Cell-Trappable BODIPY-NBD Dyad for Imaging of Basal and Stress-Induced H₂S in Live Biosystems

Abstract

H2S is a gaseous signaling molecule that is involved in many physiological and pathological processes. In general, the level of intracellular H2S (<1 μM) is much lower than that of GSH (∼1–10 mM), leading to the remaining challenge of selective detection and differentiation of endogenous H2S in live biosystems. To this end, we quantitatively demonstrate that the thiolysis of NBD amine has much higher selectivity for H2S over GSH than that of the reduction of aryl azide. Subsequently, we developed the first NBD-based cell-trappable probe 1 (AM-BODIPY-NBD) for highly selective and ultrasensitive imaging of intracellular H2S. Probe 1 demonstrates a 207-fold fluorescence enhancement at 520 nm after reaction with H2S/esterase to produce a bright BODIPY (quantum yield 0.42) and a detection limit of 15.7 nM. Probe 1 is water-soluble, cell-trappable, and not cytotoxic. Based on this excellent chemical tool, relative levels of basal H2S in different cell lines were measured to reveal a positive correlation between endogenous H2S and the metastatic potential of colon and breast cancer cells. In addition, H2S biogenesis in vivo was also validated by probe 1 both in tobacco leaves under viral infection and in zebrafish after tail amputation. It is anticipated that probe 1 will have widespread applications in imaging and for investigating different H2S-related biological processes and diseases.

Small‐Molecule Quenchers for Förster Resonance Energy Transfer: Structure, Mechanism, and Applications

“…AlexaFluor 488 (72 000; 516)/fluorescein Detect bromo-benzophenanthridinone 968 on glutaminase C (GAC) [74] /liposome [77 Detect cathepsins L, K or S, [62] caspase-3, [63] MMP-2, [65] cysteine cathepsin (CTS), [69] cathepsin S, [70] intracellular hypoxia [79] /fusion proteins [68] /pH [71] /β-Lactamase (BlaC) [73] /Glucose [82] Quantify the degradation of antibodies [91] /detect thrombin, [92] miRNA, [93] β-lactamase, [96] caspase-3 [99] /enzyme [94] /metalloprotease ADAMTS13 [95] /2-deoxyglucose (2-DG) [97] Monitor the activity of acid sphingomyelinase (ASM) [100] /ASM activity, [101] detect H 2 S [103][104][105][106][107] / influenza virus-mediated membrane fusion [109] /Phospholipase A2 (PLA2) [116] /sense the polymer conformational changes [119] DYQ Monitor the translocation of the deacetylated tRNA [125] /investigate the binding sites' changes of the N-and C-terminus of initiation factor 3 (IF3) on small ribosomal subunit (30S) platform [126] /label mRNA [127] /tRNA [128] ATTO 575Q 582 Detect retroviruses Human T-cell leukemia virus type 1 (HTLV-1) and HIV-1 simultaneously [133] /monitor minimal residual disease (MRD) [134] /rapid detection of SARS-CoV-2 [135] BMN-Q590 CBG-488/thio-BODIPY/FAM Manipulate protein functions in living cells [68] /detect GSH…”

Small‐Molecule Quenchers for Förster Resonance Energy Transfer: Structure, Mechanism, and Applications

“…AlexaFluor 488 (72 000; 516)/fluorescein Detect bromo-benzophenanthridinone 968 on glutaminase C (GAC) [74] /liposome [77 Detect cathepsins L, K or S, [62] caspase-3, [63] MMP-2, [65] cysteine cathepsin (CTS), [69] cathepsin S, [70] intracellular hypoxia [79] /fusion proteins [68] /pH [71] /β-Lactamase (BlaC) [73] /Glucose [82] Quantify the degradation of antibodies [91] /detect thrombin, [92] miRNA, [93] β-lactamase, [96] caspase-3 [99] /enzyme [94] /metalloprotease ADAMTS13 [95] /2-deoxyglucose (2-DG) [97] Monitor the activity of acid sphingomyelinase (ASM) [100] /ASM activity, [101] detect H 2 S [103][104][105][106][107] / influenza virus-mediated membrane fusion [109] /Phospholipase A2 (PLA2) [116] /sense the polymer conformational changes [119] DYQ Monitor the translocation of the deacetylated tRNA [125] /investigate the binding sites' changes of the N-and C-terminus of initiation factor 3 (IF3) on small ribosomal subunit (30S) platform [126] /label mRNA [127] /tRNA [128] ATTO 575Q 582 Detect retroviruses Human T-cell leukemia virus type 1 (HTLV-1) and HIV-1 simultaneously [133] /monitor minimal residual disease (MRD) [134] /rapid detection of SARS-CoV-2 [135] BMN-Q590 CBG-488/thio-BODIPY/FAM Manipulate protein functions in living cells [68] /detect GSH…”

Thiobenzophenones: tunable hydrolysis-based donors for intracellular H2S delivery

“…3(a), the thioketones can be directly prepared by treatment of the required ketones with LR, 12 which readily enables the access to large quantities of the donors. After preparation, the structures of thioketones were confirmed by 1 H NMR, 13 C NMR, and HRMS (see ESI † for details). The peaks of 226.2 ppm and 233.4 ppm in 13 C NMR are assigned to CQS from compounds 1 and 2, respectively.…”

“…After preparation, the structures of thioketones were confirmed by 1 H NMR, 13 C NMR, and HRMS (see ESI † for details). The peaks of 226.2 ppm and 233.4 ppm in 13 C NMR are assigned to CQS from compounds 1 and 2, respectively. The data imply that the electronic structures of thiobenzophenones can be tuned by substituent groups, which may influence the hydrolysis reactivity and H 2 S-release rate.…”

“…To investigate the H 2 S delivering capacity of donor 1 in cellular environments, the H 2 S release in HeLa cells was visualized by a cell-trappable H 2 S probe AM-BODIPY-NBD . 13 The probe can be converted into strong fluorescence BODIPY-PZ upon reaction with H 2 S and has cell-retention ability after reaction with endogenous esterase (Fig. 4(a)).…”