Two "turn on" fluorescence probes based on nitroso recognition group for detecting hydrogen sulfide.

Hydrogen sulfide is a vital signaling molecule which holds a pivotal position in numerous biological functions. In this research, two novel "OFF-ON" fluorescence probes named YNO and TNO were designed based on the nitroso recognition group to detect H2S. Both YNO and TNO performed outstanding response rate and linear relationship between the fluorescence intensity and the concentration of H2S. YNO possessed larger Stokes shift and longer emission wavelength. TNO had lower limit of detection. In addition, YNO was successful applied to sense endogenous and exogenous H2S and target endoplasmic reticulum (ER) in Hela cells.

Novel fluorescence probe for ClO- in living cells: Based on FRET mechanism.

Hypochlorous acid (HClO) as a kind of reactive oxygen species (ROS) plays a vital role in many biological processes. Organic fluorescence probes have attracted great interests for the detection of HClO, due to their relatively high selectivity and sensitivity, satisfactory spatiotemporal resolution and good biocompatibility. Constructing fluorescence probes to detect HClO with advantages of large Stokes shift, wide emission gap, near infrared emission and good water solubility is still challenging. In this work, a new ratiometric fluorescence probe (named HCY) for HClO was developed. FRET-based HCY was constructed by bonding a coumarin and a flavone fluorophore. In absence of HClO, HCY exists FRET process, however, FRET is inhibited in the presence of HClO because the conjugated double bond broke. Due to the good match of the emission spectrum of the donor and the absorption spectrum of the acceptor, the FRET system appears favorable energy transfer efficiency. HCY showed high sensitivity and rapid response time. The linearity between the ratios of fluorescence intensity and concentration of HClO was established with a low limit of detection. What's more, HCY was also applied for fluorescence images of HClO in RAW264.7 cells.

A near-infrared fluorescent probe for simultaneous detection of pH and viscosity.

In this work, we developed a ratiometric fluorescent probe (NT) based on ICT framework in near-infrared (NIR) which could detect pH and viscosity simultaneously. Long emission wavelength in NIR could protect the probe from interference of background fluorescence and improve the accuracy of the test. Due to the presence of thiazole-salt, the probe possessed good water solubility and could respond immediately to pH in water system. The pH values measured by NT in the actual samples were not much different from that measured by the pH meter, therefore, NT could give excellent accuracy. NT realized the reversible detection of pH by protonation and deprotonation. NT was used successfully to detect the pH of actual water samples, human serum and meat, as well as the viscosity variation caused by thickeners. Additionally, NT could monitor the changes of pH and viscosity in living cells. Therefore, the novel probe exhibited potential application in the fields of the environment, human health and food safety evaluation.

A highly sensitive ratiometric fluorescent probe for detecting HSO3-/SO32- and viscosity change based on FRET/TICT mechanism.

BACKGROUND: Sulfur dioxide (SO2) is a significant gas signaling molecule in organisms, and viscosity is a crucial parameter of the cellular microenvironment. They are both involved in regulating many physiological processes in the human body. However, abnormalities in SO2 and viscosity levels are associated with various diseases, such as cardiovascular disease, lung cancer, respiratory diseases, neurological disorders, diabetes and Alzheimer's disease. Hence, it is essential to explore novel and efficient fluorescent probes for simultaneously monitoring SO2 and viscosity in organisms. RESULTS: We selected quinolinium salt with good stability, high fluorescence intensity, good solubility and low cytotoxicity as the fluorophore and developed a highly sensitive ratiometric probe QQD to identify SO2 and viscosity changes based on Förster resonance energy transfer/twisted intramolecular charge transfer (FRET/TICT) mechanism. Excitingly, compared with other probes for SO2 detection, QQD not only identified HSO3-/SO32- with a large Stokes shift (218 nm), low detection limit (1.87 µM), good selectivity, high energy transfer efficiency (92 %) and wide recognition range (1.87-200 µM), but also identified viscosity with a 26-fold fluorescence enhancement and good linearity. Crucially, QQD was applied to detect HSO3-/SO32- and viscosity in actual water and food samples. In addition, QQD had low toxicity and good photostability for imaging HSO3-/SO32- and viscosity in cells. These results confirmed the feasibility and reliability of QQD for HSO3-/SO32- and viscosity imaging and environmental detection. SIGNIFICANCE: We reported a unique ratiometric probe QQD for detecting HSO3-/SO32- and viscosity based on the quinolinium skeleton. In addition to detecting HSO3-/SO32- and viscosity change in actual water and food samples, QQD could also monitor the variations of HSO3-/SO32- and viscosity in cells, which provided an experimental basis for further exploration of the role of SO2 derivatives and viscosity in biological systems.

A near-infrared fluorescent probe based FRET for ratiometric sensing of H2O2 and viscosity in live cells.

Hydrogen peroxide (H2O2) and viscosity play vital roles in the cellular environment as signaling molecule and microenvironment parameter, respectively, and are associated with many physiological and pathological processes in biological systems. We developed a near-infrared fluorescent probe, CQ, which performed colorimetric and ratiometric detection of H2O2 and viscosity based on the FRET mechanism, and was capable of monitoring changes in viscosity and H2O2 levels simultaneously through two different channels. Based on the specific reaction of H2O2 with borate ester, CQ exhibited a significant ratiometric response to H2O2 with a large Stokes shift of 221 nm, a detection limit of 0.87 µM, a near-infrared emission wavelength of 671 nm, a response time of 1 h, a wide detection ranges of 0.87-800 µM and a high energy transfer efficiency of 99.9 %. CQ could also recognize viscosity by the TICT mechanism, and efficiently detect viscosity changes caused by food thickeners. More importantly, CQ could successfully detect endogenous/exogenous H2O2 and viscosity in live HeLa cells, which was expected to be a practical tool for detecting H2O2 and viscosity in live cells.

NBD-based colorimetric and ratiometric fluorescent probe in NIR for bisulfite.

This work presented a FRET-ICT based fluorescent probe (named NTC) composed of coumarin-benzothiazole as the acceptor and 4-nitrobenzo[c][1,2,5] oxadiazole (NBD) as the donor for the detection of SO2 derivatives in NIR. Probe NTC possessed superior performance including selectivity, quickly response toward SO32-/HSO3- and high energy transfer efficiency (94 %). The test strips provided a simple and effective tool in detecting the presence of bisulfite. Besides, NTC was applied to test the sulfur dioxide derivatives in food samples and cells.

A ratiometric fluorescent probe based on the FRET platform for the detection of sulfur dioxide derivatives and viscosity.

BACKGROUND: Sulfur dioxide (SO2) is a common gaseous pollutant that significantly threatens environmental pollution and human health. Meanwhile, viscosity is an essential parameter of the intracellular microenvironment, manipulating many physiological roles such as nutrient transport, metabolism, signaling regulation and apoptosis. Currently, most of the fluorescent probes used for detecting SO2 derivatives and viscosity are single-emission probes or probes based on the ICT mechanism, which suffer from short emission wavelengths, small Stokes shifts or susceptibility to environmental background. Therefore, the development of powerful high-performance probes for real-time monitoring of sulfur dioxide derivatives and viscosity is of great significance for human health. RESULTS: In this research, we designed the fluorescent probe QQC to detect SO2 derivatives and viscosity based on FRET platform with quinolinium salt as donor and quinolinium-carbazole as acceptor. QQC exhibited a ratiometric fluorescence response to SO2 with a low detection limit (0.09 µM), large Stokes shift (186 nm) and high energy transfer efficiency (95 %), indicating that probe QQC had good sensitivity and specificity. In addition, QQC was sensitive to viscosity, with an 9.10-folds enhancement of orange fluorescence and an excellent linear relationship (R2 = 0.98) between the logarithm of fluorescence intensity at 592 nm and viscosity. Importantly, QQC could not only recognize SO2 derivatives in real water samples and food, but also detect viscosity changes caused by food thickeners and thereby had broad market application prospects. SIGNIFICANCE: We have developed a ratiometric fluorescent probe based on the FRET platform for detecting sulfur dioxide derivatives and viscosity. QQC could not only successfully detect SO2 derivatives in food and water samples, but also be made into test strips for detecting HSO3-/SO32- solution. In addition, the probe was also used to detect viscosity changes caused by food thickeners. Therefore, this novel probe had significant value in food and environmental detection applications.

Inhibition of annexin A7 suppresses senescence-associated heterochromatin foci formation and senescence through the AMPK/mTOR pathway in human dermal fibroblasts.

Senescence-associated heterochromatin foci (SAHF) is often used as a biological marker for senescent cells, but the regulation of its formation process is unclear. To find a new modulator of SAHF, we screened our chemical small molecules and found 7-amino-2,3,4,5-tetrahedrobenzo[b][1,4] oxazepin-3-ol (ABO) that was identified as an inhibitor of annexin A7 GTPase (ANXA7) dramatically suppressed the aggregation of heterochromatin protein (HP1γ), an indicator of SAHF. To understand its action mechanism, we first observed the changes in the karyoplasmic ratio of ANXA7 because HP1γ mainly located in the nucleus. The results showed that ABO elevated the protein level of ANXA7 in the nucleus. Therefore, we raised a hypothesis that ANXA7 interacted with HP1γ and regulated its phosphorylation, which is closely related to the formation of SAHF. The co-immunoprecipitation and Western blot experiment results showed that ANXA7 had no direct interaction with HP1γ, however, the phosphorylation of HP1γ was increased by ABO, which suggested that ANXA7 indirectly regulated HP1γ phosphorylation. Then, based on our previous discovery of ANXA7 interacting with AMP-activated protein kinase (AMPK), we investigated the effect of the AMPK/mammalian target of rapamycin (mTOR) signaling pathway on ABO-increased phosphorylation of HP1γ. We found that ABO decreased AMPK phosphorylation and increased the phosphorylation level and activity of mTOR. In the presence of an AMPK activator or mTOR inhibitor, ABO could not increase HP1γ phosphorylation. As a result, ABO inhibited the senescence of human dermal fibroblasts (HDFs). In this study, we found that ANXA7 was a new regulator of SAHF, it could regulate the formation of SAHF through the AMPK/mTOR pathway. The data suggested that ABO could be used as a powerful tool to inhibit the replicative senescence of HDFs.

A comparison of low- versus standard-dose bridging alteplase in acute ischemic stroke mechanical thrombectomy using indirect methods.

Background: Whether low-dose alteplase is similar to standard-dose bridging alteplase prior to endovascular mechanical thrombectomy in patients with acute ischemic stroke (AIS) remains uncertain. Aims: The aim of this study was to compare the efficacy and safety outcomes of low- versus standard-dose bridging alteplase therapy (BT) in patients with acute ischemic stroke (AIS) who are eligible for intravenous thrombolysis (IVT) within 4.5 h after onset. Methods: We conducted an indirect comparison of low- versus standard-dose bridging alteplase before mechanical thrombectomy in AIS of current available clinical randomized controlled trials (RCTs) that compared direct mechanical thrombectomy treatment (dMT) to BT. Primary efficacy outcomes were functional independence and excellent recovery defined as a dichotomized modified Rankin Scale (mRS) 0-2 and 0-1 at 90 days. Safety outcomes included symptomatic intracranial hemorrhage (sICH) and any intracranial hemorrhage (ICH). Results: We included six RCTs of 2334 AIS patients in this analysis, including one trial using low-dose bridging alteplase (n = 103) and five trials using standard-dose bridging alteplase (n = 1067) against a common comparator (dMT). Indirect comparisons of low- to standard-dose bridging alteplase yielded an odds ratio (OR) of 0.84 (95% CI 0.47-1.50) for 90-day mRS 0-2, 1.18 (95% CI 0.65-2.12) for 90-day mRS 0-1, 1.21 (95% CI 0.44-3.36) for mortality, and 1.11 (95% CI 0.39-3.14) for successful recanalization. There were no significant differences in the odds for sICH (OR 1.05, 95% CI 0.32-3.41) or any ICH (OR 1.71, 95% CI 0.94-3.10) between low- and standard-dose bridging alteplase. Conclusion: Indirect evidence shows that the effects of low- and standard-dose bridging alteplase are similar for key efficacy and safety outcomes. Due to the wide confidence intervals, larger randomized trials comparing low- and standard-dose alteplase bridging therapy are required.

Esterase D interacts with metallothionein 2A and inhibits the migration of A549 lung cancer cells in vitro.

Esterase D (ESD) is a nonspecific esterase widely distributed in various organisms. ESD plays an important role in regulating cholesterol efflux, inhibiting viral replication and lung cancer growth. MT2A (metallothionein 2A) is the most important isoform of metallothionein (MTs) in human and high expression of MT2A in tumors represents poor prognosis and metastatic behavior. However, there are no reports about the molecular mechanism of ESD in the regulation of tumor metastasis. In this study, we found for the first time that activation ESD promoted its interaction with MT2A and decreased the protein level of MT2A, which resulting in the concentration of free zinc ions up-regulated, and inhibited the migration of A549 lung cancer cells in vitro.

Dual-response and lysosome-targeted fluorescent probe for viscosity and sulfur dioxide derivatives.

Viscosity and sulfur dioxide levels are important factors to evaluate the changes of cell micro-environment because a series of diseases usually occur when they are abnormal. At present, dual-response probes that can detect both viscosity and sulfur dioxide are rare. Therefore, we developed a novel fluorescent probe CBN for simultaneous detection of sulfur dioxide and viscosity. Besides, probe CBN could target lysosome of which normal function will be disrupted by the abnormality of viscosity. Therefore, probe CBN has the potential to be served as an effective biological tool to monitor the intracellular micro-environment.

FRET-based fluorescent probe with favorable water solubility for simultaneous detection of SO2 derivatives and viscosity.

The intracellular viscosity is an important parameter of the microenvironment and SO2 is a vital gas signal molecule. At present, some dual-response fluorescence probes for simultaneous measurements of viscosity and SO2 derivatives (HSO3-/SO32-) possessed poor water solubility. In this work, we developed a water-soluble fluorescence probe CIJ (0.0864 g/100 mL of water at 20 °C) for simultaneous measurements of viscosity and SO2 derivatives. CIJ exhibited a sensitive fluorescence enhancement to environmental viscosity from 0.97 to 28.04 cP based on a twisted intramolecular charge transfer mechanism and was applied to effective measurement of viscosity in vitro and in vivo. CIJ could also respond to SO2 derivatives with a low detection limit (44 nM) and a fast response time (5 min) based on the nucleophilic addition reaction. Furthermore, CIJ was applied to monitor SO2 derivatives in ratiometric response manner in living cells.

Activation of GRP78 ATPase suppresses A549 lung cancer cell migration by promoting ITGB4 degradation.

Hypochlorous acid (HOCl) is an essential signal molecule in cancer cells. Activated GRP78 ATPase by a HOCl probe named ZBM-H inhibits lung cancer cell growth. However, the role and underlying mechanism of GRP78 ATPase in lung cancer cell migration have not been established. Here, we reported that activation of GRP78 ATPase by ZBM-H suppressed A549 cell migration and inhibited EMT process. Notably, ZBM-H time-dependently decreased the protein level of integrin ß4 (ITGB4) in A549 cells. Combinatorial treatment of 3BDO (an autophagy inhibitor) and ZBM-H partially rescued the protein level of ITGB4. Consistently, 3BDO partially reversed ZBM-H-inhibited cell migration. Furthermore, ZBM-H promoted the interaction between ANXA7 and Hsc70, which participated in the regulation of selective autophagy and degradation of ITGB4.

Intravenous thrombolysis prior to endovascular treatment for acute ischemic stroke: a meta-analysis.

OBJECTIVES: Whether intravenous thrombolysis provides additional benefits before direct endovascular treatment (dEVT) in acute ischemic stroke remains unclear. We aimed to compare the functional and safety outcomes of dEVT to endovascular treatment with bridging using intravenous thrombolysis (BT) in acute ischemic stroke. METHODS: This meta-analysis included currently available eligible randomized clinical trials (RCTs) by searching in the PubMed, EMBASE, Cochrane Central Register, and the International Stroke Conference and European Stroke Organisation Conference posted abstracts. RESULTS: The six included RCTs yielded 2334 participants (mean age, 69.8 years [SD, 11.4]; women, 44.3%; 1164 in dEVT group and 1170 in BT group). We found not significantly different 90-day functional outcomes of modified Rankin scale (mRS 0 - 2, odds ratio [OR] 0.93, 95%CI 0.79 - 1.09; mRS 0 - 1, OR 0.99, 95%CI 0.82 - 1.18), mortality (OR 1.08, 95%CI 0.86 - 1.35), and symptomatic intracranial hemorrhage (OR 0.72, 95%CI 0.49 - 1.07) for patients in dEVT and BT group. Patients treated with dEVT were less likely to experience successful recanalization (OR 0.72, 95%CI 0.57 - 0.92, p = 0.009) and any intracranial hemorrhage (OR 0.81, 95%CI 0.68 - 0.97, p = 0.02). There were no significant differences regarding procedural complications between the two groups. CONCLUSION: This meta-analysis showed no significant differences in 90-day functional outcomes or mortality between dEVT and BT, but a lower possibility of successful recanalization and intracranial hemorrhage for dEVT.

Activation of Esterase D by FPD5 Inhibits Growth of A549 Lung Cancer Cells via JAB1/p53 Pathway.

Esterase D (ESD) is widely distributed in mammals, and it plays an important role in drug metabolism, detoxification, and biomarkers and is closely related to the development of tumors. In our previous work, we found that a chemical small-molecule fluorescent pyrazoline derivative, FPD5, an ESD activator, could inhibit tumor growth by activating ESD, but its molecular mechanism is still unclear. Here, by using RNA interference (RNAi), andco-immunoprecipitation techniques, we found that ESD suppressed the nucleus exportation of p53 through reducing the interaction between p53 and JAB1. The protein level of p53 in the nucleus was upregulated and the downstream targets of p53 were found by Human Gene Expression Array. p53 inhibited the expression of CDCA8 and CDC20. Lastly, the cell cycle of A549 cells was arrested at the G0/G1 phase. Together, our data suggest that ESD inhibited the cancer cell growth by arresting the cell cycle of A549 cells via the JAB1/p53 signaling pathway. Our findings provide a new insight into how to inhibit the growth of lung cancer with the activation of ESD by FPD5.

Co-delivery of Docetaxel and Resveratrol by liposomes synergistically boosts antitumor efficiency against prostate cancer.

Combination therapy is frequently used in cancer treatments. Delivery of combined anticancer agents loaded in a nanocarrier would be a promising option for combination therapy. Here, we designed PEGylated nano-liposomes for co-delivery Docetaxel (Doc) and Resveratrol (Res) to evaluate antitumor efficiency of the combined drugs in prostate cancer. The average diameter of the liposomes was 99.67 nm with a spheral-like shape. Drug release studies showed that both drugs could synchronously leak from the liposomes in a sustained release behavior. Cellular uptake results demonstrated that liposomes could effectively deliver more cargos into cells than other formulations. Moreover, co-loaded liposomes with Doc/Res in a molar ratio of 1:2 exhibited significantly higher cytotoxicity than a mixed solution containing both drugs on cancer cells. In the study of caspase 3, we found that the combination of Doc and Res could significantly increase the activity of caspase 3 enzyme compared with Doc alone. Animal studies revealed that co-encapsulated Doc/Res in liposomes predominantly inhibited tumor growth in PC3 bearing Balb/c nude mice, as evidenced by a change in cell proliferation and apoptosis parameters. Importantly, little toxicities and prolonged survival time were observed in mice treated with liposome-loaded Doc/Res than control group exposed to liposome-free Doc/Res. These results provided evidence that loading of Doc/Res in a nano-liposome is an efficient delivery formulation for synergistic treating prostate cancer.

ZBM-H-induced activation of GRP78 ATPase promotes apoptosis via annexin A7 in A549 lung cancer cells.

Hypochlorous acid (HOCl) is an essential signal for the regulation of cancer cell fate, including autophagy and apoptosis. HOCl regulated autophagy by affecting the oxidation modification of glucose-regulated protein 78 (GRP78) and the activity of GRP78 ATPase. The mechanism of GRP78 ATPase in cell apoptosis has however not yet been clarified. Here we reported that ZBM-H, as a probe of HOCl, was able to directly bind to GRP78 in the presence or absence of ATP. Following ZBM-H treatment, the interaction between GRP78 and annexin A7 (ANXA7) was promoted, and this was accompanied by increased phosphorylation of integrin ß4 (ITGB4). In addition, ZBM-H enhanced the phosphorylation of ANXA7. ABO, an inhibitor of ANXA7, inhibited ZBM-H-induced ITGB4 phosphorylation and apoptosis, while ANXA7 activator SEC had opposite effect. Collectively, these data provide new evidence for the mechanism by which ZBM-H-induced activation of GRP78 ATPase regulates apoptosis of A549 lung cancer cells.

A novel sulfur dioxide probe inhibits high glucose-induced endothelial cell senescence.

Sulfur dioxide (SO2) is an important gas signal molecule produced in the cardiovascular system, so it has an important regulatory effect on human umbilical vascular endothelial cells (HUVECs). Studies have shown that high glucose (HG) has become the main cause of endothelial dysfunction and aging. However, the mechanism by which SO2 regulates the senescence of vascular endothelial cells induced by HG has not yet been clarified, so it is necessary to find effective tools to elucidate the effect of SO2 on senescence of HUVECs. In this paper, we identified a novel sulfur dioxide probe (2-(4-(dimethylamino)styryl)-1,1,3-trimethyl-1H-benzo [e]indol-3-ium, DLC) that inhibited the senescence of HUVECs. Our results suggested that DLC facilitated lipid droplets (LDs) translocation to lysosomes and triggered upregulation of LAMP1 protein levels by targeting LDs. Further study elucidated that DLC inhibited HG-induced HUVECs senescence by promoting the decomposition of LDs and protecting the proton channel of V-ATPase on lysosomes. In conclusion, our study revealed the regulatory effect of lipid droplet-targeted sulfur dioxide probes DLC on HG-induced HUVECs senescence. At the same time, it provided the new experimental evidence for elucidating the regulatory mechanism of intracellular gas signaling molecule sulfur dioxide on vascular endothelial fate.

Two simple but effective turn-on benzothiazole-based fluorescent probes for detecting hydrogen sulfide in real water samples and HeLa cells.

Two simple turn-on fluorescent probes, containing a benzothiazole and the 2,4-dinitrobenzenesulfonyl group, were designed for detecting H2S. Two probes exhibited good selectivity and high sensitivity, which were applied to detect the H2S in real water samples. Probe P2 with a positive charge had better solubility than probe P1 in water; therefore, probe P2 was successfully applied to detect both the endogenous and exogenous H2S in lysosomes of living HeLa cells.

Esterase D stabilizes FKBP25 to suppress mTORC1.

BACKGROUND: Esterase D (ESD) is a nonspecific esterase that detoxifies formaldehyde. Many reports have stated that ESD activity is associated with a variety of physiological and pathological processes. However, the detailed signaling pathway of ESD remains poorly understood. METHODS: Considering the advantages of the small chemical molecule, our recent work demonstrated that 4-chloro-2-(5-phenyl-1-(pyridin-2-yl)-4,5-dihydro-1H-pyrazol-3-yl) phenol (FPD5) activates ESD, and will be a good tool for studying ESD further. Firstly, we determined the interaction between ESD and FK506 binding protein 25 (FKBP25) by yeast two-hybrid assay and co-immunoprecipitation (CO-IP) and analyzed the phosphorylation levels of mTORC1, P70S6K and 4EBP1 by western blot. Furthermore, we used the sulforhodamine B (SRB) and chick chorioallantoic membrane (CAM) assay to analyze cell viability in vitro and in vivo after treatment with ESD activator FPD5. RESULTS: We screened FKBP25 as a candidate protein to interact with ESD by yeast two-hybrid assay. Then we verified the interaction between ESD and endogenous FKBP25 or ectopically expressed GFP-FKBP25 by CO-IP. Moreover, the N-terminus (1-90 aa) domain of FKBP25 served as the crucial element for their interaction. More importantly, ESD reduced the K48-linked poly-ubiquitin chains of FKBP25 and thus stabilized cytoplasmic FKBP25. ESD also promoted FKBP25 to bind more mTORC1, suppressing the activity of mTORC1. In addition, ESD suppressed tumor cell growth in vitro and in vivo through autophagy. CONCLUSIONS: These findings provide novel evidence for elucidating the molecular mechanism of ESD and ubiquitination of FKBP25 to regulate autophagy and cancer cell growth. The ESD/FKBP25/mTORC1 signaling pathway is involved in inhibiting tumor cell growth via regulating autophagy.