Near-Infrared Fluorescence Probe for Indication of the Pathological Stages of Wound Healing Process and Its Clinical Application.

Chronic wound healing is one of the most complicated biological processes in human life, which is also a serious challenge for human health. During the healing process, multiple biological pathways are activated, and various kinds of reactive oxygen species participate in this process. Hydrogen peroxide (H2O2) involves in chronic wounds and its concentration is fluctuated in different pathological stages during the wound healing process. Therefore, H2O2 may be recognized as a powerful biomarker to indicate the wound healing process. However, the pathological roles of H2O2 cannot be fully understood yet. Herein, we proposed a near-infrared fluorescent probe DCM-H2O2 for highly sensitive and rapid detection of H2O2 in living cells and scald and incision wound mice models. DCM-H2O2 exhibited a low detection limit and high specificity with low cytotoxicity for H2O2, which had great potential for its application in vivo. The probe was successfully utilized to monitor the fluctuation of endogenous H2O2 in the proliferation process of human immortalized epidermal (HACAT) cells, which confirmed that H2O2 participated in the cells' proliferation activity through a growth factor signaling pathway. In the scald and incision wound mice models, H2O2 concentration fluctuations at different pathological stages during the wound healing process could be obtained by in vivo fluorescence imaging. Finally, H2O2 concentrations in different stages of human diabetic foot tissues were also confirmed by the proposed probe. We expect that H2O2 could be a sensitive biomarker to indicate the wound healing process.

Multifunctional protein-based self-assembled nanoplatform: overcoming hypoxic tumor microenvironment for enhanced imaging-guided photodynamic therapy.

Photodynamic therapy (PDT) has emerged as a promising modality for cancer treatment, but its efficacy is often limited by tumour hypoxia. Here, we report the development of a novel protein-based, self-assembled nanoplatform, CAT-I-BODIPY NPs (CIB NPs), to address this limitation. We first design and synthesize an I-BODIPY photosensitizer based on the heavy atom effect and modification of the electron-donating group, which exhibits excellent capabilities in generating reactive oxygen species and enabling near-infrared (NIR) fluorescence imaging. The incorporation of an oxygen-producing enzyme, catalase (CAT), within these nanoassemblies enables in situ oxygen generation to counteract hypoxic constraints. Controllable self-assembly by multiple supramolecular interactions into highly ordered architecture not only guarantees CAT's catalytic activity but also leads to excellent NIR fluorescence imaging ability and enhanced PDT efficacy. Notably, the visualization of optimal accumulation of CIB NPs within tumour sites 18 h post-injection offers precise PDT application guidance. Both in vitro and in vivo studies corroborate the remarkable anti-tumour efficacy of CIB NPs under NIR illumination, providing a significant advancement in PDT. The favourable biosafety profile of CIB NPs further emphasizes their potential for clinical application in hypoxic tumour therapy.

Bioimaging of glutathione variation for early diagnosis of hepatocellular carcinoma using a liver-targeting ratiometric near-infrared fluorescent probe.

Reliable biomarkers are crucial for early diagnosis of diseases and precise therapy. Biological thiols (represented by glutathione, GSH) play vital roles in the antioxidant defense system for maintaining intracellular redox homeostasis in organisms. However, the aberrant variation in the cellular concentration of GSH correlates with diverse diseases including cancer. Here, a ratiometric near-infrared fluorescent probe CyO-Disu is constructed for the specific sensing of GSH variation in live cells and mice models of hepatic carcinoma (HCC). CyO-Disu features three key elements, a response moiety of bis(2-hydroxyethyl) disulfide, a near-infrared fluorescence signal transducer of heptamethine ketone cyanine, and a targeting moiety of D-galactose. By virtue of its liver-targeting capability, CyO-Disu was utilized for evaluating GSH fluctuations in primary and metastatic hepatoma living cells. To evaluate the efficacy of CyO-Disuin vivo, orthotopic HCC and pulmonary metastatic hepatoma mice models were employed for GSH imaging using two-dimensional and three-dimensional fluorescence molecular tomographic imaging systems. The bioimaging results offered direct evidence that GSH displayed varied concentrations during the progression of HCC. Therefore, the as-synthesized probe CyO-Disu could serve as a potential powerful tool for the early diagnosis and precise treatment of HCC using GSH as a reliable biomarker.

Flexible Designing Strategy to Construct Activatable NIR-II Fluorescent Probes with Emission Maxima beyond 1200 nm.

Activatable second near-infrared (NIR-II) fluorescent probes that can be lighted up by specific targets have attracted great attention because of their high specificity and resolution, which hold great promise in deep-tissue imaging. However, such probes were relatively rarely reported so far, and the emission maximum is still limited (mainly located at 900-1000 nm). To solve the problem, herein, we proposed a flexible strategy to modulate the emission wavelength of NIR-II fluorescent probes, and four proof-of-concept probes (WH-1, WH-2, WH-3, and WH-4) based on D-π-A molecular skeleton were obtained. These probes can be activated by H2S and the emission maximum located from 925 to 1205 nm, which was attributed to the cooperation of elongating the π-conjugated system and enhancing the electron-donating ability of the donor region. In these probes, WH-3 exhibited the combination of long excitation/emission (925/1140 nm) and moderate quantum yield as well as high sensitivity toward H2S, enabling us to track and image H2S in vivo with high contrast. We expected that such a molecular design strategy will become an important approach to developing activatable NIR-II fluorescent probes with long emission.

Drosophila P75 safeguards oogenesis by preventing H3K9me2 spreading.

Serving as a host factor for human immunodeficiency virus (HIV) integration, LEDGF/p75 has been under extensive study as a potential target for therapy. However, as a highly conserved protein, its physiological function remains to be thoroughly elucidated. Here, we characterize the molecular function of dP75, the Drosophila homolog of LEDGF/p75, during oogenesis. dP75 binds to transcriptionally active chromatin with its PWWP domain. The C-terminus integrase-binding domain-containing region of dP75 physically interacts with the histone kinase Jil-1 and stabilizes it in vivo. Together with Jil-1, dP75 prevents the spreading of the heterochromatin mark-H3K9me2-onto genes required for oogenesis and piRNA production. Without dP75, ectopical silencing of these genes disrupts oogenesis, activates transposons, and causes animal sterility. We propose that dP75, the homolog of an HIV host factor in Drosophila, partners with and stabilizes Jil-1 to ensure gene expression during oogenesis by preventing ectopic heterochromatin spreading.

Design of Activatable NIR-II Molecular Probe for In Vivo Elucidation of Disease-Related Viscosity Variations.

A clear elucidation of a disease-related viscosity change in vivo is significant yet highly challenging as well. Fluorescence imaging in the second near-infrared region (NIR-II, 1000-1700 nm) has gained increasing attention for observation in living organisms, but a viscosity-activatable fluorescent probe emitting at this region remains a vacancy. Herein, we report the first panel of a viscosity-activated NIR-II emissive fluorescent probe WD-X. By embedding different substituents into the WD-X platform and screening, we obtained an ideal probe, WD-NO2, which displayed the best combination of properties, including a 31-fold fluorescence enhancement in response to viscosity, insensitivity to environments (pH, polarity), and relatively high quantum yield (1.6% in glycerol). WD-NO2 was successfully applied to track the variation of viscosity in diabetes-induced liver injury in vivo.

Simultaneous imaging of mitochondrial viscosity and hydrogen peroxide in Alzheimer's disease by a single near-infrared fluorescent probe with a large Stokes shift.

It has been speculated that both the intracellular viscosity and H2O2 level in Alzheimer's disease (AD) brains are higher than that in healthy brains, but direct evidence from living beings is scarce. Herein, we report a NIR emissive fluorescent probe with a large Stokes shift for the associated detection of mitochondrial viscosity and H2O2 in live rat brains with AD for the first time.

Probe Type II Band Alignment in One-Dimensional Van Der Waals Heterostructures Using First-Principles Calculations.

Computational tools based on density-functional theory (DFT) enable the exploration of the qualitatively new, experimentally attainable nanoscale compounds for a targeted application. Theoretical simulations provide a profound understanding of the intrinsic electronic properties of functional materials. The goal of this protocol is to search for photocatalyst candidates by computational dissection. Photocatalytic applications require suitable band gaps, appropriate band edge positions relative to the redox potentials. Hybrid functionals can provide accurate values of these properties but are computationally expensive, whereas the results at the Perdew-Burke-Ernzerhof (PBE) functional level could be effective for suggesting strategies for band structure engineering via electric field and tensile strain aiming to enhance the photocatalytic performance. To illustrate this, in the present manuscript, the DFT based simulation tool VASP is used to investigate the band alignment of nanocomposites in combinations of nanotubes and nanoribbons in the ground state. To address the lifetime of photogenerated holes and electrons in the excited state, nonadiabatic dynamics calculations are needed.

A Robust Transposon-Endogenizing Response from Germline Stem Cells.

The heavy occupancy of transposons in the genome implies that existing organisms have survived from multiple, independent rounds of transposon invasions. However, how and which host cell types survive the initial wave of transposon invasion remain unclear. We show that the germline stem cells can initiate a robust adaptive response that rapidly endogenizes invading P element transposons by activating the DNA damage checkpoint and piRNA production. We find that temperature modulates the P element activity in germline stem cells, establishing a powerful tool to trigger transposon hyper-activation. Facing vigorous invasion, Drosophila first shut down oogenesis and induce selective apoptosis. Interestingly, a robust adaptive response occurs in ovarian stem cells through activation of the DNA damage checkpoint. Within 4 days, the hosts amplify P element-silencing piRNAs, repair DNA damage, subdue the transposon, and reinitiate oogenesis. We propose that this robust adaptive response can bestow upon organisms the ability to survive recurrent transposon invasions throughout evolution.

Hijacking Oogenesis Enables Massive Propagation of LINE and Retroviral Transposons.

Although animals have evolved multiple mechanisms to suppress transposons, "leaky" mobilizations that cause mutations and diseases still occur. This suggests that transposons employ specific tactics to accomplish robust propagation. By directly tracking mobilization, we show that, during a short and specific time window of oogenesis, retrotransposons achieve massive amplification via a cell-type-specific targeting strategy. Retrotransposons rarely mobilize in undifferentiated germline stem cells. However, as oogenesis proceeds, they utilize supporting nurse cells-which are highly polyploid and eventually undergo apoptosis-as factories to massively manufacture invading products. Moreover, retrotransposons rarely integrate into nurse cells themselves but, instead, via microtubule-mediated transport, they preferentially target the DNA of the interconnected oocytes. Blocking microtubule-dependent intercellular transport from nurse cells significantly alleviates damage to the oocyte genome. Our data reveal that parasitic genomic elements can efficiently hijack a host developmental process to propagate robustly, thereby driving evolutionary change and causing disease.

Sumoylation of SUVR2 contributes to its role in transcriptional gene silencing.

The SU(VAR)-3-9-related protein family member SUVR2 has been previously identified to be involved in transcriptional gene silencing both in RNA-dependent and -independent pathways. It interacts with the chromatin-remodeling proteins CHR19, CHR27, and CHR28 (CHR19/27/28), which are also involved in transcriptional gene silencing. Here our study demonstrated that SUVR2 is almost fully mono-sumoylated in vivo. We successfully identified the exact SUVR2 sumoylation site by combining in vitro mass spectrometric analysis and in vivo immunoblotting confirmation. The luminescence imaging assay and quantitative RT-PCR results demonstrated that SUVR2 sumoylation is involved in transcriptional gene silencing. Furthermore, we found that SUVR2 sumoylation is required for the interaction of SUVR2 with CHR19/27/28, which is consistent with the fact that SUMO proteins are necessary for transcriptional gene silencing. These results suggest that SUVR2 sumoylation contributes to transcriptional gene silencing by facilitating the interaction of SUVR2 with the chromatin-remodeling proteins CHR19/27/28.

Bright and sensitive ratiometric fluorescent probe enabling endogenous FA imaging and mechanistic exploration of indirect oxidative damage due to FA in various living systems.

As a notorious toxin, formaldehyde (FA) poses an immense threat to human health. Aberrantly elevated FA levels lead to serious pathologies, including organ damage, neurodegeneration, and cancer. Unfortunately, current techniques limit FA imaging to general comparative studies, instead of a mechanistic exploration of its biological role, and this is presumably due to the lack of robust molecular tools for reporting FA in living systems. More importantly, despite being reductive, FA, however, can induce oxidative damage to organisms, thus providing a challenge to the mechanistic study of FA using fluorescence imaging. Herein, we presented the design and multi-application of a bright sensitive ratiometric fluorescent probe 1-(4-(1H-phenanthro[9,10-d]imidazol-2-yl)phenyl) but-3-en-1-amine (PIPBA). With a π-extended phenylphenanthroimidazole fluorophore and an allylamine group, PIPBA exhibited high quantum yield (φ = 0.62) in blue fluorescent emission and selective reactivity toward FA. When sensing FA, PIPBA transformed to PIBE, which is a product capable of releasing bright green fluorescence (φ = 0.51) with its enhanced intramolecular charge transfer (ICT). Transformation of PIPBA to PIBE contributed to 80 nm of red shift in emission wavelength and a highly sensitive ratiometric response (92.2-fold), as well as a quite low detection limit (0.84 µM). PIPBA was successfully applied to various living systems, realizing, for the first time, ratiometric quantification (in cells), in vivo imaging (zebrafish), and living tissue imaging (vivisectional mouse under anaesthetic) of endogenous FA that was spontaneously generated by biological systems. Furthermore, with the aid of PIPBA, we obtained visual evidence for the oxidative damage of FA in both HeLa cells and renal tissue of a living mouse. The results demonstrated that FA exerted indirect oxidative damage by interacting with free radicals, thus producing more oxidizing species, which eventually caused aggravated oxidative damage to the organism. The indirect oxidative damage due to FA could be alleviated by an exogenous or endogenous antioxidant. The excellent behaviors of PIPBA demonstrate that a chemical probe can detect endogenous FA in cells/tissue/vivo, promising to be an effective tool for further exploration of the biological mechanism of FA in living systems.

3-(2-Bromoacetamido)-N-(9-ethyl-9H)-carbazol fluorescent probe and its application for the determination of thiophenols in rubber products by HPLC with fluorescence detection and atmospheric chemical ionization mass spectrometry identification.

A rapid, sensitive, and selective precolumn derivatization method for the simultaneous determination of eight thiophenols using 3-(2-bromoacetamido)-N-(9-ethyl-9H)-carbazol as a labeling reagent by high-performance liquid chromatography with fluorescence detection has been developed. The labeling reagent reacted with thiophenols at 50°C for 50 min in aqueous acetonitrile in the presence of borate buffer (0.10 mol/L, pH 11.2) to give high yields of thiophenol derivatives. The derivatives were identified by online postcolumn mass spectrometry. The collision-induced dissociation spectra for thiophenol derivatives gave the corresponding specific fragment ions at m/z 251.3, 223.3, 210.9, 195.8, and 181.9. At the same time, derivatives exhibited intense fluorescence with an excitation maximum at λex = 276 nm and an emission maximum at λem = 385 nm. Excellent linear responses were observed for all analytes over the range of 0.033-6.66 µmol/L with correlation coefficients of more than 0.9997. Detection limits were in the range of 0.94-5.77 µg/L with relative standard deviations of less than 4.54%. The feasibility of derivatization allowed the development of a rapid and highly sensitive method for the quantitative analysis of trace levels of thiophenols from some rubber products. The average recoveries (n = 3) were in the range of 87.21-101.12%.

A novel dual-ratiometric-response fluorescent probe for SO2/ClO- detection in cells and in vivo and its application in exploring the dichotomous role of SO2 under the ClO- induced oxidative stress.

Intracellular reactive sulfur species and reactive oxygen species play vital roles in immunologic mechanism. As an emerging signal transmitter, SO2 can be generated as the anti-oxidant, while SO2 is also a potential oxidative stress-inducer in organism. Aiming to elucidate in-depth the dichotomous role of SO2 under oxidative stress, we designed a dual-response fluorescent probe that enabled the respective or successive detection of SO2 and ClO-. The probe itself emits the red fluorescence (625 nm) which can largely switch to blue (410 nm) and green fluorescence (500 nm) respectively in response to SO2 and ClO-, allowing the highly selective and accurate ratiometric quantification for both SO2 and ClO- in cells. Moreover the ultrafast (SO2: <60 s; ClO-: within sec) and highly sensitive (detection limits: SO2: 3.5 nM; ClO-: 12.5 nM) detection were achieved. With the robust applicability, the developed probe was successfully used to quantify SO2 and endogenous ClO- in respectively the HeLa cells and the RAW 264.7 cells, as well as to visualize the dynamic of SO2/ClO- in zebrafish. The fluorescent imaging studies and flow cytometry analysis confirmed the burst-and-depletion and meanwhile the oxidative-and-antioxidative effects of intracellular SO2 under the NaClO induced oxidative stress.

A two-photon ratiometric fluorescent probe for the synergistic detection of the mitochondrial SO2/HClO crosstalk in cells and in vivo.

Known as a new member of the reactive sulphur species (RSS), endogenous sulfur dioxide (SO2) plays potentially reductive roles such as antioxidation, anti-aging, anti-inflammation, and cytoprotection; however, SO2 is also a metabolite of antioxidation. Hypochlorous acid (HClO) has powerful bio-effects on the innate immune system, and its uncontrolled production leads to adverse damage in cells. To illuminate the potential crosstalk between SO2 and HClO in redox homeostasis, we designed a two-photon ratiometric fluorescent probe for dual-response to mitochondrial SO2/HClO crosstalk in cells and in vivo. Our probe can effectively achieve ratio-type fluorescence dual-response to SO2 and HClO with desirable properties such as large two-photon absorption cross sections, rapid response times (SO2 < 50 s, HClO < 20 s), high sensitivities (limit of detection 8.0 nM for SO2 and 15.2 nM for HClO), and favorable selectivity. The probe has been successfully applied to quantitatively detect endogenous SO2 and HClO in HeLa and Raw 264.7 cells. We have verified that there exists a crosstalk between SO2 and HClO during the process of oxidative stress in mitochondria. We have also detected that SO2 can be endogenously produced through oxidation of intracellular sulfur-containing amino acids by HClO in zebrafish in real-time.

Engineering the interlayer exchange coupling in magnetic trilayers.

When the thickness of metal film approaches the nanoscale, itinerant carriers resonate between its boundaries and form quantum well states (QWSs), which are crucial to account for the film's electrical, transport and magnetic properties. Besides the classic origin of particle-in-a-box, the QWSs are also susceptible to the crystal structures that affect the quantum resonance. Here we investigate the QWSs and the magnetic interlayer exchange coupling (IEC) in the Fe/Ag/Fe (001) trilayer from first-principles calculations. We find that the carriers at the Brillouin-zone center (belly) and edge (neck) separately form electron- and hole-like QWSs that give rise to an oscillatory feature for the IEC as a function of the Ag-layer thickness with long and short periods. Since the QWS formation sensitively depends on boundary conditions, one can switch between these two IEC periods by changing the Fe-layer thickness. These features, which also occur in the magnetic trilayers with other noble-metal spacers, open a new degree of freedom to engineer the IEC in magnetoresistance devices.

Ttk69 acts as a master repressor of enteroendocrine cell specification in Drosophila intestinal stem cell lineages.

In adult Drosophila midgut, intestinal stem cells (ISCs) periodically produce progenitor cells that undergo a binary fate choice determined primarily by the levels of Notch activity that they receive, before terminally differentiating into enterocytes (ECs) or enteroendocrine (EE) cells. Here we identified Ttk69, a BTB domain-containing transcriptional repressor, as a master repressor of EE cell specification in the ISC lineages. Depletion of ttk69 in progenitor cells induced ISC proliferation and caused all committed progenitor cells to adopt EE fate, leading to the production of supernumerary EE cells in the intestinal epithelium. Conversely, forced expression of Ttk69 in progenitor cells was sufficient to prevent EE cell specification. The expression of Ttk69 was not regulated by Notch signaling, and forced activation of Notch, which is sufficient to induce EC specification of normal progenitor cells, failed to prevent EE cell specification of Ttk69-depleted progenitors. Loss of Ttk69 led to derepression of the acheate-scute complex (AS-C) genes scute and asense, which then induced prospero expression to promote EE cell specification. These studies suggest that Ttk69 functions in parallel with Notch signaling and acts as a master repressor of EE cell specification in Drosophila ISC lineages primarily by suppressing AS-C genes.

SUVR2 is involved in transcriptional gene silencing by associating with SNF2-related chromatin-remodeling proteins in Arabidopsis.

The SU(VAR)3-9-like histone methyltransferases usually catalyze repressive histone H3K9 methylation and are involved in transcriptional gene silencing in eukaryotic organisms. We identified a putative SU(VAR)3-9-like histone methyltransferase SUVR2 by a forward genetic screen and demonstrated that it is involved in transcriptional gene silencing at genomic loci targeted by RNA-directed DNA methylation (RdDM). We found that SUVR2 has no histone methyltransferase activity and the conserved catalytic sites of SUVR2 are dispensable for the function of SUVR2 in transcriptional silencing. SUVR2 forms a complex with its close homolog SUVR1 and associate with three previously uncharacterized SNF2-related chromatin-remodeling proteins CHR19, CHR27, and CHR28. SUVR2 was previously thought to be a component in the RdDM pathway. We demonstrated that SUVR2 contributes to transcriptional gene silencing not only at a subset of RdDM target loci but also at many RdDM-independent target loci. Our study suggests that the involvement of SUVR2 in transcriptional gene silencing is related to nucleosome positioning mediated by its associated chromatin-remodeling proteins.

The PRP6-like splicing factor STA1 is involved in RNA-directed DNA methylation by facilitating the production of Pol V-dependent scaffold RNAs.

DNA methylation is a conserved epigenetic marker in plants and animals. In Arabidopsis, DNA methylation can be established through an RNA-directed DNA methylation (RdDM) pathway. By screening for suppressors of ros1, we identified STA1, a PRP6-like splicing factor, as a new RdDM regulator. Whole-genome bisulfite sequencing suggested that STA1 and the RdDM pathway share a large number of common targets in the Arabidopsis genome. Small RNA deep sequencing demonstrated that STA1 is predominantly involved in the accumulation of the siRNAs that depend on both Pol IV and Pol V. Moreover, the sta1 mutation partially reduces the levels of Pol V-dependent RNA transcripts. Immunolocalization assay indicated that STA1 signals are exclusively present in the Cajal body and overlap with AGO4 in most nuclei. STA1 signals are also partially overlap with NRPE1. Localization of STA1 to AGO4 and NRPE1 signals is probably related to the function of STA1 in the RdDM pathway. Based on these results, we propose that STA1 acts downstream of siRNA biogenesis and facilitates the production of Pol V-dependent RNA transcripts in the RdDM pathway.

The association between XPC Lys939Gln gene polymorphism and urinary bladder cancer susceptibility: a systematic review and meta-analysis.

BACKGROUND: Numerous epidemiological studies have been conducted to explore the association between the Lys939Gln polymorphism of Xeroderma pigmentosum group C (XPC) gene and urinary bladder cancer susceptibility. However, the results remain inconclusive. In order to derive a more precise estimation of this relationship, a large and update meta-analysis was performed in this study. METHODS: A comprehensive search was conducted through researching MEDLINE, EMBASE, PubMed, Web of Science, China Biomedical Literature database (CBM) and China National Knowledge Infrastructure (CNKI) databases before June 2013. Crude odds ratios (ORs) with 95% confidence intervals (CIs) were calculated to estimate the strength of the association. RESULTS: A total of 12 studies with 4828 cases and 4890 controls for evaluating the XPC Lys939Gln polymorphism and urinary bladder cancer were included. Overall, there was significant associations between the XPC Lys939Gln polymorphism and urinary bladder cancer risk were found for homozygous model (OR = 1.352, 95% CL = 1.088-1.681), heterozygous model (OR = 1.354, 95% CL = 1.085-1.688), and allele comparison (OR = 1.109, 95% CL = 1.013-1.214). In subgroup analysis by ethnicity and source of controls, there were still significant associations detected in some genetic models. CONCLUSION: Our meta-analysis suggested that the XPC Lys939Gln polymorphism contributed to the risk of urinary bladder cancer. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1001118393101798.