Naphthalimide-based a highly selective two-photon fluorescent probe for imaging of hydrogen sulfide in living cells and inflamed tissue of mouse model

doi:10.1016/j.saa.2020.118886

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy

Volume 245, 15 January 2021, 118886

https://doi.org/10.1016/j.saa.2020.118886 Get rights and content

Highlights

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Naphthalimide-based a TP fluorescent probe NP-H₂S for H₂S imaging.
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NP-H₂S has high selectivity and sensitivity to H₂S over other analytes.
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Moreover, NP-H₂S can quickly image H₂S in inflamed tissues.

Abstract

Hydrogen sulfide (H₂S) is a very important third endogenously generated gaseous signaling molecule and plays a key role in physiological and pathological regulation processes of living biosystems. Although a lot of H₂S fluorescent probes have been reported, the relationship between the physiology and pathology of H₂S in inflamed tissues remains unclear. Herein, by adopting a donor-π-acceptor (D-π-A)-structured naphthalimide derivative as the two-photon (TP) fluorophore and a 4-dinitrobenzene-ether (DNB) with a strong intramolecular charge transfer (ICT) effect as the recognition moiety, we reported a novel TP bioimaging probe NP-H₂S for H₂S with improved sensitivity. The NP-H₂S exhibits very low background fluorescence in the absence of H₂S, and a significant 258-fold fluorescence intensity enhancement was observed in the presence of H₂S, resulting in a high sensitivity and selectivity to H₂S in aqueous solutions with a detection limit of 18.8 nM observed. The probe also shows a wide linear response concentration range (0–10.0 μM) to H₂S with high selectivity. All these features are favorable for direct monitoring of H₂S in complex biological samples. It was then applied for direct TP imaging of H₂S in tissues of inflammation model with satisfactory sensitivity, indicating it has the latent capability in further biological applications for investigation of the interaction H₂S with inflammation.

Graphical abstract

Introduction

Currently investigation shows H₂S is a 3rd gas signal molecule following the two well-known gaseotransmitters: nitric oxide (NO) and carbon monoxide (CO), which is catalyzed by certain enzymes (such as cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE)), and plays a very important role in regulating the physiology and pathology of biosystems [1,2]. For example, hydrogen sulfide can reduce mucosal inflammation, protect gastrointestinal mucosa from injury, augment tissue repair, etc. [[3], [4], [5], [6]], and can also effectively remove reactive oxygen/nitrogen species (ROS/RNS) [[7], [8], [9]], such as hydrogen peroxide, superoxide anion, hypochlorous acid, peroxynitrite, etc. Studies show that H₂S can effectively reduce amyloid protein (AP)-induced neurotoxicity [10]. H₂S and fasting plasma glucose (FPG) show a negative proportional relationship, and increasing the concentration of hydrogen sulfide in patients with type 2 diabetes may reduce blood glucose [11,12]. However, the disorder of H₂S in the biosystems will cause arterial and pulmonary hypertension, Alzheimer's disease, gastric mucosal damage and liver cirrhosis and other diseases [13]. Therefore, the significance of the detection of H₂S in biosystems is extremely important.

According to related studies, H₂S has a physiological and pathological relationship with inflammation [14]. Inflammation can be divided into two main groups: infectious and non-infectious, which is often manifests as redness, swelling, heat, pain, dysfunction, etc. and mainly through the defensive response of living tissues of the vascular system to injury factors. Therefore, the vascular response and leakage is the central link of the inflammatory process. Moreover, usually, inflammation is beneficial to the biosystems and is the biosystems' automatic defense response, but sometimes, inflammation is also harmful to living biosystems, such as attacks on the biosystems' own tissues, inflammation that occurs in transparent tissues, and so on. However, the physiological and pathological mechanisms of H₂S and inflammation are still unclear. Therefore, it is of great significance to study the relationship between H₂S and inflammation in living biosystems.

In the past few years, some analytical methods were developed for H₂S detection, but they are not suitable for in situ analysis [15,16]. Compared with them, small molecule fluorescent probe-based methods could maintain comparable accuracy and efficiency, provide convenience, high selectivity and sensitivity, noninvasive, and real-time imaging [17,18]. A more viable choice would adopt two-photon (TP) probe-based fluorescent imaging, which is an emerging technique that uses near-infrared light excitation to achieve deep-tissue imaging with prolonged observation time [19]. Therefore, it is of great significance to construct some TP fluorescent probes with excellent biocompatibility for the detection and imaging of H₂S in living cells and living tissues. Especially, to explore the physiology and pathology of H₂S in inflammation model is urgently needed. In this work, based-on the naphthalimide fluorophore, we constructed a simple TP fluorescent probe with good biocompatibility and excellent optical properties for detecting and imaging of H₂S in living cells and inflammation model of tissues. In this new scaffold, a naphthalimide as the TP fluorophore, and the 4-dinitrobenzene-ether (DNB) with a strong ICT effect as H₂S responsive group. The fluorescence intensity was quenched by the DNB group, and upon reaction with H₂S, inducing the cleavage of DNB moiety to be left, and resulting in the release of the TP naphthalimide fluorophore to show a “turn-on” fluorescent signal at 540 nm when excited at 400 nm, and exhibited a more than 258-fold fluorescence intensity enhancement. It also showed the probe displayed high selectivity towards H₂S over other analytes. Subsequently the probe was satisfactorily applied to the detection and imaging of gaseous signal molecule H₂S in living tissues of inflammation model.

Section snippets

Synthesis of H₂S fluorescent probe NP-H₂S

NP-H₂S was synthesized following the procedure shown in Fig. 1A. 2 and 3 were synthesized according to previously reported methods [20]. Synthesis of NP-H₂S: A mixture of 3 (1.0 mmol, 372 mg), commercial 2,4-dinitrofluorobenzene (1.0 mmol, 186 mg), 3 mL triethylamine, and 40 mL CH₂Cl₂ (DCM) was added to a round bottom flask with reflux condenser and stirred at 50 °C for 4 h. After the reaction mixture was cooled to room temperature, the solvent was removed with a rotary evaporator, which was

Design and synthesis of TP fluorescent probe NP-H₂S

In order to study the imaging study of H₂S in the mouse of the inflamed tissue model. Herein, we intended to design and synthesize a new TP fluorescence probe NP-H₂S monitoring H₂S in living mouse of the inflamed tissue, as showed in Fig. 1. Specifically, a naphthalimide derivative as the TP fluorophore and a DNB with a strong ICT effect as H₂S “turn-on” responsive group. Such a scaffold affords this new TP fluorescent probe NP-H₂S is an effective probe for staining inflamed tissues with NIR

Conclusions

In summary, we have constructed a new TP fluorescent probe, termed NP-H₂S. For this probe, naphthalimide acts as a TP signaling group (fluorophore) with a hydroxyl group, when the -OH was modified with a DBN moiety as a H₂S recognition site, in this new scaffold affords the probe NP-H₂S response fast, sensitively and high selectivity towards H₂S in living cells and tissues. More importantly, NP-H₂S could be used for H₂S imaging in mouse inflamed tissue with TP excitation. Therefore, we expect

CRediT authorship contribution statement

Dr. Pinghua Ou received her Master degree in 2009 from Capital Medical University. Currently, he works as a doctor in department of stomatology, the third Xiangya hospital, Central South University, China. Her research interests focus mainly on the applied research of new materials and new methods.

Yali Wang she works as a professor in a doctor in department of stomatology, the third Xiangya hospital, Central South University, China. Her research interests focus mainly on the stomatology.

Cong Hao

Declaration of competing interest

We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of, the manuscript entitled.

Acknowledgments

This work was supported by the Natural Science Foundation of Hunan Province (No. 2020JJ5862) and Key Research and Development Project of Hunan Province (2019NK2101).

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Naphthalimide-based a highly selective two-photon fluorescent probe for imaging of hydrogen sulfide in living cells and inflamed tissue of mouse model

Highlights

Abstract

Graphical abstract

Introduction

Section snippets

Synthesis of H2S fluorescent probe NP-H2S

Design and synthesis of TP fluorescent probe NP-H2S

Conclusions

CRediT authorship contribution statement

Declaration of competing interest

Acknowledgments

Biosens. Bioelectron.

Nitric Oxide

Biochim. Biophys. Acta

Chem. Sci.

Sensors Actuators B Chem.

J. Hazard. Mater.

A lysosome-targetable fluorescent probe for imaging hydrogen sulphide in living cells

Org. Lett.

Mitochondrial H2S production in Parkinson’s disease gene knockout astrocytes

J. Am. Chem. Soc.

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J. Mol. Med.

Design of a novel composite H2S-releasing hydrogel for cardiac tissue repair

Macromol. Biosci.

Synthesis of H₂S fluorescent probe NP-H₂S

Design and synthesis of TP fluorescent probe NP-H₂S

Design of a novel composite H₂S-releasing hydrogel for cardiac tissue repair