity in live cells is dynamically regulated by small-molecule transmitters for maintaining normal physiological functions. A few probes have been devised to measure intracellular enzyme activities by fluorescent imaging, but the study of the regulation of enzyme activity via gasotransmitters in situ remains a long-standing challenge. Herein, we report a three-channel imaging correlation by a single dual-reactive fluorescent probe to measure the dependence of phosphatase activity on the H2 S level in cells. The two sites of the probe reactive to H2 S and phosphatase individually produce blue and green fluorescent responses, respectively, and resonance energy transfer can be triggered by their coexistence. Fluorescent analysis based on the three-channel imaging correlation shows that cells have an ideal level of H2 S to promote phosphatase activity up to its maximum. Significantly, a slight deviation from this H2 S level leads to a sharp decrease of phosphatase activity. The discovery further strengthens our understanding of the importance of H2 S in cellular signaling and in various human diseases.

Membrane-Penetrating Carbon Quantum Dots for Imaging Nucleic Acid Structures in Live Organisms.

The dynamics of DNA and RNA structures in live cells are important for understanding cell behaviors, such as transcription activity, protein expression, cell apoptosis, and hereditary disease, but are challenging to monitor in live organisms in real time. The difficulty is largely due to the lack of photostable imaging probes that can distinguish between DNA and RNA, and more importantly, are capable of crossing multiple membrane barriers ranging from the cell/organelle to the tissue/organ level. We report the discovery of a cationic carbon quantum dot (cQD) probe that emits spectrally distinguishable fluorescence upon binding with double-stranded DNA and single-stranded RNA in live cells, thereby enabling real-time monitoring of DNA and RNA localization and motion. A surprising finding is that the probe can penetrate through various types of biological barriers in vitro and in vivo. Combined with standard and super-resolution microscopy, photostable cQDs allow time-lapse imaging of chromatin and nucleoli during cell division and Caenorhabditis elegans (C. elegans) growth.

Two-Photon-Active Organotin(IV) Complexes for Antibacterial Function and Superresolution Bacteria Imaging.

Antibacterial agents with two-photon absorption are expected to play a significant role in biomedical science. Herein, two novel organotin complexes, HLSn1 and HLSn2, based on coumarin were designed, synthesized, and systematically investigated. It was found that these complexes possessed suitable two-photon-active cross sections in the near-infrared region. Moreover, complex HLSn1 could efficiently inhibit the growth of Gram-negative Escherichia coli and Gram-positive Bacillus subtilis, especially the latter with a minimum inhibitory concentration (MIC; 90%) of 2 ± 0.14 µg mL-1, which is lower than that of Kanamycin (Kana, 8 ± 0.42 µg mL-1). Importantly, two-photon imaging and superresolution development of bacterial stain revealed that complex HLSn1 can react with bacterial membranes, producing reactive oxygen species (ROS) and leading to cell death. These outcomes provide promising applications in the superresolution bacteria imaging, diagnostics, and treatment of bacterial infectious.

A Series of Zn(II) Terpyridine-Based Nitrate Complexes as Two-Photon Fluorescent Probe for Identifying Apoptotic and Living Cells via Subcellular Immigration.

Two-photon active probe to label apoptotic cells plays a significant role in biological systems. However, discrimination of live/apoptotic cells at subcellular level under microscopy remains unachieved. Here, three novel Zn(II) terpyridine-based nitrate complexes (C1-C3) containing different pull/push units were designed. The structures of the ligands and their corresponding Zn(II) complexes were confirmed by single-crystal X-ray diffraction analysis. On the basis of the comprehensive comparison, C3 had a suitable two-photon absorption cross section in the near-infrared wavelength and good biocompatibility. Under two-photon confocal microscopy and transmission electron microscopy, it is found that C3 could target mitochondria in living cells but immigrate into the nucleolus during the apoptotic process. This dual-functional probe (C3) not only offers a valuable image tool but also acts as an indicator for cell mortality at subcellular level in a real-time manner.