EGF activates TTP expression by activation of ELK-1 and EGR-1 transcription factors

<p>Abstract</p> <p>Background</p> <p>Tristetraprolin (TTP) is a key mediator of processes such as inflammation resolution, the inhibition of autoimmunity and in cancer. It carries out this role by the binding and degradation of mRNA transcripts, thereby decreasing their half-life. Transcripts modulated by TTP encode proteins such as cytokines, pro-inflammatory agents and immediate-early response proteins. TTP can also modulate neoplastic phenotypes in many cancers. TTP is induced and functionally regulated by a spectrum of both pro- and anti-inflammatory cytokines, mitogens and drugs in a MAPK-dependent manner. So far the contribution of p38 MAPK to the regulation of TTP expression and function has been best described.</p> <p>Results</p> <p>Our results demonstrate the induction of the gene coding TTP (<it>ZFP36</it>) by EGF through the ERK1/2-dependent pathway and implicates the transcription factor ELK-1 in this process. We show that ELK-1 regulates <it>ZFP36 </it>expression by two mechanisms: by binding the <it>ZFP36 </it>promoter directly through ETS-binding site (+ 883 to +905 bp) and by inducing expression of EGR-1, which in turn increases <it>ZFP36 </it>expression through sequences located between -111 and -103 bp.</p> <p>Conclusions</p> <p>EGF activates TTP expression via ELK-1 and EGR-1 transcription factors.</p

ADAM17 Silencing in Mouse Colon Carcinoma Cells: The Effect on Tumoricidal Cytokines and Angiogenesis

ADAM17 (a disintegrin and metalloprotease 17) is a major sheddase for numerous growth factors, cytokines, receptors, and cell adhesion molecules and is often overexpressed in malignant cells. It is generally accepted that ADAM17 promotes tumor development via activating growth factors from the EGF family, thus facilitating autocrine stimulation of tumor cell proliferation and migration. Here we show, using MC38CEA murine colon carcinoma model, that ADAM17 also regulates tumor angiogenesis and cytokine profile. When ADAM17 was silenced in MC38CEA cells, in vivo tumor growth and in vitro cell motility were significantly diminished, but no effect was seen on in vitro cell proliferation. ADAM17-silencing was accompanied by decreased in vitro expression of vascular endothelial growth factor-A and matrix metalloprotease-9, which was consistent with the limited angiogenesis and slower growth seen in ADAM17-silenced tumors. Among the growth factors susceptible to shedding by ADAM17, neuregulin-1 was the only candidate to mediate the effects of ADAM17 on MC38CEA motility and tumor angiogenesis. Concentrations of TNF and IFN gamma, cytokines that synergistically induced proapoptotic effects on MC38CEA cells, were significantly elevated in the lysates of ADAM17-silenced tumors compared to mock transfected controls, suggesting a possible role for ADAM17 in host immune suppression. These results introduce new, complex roles of ADAM17 in tumor progression, including its impact on the anti-tumor immune response

aPTT in plasma of Wistar rats mixed with different concentrations of Dex40-GTMAC.

<p>a-P<0.05, b-P<0.01 vs. vehicle, Mann-Whitney test. Results are shown as mean ± SD, n = 5.</p

Reversal of the effects of UFH on the venous thrombosis and aPTT in mice.

<p>Dry thrombus weight <b>(A)</b> and the aPTT <b>(B)</b> in mice treated with vehicle (PBS), UFH (300 U·kg^-1) alone or followed by Dex40-GTMAC3 (7.5 mg·kg^-1), and protamine (3.0 mg·kg^-1), b-P<0.01, c-P<0.001 vs. vehicle; e-P<0.01 vs. UFH 300 U·kg^-1; h-P<0.01 vs. UFH 300 U·kg^-1 + Protamine 3.0 mg·kg^-1, Mann-Whitney test. Results are shown as mean ± SD n = 5–7.</p

Evaluation of the immune response to the cationic polymers.

<p>Humoral immune response evaluation by ELISA on day 36 of the experiment. The levels of IgG specific toward different UFH inhibitors in individual mice presented as values of absorbance in ELISA test. The inserted graph shows mean ELISA signals corresponding to the levels of anti-protamine-, anti-Dex40-GTMAC2- or Dex40-GTMAC3-IgG present in sera of mice immunized with matching UFH-antidotes (gray bars) compared to the signals from sera of control mice treated with UFH alone (black bars). All sera applied in 1:100 dilution, b-P<0.01, Mann-Whitney test. Results are shown as mean ± SD, n = 5.</p

Influence of the cationic polymers on aPTT in rats.

<p>aPTT in Wistar rats treated with vehicle (PBS), UFH (300 U·kg^-1) alone or followed by Dex40-GTMAC2 (4.2 mg·kg^-1), Dex40-GTMAC3 (2.5 and 7.5 mg·kg^-1), Dex6-GTMAC (9.6 mg·kg^-1), GCD-GTMAC2 (10.8 mg·kg^-1), and protamine (3.0 mg·kg^-1), c-P<0.001 vs. vehicle; f-P<0.001 vs. UFH 300 U·kg^-1; i-P<0.001 vs. UFH 300 U·kg^-1 + Protamine 3.0 mg·kg^-1; l-P<0.001 vs. UFH 900 U·kg^-1, Mann-Whitney test. Results are shown as mean ± SD, n = 8–10.</p

Characteristics of the polymers.

<p>Dex—dextran, Pul—pullulan, GCD—γ-cyclodextrin, HPC—hydroxypropylcellulose, GTMAC—ammonium group of glycidyltrimethyl-ammonium chloride, APTMAC—ammonium group of N-acrylamidopropyl-N,N,N-trimethylammonium chloride, Spm—primary and secondary amine groups of N,N′-Bis(3-aminopropyl)-1,4-diaminobutane (Spermine), PAH—primary amine group of poly(allylamine hydrochloride), PAH-Arg—primary amine group of poly(allylamine hydrochloride), guanidyl group of arginine,</p><p>^a MW—weight average molecular weight, for GCD exact molecular weight is given</p><p>^b ξ—zeta potential</p><p>^c degree of substitution defined as the number of the cationic groups per a glucose unit, as found from elemental analysis</p><p>^d UFH binding—ratio of polymer mass to UFH mass required for binding 90% of UFH</p><p>^e the mass of the polymer required to bind UFH was at least 8 times higher than that of protamine</p><p>^f depending on concentration</p><p>^g degree of substitution defined as the number of arginine groups per PAH repeating unit</p><p>Characteristics of the polymers.</p

Reversal of plasma anti-fXa activity by the cationic polymers.

<p>Anti-fXa activity in Wistar rats treated with vehicle (PBS), UFH (300 U·kg^-1) alone or followed by Dex40-GTMAC2 (4.2 mg·kg^-1), Dex40-GTMAC3 (2.5 and 7.5 mg·kg^-1), Dex6-GTMAC (9.6 mg·kg^-1), GCD-GTMAC2 (10.8 mg·kg^-1), and protamine (3.0 mg·kg^-1), c-P<0.001 vs. vehicle; f-P<0.001 vs. UFH 300 U·kg^-1; i-P<0.001 vs. UFH 300 U·kg^-1 + Protamine 3.0 mg·kg^-1, Mann-Whitney test. Results are shown as mean ± SD, n = 8–10.</p

Body weights and complete blood count results after arterial thrombosis stimulation in rats, means ± SD.

<p>a-P<0.05,</p><p>b-P<0.01,</p><p>c-P<0.001 vs. vehicle;</p><p>d-P<0.05,</p><p>e-P<0.01,</p><p>f-P<0.001 vs. UFH 300 U·kg^-1;</p><p>g-P<0.05,</p><p>h-P<0.01,</p><p>i-P<0.001 vs. UFH 300 U·kg^-1 + Protamine 3.0 mg·kg^-1, Mann-Whitney test.</p><p>WBC: white blood cells, RBC: red blood cells, HGB: hemoglobin, HCT: hematocrit, MCV: mean corpuscular volume, MCH: mean corpuscular hemoglobin, MCHC: mean corpuscular hemoglobin concentration, PLT: blood platelets. Results are shown as mean ± SD, n = 8–10.</p><p>Body weights and complete blood count results after arterial thrombosis stimulation in rats, means ± SD.</p