text:
The involvement of P2Y receptors, which are activated by extracellular nucleotides, in proliferative regulation of human lung epithelial cells is unclear. Here we show that extracellular ATP and UTP stimulate bromodeoxyuridine (BrdU) incorporation into epithelial cell lines. The nucleotide efficacy profile [ATP = ADP > UDP ≥ UTP > adenosine ≥ 2-methylthioadenosine-5′-diphosphate, with α,β-methylene adenosine 5′-triphosphate, 2′,3′-O-(4-benzoylbenzoyl)adenosine 5′-triphosphate, AMP, UMP, and ATPαS inactive] and PCR analysis indicate involvement of P2Y2 and P2Y6 receptors. The signal transduction pathway, which, via the P2Y2 receptor, transmits the proliferative activity of ATP or UTP in A549 cells downstream of phospholipase C, depends on Ca2+/calmodulin-dependent protein kinase II and nuclear factor-κB, but not on protein kinase C. Signaling does not involve the mitogen-activated protein kinases extracellular signal-regulated kinases-1 and -2, the phosphatidylinositol 3-kinase pathway, or Src kinases. Thus nucleotides regulate proliferation of human lung epithelial cells by a novel pathway. The stimulatory effect of UTP, but not ATP, in A549 cells is attenuated by preincubation with interleukin-1β and interleukin-6, but not tumor ncerosis factor-α. This indicates an important role for the pyrimidine-activated P2Y receptor in the inflammatory response of lung epithelia. ATP antagonizes the antiproliferative effect of the anticancer drugs paclitaxel and etoposide, whereas it enhances the activity of cisplatin about fourfold. Thus pathways activated by extracellular nucleotides differentially control proliferation of lung epithelial tumor cells.
- purinergic receptors
- cancer
- cytokines
- extracellular nucleotides
extracellular nucleotides activate epithelial cells in many organs and tissues under physiological and pathological conditions (2, 37, 46). The nucleotides ATP and UTP are released from airway epithelial cells on the apical and basolateral sides in response to cell swelling (43) or mechanical stimuli (11, 27, 48). The released ATP or UTP, in an autocrine or paracrine fashion, controls cell volume through Cl- and fluid secretion (17, 22, 41, 43), mucociliary clearance (mucin, salt, and water secretion) (28, 33), or intracellular Ca2+ oscillations (8) and coordinates responses of distant airway cells by modulating intercellular communication (12, 18) and ciliary beat frequency (35).
The effects of ATP and UTP are mediated through the metabotropic P2Y receptor family. The subtypes, P2Y1, P2Y2, P2Y4, P2Y6, and P2Y13 receptors, are expressed in human lung (37, 50). In airway epithelial cells, extracellular nucleotides regulate physiological functions via phospholipase C activation and subsequent inositol 1,4,5-trisphosphate formation, mainly through P2Y2 and P2Y1 receptors (8, 12). The P2Y2 receptor, which responds equally to extracellular UTP and ATP, is expressed at the apical side of human airway epithelial cells (26). Moreover, diadenosine polyphosphates can activate a Ca2+ increase in human lung epithelial cells via purinergic receptors (24, 25). There is no direct evidence of a role for P2X receptors in regulation of airway ion transport activities. However, it has been proposed that, in tubes and sacs of gallbladders and in the lung, tension leads to release of ATP from the lining epithelium, which then acts on P2X2 and P2X3 receptors of subepithelial sensory nerves to convey information to the central nervous system (2).
Recently, extracellular nucleotides have emerged as regulators of tumor proliferation and possible effectors of neoplastic transformation. In carcinoma tissues, not only is ATP released during chemotherapy (23), it also controls proliferation of various tumor cells. Depending on the subtype(s) of P2Y receptor expressed, P2Y receptors facilitate proliferation, as in C6 glioma cells (44) and breast cancer cells (47), or inhibit cell proliferation, as reported for endometrial cells (20), colon (13), or esophageal cancer cells (31). However, the mitogenic actions of extracellular nucleotides on airway epithelial cells have not been investigated. This is especially important for understanding lung tumorigenesis. Moreover, the actions of the extracellular nucleotides, in concert with cytokines and epidermal growth factor (EGF), which are known to tightly control proliferation of tumor cells, remain to be clarified.
Extracellular nucleotides have been reported to regulate proliferation of several cancer cell lines (13, 31, 47). ATP inhibits weight loss in tumor-bearing murine hosts (38). Moreover, clinical studies have indicated that, in patients with advanced lung cancer, ATP 1) may inhibit weight loss and suppress tumor growth and 2) displays protective effects against radiation tissue damage (reviewed in Ref. 3).
We therefore examined the effect of extracellular nucleotides on proliferation of tumorigenic and nontumorigenic human lung epithelial cell lines. Proliferation of the tumorigenic A549 cells and the nontumorigenic BEAS-2B cells is stimulated by the purine nucleotide ATP and the pyrimidine nucleotides UTP and UDP through activation of the P2Y receptor subtypes P2Y2 and P2Y6 receptors. P2Y2 receptor-induced A549 tumor cell proliferation depends on activation of phospholipase C, Ca2+/calmodulin-dependent kinase II (CaMKII), nuclear factor-κB(NF-κB), and, possibly, EGF receptor (EGFR) tyrosine kinase. This new pathway directly stimulates cell cycle progression, as indicated by the specific interference of ATP with the cytostatic actions of paclitaxel, etoposide, and cisplatin.
Cell culture and reagents. The human lung cancer cell line A549 was obtained from the German Collection of Micro-organisms and Cell Cultures (DSMZ, Braunschweig, Germany), and the noncarcinogenic human lung cell line BEAS-2B was kindly provided by Dr. F. Bühling (Otto-von-Guericke University). A549 human non-small cell lung cancer cells were cultured in DMEM (Biochrom) supplemented with 5% fetal calf serum, 100 IU/ml penicillin, and 100 μg/ml streptomycin in a 10% CO2 incubator at 37°C. The BEAS-2B cells were cultured in DMEM supplemented with 10% fetal calf serum, 100 IU/ml penicillin, and 100 μg/ml streptomycin in a 10% CO2 incubator at 37°C.
The anticancer drugs paclitaxel, etoposide, and cisplatin were purchased from Calbiochem (Bad Soden, Germany). U-73122 and its inactive enantiomer U-73343, N-nitro-l-arginine, chelerythrine, SB-203580, wortmannin, AG-1478, NS-398, 8-cyclopentyl-1,3-dipropylxanthine, CGS-15943, alloxazine, suramin, reactive blue-2 (RB-2), pyridoxal phosphate-6-azophenyl-2′,4′-disulfonic acid (PPADS), ARL-67156, adenosine 3′-phosphate 5′-phosphate (A3P5P; Sigma), KN-62, GF-109203X, PD-908059, PP-2, methyl arachidonyl fluorophosphonate (Bio-Trend), genistein, SKF-96365, indomethacin, and NF-κB SN-50 (Calbiochem) were dissolved according to the recommendations of the manufacturer in dimethylsulfoxide or in phosphate-buffered saline to give a stock concentration of 10 or 100 mM. The cytokines interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α) were obtained from Roche.
RNA extraction and RT-PCR for detection of P2Y receptor subtypes. Total RNA was isolated from A549 and HEK-293 cells with the RNeasy kit (Qiagen). HEK-293 cells were used to verify the usefulness of the primers chosen for PCR and to control the RT-PCR conditions, because HEK-293 cells endogenously express P2Y1, P2Y2, and P2Y4 receptors. The isolation included a DNase treatment to remove contaminating DNA. RT was performed with 1 μg of total RNA using oligo(dT) primer with the Omniscript kit (Qiagen). For amplifying the P2Y receptor cDNAs, sets of specific oligonucleotide primers were synthesized on the basis of the published sequences for P2Y1, P2Y2, P2Y4, and P2Y6 receptors (see below). Because the P2Y4 receptor has only one exon and the P2Y6 receptor has two exons, primers were defined, flanking the intron of P2Y6 (exon 1 = 156–242, exon 2 = 243–1,563). PCR without the RT step was used as control to exclude amplification of genomic DNA. Amplification was performed with 1 μl of cDNA, and the conditions were as follows: 98°C for 2 min, 94°C for 1 min, 55°C for 90 s, 72°C for 2 min, and a final extension step of 10 min at 72°C for 30 cycles.
The primers were 5′ GCC GCC GTC TCC TCG TCG TT 3′ (forward, position 146–166) and 5′ CCA TTC TGC TTG AAC TCA G 3′ (reverse, position 1133–1152) with 1,006 bp for hP2Y1 (accession no. U42029), 5′ CGA GGA CTT CAA GTA CGT GC 3′ (forward, position 329–349) and 5 ′GTT CTC GCT ACC AGC CGG CG 3′ (reverse, position 1338–1358) with 1,030 bp for hP2Y2 (accession no. U07225), 5′ GCT GCC TGT GAG CTA TGC AG 3′ (forward, position 110–140) and 5′ GAG CTG ACG GAG CTG ACG TCG′ (reverse, position 939–960) with 850 bp for hP2Y4 (accession no. X91852), and 5′ GTT CAG GCT GAG GAG ATG 3′ (forward, position 201–219) and 5′ GGA AGC TGA TGC AGG TGA 3′ (reverse, position 623–643) with 442 bp for hP2Y6 (accession no. U52464).
Determination of cell proliferation. Cell proliferation was measured by luminometric immunoassay based on BrdU incorporation during DNA synthesis using a cell proliferation enzyme-linked immunosorbent assay BrdU kit (Roche, Mannheim, Germany) according to the manufacturer's protocol. Briefly, A549 or BEAS-2B cells were seeded at a density of 3 × 103 cells/well on 96-well plates and incubated for 24 h. Cells were then incubated with the different nucleotides or nucleotide analogs at 100 μM (standard conditions) for 1 h in a final volume of 100 μl/well. For each concentration as well as the controls, three wells were used. Subsequently, 10 μl of BrdU-labeling solution were added to each well, and the cells were incubated again for 12 h. The different antagonists were added to the cells 1 h before addition of the nucleotides. After removal of the BrdU-labeling solution, cells were washed with 100 μl of phosphate-buffered saline, dried for 1 h at 60°C, and then incubated with the kit's FixDenat solution for 45 min at room temperature. Denaturation of the DNA is necessary to improve the accessibility of the incorporated BrdU for detection by antibody. Samples were incubated for 90 min with peroxidase-conjugated anti-BrdU antibody, which binds to BrdU incorporated into newly synthesized cellular DNA. After the unbound peroxidase-conjugated anti-BrdU-antibody was washed off the samples, 100 μl of the luminescence substrate was added, the generation of the signal was allowed to develop for 3–5 min under gentle shaking, and the luminescence was quantified using a microplate luminometer (Spectrafluorplus, Tecan).
Data analysis. Proliferation data refer to the chemiluminescence values of BrdU-labeled cellular DNA content per well. Stimulation is expressed as x-fold proliferation over basal value, where basal growth of the untreated control was set at 1.0, or as percentage of nucleotide-stimulated proliferation. In the latter case, the difference between the values of the untreated control and the values of the nucleotide-stimulated samples was taken as 100%. Unless indicated otherwise, values are means ± SD from at least three independent experiments.
Regulation of epithelial lung tumor cell proliferation by extracellular nucleotides. To investigate the effects of extracellular nucleotides on proliferation of human lung tumor cells, proliferating A549 cells were pulse stimulated with ATP or UTP. Then cell proliferation was measured by monitoring BrdU incorporation into newly synthesized DNA for 12 h after addition of nucleotides. ATP and UTP induced a concentration-dependent increase in proliferation of A549 cells. The maximal stimulation, which was seen at 100 μM nucleotide, was 3.2-fold for ATP and 1.8-fold for UTP (Fig. 1A). Despite the difference in maximal stimulation, the potency of both nucleotides to stimulate proliferation was very similar, with EC50 of 26.0 ± 2.8 μM(n = 3) for ATP and 31.3 ± 4.1 μM (n = 3) for UTP.
The effect of the nucleotides on BrdU incorporation into A549 cells was time dependent. The maximal stimulatory effect by ATP and UTP was reached at 1 h after addition of the nucleotides and lasted for 3–6 h (Fig. 1B). Incorporation of BrdU into newly synthesized DNA rapidly declined to baseline values at >6 h after nucleotide addition. This indicates that extracellular nucleotides stimulate DNA synthesis by direct interaction with cell cycle progression.
Besides ATP and UTP, the diphosphate nucleotides ADP and UDP also stimulated BrdU incorporation into proliferating A549 cells with an efficacy nearly identical to that of the respective triphosphate nucleotides (Fig. 2A). At 100 μM, ADP and UDP enhanced proliferation of A549 cells 3.1- and 2.2-fold, respectively.
To determine whether the stimulatory effect of extracellular nucleotides on proliferation of A549 lung tumor cells was associated with neoplastic transformation of the cells, we also investigated the effect of nucleotides on proliferation of the nontumorigenic human epithelial lung cell line BEAS-2B. BEAS-2B and A549 cells serve as in vitro models for bronchiolar and alveolar cells, respectively.
Enhancement of proliferation by extracellular nucleotides was even stronger in the BEAS-2B cells than in the A549 cells. Here, 100 μM ATP and 100 μM ADP stimulated BrdU incorporation into newly synthesized DNA 7.1- and 6.5-fold, respectively (Fig. 2B). The pyrimidine nucleotides UTP and UDP exhibited identical efficacy and induced a 3.2- and 3.4-fold stimulation, respectively (Fig. 2B). Thus purine and pyrimidine nucleotides may regulate human lung epithelial cell proliferation in vivo. Moreover, control of proliferation by nucleotides is not due to transformation of lung epithelial cells, because proliferation of tumorigenic and nontumorigenic airway cells is equally affected.
To evaluate whether the trophic actions of the extracellular nucleotides on the human lung cells were mediated by the P2Y or the P2X receptors, proliferative regulation of A549 and BEAS-2B cells was investigated by using different nucleotides, nucleotide analogs, and P2 receptor antagonists. Characterization of A549 cell proliferation revealed that only ATP, ADP, UTP, and UDP, which stimulate different P2Y receptor subtypes, were active, but the P2Y1 receptor-selective analogs 2-methylthioadenosine 5′-diphosphate (2-MeS-ADP) and ATPαS and the P2X receptor-activating nucleotides α,β-methylene adenosine 5′-triphosphate (α,β-MeATP) and 2′,3′-O-(4-benzoylbenzoyl)adenosine 5′-triphosphate (BzATP) were inactive (Fig. 2). AMP and UMP at up to 100 μM also had no effect on proliferation (data not shown).
The efficacy of the different nucleotide ligands was as follows: ATP = ADP > UDP ≥ UTP > 2-MeS-ADP, whereas α,β-MeATP, BzATP, AMP, UMP, and ATPαS were inactive. This clearly shows that stimulation of A549 cell proliferation is mediated by P2Y, but not by P2X, receptors, inasmuch as α,β-MeATP and BzATP, which, in addition to ATP, can activate P2X receptors (P2X1, P2X2, P2X3, P2X7) (21), have no influence on proliferation (Fig. 2). In addition, the lack of activity of 2-MeS-ADP and ATPαS indicates that in A549 cells no functional P2Y1 receptors are involved. However, the pharmacological profile suggests involvement of more than one P2Y receptor in stimulation of A549 cell proliferation, because the profile is not consistent with the properties of a single P2Y receptor subtype (46).
We further attempted to identify the P2Y receptor subtypes involved by applying the P2 receptor-selective antagonists PPADS, suramin, RB-2, and ARL-67156. Preincubation of A549 cells with 100 μM suramin or RB-2 inhibited ATP-stimulated proliferation to 37 ± 8.4% (n = 4) and 45 ± 4.5% (n = 3), respectively (Table 1). There was no significant inhibition of ATP-stimulated proliferation by PPADS, which predominantly inhibits the human P2Y1 and P2Y4 receptors or the P2Y1 receptor-selective antagonists ARL-67156 and A3P5P (Table 1). Thus this approach also did not identify a single P2Y receptor subtype (see discussion).
Adenosine did not mimic ATP-stimulated proliferation. At 100 μM, where ATP is maximally effective, adenosine enhanced proliferation only marginally by 35 ± 4.3% and 57 ± 12.1% in A549 and BEAS-2B cells, respectively (Fig. 2), excluding the possibility that the stimulatory effect of ATP was caused by the metabolic breakdown product adenosine. Furthermore, to rule out the possibility that adenosine-mediated signaling partly contributed to the ATP-stimulated proliferative response, the effect of different adenosine receptor antagonists (i.e., P1 antagonists) on ATP-stimulated proliferation of A549 cells was investigated (Table 1). Neither the nonselective adenosine receptor antagonist CGS-15943 nor the A1 and A2B receptor-selective antagonists 8-cyclopentyl-1,3-dipropylxanthine and alloxazine, respectively, inhibited ATP-stimulated proliferation. We did not test specific inhibitors of A3 receptors, inasmuch as this receptor subtype is not expressed in A549 cells (42). Thus, clearly, P1 receptors are not contributing to the signaling of extracellular nucleotides in the proliferative response of A549 cells. Therefore, characterization of A549 cell proliferation using agonists and antagonists of P2 and P1 receptors reveals that only P2Y receptors, but not P2X or P1 receptors, mediate stimulation of lung epithelial cell proliferation.
Furthermore, we examined expression of different P2Y receptor subtypes (P2Y1, P2Y2, P2Y4, and P2Y6 receptors) by RT-PCR. In A549 cells, only mRNA specific for the P2Y2 receptor, which is activated with identical potency by ATP and UTP, and the UDP-selective P2Y6 receptor was detected by RT-PCR, whereas no transcripts for the P2Y1 or P2Y4 receptor could be found (Fig. 3). In the same experiment, we employed HEK-293 cells to control the RT-PCR conditions. HEK-293 cells endogenously express P2Y1, P2Y2, and P2Y4 receptors. These results indicate that primarily the P2Y2 and P2Y6 receptors mediate nucleotide-stimulated proliferation of A549 cells.
Signal transduction pathways of nucleotide-stimulated proliferation of A549 cells. We next investigated the signaling pathways involved in P2Y receptor-stimulated proliferation. A549 or BEAS-2B cells were pretreated with different inhibitors of signaling cascades that mediate proliferation of other cell types. Basal DNA synthesis of proliferating cells was not blocked by these inhibitors at the concentrations used. ATP-stimulated proliferation of A549 cells was almost completely attenuated by preincubation for 1 h with U-73122 (10 μM), an inhibitor of phospholipase C, whereas the inactive control derivative U-73343 was not effective (Fig. 4A). Also, inhibition of CaMKII activity with KN-62 (10 μM) and of the influx of extracellular Ca2+ through receptor-operated Ca2+ channels with SKF-96365 (50 μM) strongly decreased ATP-stimulated BrdU incorporation to 34.6 ± 8.7% (n = 6) and 33.3 ± 5.2% (n = 5), respectively (Fig. 4A). However, the following drugs did not influence the ability of ATP or UTP to stimulate proliferation of A549 cells (Table 2): chelerythrine (20 μM) and GF-109203X (1 μM), which inhibit different protein kinase C (PKC) isoforms, the mitogen-activated protein kinase (MAPK) kinase (MEK1/2) inhibitor PD-98059 (20 μM), the p38 kinase inhibitor SB-203580 (20 μM), the phosphatidylinositol 3-kinase pathway inhibitors wortmannin (0.1 μM) and LY-294002 (25 μM), and Nω-nitro-l-arginine (10 μM), which blocks generation of nitric oxide.
To identify components of the signal transduction pathway resulting in DNA synthesis, we evaluated pathways that can be activated by CaMKII or tyrosine kinases. Pretreatment of A549 cells with an inhibitory peptide specific for the p50 form of NF-κB (NF-κB SN-50; 20 μM) or with AG-1478 (500 nM), specifically inhibiting EGFR tyrosine kinase activity, significantly reduced ATP stimulation of DNA synthesis to 42.8 ± 5.9% and 43.0 ± 4.6%, respectively (Fig. 4B, Table 2). Inhibition of Src kinase, which can also be activated by G protein-coupled receptors, with PP-2 (500 nM), inhibition of general tyrosine kinase activity (genistein at 100 μM), inhibition of cytosolic phospholipase A2 (methyl arachidonyl fluorophosphonate, 10 μM), or inhibition of cyclooxygenases with indomethacin (20 μM) or NS-398 (10 μM) did not abolish the effect of ATP and UTP on proliferation of A549 cells (Fig. 4B, Table 2).
Interestingly, similar results were derived for BEAS-2B cells. In the analysis of signal transduction, we obtained identical inhibition by the various drugs for the bronchial epithelial BEAS-2B cells, except for genistein, which only in BEAS-2B cells decreased ATP- and UTP-stimulated proliferation to 60 and 68%, respectively (Table 2). This indicates that, in the immortalized bronchial BEAS-2B cells, additional tyrosine kinases are involved in the proliferative regulation.
Modulation of nucleotide-regulated proliferation of lung tumor cells by cytokines. Airway epithelial cells are a major source of proinflammatory cytokines and, thus, play an important role in the pathophysiology of lung diseases. Therefore, we investigated the effect of cytokines on nucleotide-regulated proliferation. Moreover, cytokines had also been found to upregulate P2Y2 receptors in vascular smooth muscle cells (15). Preincubation of A549 cells for 24 h with IL-1β (10 ng/ml) or IL-6 (50 ng/ml), at concentrations that are detected under inflammatory conditions, attenuated UTP-stimulated proliferation to 59.7 ± 10.3% and 34.7 ± 12.1%, respectively, compared with the untreated stimulation (Fig. 5, Table 3). However, proliferation of A549 cells induced by the purine nucleotide ATP was clearly not affected by pretreatment with the cytokines. TNF-α had no effect at concentrations up to 50 ng/ml. Moreover, pretreatment of BEAS-2B cells with the cytokines IL-1β, IL-6, and TNF-α did not influence ATP- or UTP-stimulated proliferation (Table 3).
Interaction of anticancer drugs with nucleotide-stimulated proliferation of lung tumor cells. Inasmuch as control of proliferation is a key target in cancer therapy, we also examined the effect of anticancer drugs on the nucleotide-regulated proliferation. BrdU incorporation into proliferating A549 cells was inhibited dose dependently by the anticancer drugs cisplatin, etoposide, and paclitaxel, with IC50 of 2.7 ± 1.1 μM (n = 5), 19.8 ± 2.2 μM (n = 5), and 17.6 ± 3.8 nM (n = 4), respectively (Fig. 6). The simultaneous addition of 100 μM ATP enhanced the antiproliferative potency of cisplatin 3.6-fold, shown by reduction of IC50 to 0.75 μM (Fig. 6). The effect of ATP was most obvious at low concentrations of the anticancer drugs. At 1 μM, cisplatin was twice as effective in reducing proliferation in the presence of ATP than in its absence.
The antiproliferative activity of etoposide and paclitaxel was antagonized by the simultaneous presence of 100 μM ATP, resulting in 1.6-fold (IC50 = 31 ± 2.9 μM, n = 5) and 2.6-fold (IC50 = 46.2 ± 4.8 nM, n = 4) lower inhibitory potency, respectively (Fig. 6). ATP still stimulated proliferation 80% above control in the presence of 1 μM etoposide. Addition of 100 μM UTP did not change the potency of the three anticancer drugs (data not shown). These results support the notion that ATP regulates proliferation of human lung tumor cells by direct control of the cell cycle.
This is the first study showing that extracellular nucleotides stimulate proliferation of human lung epithelial cells by activation of the P2Y2 and P2Y6 receptors, which are detected by RT-PCR to be expressed in A549 cells. The identical EC50 values for ATP- and UTP-stimulated proliferation of A549 cells indicate that both nucleotides stimulate A549 cell proliferation through activation of P2Y2 receptors, because this receptor is equally sensitive to ATP and UTP. This conclusion is also supported by the activity profile of the P2 receptor antagonists. The significant, but not complete, inhibition of ATP- and UTP-stimulated proliferation by 100 μM suramin or 100 μM RB-2 and the lack of activity of 100 μM PPADS are characteristic of the human P2Y2 receptor (46).
Furthermore, our results showing that UTP and UDP stimulate BrdU incorporation in A549 cells with equal efficacy indicate that, in addition to the P2Y2 receptor, the P2Y6 receptor can also mediate stimulation of proliferation. At the human P2Y6 receptor, UDP is ∼100-fold more potent than UTP (46). Thus, as in vascular smooth muscle cells, where for the first time UDP-stimulated growth via the P2Y6 receptor had been reported (14), the P2Y6 receptor is likely to be involved in the control of proliferation in lung epithelial cells.
However, the efficacy profile (ATP = ADP > UDP ≥ UTP) found for A549 cells indicates involvement of some additional receptor, other than the P2Y2 or P2Y6 receptor subtypes (46). ADP activated proliferation of A549 cells with a potency similar to that of ATP. The identity of the receptor, which mediates the action of ADP, is not known. ADP has been found to be considerably less active at the P2Y2 and P2Y6 receptors than the preferred agonists ATP/UTP and UDP, respectively (46). ADP preferentially activates the P2Y1 receptor, which, however, is not expressed in A549 cells. The absence of P2Y1 receptor expression in A549 cells is also pharmacologically underlined by the inactivity of 2-MeS-ADP. 2-MeS-ADP activates the P2Y1 receptor to an even greater degree than ADP.
An alternative possibility explaining the effectiveness of ADP implies that ADP may be converted to ATP by an ectonucleoside diphosphokinase or by an extracellular adenylate kinase, similar to cardiac endothelial cells (4) and nasal airway cells (7), thereby activating the P2Y2 receptor. Both kinases are secreted from human nasal epithelia and are suggested to play an important role in the interconversion and metabolism of extracellular nucleotides on airway surfaces (7). The mechanism in the proliferative response of ADP in A549 cells remains to be examined.
The two lung cell lines used in our study, bronchiolar BEAS-2B cells and alveolar A549 epithelial airway cells, originate from two distinct lung regions. These regions respond distinctly to different stimuli, such as steroids, inflammatory cytokines, or chemotherapy. Nevertheless, ATP/UTP and UDP stimulated proliferation of both cell lines, suggesting a common mechanism of action in the physiological responses of human lung cells to extracellular nucleotides. The maximal concentrations of extracellular nucleotides achieved on stimulation of epithelial cells were calculated to be 5–10 μM (see Ref. 40 and references therein). These concentrations, determined for the release of ATP from resting cells, appear to be sufficient to stimulate proliferating epithelial cells, as detected in our experiments. It is conceivable that even higher ATP concentrations, i.e., 50–100 μM, can be reached under conditions where the epithelial cell layer is disrupted (ventilation injury) or under inflammatory conditions, where proliferation of these cells is necessary to regain an intact barrier. BEAS-2B and A549 cells serve as in vitro models for bronchiolar and alveolar cells, respectively. Therefore, a further in-depth analysis, similarly studying the regulation of BEAS-2B cell proliferation by extracellular nucleotides, will be important for understanding bronchiolar physiology.
The P2Y2 receptor has been described to have an important role in control of proliferation of different cell types. Interestingly, activation of the P2Y2 receptor has been reported to exert contrasting effects. It can lead to proliferation, as in MCF-7 breast cancer cells (47) and C6 glioma cells (44), or to apoptosis, as in colon carcinoma cells (13), endometrial cells (20), and esophageal cancer cells (31). In these cells, the P2Y2 receptor signals through the MAPK pathway.
Our results indicate that, in A549 and BEAS-2B human lung cells, the Gq protein-initiated signaling by nucleotides eliciting proliferation is via NF-κB activation and does not involve the classical Ras-Raf-MAPK pathway, as in astrocytes or C6 glioma cells (36, 44), because the specific MEK1/2 inhibitor PD-98059 did not block nucleotide-mediated proliferation. In addition, inhibition of the phosphatidylinositol 3-kinase pathway with wortmannin or LY-294002 also did not affect nucleotide-stimulated lung cell proliferation. This pathway had been shown to play an important role in P2Y2 receptor-mediated activation of MAPK in human monocytes (39) and renal mesangial cells (16).
Furthermore, in A549 cells, P2Y2 receptor-mediated proliferation is not dependent on PKC activation and general tyrosine kinase activity, because the PKC inhibitors chelerythrine and GF-109203X and the tyrosine phosphorylation inhibitor genistein were ineffective. In A549 cells, a Ca2+-dependent activation of NF-κB or CaMKII-triggered phosphorylation of the EGFR may be one of the main signaling pathways. Alternatively, CaMKII may directly translocate to the nucleus, thereby acting as a trigger for centrosome duplication or coordinating centrosome formation and DNA replication (34). CaMKII and NF-κB activation in A549 cells, however, do not involve PKC activation or an intermediate tyrosine kinase, which has been found for the P2Y2 receptor-stimulated proliferation of C6 glioma cells (44).
Signaling of the P2Y receptor through activation of NF-κB or CaMKII during regulation of proliferation has not been reported. CaMKII has only been found to be involved in the P2Y receptor-mediated enhancement of lipopolysaccharide-induced activation of IκB kinase in macrophages (5). Involvement of NF-κB in P2Y receptor signaling was detected in UTP-induced activation of nuclear factor of activated T cells, initiating IL-6 expression in vascular smooth muscle cells (1), and in UTP-initiated prostaglandin E2 release from macrophages (5). NF-κB activation was found for P2X7 receptor-mediated apoptosis of endothelial cells (45) and microglia cells (9). Thus the extracellular nucleotides ATP and UTP stimulate proliferation via a novel pathway in A549 and BEAS-2B cells. EGFR transactivation, suggested by the inhibitory effect of AG-1478, may also contribute to ATP-stimulated proliferation by activation of NF-κB. It is important to note that our results were obtained with proliferating cells. Whether this mechanism is also active in resting cells remains to be examined.
To further understand the mechanism of control of proliferation of lung epithelial cells by nucleotide receptors, we investigated the effects of the cytostatic drugs cisplatin, etoposide, and paclitaxel. These substances inhibit cell cycle progression and form the basis of current chemotherapeutic regimens in lung cancer treatment. The concentrations of the three anticancer drugs used in our study match those used for investigation of cell cycle inhibition of A549 cells (6). Paclitaxel and etoposide cause cell cycle arrest in A549 cells at the G2/M phase through interference with disassembly of microtubules (paclitaxel) or inhibition of spindle formation and topoisomerase II (etoposide) (6, 30). A consistent block in 40% of the cells at the G2/M phase after 24 h of application was found with 10–30 nM paclitaxel, and an induction of apoptotis was found in 20% of the cells with the higher concentration (30). Etoposide at 5–30 μM exerted a block at the G2 phase in A549 cells without significant apoptosis (6). In contrast, cisplatin does not induce G2 arrest in A549 cells but cross-links with DNA, forming intra- and inter- strand adducts (6). Cell death is believed to be mediated by recognition of the cisplatin-DNA adducts by cellular proteins (19).
We found a specific interaction of ATP, but not UTP, with control of proliferation by the anticancer drugs in A549 cells. ATP selectively potentiated the effect of cisplatin and most effectively antagonized the cell cycle-blocking activity at low paclitaxel and etoposide concentrations, where no significant apoptotic or cytotoxic activity of the anticancer drugs at A549 cells was found (6, 30). The antagonistic effects of ATP on the action of paclitaxel/etoposide are consistent with the assumption that ATP stimulates growth by directly promoting cell cycle progression from the G1/S phase to beyond the G2/M phase. Furthermore, stimulation of cells to progress into the S phase leads to more DNA in the uncoiled state, which then forms adducts with cisplatin. This might explain enhancement of the antiproliferative activity of cisplatin by ATP. The present results using these drugs are supportive but do not provide definitive evidence for direct control of cell cycle progression by extracellular nucleotides in A549 cells. The molecular mechanisms or the proteins mediating the action of ATP are not known. Possible candidates, which are known to be regulated in A549 cells by low concentrations of paclitaxel or etoposide, include proteins regulated by the tumor suppressor protein p53 and c-jun (p21WAF1 and cyclin B1) and the metastasis suppressor gene KAI1 (6, 32).
Cytokines influence UTP-stimulated, but not ATP-stimulated, proliferation of A549 cells. Thus pyrimidine nucleotides preferentially interact with the signal transduction of inflammatory cytokines. Different interactions of extracellular nucleotides with cytokines have been described for macrophages, monocytes, and dendritic cells, where cytokines regulate inflammatory or immune responses. In other cells, such as fetal human astrocytes, P2 receptors modulate IL-1β- and TNF-α-stimulated activation of NF-κB and activator protein-1 (29), supporting a regulatory role for the P2 receptors in inflammatory reactions in the human central nervous system. Moreover, in human bronchial epithelia, activation of Cl- channels by UTP is enhanced by IL-4 (10), and in vascular smooth muscle cells, IL-1β induces upregulation of the P2Y2 receptor (15).
In conclusion, our results, for the first time, show that extracellular nucleotides exert mitogenic effects on human lung epithelial cells through ATP/UTP-mediated activation of the P2Y2 receptor and the UDP-activated P2Y6 receptor. Stimulation of the P2Y2 receptor leads to activation of CaMKII and NF-κB. This novel pathway is modulated by extracellular Ca2+ and, possibly, involves EGFR transactivation. Understanding the mechanisms of interactions of extracellular nucleotides with anticancer drugs, growth factors, and cytokines will be an important task, because non-small cell lung cancer cells, such as A549 cells, rapidly develop resistance to conventional chemotherapy.
This work was supported by Deutsche Krebshilfe Project 10-1754, Land Sachsen-Anhalt, and Fonds der Chemischen Industrie.
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
- Copyright © 2003 the American Physiological Society


text:
1637 m² Land available in Paradise
Hazel Eksteen
Lew Geffen Sotheby's International Realty-Woodmill Lane Knysna
1637 m² Land available in Paradise
JOINT MANDATE. This rare and unique plot for sale in Knysna is situated in Paradise, against the hill, full of beautiful indigenous trees. The trees in the street past the house forms a wonderful green canopy. Due to the size and shape of the plot, one can create a very private retreat and live in this fantastic environment in peace and quiet.
The property is just above town and the Knysna CBD, near shops, schools and entertainment areas but still secluded and away from any traffic. Neighbouring houses are already built and established so the future house on this property can be placed in the ideal spot to get the best view through the trees of Knysna town and lagoon. A design concept has been drawn up but there is huge potential to design a unique house here. Excellent property in one of the most beautiful and sought after areas in Knysna where you can build your dream house
Email the Agent or request the agent Call You Back should you require any further details.
Share this Property
Need a Home Loan?
|Loan Value (Capital)
|Total Interest
|Total Repayment
Experience Knysna
Nestled between the Knysna estuary, the Indian Ocean and a vast indigenous forest, this town is full of natural beauty. With three magnificent golf courses, it is a haven for golfers. The waterfront is a popular tourist destination for great food and shopping. Known for its oysters, this town has festivals and events for all kinds of interests. An ideal holiday destination or retirement place, experience tranquility and a safe, small-town life.Find out more about Knysna
Need a Home Loan?
3D View
Video Tour
Apply for a Home Loan
Contact Agent

subset: commoncrawl


text:
2316 W Garvey Avenue, West Covina CA, 91790 (Land Lot)
Sold Price:
Status: Sold
Last Updated: 11/20/2017 3:19 PM
Description for 2316 W Garvey Avenue West Covina, CA 91790
Sold Price:
Sold Date:09/16/2015
Los Angeles
Listing Information
Room Information
Exterior / Lot Features
Raw La
Financial Considerations
Listing Price History
West Covina Real Estate
West Covina is located in California. West Covina, California 91790 has a population of 108,484.
The median household income in West Covina, California 91790 is $67,069. The median household income for the surrounding county is $55,870 compared to the national median of $53,482. The median age of people living in West Covina 91790 is 36.6 years.
Loading nearby sales and history ...
Ask a Question?
Start a no spam email conversation
My Drive Time
Enter your work address to view actual drive times with traffic for this property
Here is the weekday INRIX Drive Time from this property to .
When you register, you will receive ...
Weekday drive times
Plus all these great features
Register or login today to take advantage of Drive Time
West Covina Real Estate
Nearby Communities
Nearby homes for sale
Recent sales nearby


text:
With First Tutors you can find a private french tutor in Choppington and anywhere in the UK. Local french tuition is an excellent way to boost confidence while improving grades and doesn't have to be boring! Please use our service to search for Choppington french tutors and select the one who best suits your needs.
First Tutors is the best place to discover through one search the most suitable Choppington french tutors for your needs, enabling you to find a local Choppington french teacher for any subject, for example, GCSE English, A-Level Maths or even university Physics. We also help with language tuition, so if you want to learn Spanish, take a Russian course or prepare yourself for Spanish life, we can help - and all in Choppington! Our search will order french Choppington teachers in your area according to their french tuition charges and distance from you.
If you are a Choppington french tutor wishing to offer your local french tuition services in Choppington or the UK, please register with First Tutors or find out more about us here. Home french tuition is a fantastic way to re-affirm your own french knowledge while helping someone else, all in your local Choppington area!


text:
Notes on the Bankruptcy Problem: an Application of Hydraulic Rationing
We offer a new approach to the well-known bankruptcy problem based on Kaminski's idea. With the help of hydraulic rationing we give a proof to Aumann and Maschlers theorem i.e. the consistent solution of a bankruptcy problem is the nucleolus of the corresponding game. We use a system of vessels and water and the principles of mechanics to show this fact. The proof is not just simple and demonstrative but also provides an insight how the nucleolus is constructed in such games.
If you experience problems downloading a file, check if you have the proper application to view it first. In case of further problems read the IDEAS help page. Note that these files are not on the IDEAS site. Please be patient as the files may be large.
|18 pages
|Date of creation:
|May 2011
|Contact details of provider:
| Postal: 1112 Budapest, Budaorsi ut 45.
Phone: (+36-1) 309-2652
Fax: (36-1) 319-3136
Web page: http://econ.core.hu
More information through EDIRC
References listed on IDEAS
Please report citation or reference errors to , or , if you are the registered author of the cited work, log in to your RePEc Author Service profile, click on "citations" and make appropriate adjustments.:
- Moreno-Ternero, Juan D. & Villar, Antonio, 2004.
"The Talmud rule and the securement of agents' awards,"
Mathematical Social Sciences,
Elsevier, vol. 47(2), pages 245-257, March.
- Juan de Dios Moreno Ternero & Antonio Villar Notario, 2003. "The Talmud Rule And The Securement Of Agents? Awards," Working Papers. Serie AD 2003-05, Instituto Valenciano de Investigaciones Económicas, S.A. (Ivie).
This item is not listed on Wikipedia, on a reading list or among the top items on IDEAS.
When requesting a correction, please mention this item's handle: RePEc:has:discpr:1123. See general information about how to correct material in RePEc.
For technical questions regarding this item, or to correct its authors, title, abstract, bibliographic or download information, contact: (Adrienn Foldi)
If you have authored this item and are not yet registered with RePEc, we encourage you to do it here. This allows to link your profile to this item. It also allows you to accept potential citations to this item that we are uncertain about.
If references are entirely missing, you can add them using this form.
If the full references list an item that is present in RePEc, but the system did not link to it, you can help with this form.
If you know of missing items citing this one, you can help us creating those links by adding the relevant references in the same way as above, for each refering item. If you are a registered author of this item, you may also want to check the "citations" tab in your profile, as there may be some citations waiting for confirmation.
Please note that corrections may take a couple of weeks to filter through the various RePEc services.


text:
Status: Closed
Know more about this business than we do? Cool! Please submit any corrections or missing details you may have.Help us make it right
We are a well established General Dentistry practice, with specializations in Oral Surgery and Endodontics. We are known for our quick and superb work, with very little wait time. We offer emergency 24-hour assistance for our patients and we can almost always make same day appointments for those that need it.
Be the first OregonLive.com user to add photos or videos of Soft Touch Dental
I am EXTREMELY fearful of going to the dentist and put it off until I'm in so much pain I can't stand it. That's what brought me to Soft Touch Dental of Woodburn. I discovered they have about 6...
Excellent service, and wonderful staff. I love how everyone is always smiling and and always cheerful. They make me feel comfortable and secure. Thank you for the wonderful work and support.
Bailey Will DDS can be found at N Cascade Dr 980. The following is offered: Doctors & Clinics. The entry is present with us since Sep 9, 2010 and was last updated on Nov 14, 2013. In Woodburn there are 22 other Doctors & Clinics. An overview can be found here.
Woodburn Dental Images is located at 980 N Cascade Dr, Woodburn, OR. This business specializes in Dentistry.


text:
prosessing silver ore
silver ore processing mills in the usa in the 19th century ...
Silver mining process. Silver is commonly extracted from ore by crushing, grinding, smelting or chemical leaching. Gulin crushers have a very important role in silver ores mining, especially in silver ores crushing process.Read More
processing silver ore at home
Smelting and Roasting Ores to recover gold, silver and other metals. The process works best when the ore is crushed to small pieces before roasting. ...Read More
home processing silver ore
processing silver ore at home. silver processing Encyclopedia Britannica. silver processing, preparation of the ore for use in various products.Read More
Occurrence and Production,Silver
Silver is usually extracted from its ores by the flotation process, and then refined by smelting. Silver is sometimes obtained from its ores by the cyanidation process and the amalgamation process. Cyanidation Process. Crushed ore is treated with a dilute cyanide solution to dissolve the silver.Read More
processing silver ore at home,BINQ Mining
processing gold ore at home Grinding Mill China. Home » Basic Ore Processing At the Gold ore Processing Plant, gold and silver are extracted from the ore.Read More
Processing silver ore at home,Gulin Machinery
Processing silver ore at home. Posted at: September 14, 2013 Dummies Guide to EASY silver bullion refining at home as a long Feb 19, 2012 Dummies Guide to EASY silver bullion refining at home as a long termRead More
Society of American Silversmiths,Silver Processing
For the recovery of New World silver, the Patio process was employed. Silver-bearing ore was ground and then mixed with salt, roasted copper ore, and mercury. The mixing was accomplished by tethering mules to a central post on a paved patio (hence the name of the process) and ...Read More
silver ore processing products
Silver Ore Processing, process crusher, mining equipment exports . Silver Ore Processing 56 Views. The Zenith is the professional mining equipments manufacturer in the world, located in China,India, along with other Asian ...Read More
Silver refining,YouTube
A micro-scale experiment explaining some principles related to the refining process of silver. Some Questions and answers: -Why use such a small container? =...Read More
how to extract silver from ore,Coal Surface Mining,SAMAC
how to extract silver from ore Description : Silver mining Wikipedia, the free encyclopedia Silver mining is the resource extraction of silver by mining.Read More
How to extract silver from mine,YouTube
How to extract silver from mine,YouTube ... FantasticRead More
silver processing,Britannica.com
silver processing, preparation of the ore for use in various products. Silver has long been valued for its white metallic lustre, its ability to be readily worked, and its resistance to the corrosive effects of moisture and oxygen.Read More
processing silver ore chlorargyrite
News. Chlorargyrite,Wikipedia, the free encyclopedia. Chlorargyrite is the mineral form of silver chloride (AgCl). Chlorargyrite occurs as a secondary mineral phase in the oxidation of silver mineral deposits.Read More
Smelting and Roasting Ores to recover gold, silver and other ...
Interested in The processes of smelting or roasting your ore to recover its gold and silver? Some fairly rich ores require a lot of work to get all the gold and silver out them.Read More
Small scale silver mining processing,CGM Mining Solution
Small scale gold ore processing equipment for mining. Small scale gold ore processing equipment. Small... Small stone crusher has a wide range of application as mining equipment to crush hard gold ore, silver ore,Read More
home processing silver ore,MTM Crusher
Gold & Silver Ore Processing,SME. Gold & Silver Ore Processing. This course incorporates theory and practice, building from the knowledge and practical experiences that the presenters haveRead More
Gold and Silver Ore
Buy Gold Ore, Silver Ores, & Gold Refining Equipment. We sell gold and silver ore samples.Read More
processing silver ore,FAQ About Stone Crusher
Stock Footage Extracted ore of silver is transported, crushed and View processing of ore vintage historic stock footage in Ontario Canada, 1920.Read More
Ore Processing » Newmont Waihi Gold
Ore processing. At the processing plant, gold and silver are extracted from the ore. The processing plant alternates between treating ore from the Martha Mine open pit and from the Favona underground mine.Read More
How silver is made,material, making, history, used ...
Silver was first obtained in sixteenth-century Mexico by a method called the patio process. It involved mixing silver ore, salt, copper sulphide, and water. The resultant silver chloride was then picked up by adding mercury. This inefficient ...Read More
Fote Machinery: Silver Ore Processing Plant,Silver Ore ...
Silver ore processing plant or silver ore beneficiation plant is made up of jaw crusher, impact crusher, cone crusher, ball mill, tube mill, magnetic separator, mine flotation machine, drum screen, high frequency screen, spiral classifier, feeder, rotary dryer and mine mixing barrel.Read More
Next Post: Electrical Machines Used In Coal Mining


text:
A conglomeration of hospitals in one campus, including: the Sparsh Hospital for Orthopedics & Trauma, the Mazumdar Shaw Cancer Center, the Narayana Hrudayalaya Heart Hospital, the Narayana Nethralaya Eye Hospital, as well as a full fledged multi specialty hospital.
Junior Consultant- Dept of General Surgery
Apollo Bangalore is a modern, JCI accredited, multi-specialty hospital, capable of treating the most complex medical cases, including neurosurgy, bone marrow transplants, oncology and cardiac surgery. Apollo provides a full range of services for foreign patients.
Transplant Surgery
This information is intended for general information only and should not be considered as medical advice on the part of Health-Tourism.com. Any decision on medical treatments, after-care or recovery should be done solely upon proper consultation and advice of a qualified physician.
What Is Laparoscopy?
This is a minimally invasive surgical procedure used to examine the abdominal organs using a laparoscope. It does not necessitate large incisions in the skin and allows the doctor to examine the inside of your body without open surgery. A laparoscope is a long thin tube that has a high-resolution camera and high-intensity light attached at the front.
It is used to identify and diagnose pelvic or abdominal pain. Laparoscopy is usually performed when noninvasive tests such as ultrasound MRI and CT scan don't provide adequate information for a diagnosis. A biopsy may also be taken during the procedure.
Copyright © 2008 - 2017 Health-Tourism.com, All Rights Reserved


text:
Is it a good idea for a supervisor to date his or her subordinate? Of course not! Too many potential complications.
And yet … in a recent survey for CareerBuilder, the online job board, 16% of the 4,000+ respondents said they'd dated their boss. (Beyond those who 'fess up, you can be pretty sure there are more who have but don't want to say.)
From a supervisor's perspective, lots can go wrong if he or she – more likely he, according to the survey – romances an underling. Among the consequences:
- quid pro quo harassment complaints if the relationship goes south
- demoralizing perceptions of favoritism among co-workers, and
- damage to the supervisor's career if it leaks out that he/she is violating any policy on intra-office dating.
Notify if necessary
The point for supervisors is clear: Resist the siren song of a subordinate's attractions.
But if you can't – if the earth moves every time you see him or her – let your own superiors know about the relationship, and be prepared to accept a transfer of yourself, your co-romantic lead or both. That's the solution many organizations opt for.
Subscribe to the Leadership Blog
Get the latest research on workplace learning with weekly posts delivered to your inbox


text:
Loading...You are here: Home > 'complain'
University Election: An Untold Vendetta
In the wake of the brutal assault student leader of UKM, an alarming indicator has been reverberating throughout the nation on where the youth politics are leading nowadays. Just to keep you in the loop, local universities in Malaysia are entering election phase this month. With opposite factions of groups spearing contradicting ideologies, they determined […]Read More →


text:
Discussion content should not be viewed as an endorsement of the views of Tulsa City County Library. TCCL reserves the right to edit or refuse to post any material, in whole or in part, that does not meet these guidelines. TCCL reserves the right to make changes to the guidelines at any time.
See the Comment Guidelines for more information.
Join us for this upcoming workshop sponsored by Infogroup.
Wednesday May 20
1:30-3:30 p.m.
Hardesty Regional Library
8316 E. 93rd St.
Entrepreneurs will learn how to use ReferenceUSA, the leading provider of business and consumer research, to create marketing plans, conduct competitive analysis, find investors, source new employees and much more!
Class is limited to 18 on a first-come, first-served basis.
Registration is required. Call 918-549-7422 to register.
Tulsa's Young Professionals (TYPros) Foundation is seeking applications for grants from individuals and organizations for festivals, programs, events--something you think would make Tulsa and even more exciting place to be.
I don't know about you, but my "to read" list is completely out of control. I keep adding titles to the list (which I keep on my "for later" list in TCCL's catalog—you should try it if you aren't already taken advantage of that terrific feature), but never manage to actually read them. Oh, well, it makes me feel better just seeing them there, knowing that I've captured them online and that they can't easily escape me.
Sodexo Hiring Event*
· Seeking Baker III "Supervisory position", Cook I, Food Service workers, Cashiers, Custodian I opportunity
· Monday 5/18 @9:30
Tuesday 5/19, Wednesday 5/20, Thursday 5/21 @ 1:00pm
· Workforce Tulsa at Eastgate Metroplex
14002 E. 21st St., #1030, Tulsa, OK
· Please bring 2 forms of ID (Driver's License, Social Security card) and a resume, if available
Laser Quest is Hiring*
· Laser Quest is needing an energetic and fun Crew Member and an experienced Assistant Manager.
o Crew Member:
o Must be 16 years or older
o Energetic, outgoing and fun personality
o 15 hours/week with varying shift times
o Available all weekends
o $7.25/hour
o Assistant Manager/Shift Supervisor:
o 25 hours/week with varying shift times
o Open availability is required
o Experience managing a team
Library Apps TCCL Careers
400 Civic Center ~ Tulsa, OK 74103 ~ 918.549.7323


text:
Carol Cypher
Having studied,
explored and taught traditional, if not universal, techniques, provides
solid footing for venturing off on my own adventures. Adventures such as
pairing my two favored mediums, and sharing these adventures in my book,
Hand Felted Jewelry and Beads: 25 Artful Designs.
Benefiting by the
experience of teaching hundreds of beading classes and wishing to reach
more beaders than can fit in a classroom, I wrote Mastering Beadwork: A
Comprehensive Guide to Off-Loom Techniques.
Inspired by
Barbara Albright's knitting books, compilations of projects by several
designers and, seeking to feature the artful and elegant side of
felt making, drove me to write: How We Felt: Designs and Techniques
from Contemporary Felt Artists, also Interweave Press, and published in
October 2007.
Now that the
books are done,; I have the luxury of time. Time to felt and bead and,
of course, teach. Australia and Japan are recent additions to my
international teaching schedule. I continue to teach regularly scheduled
bead classes locally.
Wherever and
whenever we meet, I look forward to beading and felting with you.
Upcoming classes by
Take 2
half days
Take 1 1/2 days ~ $140
Take 2 days ~ $185
Take 2 1/2days ~ $230
Take 3 days ~ $275.00
Take 3 1/2 days ~ $325
Take all 4 days ~ $360


text:
Cillian's Return has gradually improved and gave supporters of the odds-on Black Hercules a fright at Punchestown. She'll be very difficult to beat though may be up against a potentially useful newcomer in VIGIL. Dermot Weld's charge is a son of Dansili out of a full sister to Yeats so has plenty to live up to and will hopefully be good enough. Gordon Elliott's pair Drunken Counsel and Sheemore Lad haven't shown much while Mick Spadoun has a fine chasing pedigree and will probably be seen to better effect later.
Calau (GER) 12-1 (11-7) Always behind, no impression from 3f out, never a factor, last of 11, 25l behind Mr Boss Man (11-7) at Punchestown 2m N H flat in Nov.
Texas Ray (IRE) Beneficial gelding out of Dusty Melody.
Cillian's Return (IRE) 10-1 (11-4) Mid-division, 6th halfway, headway into 3rd 4f out, ridden to challenge 1 1/2f out, soon under pressure in 3rd, 2nd 1f out, kept on same pace final furlong, not reach winner, 2nd of 12, 2 1/2l behind Black Hercules (11-9) at Punchestown 2m 4yo nhf in Dec.
Drunken Counsel (IRE) 33-1 (11-7) Mid-division, 16th 4 out, improved into 7th and ridden 2 out, no extra, 8th of 30, 26l behind Vautour (11-7) at Navan 2m 4yo mdn hdl in Dec.
Ekbaal Red Ransom gelding out of Tamazug.
Mick Spadoun (IRE) Spadoun gelding out of Mick Monika.
Sheemore Lad (IRE) 25-1 (11-2) Chased leader in 5th, ridden in 7th and no impression on clear leader entering straight, no extra from 2f out, last of 9, 20l behind Value At Risk (11-4) at Fairyhouse 2m 4yo nhf in Dec.
Vigil (IRE) Dansili gelding out of Magnolia Lane.


text:
Last evening, I hung my new collage creation on the fence in the backyard. I've been so delighted by my subconscious choice to plant a vine and a fig tree back there, and how this means I'm living into Micah's vision of the good stuff. Swords into plowshares and all that. No more war. Instead: vines and fig trees.
I used one of the nasty boards from the pile behind the shed--a piece of old trim that we tore out of the house. With all that ModPodge on there, it the grain looks pretty. New life, I suppose.
Now, I'm inspired to make another, but need to figure out what it should say or depict...
In other backyard news, I fashioned a set of four marshmallow roasters, with bent wire and bead decoration on the handles, too. They're hanging on a lovely holder. Now, the next time folks gather around our recycled brick firepit, and we want to make s'mores, we don't have to rummage for more wire hangers. They're already there and waiting.
Last weekend, I tore out most of the things that had been growing through the "cold season." (One has to write "cold season" with a little irony when she lives in San Diego.) So, I have lots of bare soil, waiting for things to sprout through it. But this corner, home to rhubarb, four tomatoes, strawberries growing full-tilt and some carrots I really ought to harvest, is GREEN. In the pot is a "yuca"--manioc. I planted one from the grocery store to see what would happen. It's growing!!
I continue to wait in anticipation for summer tomatoes. (If you remember those newspaper-pot tomatoes of long ago--they've gone back to the soil. But, thankfully, others grow on.)
And, a columbine plant is blooming. The poppies are still lovely. And, happily, the shortage of bees hasn't come to my yard. They're happily pollinating away here.
(Do you think Matt would let me keep a hive? He didn't go for the keeping-chickens plan...)
Thank God for growing things.
Friday, June 01, 2007
how my (kingdom of god?) garden grows
Posted by Molly Vetter at 10:03 AM
Labels: garden, garden art, theology
1 comment:
Subscribe to: Post Comments (Atom)


text:
The Sept. 27 news story "Hezbollah becomes wild card in Syria" failed to mention that Hezbollah is a terrorist group and was responsible for killing more U.S. citizens, prior to Sept. 11, 2001, than any other group. Specifically, among its many acts, Hezbollah was responsible for the 1983 bombings of the U.S. Embassy and U.S. and French barracks in Beirut that collectively killed more than 360 people.
In calling Hezbollah a "militant group," The Post is softening the image of a terrorist organization and failing to provide readers an understanding of what it is.
Hezbollah is a bunch of opportunist thugs who have received considerable funding from the governments of Iran and Syria and illegal activities around the world and who have killed and kidnapped many people who have opposed their views.
Richard Williams, Falls Church


text:
Only worn a couple of times for shoots.
It measures 21 inches and so would fit a waist of around 25 inches, I'd imagine more if you are waist trained. My waist was about 26 inches when I bought this so I guess you could make your judgement on that!
It's super comfy and gives amazing shape. I don't want to sell it at all, to be honest! But, alas, I'm saving up to go away.
I'm asking for £50 with free shipping to the UK, or £4 shipping to the rest of the world (where shipping usually costs me £4 to the UK and £9 to the rest of the world).
Any questions? Feel free to ask!
Jul 10 12 05:01 am Link
Any offers? Maybe £55?
Jul 11 12 04:15 pm Link
Posts: 5617
Marion, Iowa, US
Does the model come with?
Jul 11 12 05:07 pm Link
Haha well thank you! ;-)
Jul 12 12 05:34 am Link
Just reduced this! Still considering offers!
Jul 14 12 03:59 am Link
Posts: 5513
Saint Louis, Missouri, US
Damn! I'd buy it if I had a specific model in mind - and it was her size.
Jul 14 12 04:17 am Link
Posts: 42
Adelaide, South Australia, Australia
How much would it be in Australian dollars?
Jul 16 12 10:08 pm Link
Jul 21 12 04:08 am Link
Posts: 538
Warren, Ohio, US
Groovyness, bump!
Jul 26 12 09:26 pm Link
Posts: 185
Birmingham, Alabama, US
do you have any pics of it without the latex outfit (which I absolutely love by the way).
How much is that in US $? I just got 2 tiny packages from the UK this week - nice surprises from a friend.
Jul 26 12 10:04 pm Link
Posts: 185
Birmingham, Alabama, US
Aug 16 12 11:01 am Link
Posts: 23
Oakland, California, US
Tilcia wrote:
What she said. Pictures aren't loading, and the non-reply makes me think this item is no longer available..?
Sep 28 12 11:20 pm Link
Posts: 5
Toronto, Ontario, Canada
do you have pictures of it? the pictures won't load
Oct 16 12 09:51 am Link


text:
The picture shows the standing ovation given on the welcome of Rifleman Tull bahadur Pun VC, 6th Gurkha Rifles, to the UK by the Gurkha community, members of ARRSE and other military bulletin boards. A number of members of the public, unconnected with the armed forces, but who, nonetheless felt moved to action by the ARRSE campaign, joined in welcoming Tull bahadur Pun VC on 4th July 2007.
Pun VC Standing Ovation
Click on the photo to start tagging. Done Tagging
In This Album
XenGallery by sonnb


text:
When he was twelve years old, Adam Ryan went playing in the woods one day with his two best friends. He never saw them again. Their bodies were never found, and Adam himself was discovered with his back pressed against an oak tree and his shoes filled with blood. He had no memory of what had happened. Twenty years on, Rob Ryan - the child who came back - is a detective in the Dublin police force. He's changed his name. No one knows about his past. Then a little girl's body is found at the site of the old tragedy and Rob is drawn back into the mystery. Knowing that he would be thrown off the case if his past were revealed, Rob takes a fateful decision to keep quiet but hope that he might also solve the twenty-year-old mystery of the woods.
We've sent an email with your order details. Order ID #:
Listen on the Audible app or these devices.Launch Audible App
-or-Download Free Audible App
To access this title, visit your library in the app or on the desktop website.
Listen on the Audible app or these devices.Launch Audible App
-or-Download Free Audible App
Audible does not currently have the rights to sell this title in your country.
Please consider another book.Continue Browsing
Please add a payment method to continue.Add a Payment Method
Please edit your payment details or add a new card.
If the problem persists, contact customer service at (888) 283-5051Review My Payment Method
If the problem persists, contact customer service at (888) 283-5051
Please add your billing address to continue.Add My Billing Address
Please select a default credit card to continue.Select Credit Card
It looks like you've already used the specified payment method for a free trial.
Please contact customer service if you think we have made a mistake. (888) 283-5051Start Your $14.95 Membership
If the problem persists, contact customer service at (888) 283-5051
If the problem persists, contact customer service at (888) 283-5051
Audible does not currently have the rights to sell this title in your country.
Please consider another book.Continue Browsing
Don't miss this one, literary crime lovers...
In the Woods is one of the best audiobooks I've listened to - partly because I love the story & the writing (I've actually read it before) - but also because it's very well read.
The protagonist's back story is gripping & informs the present-day plot & main character's actions. Without giving anything away, the main character starts to lose the plot as past & present come together...
I usually prefer reading books but love to listen to audiobooks while driving or walking the dogs. One thing I don't like when listening to an audiobook is the (more old-fashioned) over-the-top storytelling style that quite a few readers have - almost like they're reading to children.
John McCormack reads in a more realist style, more like he's telling you the facts in a one-sided conversation. It draws you in. At the same time his voice is very clear & pleasant to listen to.
At first I thought his BBC accent was some sort of horrible mistake, as the character (told in first person) is Irish, but it emerges that he in fact (for an important reason) does have an English accent. It's a brilliant element of John McCormack's reading that his accent very subtly slides into a more Irish one as events unfold.
Many moving moments - & John McCormack doesn't overplay them - he's a subtle reader. Very effective.
The ending was almost ruined. As the narrator says his last words - voice low, atmospheric - a loud voice comes almost over his voice & intones 'Audible hopes you've enjoyed this program'. I needed a few beats to process what the ending was telling me - frustrating, & completely at odds with the style of book & the reading.
- Amazon Customer
A nice start to the series
Well worth listening.
The quality of writing continued to impress me.
John McCormack was a good, not great, narrator (but I am fussy). Make sure you get this version rather than the one narrated by Steven Crossley if chronic mouth clicks and whistles give you the creeps.
The lovely Marian Keyes (via Twitter) often heaps praise on Tana French, so I thought I should give her a go. I wasn't disappointed. BUT although this is a series, the next books are about different characters, so there isn't an on-going story arc as such. The second book lost me with its rather far fetched story. This one, though, is very enjoyable, but beware, not every loose end is tied up.
- Margaret


text:
Comedy night at Arts Centre
Begin your New Year with comedy on Friday, January 29, at Wexford Arts Centre with the Stars of Irish Comedy with Pat McDonnell, Danny O'Brien and Tom O'Mahony.
These are three of Ireland's best comics and they take to the stage for a brilliant two hour comedy show. Featuring a comedian who has performed in three of Ireland's most popular TV shows, Pat McDonnell, star of Channel 4's Father Ted (Eoin McLove), RTE's The Savage Eye, and Sky 1's Moone Boy. Tom O Mahony, who for many years has been a regular and highly-lauded stand-up fixture at the Edinburgh Fringe Festival.
More recently, he has gained much notoriety for his role as 'Tarquin', the intolerable toff in the RTE television series Damo and Ivor and finally Danny O Brien completes this line-up. Danny has been performing stand-up comedy for over six years, performing regularly at top comedy clubs and festivals in Ireland, the UK, Europe and the US.
Wexford People


text:
Heirloom Wooden Toys » Music Maker
Anyone can make beautiful music with no experience - really! Just slide a songsheet under the strings and pluck the string above each note. It's that easy! Our daughter Julia has had one since she was about 6 and still uses it at 15. Dad Alan, a professional musician, loves the sweet tones and immediate rewards. Includes pick, tuner, wire and 12 songs (using real musical notations). Songs are a mix of classical, folk songs, lullabies and children's songs. Made in Mexico.


text:
High Tech Fetish (NSFW)
Photographer Wolfgang Zajc created this series as part of an advertising campaign for an Austrian electronics website. If Best Buy ever ran a campaign like this I might actually shop there.
This post was created by a user and has not been vetted or endorsed by BuzzFeed's editorial staff. BuzzFeed Community is a place where anyone can post awesome lists and creations. Learn more or post your buzz!
Preview Your Response
Hot Buzz
Now Buzzing


text:
Competency-based education bill clears committee
worf Mcallen, TXMarch 8, 2013 3:37 p.m.
How's that standardized tests working out?
worf Mcallen, TXMarch 8, 2013 3:30 p.m.
Wow! 80% percent of New York graduates can't read---CBS News March 7,2013
How can teachers be blamed when law makers make the strategies. Competency-based education?
Let the teachers do their job, and quit trying to do it for them.
Seek to understand Sandy, UTMarch 6, 2013 12:00 a.m.
This bill is a vision of the future. We can't afford to force students into "seat time" for all of their credits. In a state that is so tight with funding, we need to be able to allow students to gain credits if they prove mastery in a subject, and not force them to take up a seat for a year in a class.
Supt. Menlove's comment makes me chuckle - yes, the State Board sets the standards, but there is no accountability to them now, so why the fuss? The State Board and USOE has no requirement that students are at mastery before they give credits now. Students often fail the CRT but are given the Algebra credit, or the Science or Language Arts credit. So to act as if there is this great system where the state sets accountability and it means something is a joke.
Schools can determine competency to the standards using their own criteria and measuring tools, and districts should be allowed to award credit if they deem a student competent using those measurements.


text:
Case No. 12-90-GA Notice Issued: March 22, 2013 James M. O'Briant, P 41156, East Lansing, Michigan, by the Attorney Discipline Board Ingham County Hearing Panel #1. 1. Reprimand 2. Effective March 22, 2013 The respondent and the Grievance Administrator filed ...Read More »
Public Utilities – DEQ permit to allow city to install new electrical unit upheld
The Department of Environmental quality adequately analyzed the types of fuels that could be used in a new electrical generating unit a city proposed to install in an existing power plant. As a result, the trial court's decision to uphold ...Read More »
Family Law – Parental rights to Indian child properly terminated
Respondent's parental rights were properly terminated because proper notice was given as required by the Indian Child Welfare Act, 25 USC 1901 et seq. Further, the trial court correctly determined that the ICWA does not apply to the child-custody proceeding ...Read More »
Criminal Law – Lay opinion testimony about video evidence properly admitted
Defendant's convictions of felony murder, armed robbery and carjacking were not tainted by the admission of lay opinion testimony about the identity of certain individuals depicted in a surveillance video, and in still-frame photos captured from the video. Contrary to ...Read More »
Criminal Law – Felon with firearm can assert self-defense
A felon possessing a firearm can assert self-defense if the evidence otherwise justifies a self-defense claim. In this case, the evidence shows that defendant was the initial aggressor, so the trial court properly refused to instruct the jury on self ...Read More »
Insurance – Hospital's antitrust settlement is 'loss' under policy provision
A hospital is entitled to indemnification for a settlement paid to nurses who sued the hospital for antitrust violations regarding their pay. The settlement is best understood as a loss covered by a policy defendant issued to the hospital. The ...Read More »
Civil Rights – Jury to decide whether traffic stop violated First Amendment
A plaintiff can get to the jury on a claim that defendant sheriff's deputy pulled him over in retaliation for displaying a political campaign sign on his vehicle's rear window. Hazel was driving his car, which had a large Ron ...Read More »
Insurance – Business use exclusion prevents claim against no-fault policy
A pizza delivery driver and her employer are not entitled to coverage and indemnification from the driver's no-fault insurer for any damages caused when the driver struck a bicyclist. A business use exclusion in the insurance policy applies even though ...Read More »
Van Bolt announces retirement from ADB
John Van Bolt, the executive director and general counsel of the Attorney Discipline Board, has announced his retirement effective Oct. 1, 2013. Van Bolt has led the ADB since 1986, and has been with Michigan's attorney discipline system since 1980. ...Read More »
Construction Law – Owners' intent to reside in home key dispute in lien foreclosure
The trial court incorrectly granted a bank summary disposition of a contractor's suit to foreclose a construction lien he had placed on property after not being paid for a seawall and other improvements he constructed. The work was done without ...Read More »


text:
Marketing Program Lead
Crossroads Evangelical Covenant Church
Woodbury, Minnesota, United States
Job Title: Marketing Program Lead
About Us: Crossroads Church is a rapidly growing, outward-looking, fast-paced, multi-site congregation that wants to be a church that unchurched people would want to attend, a church where church people really want to reach un-churched people who are distracted and disinterested in Jesus.
About You: As a marketing program lead you are solutions minded, you have room to grow professionally and spiritually, and you are an innovative and creative thinker. You enjoy working independently while still working well and collaborating within a team. The marketing program lead position stresses adaptability due to the fast paced, changing environment of Crossroads Church. The person who fills this role must profess a belief in Christ that is demonstrated by actively living out that faith.
About the Role: The Marketing Program Lead is a full-time, non-exempt role for Crossroads, and it offers a unique opportunity to be part of the development and ownership of the digital communication program. The Marketing Program Lead has
the opportunity to influence the look, feel, content, and format for all aspects of media, communication, and marketing for the internal and external constituents of Crossroads Church.
Key responsibilities include:
- Marketing program development: Develops program workflows, goals, and strategy for web, social media, and internal branded templates in partnership with the Executive Pastor of Church Development.
- Marketing and branding program delivery: Drives program success and accomplishes goals within time and budget as outlined in the strategic plan, this includes creating and providing contextual and targeted reporting to the organization.
- Marketing project management: Lead marketing projects based on needs as prioritized and outlined in partnership with the Executive Pastor of Church Development. Fun projects include website redesign.
- Brand management: Safeguards integrity and consistency of the Crossroads Church brand across all platforms. Additionally, the Marketing Program Lead ensures the brand is relevant and continues to grow.
- Vendor relationships: Lead vendor relationships according to program goals and deliverables as outlined in partnership with the Executive Pastor of Church Development. Central to this function is the ability to measure vendor performance against established key performance indicators.
Minimum Requirements:
- Bachelor degree in Marketing/Communications plus 2-4 years of business experience. Professional experience will be evaluated and considered in lieu of academic preparation.
- Proficient in MS Office, collaboration tools (e.g., SharePoint), social media platforms (e.g., Facebook, Twitter, and Instagram), web design applications (e.g., HTML and Concrete5), and graphics design applications (e.g., Creative Suite).
- Proven ability to drive projects from inception to completion.
Learn more about us: www.crossroadschurch.cc
Posting closes August 14th
You Also May Be Interested In:
Executive AssistantNovember 17
Cherry Hills Community ChurchHighlands Ranch, Colorado
Office AdministratorNovember 16
Huntertown UMC/LifehouseHuntertown, Indiana
Administrative AssistantNovember 15
West Ridge ChurchEau Claire, Wisconsin
Director of Men's ProgramsNovember 15
Grace Centers of HopePontiac, Michigan
Campus Children's Pastor/DirectorNovember 15
Real LifeOrlando, Florida
About Crossroads Evangelical Covenant Church
A growing and dynamic evangelical church in the Minneapolis/St. Paul area, with 2500+ in weekly attendance at multiple campuses.More...


text:
Xolo Mobile Service Center in Bhiwani
Xolo Mobile Service Center in Bhiwani, Haryana – Address, Contact Number are listed below. If you have any problem with your Xolo mobile, kindly contact the service center officials. There is 1 Xolo authorized mobile service center in Bhiwani
List of Xolo Authorized Mobile Service Centers in Bhiwani:
Xolo mobile service center in Bhiwani
Name: MOBILE CREATORS
Address: Shop No-12, Halwasia Mall, Opp. K.M School Hansi Gate,Bhiwani
City: Bhiwani
State: Haryana
Pincode: 127021
Contact Person: Parvat Raj Sharma
Contact Number: 8930000022
Email: firstname.lastname@example.com
Working Hours: 10:00 AM to 6:00 PM
Working Days: Monday to Saturday [Sunday Holiday]
* Call the XOLO Service Center before planning your visit, to ensure that the service center address is correct and is open for service.
** Please comment below if you find any of the above information incorrect, we will fix it immediately. Thanks in advance


text:
- select colour
it's an internet thing!
it's an internet thing!
18:47 04 March 2010
'Desperate Housewives' actress Drea De Matteo will not be returning to the series.
Producers of hit TV drama have confirmed she will not return. However, they denied reports she was fired due to falling ratings.
The former Emmy-winning 'Sopranos' star joined housewives Eva Longoria, Teri Hatcher, Marcia Cross and Felicity Huffman as fugitive Angie.
Rumours surfaced of the dismissal after De Matteo left the cast of Wisteria Lane, following only five episodes.
But show bosses have insisted it was merely due to her request to quit the show, to spend more time with her fianc, singer Shooter Jennings, and their two-year-old baby girl, Alabama.
Executive producer Marc Cherry said the reports were completely untrue, saying: "When I heard that stuff, I was stunned. We adore her. She is the sweetest gal in the world.
"Part of the deal when [we]) hired Drea was she was only interested in doing one season. She has a baby and is eager to get back to her life in New York," Cherry explained.
Speaking of how many fans had failed to warm to the Italian matriarch's icy character, Cherry said that had more to do with the storyline than the actress: "We shielded parts of her character because of the mystery.
"As we build toward the end and we start undoing the many layers of her, everyone will get how brilliant she is."
Producers declined to reveal how Angie will exit Wisteria Lane, but teased that the arrival in April of 'Torchwoods' John Barrowman as the man she has been on the run from will result in an "explosive finale" where "all the mysterious parts of her past will be revealed."
De Matteo's last episode is scheduled to air in May.
Disclaimer: Supanet is not responsible for, and disclaims any and all liability for the content of comments written by contributors to this website


text:
If you would like a self contained private family or couples villa then this is the place to go. It's centrally located lots to do there and around this wonderful place. The staff are so welcoming and caring its great they get out and talk to anyone. We meet a few people that stayed a couple of places away from us at warrick le lagon they said it was great but so expensive that they didn't eat there well we never had a problem we bought food and drink at the supermarket and cooked a few nights and went out to different places across the island. I can't speak highly of this place and also how clean it was. I will be going back as I think it was the best place to stay after looking at a few other places I was going to book and lily and Chris made everyone fill so welcome and even came out to have a cricket match and yes lily you should play in the women's Vanuatu team great batting and throwing. Anyway we will see you soon with the rest of the family.
Kristy, Nick, Emily and Heather
- Official Description (provided by the hotel):
- On the water's edge in a peaceful garden setting Pacific Lagoon Apartments is conveniently located, a little out of town, at the mouth of picturesque Erakor Lagoon, near other resorts and 4 kilometres from Port Vila town centre. Our eleven 2 bedroom apartments are fully serviced, have well equipped kitchens, satellite TV, in-house DVDs, kayaks and barbecue. Our friendly staff will look after you and PLA can be your home away from home. ... more less
- Reservation Options:
- TripAdvisor is proud to partner with Booking.com, Priceline, Hotels.com, Expedia, Travelocity, Cancelon, TripOnline SA and Hotwire so you can book your Pacific Lagoon Apartments reservations with confidence. We help millions of travelers each month to find the perfect hotel for both vacation and business trips, always with the best discounts and special offers.
- Also Known As:
- Pacific Lagoon Apartments Hotel Port Vila
- Pacific Lagoon Apartments Vanuatu/Port Vila


text:
Join Mature Nl! Download full-length 15min 39sec movie now! Please leave the link to our site! Thank you.
Redhead mature Evetta likes to spread her legs for us. This filthy whore has a huge sex drive so here she is taking care of it all alone on the bed. Damn the bitch likes masturbating! Let's keep her some company as she rubs and fingers that shaved cunt between her sensual milky white thighs


text:
Posted by: Jacques Desrosiers« on: May 30, 2014, 18:26:16 »
You write: "Nevertheless, it is doubtful just how much sense a touchpad makes in a conventional laptop."
But you mean: "touchscreen", don't you?
But you mean: "touchscreen", don't you?
Working For Notebookcheck
Are you a loyal reader of notebookcheck? Are you a techie who knows how to write? Then join our Team!
Especially wanted:
German-English-Translator - Details here
Review Editor - Details here
News Editor - Details here
See our Top 10 Notebooks:
» Multimedia, » Gaming, » Lightweight Gaming
» Budget Office, » Business, » Workstations
» Subnotebooks, » Ultrabooks, » Convertibles
» Top 10 under 300 Euros, » under 500 Euros
» Best Notebooks for University Students
» Top Windows alternatives to Apple's MacBook series
» Best Notebook Displays
Top 10 Tablets / Smartphones:


text:
TCP, HTTP and SPDY
Webinar: 4.61 Instructor: 4.88 Materials: 4.79 more …
Register to access this free webinar
Webinar details
TCP and HTTP, the transport and presentation-layer protocols used by almost all web-based applications, could have significant impact on web application responsiveness and page load time.
This webinar documents the characteristics of TCP and HTTP, and describes SPDY, a novel protocol that is currently the basis for HTTP 2.0.
Target Audience
If you're involved with web application development, deployment or operation, or have to plan or design underlying networking infrastructure, you simply have to attend this webinar.
The Problem: This section describes typical modern web application behavior, numerous HTTP requests hidden behind the scenes, and the impact of page load time on perceived application responsiveness.
TCP and HTTP 101 section focuses on TCP and HTTP characteristics that cause delays in web page load process, including 3-way TCP session establishment handshake, TCP slow start, and request-response nature of HTTP.
TCP and HTTP improvements section describes mechanisms used by web browsers (parallel TCP sessions) and servers (increased initial congestion window) to alleviate TCP drawbacks, as well as speedup mechanisms of HTTP (persistent sessions and pipelining).
SPDY: the last section of the webinar introduces SPDY, Google's new session-layer protocol. SPDY is already available on popular web servers and browsers, and used by some high-volume web sites. It has also been adopted by IETF for use in next-generation HTTP standardization efforts.
About the Author
Ivan Pepelnjak, CCIE#1354 Emeritus, is the chief technology advisor at NIL Data Communications. He has been designing and implementing large-scale service provider and enterprise networks as well as teaching and writing books about advanced technologies since 1990. He's author of several Cisco Press books, prolific blogger and writer, occasional consultant, and author of a series of highly successful webinars.
You'll get
- Recording of virtual classroom session
- Downloadable videos in MP4 format (list)
Webinar roadmaps
- SDN, NFV and OpenFlow
- Network Automation
- Cloud Infrastructure and SDDC
- Data Center Infrastructure
- Containers and Docker
- Virtualization and Virtual Networks
- IPv6
- Security
- Virtual Private Networks (VPN)
Custom webinars
To get more information about customized versions of this webinar, on-site workshops, pricing or scheduling details, please contact us.
Upcoming webinars


text:
His quizzes
User hasn't taken any quizzes
Profile link:
Story about Саша Осипчук
My name is Саша Осипчук, I'm male and 29 Years old. My Sign of Zodiac is aries. I live in Kostopil', Ukraine. On Flirtic.com I've been registered for 09.12.2012 years.
If our iterests match, you can send me a message, make a gift or invite for a date.
I speak russian. My mother tongue is russian.
Саша Осипчук
Was online 30.04.2014
He is not a VIP user. Send him VIP as a gift »
|Aim of acquaintance:
|Yet to be answered
|Interested in:
|Native language:
|Response rate
|very low
Do not participate in Top 100. Reason:
Has no new profile pictures


text:
Paperless billing
Every month, over 214,000 invoices are printed and delivered to Direct Energy Business customers just like you. Help reduce the paper clutter and your carbon footprint by signing up for paperless billing today. Enrollment is quick and simple.
To enroll:
- Log in to My Account
- Go to Preferences
- On the Invoice Preferences tab, check the "I have read and agree with the Paperless Billing terms and conditions" checkbox.
- All of your accounts will be listed in the table. Select the checkbox in the Paperless Billing column for each account you would like to enroll.
- Click the Save Invoice Preferences button
As a part of our commitment to corporate responsibility, we plan to reduce paper statements to conserve the following resources in just one year. Together, we can help make an environmental impact by saving:
- 3,595 pounds of paper
- 13,277 pounds of wood
- 15,723 miles driven in your car
- 39,941 gallons of wastewater from discharging into lakes, streams and rivers
- 172 household garbage bags filled with waste from paper production
You will receive monthly email reminders when your bill is ready for viewing through My Account. With paperless billing, you can view, download or print your statement online anytime at your convenience. After enrollment in paperless billing, you should start receiving paperless statements in your next billing cycle.


text:
At Uniplaces you can choose to rent your property in 3 different ways: the entire property, by bedroom or by bed.
- Entire property, if you want to rent your property as a whole.
- By bedroom, if you want to rent your properties' bedrooms one at a time.
- By bed, if your bedrooms have more than one bed, you can rent per bed. Each property can only have one rental type associated with it.
If you want to change the rental type of your property, contact us here.


text:
You have Adblock enabled. Adblock has been known to cause issues with site functionality. If you experience any difficulties, please try disabling Adblock. Close
Add to Favorite List:
Create a new folder:
Domme in nurses outfit strips and masturbates with dildo
Recognize a pornstar in this video?
Help make pornstars easier to find on Tube8 by telling us who is in this video.
Thanks! The pornstar names will be added to this video when enough users submit the same names.
You must enter a pornstar name that already exists in our database.
- Category: Amateur
- Views: 1,360
- Added: 3 years ago
- Description: Vixen brunette in red nurses outfit and stockings strips naked and masturbates. She fucks her wet pussy and asshole with her purple dildo and gets her men desiring her body as she dominates them in bed.
- Tags: kink kinky
- Pornstars: Add Pornstar
- Favorite by: 0
- Report This Video
Video Comments
All Video Comments (0)
Video Results For:
Sort by:
Sorry, we couldn't find any pages containing
Watch some of our most popular Tube8.com porn videos: Homepage or Category page


text:
If I could I would is a fast-paced piece of physical theatre featuring virtuoso acrobatics, imagination and humour that recognises it sometimes takes a superhuman effort to rise above the relentless demands of everyday urban life.
An all-female cast play an array of familiar characters in a day where the coffee is spilled, the commute is a trial, street-life is a threat and office politics get under the skin. From every woman to superwoman, If I could I would delivers a heart-warming message for everyone about resilience in the face of the everyday grind. If you could, what would you change?
The show is self-contained family friendly performance, created for outdoor festivals and events, with a small transformable set and cast of three performers.
Artistic Team
Artistic Director: Lina Johansson
Originally performed & devised: Silvia Fratelli, Alcina Mendes, Martha Harrison
Performed for tour 2017: Alcina Mendes, Martha Harrison, Rosie Roberts
Set Designer: Michalis Kokkoliadis
Costume Designer: Bettina John
Sound designer & composer: Isa Suarez
This piece was created using sounds from freesound, please find the full list at the bottom of the page.
If I could I would was created in 2016 and is available for bookings.
The audience say:
'You've inspired my little girl, she started running around and jumping into my arms!'
'That was great! Thank you for the fantastic show, we were impressed and laughed very much!'
'Very impressed! Loved the mix of acrobatics, kung fu, contemporary dance, capoiera. Silvia is hilarious.'
We've also been featured on Lucy Loves Circus' blog, you can read all about the show there too.
Supported by Arts Council England
Photo credits: Eric Richmond, Jamie Currey, Steve Eggleton, Mimbre
If I Could I Would uses these sounds from freesound:
Radio Zapping by Beskhu, Ambulance by Saphe, London Bus Approaches Leaves by icmusic, Piccadilly Circus Ambience by habbis92, Charing Cross Station Walk by Onj, Ambulance Siren by LanDub, Photocopy Machine by Tomlija, Stapler Usage by Callum_Sharp279, Keyboard Typing by Trollarch2, Post Office Walk 01 by Corsica_S, Post Office Ambience by akx, Walking Through Busy Streets by cusconauta, Phone by infobandit, Digital Alarm by Tempouser


text:
We recommend upgrading to the latest Internet Explorer, Google Chrome, or Firefox.
If you are using IE 9 or later, make sure you turn off "Compatibility View".
Handpicked brands for you
We've been stoked on @maxdubler's article on @skatehousemedia since last week: 6 things men in longboarding need to stop doing. Head to skatehousemedia.com to check it out. Awesome, Max. In the photo @emilylongboards being her rad self. Max Dubler...


text:
Race one
I started 5th while Connor started 8th.
The track was pretty good but very wet near the pole line.
I finished 1st while Connor finished 5th.
In the second race I started on pole while Connor started 4th.
I got 1st, While Connor did bloody well and got 3rd.
In the last race i started 9th and Connor started 3rd.
Connor managed to get to the front by the first corner but then got hit and spun in the first corner, he managed to fire it back up and got a few positions back. I finished 2nd.
Over all
1st Liam
2nd Clayton
3rd Cody
4th Connor
Unfortunatly The gopro decided to currupt the videos so the will be none from this meeting but still have to upload vi


text:
I want my entries to be fully justified both left and right.
sort by best latest
Doesn't appear to be possible, FT. Every time I try to enter HTML tags to align the text the capsule editor strips them out. It also doesn't seem keen on preformatted text that is centered or justified.
Although that may be the capsule editor not working to well with Open Office or Linux. I can only suggest you give it a try and see what happens for yourself, depending on what software you are using.
The only clumsy work-around that I can see is to write it as you want it on your computer and then take a screen grab of that and put that in a picture capsule.
Unless anyone else knows a way to make it happen.
You can help the HubPages community highlight top quality content by ranking this answer up or down.


text: For one of my contacts if I selected 'Edit' there are three Google icons and one Phone icon at the top of the screen. When I click the Phone icon at the top, it contains all of the info I want (and none of the info I don't want) for that contact. How do I remove the three Google icons, and what I am guessing are superfluous data due to some sort of name matching? Thx


text:
Former England batsman Pietersen gave insight into his communications with Amla and decorated the facts that make him one of the very best cricketers around.
Pietersen was quoted as saying by South Africa Cricket Magazine, "I remember him in a match in Hobart, when the Australian were well on top at tea. He came in and in his 60 or 70 balls, scored a forty and calmed the dressing room down. He comes in at No 3, does his job, occupies the crease and lets the rest of the order to play,"
Former Australian captain Michael Clarke said that Amla was not easy batsman to counter and he unloved captaining against him. "I hated captaining against him because he was so hard to set fields to," Clarke said. "He had a great defence and based his game around that but he found a way to be successful across all three formats."
"Like anyone who has played a hundred Tests, it has been a roller-coaster ride. The game of cricket is hard and there going to be ups and down and tough times. But it shows your character the way you get through that and turn things around. It's not always about statistics, sometimes it's about the way you played and the way you helped your team be successful, but his statistics show what a great player he is," He said.
Former West Indies spearhead Michael Holding wasn't faraway either from congratulating Amla on a amazing career. Amla have to be a great player to play a 100th Test matches because you are under analysis every time you go out there.
"What is important for a test cricketer is your commitment, your willingness to do hard work and your skill, and Hashim has all of those … you can see it in the way he approaches the game. He can take the attack to bowlers and target an area; he can adapt. That what makes him such a great cricketer."


text:
It's still Winter; it's still pretty cold. And, although all of our six inches of snow from last week melted away because of our recent "heat wave" (it was 45, don't worry), I know it's still a bit colder and snowy-er (yes, technical term) up North.
Zeidler Photography, a St. Paul/Minneapolis photographer, recently blogged an engagement session she shot, with some of the images indoors and others out in the snow. I couldn't resist not sharing with you. Many brides-to-be are none to thrilled to have a winter engagement session, but you can really have some fun with it. I urge more couples to go the Winter route!
I appreciate that they used an older sled. There are too many of those plastic ones out there (I know, you can go fast on them, I get it). Something about the vintage-y sled just gives those pictures character.
I LOVE hot chocolate, so of course I think those shots are adorable!
Images courtesy of Zeidler Photography
8 Comments
7:17 am on January 25th, 2010
9:00 am on January 25th, 2010
absolutely adorable!!
9:13 am on January 25th, 2010
glad you were able to post this! 😉 My favorite is the hot chocolate and sled shots!
12:56 pm on January 25th, 2010
I really like the hot chocolate shots too!
1:47 pm on January 25th, 2010
Love the shots with the mugs. So cute!
8:15 pm on January 25th, 2010
These are super cute!
9:44 am on January 26th, 2010
Makes me miss winter.
10:37 pm on January 26th, 2010
Love the sledding shots


text:
AvMap's EKP (electronic kneepad) is one of the most successful flight navigation systems sold worldwide.
20 years of expertise: In a market that is overcrowded with new players, AvMap distinguishes itself as one of the pioneers of GPS navigation.
The AvMap EngiBOX is a stand-alone Engine Monitoring System, displaying EGT, CHT / Coolant Temp, Oil / Air Temp, Oil pressure, RPM, MAP.
AvMap presents EKP V the next step in the evolution of aeronautical GPS, redefining the role of the GPS inside the cockpit.
Standalone EFIS with a 3.5" Display


text:
BEHA partner – Cranfield University
When the BEHA concept was born, there was only one University that instantly came to mind as the most likely place for us to be able to receive world class assistance in initial design analysis and that was Cranfield. As a startup company, we didn't have all the aerospace engineering resources that we would have liked in terms of personnel and equipment and so we turned to Cranfield University and the College of Aeronautics to help us in the initial analysis of our design.
With the assistance of an Innovate UK grant, we began work with the University, studying our innovative wing configuration along with a number of alternative configurations as a comparison. The majority of this work was completed via Computational Fluid Dynamics (CFD) to better understand the fluid mechanics of our proposed design. Over 65 simulations were run and we were absolutely delighted with our initial design performance, which we fine tuned with wing profile selection and stagger. We will continue to work with the University as we develop the BEHA design.
Cranfield is a postgraduate university, specialising in technology and management.
The theme running through University's 70-year history is a commitment to and passion for applied research which makes a difference to the world in the fields of science, technology and management.
The forerunner of Cranfield University was the College of Aeronautics, created in 1946 and based at the RAF station in Cranfield, Bedfordshire. Through the 1950s and 1960s, the development of many aspects of aircraft research and design led to considerable growth and diversification into other areas such as manufacturing and management.
In 1967, the School of Management was founded, so 2017 marks its 50th anniversary.
In 1969, the College of Aeronautics became The Cranfield Institute of Technology incorporated by Royal Charter with full degree-awarding powers.