Seeking eye protection from biomass: Carbon dot-based optical blocking films with adjustable levels of blue light blocking.

The intensity of blue light in white light emitting diodes is typically higher than that of the green and red-light components in screen displays and lighting systems. To reduce the potential harm of in white light emitting diodes to the eyes, in this paper, we have used microcrystalline cellulose to synthesize biomass-based carbon dots (Bio-CD), which not only absorb short wavelength light to produce longer wavelength emissions, but also show concentration-dependent maximum excitation and maximum emission. The Bio-CDs were mixed with polyvinyl alcohol (PVA) to produce optical blocking films (OBF) that preferentially block blue light. OBFs have good transparency and also block blue light effectively. With OBFs containing 9.9% of Bio-CDs, the film blocked 99.6% and 98.6% of 395 nm light and 450 nm light respectively, and also blocked 93.4% and 97%, respectively, of the blue light emitted by computers and mobile phone screens. OBFs containing more than 9.9% Bio-CDs block blue light more than commercially available blue light blocking glasses. By adjusting the amount of Bio-CDs in the OBFs, it is possible to produce films with different degrees of blue light blocking to meet the requirements of different applications.

Biomass-Based Polymer Nanoparticles With Aggregation-Induced Fluorescence Emission for Cell Imaging and Detection of Fe3+ Ions.

Polymeric nanoparticles, which show aggregation-induced luminescence emission, have been successfully prepared from larch bark, a natural renewable biomass resource, in a simple, rapid ultrasonic fragmentation method. The structure, element, particle size and molecular weight distribution of larch bark extracts (LBE) were studied by FTIR, XPS, TEM, XRD and linear mode mass spectrometry, respectively. LBE was found containing large numbers of aromatic rings, displaying an average particle size of about 4.5 nm and mainly presenting tetramers proanthocyanidins. High concentration, poor solvent, low temperature and high viscosity restricted the rotation and vibration of the aromatic rings in LBE, leading to the formation of J-aggregates and enhancing the aggregation-induced fluorescence emission. LBE possessed good resistance to photobleaching under ultraviolet light (200 mW/m2). Cytotoxicity experiments for 24 h and flow cytometry experiments for 3 days proved that even the concentrations of LBE as high as 1 mg/mL displayed non-toxic to MG-63 cells. Therefore, LBE could be employed for MG-63 cell imaging, with similar nuclear staining to the DAPI. The effects of different metal ions on the fluorescence emission intensity of LBE were analyzed and exhibited that Fe3+ owned obvious fluorescence quenching effect on LBE, while other metal ions possessed little or weak effect. Furthermore, the limit of detection (LOD) of Fe3+ was evaluated as 0.17 µM.

Preparation of Biomass-Based Carbon Dots with Aggregation Luminescence Enhancement from Hydrogenated Rosin for Biological Imaging and Detection of Fe3.

Fluorescent carbon dots (CDs) have numerous important applications, but enhancing the fluorescence emission and overcoming fluorescence quenching are still big challenges. Here, fluorescence-enhanced carbon dots (named hr-CDs) were prepared from sustainable hydrogenated rosin, using a simple hydrothermal method in a water solvent. The hr-CDs were mainly composed of graphitized carbon cores with surface functional groups. With the increase in the concentration to hr-CDs aqueous solutions, the distance between the carbon cores decreased, which resulted in the formation of J aggregates and the enhanced blue fluorescence emission. Even in the solid state, the hr-CDs show fluorescence emission because the surface functional groups could prevent π-π stacking interactions between the carbon cores. The hr-CDs show excellent resistance to photobleaching under intense ultraviolet light (200 mW/cm2). Vibrations and rotations of graphitized carbon core are restricted by low temperature and high viscosity, leading to increased radiative transition and thus increase in fluorescence intensity. The pH value in the range of 3.99-9.87 and anions have little effect on the fluorescence emission of hr-CDs. The fluorescence emission of the hr-CDs was selectively quenched by Fe3+ and can thus be used to detect Fe3+. The hr-CDs also have good biocompatibility and show the same ability in cell nuclear staining as 4',6-diamidino-2-phenylindole (DAPI).

NPM1c impedes CTCF functions through cytoplasmic mislocalization in acute myeloid leukemia.

Normal cytogenetic acute myeloid leukemia (AML) frequently harbor a TCTG insertion in exon 12 of Nucleophosmin 1 (NPM1); the resulting frameshift creates a nuclear export signal (NES) and cytoplasmic localization of NPM1c. However, how NPM1c causes AML is not completely understood. NPM1 participates in multiple protein-protein interactions one of which involves the CCCTC-binding factor (CTCF). Through binding of CTCF binding sites (CBS), CTCF mediates nuclear functions including DNA looping, regulation of gene expression, and RNA splicing. We hypothesized that mislocalization of CTCF into the cytoplasm by NPM1c reduces the functional level of nuclear CTCF and so alters gene expression. We verified the interaction of CTCF with NPM1 and showed that CTCF interacts with NPM1c, with redistribution of CTCF into the cytoplasm. The interaction of CTCF and NPM1c involves the amino terminus of CTCF and the last 50 amino acids of NPM1. By interfering with the interaction of CTCF and NPM1c, CTCF becomes relocalized into the nucleus.

SubID, a non-median dichotomization tool for heterogeneous populations, reveals the pan-cancer significance of INPP4B and its regulation by EVI1 in AML.

Our previous studies demonstrated that INPP4B, a member of the PI3K/Akt signaling pathway, is overexpressed in a subset of AML patients and is associated with lower response to chemotherapy and shorter survival. INPP4B expression analysis in AML revealed a right skewed frequency distribution with 25% of patients expressing significantly higher levels than the majority. The 75% low/25% high cut-off revealed the prognostic power of INPP4B expression status in AML, which would not have been apparent with a standard median cut-off approach. Our identification of a clinically relevant non-median cut-off for INPP4B indicated a need for a generalizable non-median dichotomization approach to optimally study clinically relevant genes. To address this need, we developed Subgroup Identifier (SubID), a tool which examines the relationship between a continuous variable (e.g. gene expression), and a test parameter (e.g. CoxPH or Fisher's exact P values). In our study, Fisher's exact SubID was used to reveal EVI1 as a transcriptional regulator of INPP4B in AML; a finding which was validated in vitro. Next, we used CoxPH SubID to conduct a pan-cancer analysis of INPP4B's prognostic significance. Our analysis revealed that INPP4Blow is associated with shorter survival in kidney clear cell, liver hepatocellular, and bladder urothelial carcinomas. Conversely, INPP4Blow was shown to be associated with increased survival in pancreatic adenocarcinoma in three independent datasets. Overall, our study describes the development and application of a novel subgroup identification tool used to identify prognostically significant rare subgroups based upon gene expression, and for investigating the association between a gene with skewed frequency distribution and potentially important upstream and downstream genes that relate to the index gene.

Suspected leukemia oncoproteins CREB1 and LYL1 regulate Op18/STMN1 expression.

Stathmin (STMN1) is a microtubule destabilizing protein with a key role in cell cycle progression and cell migration that is up-regulated in several cancers and may contribute to the malignant phenotype. However, the factors that regulate its expression are not well understood. Loss as well as gain-of-function p53 mutations up-regulate STMN1 and in acute myelogenous leukemia where p53 is predominantly wild-type, STMN1 is also over-expressed. Here we show regulatory control of STMN1 expression by the leucine zipper transcription factor (TF) CREB1 and the basic helix-loop-helix TF LYL1. By ChIP-chip experiments we demonstrate in vivo the presence of LYL1 and CREB1 in close proximity on the STMN1 promoter and using promoter assays we reveal co-regulation of STMN1 by CREB1 and LYL1. By contrast, TAL1, another suspected oncoprotein in leukemia and close relative of LYL1, exerts no regulatory effect on the STMN1 promoter. NLI, LMO2 and GATA2 are previously described co-activators of Tal1/Lyl1-E47 transcriptional complexes and potentiate Lyl1 activation of the STMN1 promoter while having no effect on TAL1 transactivation. Promoter mutations that abrogate CREB1 proximal binding or mutations of the DNA-binding domain of CREB1 abolish LYL1 transcriptional activation. These results show that CRE and Ebox sites function as coordinated units and support previous evidence of joint CREB1-and LYL1 transcription events activating an aberrant subset of promoters in leukemia. CREB1 or LYL1 shRNA knock-down down-regulate STMN1 expression. Because down-regulation of STMN1 has been shown to have anti-proliferative effects, while CREB1 and LYL1 are suspected oncoproteins, interference with CREB1-LYL1 interactions may complement standard chemotherapy and yield additional beneficial effects.

FCHSD2 predicts response to chemotherapy in acute myeloid leukemia patients.

Acute myeloid leukemia (AML) is characterized by the growth and accumulation of cells blocked in their ability to differentiate, and blocks production of normal blood cells. The response to induction chemotherapy is heterogeneous, therefore biomarkers that predict for the outcome of such treatment are of potential value. FCHSD2 in a sensitized screen was identified as a potential chemo-protector (TW Mak, unpublished). In the present study, we found that FCHSD2 knockdown by shRNA could enhance chemosensitivity of U937 cells. This coincided with the increased expression of p21 and PUMA as well as the decreased expression of Bcl-2, c-myc and hTERT. In contrast, over-expression of FCHSD2 significantly increased cellular chemoresistance. To see if there was potential clinical relevance of FCHSD2 expression in leukemia we used qRT-PCR to assess FCHSD2 expression levels in peripheral blood or bone marrow blasts of 71 AML patients. There was an inverse correlation between the level of FCHSD2 with overall survival time (r=-0.7647, p<0.0001) and relapse free time (r=-0.8165, p<0.0001). By dividing patients into high and low expression groups using a FCHSD2 expression threshold value of 0.001, the median survival of the high expression group (72 days) was shorter than in the low expression group (2472 days). The average FCHSD2 expression level in 41 patients with complete remission was significantly lower than that in 30 non-responder patients (p<0.0001). Moreover, in 32 de novo AML patients receiving initial remission-induction chemotherapy, we confirmed that the patients with lower FCHSD2 expression prior to the treatment had an increased chance of attaining remission compared to patients with high level FCHSD2. In conclusion, our study, for the first time, demonstrates FCHSD2 as a predictor of outcome for AML patient. The determination of FCHSD2 expression at the time of diagnosis could help to identify the responses of AML patients to chemotherapy.

Wilms' tumor 1 suppressor gene mediates antiestrogen resistance via down-regulation of estrogen receptor-alpha expression in breast cancer cells.

The antiestrogen tamoxifen has been used in the treatment of hormone-responsive breast cancer for over a decade. The loss of estrogen receptor (ER) expression is the most common mechanism for de novo antiestrogen resistance. Wilms' tumor 1 suppressor gene (WT1) is a clinically useful marker that is associated with poor prognosis in breast cancer patients; its high level expression is frequently observed in cases of breast cancer that are estrogen and progesterone receptor negative. The lack of expression of these receptors is characteristic of tumor cells that are not responsive to hormonal manipulation. To determine whether there is a linkage between WT1 expression and antiestrogen resistance in breast cancer cells, we studied the effect of WT1 on tamoxifen responsiveness in ERalpha-positive MCF-7 cells. We found that overexpression of WT1 in MCF-7 markedly abrogated tamoxifen-induced cell apoptosis and 17beta-estradiol (E(2))-mediated cell proliferation. The expressions of ERalpha and its downstream target genes were significantly repressed following overexpression of WT1, whereas the down-regulation of WT1 by WT1 shRNA could enhance ERalpha expression and the sensitivity to tamoxifen treatment in ERalpha-negative MDA468 and HCC1954 cells that express high levels of WT1. Furthermore, we have confirmed that the WT1 protein can bind to endogenous WT1 consensus sites in the proximal promoter of ERalpha and thus inhibit the transcriptional activity of the ERalpha promoter in a WT1 site sequence-specific manner. Our study clearly implicates WT1 as a mediator of antiestrogen resistance in breast cancer through down-regulation of ERalpha expression and supports the development of WT1 inhibitors as a potential means of restoring antiestrogen responsiveness in breast cancer therapy.

Lyl1 interacts with CREB1 and alters expression of CREB1 target genes.

The basic helix-loop-helix (bHLH) transcription factor family contains key regulators of cellular proliferation and differentiation as well as the suspected oncoproteins Tal1 and Lyl1. Tal1 and Lyl1 are aberrantly over-expressed in leukemia as a result of chromosomal translocations, or other genetic or epigenetic events. Protein-protein and protein-DNA interactions described so far are mediated by their highly homologous bHLH domains, while little is known about the function of other protein domains. Hetero-dimers of Tal1 and Lyl1 with E2A or HEB, decrease the rate of E2A or HEB homo-dimer formation and are poor activators of transcription. In vitro, these hetero-dimers also recognize different binding sites from homo-dimer complexes, which may also lead to inappropriate activation or repression of promoters in vivo. Both mechanisms are thought to contribute to the oncogenic potential of Tal1 and Lyl1. Despite their bHLH structural similarity, accumulating evidence suggests that Tal1 and Lyl1 target different genes. This raises the possibility that domains flanking the bHLH region, which are distinct in the two proteins, may participate in target recognition. Here we report that CREB1, a widely-expressed transcription factor and a suspected oncogene in acute myelogenous leukemia (AML) was identified as a binding partner for Lyl1 but not for Tal1. The interaction between Lyl1 and CREB1 involves the N terminal domain of Lyl1 and the Q2 and KID domains of CREB1. The histone acetyl-transferases p300 and CBP are recruited to these complexes in the absence of CREB1 Ser 133 phosphorylation. In the Id1 promoter, Lyl1 complexes direct transcriptional activation. We also found that in addition to Id1, over-expressed Lyl1 can activate other CREB1 target promoters such as Id3, cyclin D3, Brca1, Btg2 and Egr1. Moreover, approximately 50% of all gene promoters identified by ChIP-chip experiments were jointly occupied by CREB1 and Lyl1, further strengthening the association of Lyl1 with Cre binding sites. Given the newly recognized importance of CREB1 in AML, the ability of Lyl1 to modulate promoter responses to CREB1 suggests that it plays a role in the malignant phenotype by occupying different promoters than Tal1.

The zinc finger domain of Wilms' tumor 1 suppressor gene (WT1) behaves as a dominant negative, leading to abrogation of WT1 oncogenic potential in breast cancer cells.

INTRODUCTION: There is growing evidence that the Wilms' tumor 1 suppressor gene (WT1) behaves as an oncogene in some forms of breast cancer. Previous studies have demonstrated that the N-terminal domain of WT1 can act as a dominant negative through self-association. In the studies presented here we have explored the potential for the zinc finger domain (ZF) of WT1 to also have dominant-negative effects, and thus further our understanding of this protein. METHODS: Using full-length and ZF-only forms of WT1 we assessed their effect on the WT1 and c-myc promoter using luciferase and chromatin immunoprecipitation assays. The gene expression levels were determined by quantitative real-time RT-PCR, northern blot and western blot. We also assessed the effect of the ZF-only form on the growth of breast cancer cell lines in culture. RESULTS: Transfection with WT1-ZF plasmids resulted in a stronger inhibition of WT1 promoter than full-length WT1 in breast cancer cells. The WT1-ZF form lacking the lysine-threonine-serine (KTS) insert (ZF - KTS) can bind to the majority of WT1 consensus sites throughout the WT1 promoter region, while the ZF containing the insert (ZF + KTS) form only binds to sites in the proximal promoter. The abundances of endogenous WT1 mRNA and protein were markedly decreased following the stable expression of ZF - KTS in breast cancer cells. The expressions of WT1 target genes, including c-myc, Bcl-2, amphiregulin and TERT, were similarly suppressed by ZF - KTS. Moreover, WT1-ZF - KTS abrogated the transcriptional activation of c-myc mediated by all four predominant isoforms of WT1 (including or lacking alternatively spliced exons 5 and 9). Finally, WT1-ZF - KTS inhibited colony formation and cell division, but induced apoptosis in MCF-7 cells. CONCLUSION: Our observations strongly argue that the WT1-ZF plasmid behaves as a dominant-negative regulator of the endogenous WT1 in breast cancer cells. The inhibition on proliferation of breast cancer cells by WT1-ZF - KTS provides a potential candidate of gene therapy for breast cancer.

Transcriptional activation of c-myc proto-oncogene by WT1 protein.

The Wilms' tumor 1 gene (WT1) plays an essential role in urogenital development and malignancy. Through DNA binding, WT1 can either enhance or repress transcription depending on the context of the DNA-binding sites or the cell type in which it is expressed. WT1 is overexpressed in a variety of human cancers, including leukemia and breast cancer; in these diseases, the expression of WT1 is associated with a poor prognosis. To determine how WT1 affects c-myc expression in the context of breast cancer cells, we have examined the ability of both endogenous and exogenous WT1 proteins in breast cancer cells to bind to the c-myc promoter in vivo. Using c-myc-promoter-driven luciferase constructs, we found that different forms of WT1 could enhance the expression of the reporter. Unlike other studies where WT1 is reported to be a negative regulator of c-myc, we found that both the - and + KTS forms of WT1 could act to enhance c-myc expression, depending on the cell type. The WT1-binding site near the second major transcription start site of the c-myc promoter was confirmed to be involved in upregulation of human c-myc by WT1. Finally, we demonstrated that overexpression of WT1 induced a significant increase in the abundance of endogenous c-myc protein in breast cancer cells, consistent with the upregulation of c-myc transcription following WT1 induction. These observations strongly argue that in the case of breast cancer WT1 is functioning as an oncogene in part by stimulating the expression of c-myc.