Chunchang Zhao

Förster Resonance Energy Transfer Switchable Self-Assembled Micellar Nanoprobe: Ratiometric Fluorescent Trapping of Endogenous H2S Generation via Fluvastatin-Stimulated Upregulation

H2S produced in small amounts by mammalian cells has been identified in mediating biological signaling functions. However, the in situ trapping of endogenous H2S generation is still handicapped by a lack of straightforward methods with high selectivity and fast response. Here, we encapsulate a semi-cyanine-BODIPY hybrid dye (BODInD-Cl) and its complementary energy donor (BODIPY1) into the hydrophobic interior of an amphiphilic copolymer (mPEG-DSPE), especially for building up a ratiometric fluorescent H2S nanoprobe with extraordinarily fast response. A remarkable red-shift in the absorption band with a gap of 200 nm in the H2S response can efficiently switch off the Förster resonance energy transfer (FRET) from BODIPY1 to BODInD-Cl, subsequently recovering the donor fluorescence. Impressively, both the interior hydrophobicity of supramolecular micelles and electron-withdrawing nature of indolium unit in BODInD-Cl can sharply increase aromatic nucleophilic substitution with H2S. The ratiometric strategy based on the unique self-assembled micellar aggregate NanoBODIPY achieves an extremely fast response, enabling in situ imaging of endogenous H2S production and mapping its physiological and pathological consequences. Moreover, the amphiphilic copolymer renders the micellar assembly biocompatible and soluble in aqueous solution. The established FRET-switchable macromolecular envelope around BODInD-Cl and BODIPY1 enables cellular uptake, and makes a breakthrough in the trapping of endogenous H2S generation within raw264.7 macrophages upon stimulation with fluvastatin. This study manifests that cystathione γ-lyase (CSE) upregulation contributes to endogenous H2S generation in fluvastatin-stimulated macrophages, along with a correlation between CSE/H2S and activating Akt signaling pathway.

Development of an Indole-Based Boron-Dipyrromethene Fluorescent Probe for Benzenethiols

Discrimination between chemically related benzenethiols and aliphatic thiols represents a big problem. In this paper, a fluorescent probe, Bodipy-1, containing an indole-based Bodipy as a fluorophore and a 2,4-dinitrobenzenesulfonyl group as a recognition unit was constructed to achieve the selectivity between them. The Bodipy group in the prepared probe was selectively released through aromatic nucleophilic substitution by thiolate anions from benzenethiols, resulting in blue-red switching in the emission spectra in buffer solutions; that is, two new peaks of the phenol/phenolate state of Bodipy-2 at 565 and 629 nm appeared in emission spectra. By varying the pH value from 6.6 to 8.8, the intensity ratio of I(565)/I(629) varies from 2.0 to 0.3 after complete conversion to Bodipy-2, a ca. 7-fold emission ratio change. This ratiometric emission property by varying the pH value makes Bodipy-1 a promising probe to discriminate benzenethiols from aliphatic thiols by careful selection of the reaction pH.

Fluorescent In Situ Targeting Probes for Rapid Imaging of Ovarian‐Cancer‐Specific γ‐Glutamyltranspeptidase

γ-Glutamyltranspeptidase (GGT) is a tumor biomarker that selectively catalyzes the cleavage of glutamate overexpressed on the plasma membrane of tumor cells. Here, we developed two novel fluorescent in situ targeting (FIST) probes that specifically target GGT in tumor cells, which comprise 1) a GGT-specific substrate unit (GSH), and 2) a boron-dipyrromethene (BODIPY) moiety for fluorescent signalling. In the presence of GGT, sulfur-substituted BODIPY was converted to amino-substituted BODIPY, resulting in dramatic fluorescence variations. By exploiting this enzyme-triggered photophysical property, we employed these FIST probes to monitor the GGT activity in living cells, which showed remarkable differentiation between ovarian cancer cells and normal cells. These probes represent two first-generation chemodosimeters featuring enzyme-mediated rapid, irreversible aromatic hydrocarbon transfer between the sulfur and nitrogen atoms accompanied by switching of photophysical properties.

A colorimetric and ratiometric NIR fluorescent turn-on fluoride chemodosimeter based on BODIPY derivatives: high selectivity via specific Si–O cleavage

A colorimetric and fluorescent turn-on chemodosimeter for fluoride with high selectivity was developed on the basis of the specific reaction of F− with BODIPY-OSi, displaying a dramatic color change and distinct near-infrared (NIR) fluorescence enhancement at 676 nm.

Development of a BODIPY-based ratiometric fluorescent probe for hypochlorous acid and its application in living cells

A BODIPY-based ratiometric fluorescent probe for HOCl has been designed based on the transduction of thioether to sulfoxide function. This probe features a marked absorption and emission blue-shift upon the HOCl-promoted rapid transduction, enabling the highly selective and ratiometric detection. In addition, the probe works excellently within a wide pH range of 4-10, addressing the existing pH dependency issue. Living cells studies demonstrate that the probe is cell membrane permeable and can be employed successfully to image endogenous HOCl generation in macrophage cells.

Construction of a fluorescence turn-on probe for highly discriminating detection of cysteine

It is still a challenge to construct probes for discriminating thiols due to the similar structure and reactivities of thiol-containing molecules. We have here developed a Cys specific probe by utilizing the remarkable difference in reactivity toward Cys, Hcy and GSH. The reaction between the designed probe and Cys produces an amino-substituted BODIPY, giving a yellow fluorescence turn-on response. The response to Hcy or GSH shows a red fluorescence turn-on signal, due to the formation of sulfenyl-substituted BODIPY. These distinct fluorescence turn-on responses allow Cys to be distinguished from Hcy and GSH. This probe was also utilized for detection of Cys in living cells and monitoring cystathionine γ-lyase activity in vitro.

A Ratiometric Fluorescent Probe for Monitoring Leucine Aminopeptidase in Living Cells and Zebrafish Model

Leucine aminopeptidase (LAP) is an important cancer-related biomarker, which shows significant overexpression in malignant tumor cells like liver cancer. Developing an effective method to monitor LAP in tumor cells holds great potential for cancer diagnosis, treatment, and management. In this work, we report a novel BODIPY-based fluorescent probe (BODIPY-C-Leu) capable of monitoring LAP in vitro and in vivo in both ratiometric and turn-on model. BODIPY-C-Leu contains an asymmetrical BODIPY dye for fluorescent signaling and a dipeptide (Cys-Leu) as the triggered moiety. Activation occurs by cleavage of the amide bond in dipeptides and subsequently an intramolecular S → N conversion to convert sulfur-substituted BODIPY to amino-substituted BODIPY, resulting in a dramatic fluorescence variation to realize the detection of LAP. Furthermore, we have successfully employed BODIPY-C-Leu to monitor LAP activity in different cancer cells, indicating that HeLa cells have a higher level of LAP activity than A549 cells. Importantly, we demonstrated the capability of the probe for real-time monitoring the drug-induced LAP level changes in zebrafish.

Realizing highly chemoselective detection of H 2 S in vitro and in vivo with fluorescent probes inside core-shell silica nanoparticles

Hydrogen sulfide (H2S) is an appealing signaling molecule that plays fundamental roles in health and disease. However, H2S-mediated selective chemical transformations for the construction of imaging probes are limited, retarding the interrogation of H2S-related biological processes. Here, we present an alternative approach for engineering a new generation of efficient probes with a nonchemoselective moiety as a building block. To demonstrate our design concept, we developed a sulfoxide-functionalized BODIPY that exhibited a substantial redshift in its absorption and emission spectra upon reduction with H2S. However, such a probe also showed reactivity toward various competing biothiols under aqueous buffer conditions. To achieve high chemoselectivity, we used core-shell silica nanoparticles as an encapsulation matrix to confine the designed molecule probe within their interiors. The inherent molecular-size sieving character of the porous silica shell was capable of impeding competing biothiols from accessing the molecule probe within the core while allowing the specific reaction with the small target H2S. Thus, this strategy avoided disturbance from coexisting biothiols and achieved highly chemoselective detection in ratiometric and near-infrared (NIR) turn-on fluorescence modes. In light of these promising features, together with fast responsiveness and favorable cellular uptake, such a silica nanocomposite was successfully used to detect the endogenous production of H2S in estrogen-induced cardiomyocytes and living mouse model. To our knowledge, the approach reported here is the first to exploit the usefulness of common thiol-sensitive moieties for building chemoselective probes.

Fine Regulation of Porous Architectures of Core–Shell Silica Nanocomposites Offers Robust Nanoprobes with Accelerated Responsiveness

Probes bearing good aqueous solubility and biocompatibility as well as fast response can serve as ideal tools for evaluating the underlying molecular mechanism of endogenous production of H2S caused by drugs; however, they are still lacking but highly desirable. Here, we demonstrate a novel strategy for constructing highly efficient H2S nanoprobes through locking Förster resonance energy transfer borondipyrromethene (BODIPY) pairs in water-dispersible core-shell silica nanoparticles. Importantly, these nanocomposites can effectively confine complementary guests within the same cores due to the existence of a shield, thus guaranteeing efficient Förster resonance energy transfer. Interestingly, the interior microenvironment of such nanoparticles could be tuned by silylation agents. In this way, an ideal probe for rapid and ratiometric detection of H2S within 15 s is established by optimizing the amount of silylation agent with a polar organic group. Obviously, the silylation agents are explored to serve as a platform not only for establishment of robust structures but also for optimizing the microenvironment of the interior to afford an ideal probe. These silica nanocomposites have also been successfully employed in disclosing the endogenous production of H2S induced by estrogen in cardiomyocytes.

Hydrogen Sulfide-Activatable Second Near-Infrared Fluorescent Nanoassemblies for Targeted Photothermal Cancer Therapy

Near-infrared (NIR)-II fluorescence agents hold great promise for deep-tissue photothermal therapy (PTT) of cancers, which nevertheless remains restricted by the inherent nonspecificity and toxicity of PTT. In response to this challenge, we herein develop a hydrogen sulfide (H2S)-activatable nanostructured photothermal agent (Nano-PT) for site-specific NIR-II fluorescence-guided PTT of colorectal cancer (CRC). Our in vivo studies reveal that this theranostic Nano-PT probe is specifically activated in H2S-rich CRC tissues, whereas it is nonfunctional in normal tissues. Activation of Nano-PT not only emits NIR-II fluorescence with deeper tissue penetration ability than conventional fluorescent probes but also generates high NIR absorption resulting in efficient photothermal conversion under NIR laser irradiation. Importantly, we establish NIR-II imaging-guided PTT of CRC by applying the Nano-PT agent in tumor-bearing mice, which results in complete tumor regression with minimal nonspecific damages. Our studies thus shed light on the development of cancer biomarker-activated PTT for precision medicine.