Active Stat3 is required for survival of human squamous cell carcinoma cells in serum-free conditions

BACKGROUND: Squamous cell carcinoma (SCC) of the skin is the most aggressive form of non-melanoma skin cancer (NMSC), and is the single most commonly diagnosed cancer in the U.S., with over one million new cases reported each year. Recent studies have revealed an oncogenic role of activated signal transducer and activator of transcription 3 (Stat3) in many human tumors, especially in those of epithelial origin, including skin SCC. Stat3 is a mediator of numerous growth factor and cytokine signaling pathways, all of which activate it through phosphorylation of tyrosine 705. RESULTS: To further address the role of Stat3 in skin SCC tumorigenesis, we have analyzed a panel of human skin-derived cell lines ranging from normal human epidermal keratinocytes (NHEK), to non-tumorigenic transformed skin cells (HaCaT), to highly tumorigenic cells (SRB1-m7 and SRB12-p9) and observed a positive correlation between Stat3 phosphorylation and SCC malignancy. We next determined the role of Stat3 activity in cell proliferation and viability under serum-free culture conditions. This was accomplished by suppressing Stat3 activity in the SRB12-p9 cells through stable expression of a dominant negative acting form of Stat3β, which contains a tyrosine 705 to phenylalanine mutation (S3DN). The S3DN cells behaved similar to parental SRB12-p9 cells when cultured in optimal growth conditions, in the presence of 10% fetal calf serum. However, unlike the SRB12-p9 cells, S3DN cells underwent apoptotic cell death when cultured in serum-free medium (SFM). This was evidenced by multiple criteria, including accumulation of sub-G1 particles, induced PARP cleavage, and acquisition of the characteristic morphological changes associated with apoptosis. CONCLUSION: This study provides direct evidence for a role for Stat3 in maintaining cell survival in the conditions of exogenous growth factor deprivation produced by culture in SFM. We also propose that delivery of the S3DN gene or protein to tumor cells could induce apoptosis and/or sensitize those cells to the apoptotic effects of cancer therapeutic agents, raising the possibility of using S3DN as an adjunct for treatment of skin SCC

HGF/c-met/Stat3 signaling during skin tumor cell invasion: indications for a positive feedback loop

<p>Abstract</p> <p>Background</p> <p>Stat3 is a cytokine- and growth factor-inducible transcription factor that regulates cell motility, migration, and invasion under normal and pathological situations, making it a promising target for cancer therapeutics. The hepatocyte growth factor (HGF)/c-met receptor tyrosine kinase signaling pathway is responsible for stimulation of cell motility and invasion, and Stat3 is responsible for at least part of the c-met signal.</p> <p>Methods</p> <p>We have stably transfected a human squamous cell carcinoma (SCC) cell line (SRB12-p9) to force the expression of a dominant negative form of Stat3 (S3DN), which we have previously shown to suppress Stat3 activity. The <it>in vitro </it>and <it>in vivo </it>malignant behavior of the S3DN cells was compared to parental and vector transfected controls.</p> <p>Results</p> <p>Suppression of Stat3 activity impaired the ability of the S3DN cells to scatter upon stimulation with HGF (c-met ligand), enhanced their adhesion, and diminished their capacity to invade <it>in vitro </it>and <it>in vivo</it>. Surprisingly, S3DN cells also showed suppressed HGF-induced activation of c-met, and had nearly undetectable basal c-met activity, as revealed by a phospho-specific c-met antibody. In addition, we showed that there is a strong membrane specific localization of phospho-Stat3 in the wild type (WT) and vector transfected control (NEO4) SRB12-p9 cells, which is lost in the S3DN cells. Finally, co-immunoprecipitation experiments revealed that S3DN interfered with Stat3/c-met interaction.</p> <p>Conclusion</p> <p>These studies are the first confirm that interference with the HGF/c-met/Stat3 signaling pathway can block tumor cell invasion in an <it>in vivo </it>model. We also provide novel evidence for a possible positive feedback loop whereby Stat3 can activate c-met, and we correlate membrane localization of phospho-Stat3 with invasion <it>in vivo</it>.</p

Memory-Relevant Mushroom Body Output Synapses Are Cholinergic

Memories are stored in the fan-out fan-in neural architectures of the mammalian cerebellum and hippocampus and the insect mushroom bodies. However, whereas key plasticity occurs at glutamatergic synapses in mammals, the neurochemistry of the memory-storing mushroom body Kenyon cell output synapses is unknown. Here we demonstrate a role for acetylcholine (ACh) in Drosophila. Kenyon cells express the ACh-processing proteins ChAT and VAChT, and reducing their expression impairs learned olfactory-driven behavior. Local ACh application, or direct Kenyon cell activation, evokes activity in mushroom body output neurons (MBONs). MBON activation depends on VAChT expression in Kenyon cells and is blocked by ACh receptor antagonism. Furthermore, reducing nicotinic ACh receptor subunit expression in MBONs compromises odor-evoked activation and redirects odor-driven behavior. Lastly, peptidergic corelease enhances ACh-evoked responses in MBONs, suggesting an interaction between the fast- and slow-acting transmitters. Therefore, olfactory memories in Drosophila are likely stored as plasticity of cholinergic synapses

The origin of slow electron recombination processes in dye-sensitized solar cells with alumina barrier coatings

We investigate the effect of a thin alumina coating of nanocrystalline TiO2 films on recombination dynamics of dye-sensitized solar cells. Both coated and uncoated cells were measured by a combination of techniques: transient absorption spectroscopy, electrochemical impedance spectroscopy, and open-circuit voltage decay. It is found that the alumina barrier reduces the recombination of photoinjected electrons to both dye cations and the oxidized redox couple. It is proposed that this observed retardation can be attributed primarily to two effects: almost complete passivation of surface trap states in TiO2 that are able to inject electrons to acceptor species, and slowing down by a factor of 3–4 the rate of interfacial charge transfer from conduction-band [email protected]

Coastal Geology and Geomorphology of Cape Cod - An Aerial and Ground View

Guidebook for field trips to the Boston area and vicinity : 68th annual meeting, New England Intercollegiate Geological Conference, October 8-10, 1976: Trip A-10; B-1

Identification of the B-Raf/Mek/Erk MAP kinase pathway as a target for all-trans retinoic acid during skin cancer promotion

<p>Abstract</p> <p>Background</p> <p>Retinoids have been studied extensively for their potential as therapeutic and chemopreventive agents for a variety of cancers, including nonmelanoma skin cancer (NMSC). Despite their use for many years, the mechanism of action of retinoids in the prevention of NMSC is still unclear. In this study we have attempted to understand the chemopreventive mechanism of all-<it>trans </it>retinoic acid (ATRA), a primary biologically active retinoid, in order to more efficiently utilize retinoids in the clinic.</p> <p>Results</p> <p>We have used the 2-stage dimethylbenzanthracene (DMBA)/12-<it>O</it>-tetradecanoylphorbol-13-acetate (TPA) mouse skin carcinogenesis model to investigate the chemopreventive effects of ATRA. We have compared the gene expression profiles of control skin to skin subjected to the 2-stage protocol, with or without ATRA, using Affymetrix 430 2.0 DNA microarrays. Approximately 49% of the genes showing altered expression with TPA treatment are conversely affected when ATRA is co-administered. The activity of these genes, which we refer to as 'counter-regulated', may contribute to chemoprevention by ATRA. The counter-regulated genes have been clustered into functional categories and bioinformatic analysis has identified the B-Raf/Mek/Erk branch of the MAP kinase pathway as one containing several genes whose upregulation by TPA is blocked by ATRA. We also show that ATRA blocks signaling through this pathway, as revealed by immunohistochemistry and Western blotting. Finally, we found that blocking the B-Raf/Mek/Erk pathway with a pharmacological inhibitor, Sorafenib (BAY43-9006), induces squamous differentiation of existing skin SCCs formed in the 2-stage model.</p> <p>Conclusion</p> <p>These results indicate that ATRA targets the B-Raf/Mek/Erk signaling pathway in the 2-stage mouse skin carcinogenesis model and this activity coincides with its chemopreventive action. This demonstrates the potential for targeting the B-Raf/Mek/Erk pathway for chemoprevention and therapy of skin SCC in humans. In addition our DNA microarray results provide the first expression signature for the chemopreventive effect of ATRA in a mouse skin cancer model. This is a potential source for novel targets for ATRA and other chemopreventive and therapeutic agents that can eventually be tested in the clinic.</p

Metric for Measuring the Effectiveness of Clustering of DNA Microarray Expression

BACKGROUND: The recent advancement of microarray technology with lower noise and better affordability makes it possible to determine expression of several thousand genes simultaneously. The differentially expressed genes are filtered first and then clustered based on the expression profiles of the genes. A large number of clustering algorithms and distance measuring matrices are proposed in the literature. The popular ones among them include hierarchal clustering and k-means clustering. These algorithms have often used the Euclidian distance or Pearson correlation distance. The biologists or the practitioners are often confused as to which algorithm to use since there is no clear winner among algorithms or among distance measuring metrics. Several validation indices have been proposed in the literature and these are based directly or indirectly on distances; hence a method that uses any of these indices does not relate to any biological features such as biological processes or molecular functions. RESULTS: In this paper we have proposed a metric to measure the effectiveness of clustering algorithms of genes by computing inter-cluster cohesiveness and as well as the intra-cluster separation with respect to biological features such as biological processes or molecular functions. We have applied this metric to the clusters on the data set that we have created as part of a larger study to determine the cancer suppressive mechanism of a class of chemicals called retinoids. We have considered hierarchal and k-means clustering with Euclidian and Pearson correlation distances. Our results show that genes of similar expression profiles are more likely to be closely related to biological processes than they are to molecular functions. The findings have been supported by many works in the area of gene clustering. CONCLUSION: The best clustering algorithm of genes must achieve cohesiveness within a cluster with respect to some biological features, and as well as maximum separation between clusters in terms of the distribution of genes of a behavioral group across clusters. We claim that our proposed metric is novel in this respect and that it provides a measure of both inter and intra cluster cohesiveness. Best of all, computation of the proposed metric is easy and it provides a single quantitative value, which makes comparison of different algorithms easier. The maximum cluster cohesiveness and the maximum intra-cluster separation are indicated by the metric when its value is 0. We have demonstrated the metric by applying it to a data set with gene behavioral groupings such as biological process and molecular functions. The metric can be easily extended to other features of a gene such as DNA binding sites and protein-protein interactions of the gene product, special features of the intron-exon structure, promoter characteristics, etc. The metric can also be used in other domains that use two different parametric spaces; one for clustering and the other one for measuring the effectiveness

Necessary fictions: indigenous claims and the humanity of rights

Indigenous right insistently challenges the surpassing arrogations of sovereign right. In so doing, it affirms dimensions of being-together denied or stunted in sovereign modes of political formation. This force of Indigenous right is amplified here through legal and literary instantiations. These, in turn, uncover the continuously created and fictional quality of rights, revealing them to be necessary fictions