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Androgen receptor (AR) transactivation is known to enhance prostate cancer cell survival. However, the precise effectors by which the prosurvival effects of androgen and AR drive prostate cancer progression are poorly defined. Here, we identify a novel feed-forward loop involving cooperative interactions between ligand-activated AR and heat-shock protein 27 (Hsp27) phospho-activation that enhance AR stability, shuttling, and transcriptional activity, thereby increasing prostate cancer cell survival. Androgen-bound AR induces rapid Hsp27 phosphorylation on Ser78 and Ser82 residues in an AR- and p38 kinase–dependent manner. After this androgen-induced, non-nuclear phospho-activation, Hsp27 displaces Hsp90 from a complex with AR to chaperone AR into the nucleus and interact with its response elements to enhance its genomic activity. Inhibition of Hsp27 phosphorylation, or knockdown using the antisense drug OGX-427, shifted the association of AR with Hsp90 to MDM2, increased proteasome-mediated AR degradation, decreased AR transcriptional activity, and increased prostate cancer LNCaP cell apoptotic rates. OGX-427 treatment of mice bearing LNCaP xenografts transfected with an androgen-regulated, probasin-luciferase reporter construct resulted in decreased bioluminescence and serum PSA levels as pharmacodynamic readouts of AR activity, as well as AR, Hsp27, and Hsp90 protein levels in LNCaP tumor tissue. These data identify novel nongenomic mechanisms involving androgen, AR, and Hsp27 activation that cooperatively interact to regulate the genomic activity of AR and justify further investigation of Hsp27 knockdown as an AR disrupting therapeutic strategy in prostate cancer. [Cancer Res 2007;67(21):10455–65]
- Hsp27
- androgen receptor
- chaperone
- p38
- antisense
- prostate cancer
The androgen receptor (AR), a ligand-dependent transcription factor and member of the class I subgroup of the nuclear receptor superfamily, plays a key role in prostate carcinogenesis and progression. The classic model of androgen-regulated AR transcriptional activity has not fully defined the many diverse effects of androgens on prostate cancer cell survival and growth. In response to androgen, cytoplasmic AR rapidly translocates to the nucleus and interacts with sequence-specific androgen response elements (ARE) in the transcriptional regulatory regions of target genes ( 1, 2). In addition to this transcriptional genomic action, androgens and other steroid hormones like progesterone and estrogen can exert rapid nongenomic effects that are not mediated through nuclear receptors, but rather initiated at the plasma membrane, presumably through surface receptors ( 3, 4).
Like progesterone (PR) and estrogen (ER) receptors, AR can interact with the intracellular tyrosine kinase c-Src to activate the mitogen-activated protein kinase (MAPK) pathway ( 5, 6). In androgen-sensitive LNCaP prostate cancer cells, AR interacts with the SH3 domain of Src within minutes of androgen treatment to promote cell survival, proliferation, and differentiation ( 5, 6). Androgens also rapidly stimulate Raf-1 and Erk-2, both components of the MAPK signaling cascade ( 5), suggesting that androgen-stimulated MAPK activation occurs via nongenomic mechanisms. This nongenomic action of androgen can also influence classic genomic AR activity, including modulation of AR by coactivators ( 7). Phosphoinositide-3-kinase (PI3K)/Akt is another pathway involved in nongenomic activity of ER and PR ( 8, 9), as well as AR ( 3, 10). In fact, androgen-activated PI3K and Akt enhance cell growth and survival in AR-positive cells, which can be inhibited by PI3K inhibitors or dominant-negative Akt. AR is found in a large protein complex with p85α-PI3K and Src, both required for androgen-stimulated PI3K/Akt activation ( 3, 10).
Molecular chaperones are involved in processes of folding, activation, trafficking, and transcriptional activity of most steroid receptors, including AR. In the absence of ligand, AR is predominantly cytoplasmic, maintained in an inactive but highly responsive state by a large dynamic heterocomplex composed of heat-shock proteins (Hsp), co-chaperones, and tetratricopeptide repeat (TPR)–containing proteins. Ligand binding leads to a conformational change in the AR and dissociation from the large Hsp complex. Subsequently, the AR translocates to the nucleus, interacts with coactivators, dimerizes, and binds to its ARE to transactivate target gene expression ( 11). Dissociation of the AR-chaperone complex after ligand binding is viewed as a general regulatory mechanism of AR signaling ( 11). Molecular chaperones remain important players in the events downstream of receptor activation and throughout the life cycle of the AR. For example, the Hsp90 inhibitor, geldanamycin, destabilizes AR and increases its proteasomal degradation, thereby decreasing the expression of AR-regulated genes ( 12). Recent reports further highlight the important roles of other co-chaperones on AR activation. Bag-1L is overexpressed in hormone refractory prostate cancer ( 13) and enhances the transactivation of the AR by using its NH2 and COOH-terminal domains to bind to COOH- and NH2-terminal sequences of AR ( 14). Another co-chaperone, FKBP52, also increases AR transactivation activity ( 15, 16), and FKBP52 knock-out mice exhibit defects in male reproductive tissues, including ambiguous external genitalia and defects in prostate and seminal vesicle development.
Another class of Hsps that complex with ER and glucocorticoid receptors (GR) are ATP independent and include Hsp27 ( 17). Hsp27 is a cytoprotective chaperone expressed in response to many stress signals to regulate key effectors of the apoptotic machinery, including the apoptosome, the caspase activation complex ( 18, 19), and proteasome-mediated degradation of apoptosis-regulatory proteins ( 20, 21). Antisense knockdown of Hsp27 delays prostate cancer xenograft growth and androgen-independent progression ( 22, 23). Although Hsp27 is induced by estrogens and glucocorticoids ( 24, 25), its relationship with AR and androgens is undefined. Here, we identify a feed-forward loop whereby androgen-bound AR induces rapid Hsp27 phosphorylation that, in turn, cooperatively facilitates genomic activity of the AR, thereby enhancing prostate cancer cell survival.
Materials and Methods
Cell culture and antisense oligonucleotide transfection. LNCaP cells are maintained in RPMI with 5% fetal bovine serum (FBS; ref. 26) and treated with Hsp27 antisense oligonucleotide (ASO) designated OGX-427 (OncoGenex Technology, Inc.) or mismatch control oligonucleotide (ODN) with OligofectAMINE in serum-free OPTI-MEM (Invitrogen Life Technologies, Inc.) for 20 min. Four hours later, 5% FBS was added. Cells were treated once daily for 2 successive days and harvested 48 h after the second treatment.
Plasmids, reagents, and antibodies. An Hsp27 wild type (wt) was subcloned into pcDNA3.1-GFP (Invitrogen-Life Technologies). The Hsp27 triple mutant (TM) was generated by introducing direct mutagenesis, replacing Ser15, Ser78, and Ser82 with alanine using QuikChange® II XL, site-directed mutagenesis kit according to the manufacturer's instructions (Stratagene). R1881 (Perkin-Elmer) was dissolved in 100% ethanol. Cyclohexamide, MG-132, SB 203580, and LY 294002 were purchased from Calbiochem; Hsp27, pHsp27, and Hsp90 were from StressGen; MDM2, AR N-20, and AR-441 were from Santa Cruz Biotechnology, Inc.; and pAkt/Akt, pp38/p38, and poly(ADP ribose)polymerase (PARP) were from Cell Signaling Technology.
Cell proliferation and apoptosis assays. LNCaP cells were plated in RPMI with 5% FBS and switched to charcoal-stripped serum (CSS) the next day with or without 10 nmol/L R1881. After a time course exposure, LNCaP cell growth was measured using the crystal violet assay as previously described ( 23). Detection and quantitation of apoptotic cells were done by flow-cytometric analysis as previously described ( 23). Each assay was done six times.
Western blot analysis and immunoprecipitation. Total proteins (500 μg) were pre-cleared with protein-G sepharose (Invitrogen Life Technologies) for 1 h at 4°C and immunoprecipitated with 2 μg of anti-Hsp27, anti-AR, or immunoglobulin G (IgG) as a control overnight at 4°C. The immune complexes were recovered with protein-G sepharose for 2 h and then washed with radioimmunoprecipitation assay buffer (RIPA) at least thrice, centrifuged, and submitted to SDS-PAGE, followed by Western blotting.
Immunofluorescence. LNCaP cells were grown on coverslips and treated ±R1881 for 15 min. For Hsp27 knockdown, cells were treated with 50 nmol/L Hsp27 ASO as described above. After treatment, cells were fixed in ice-cold methanol completed with 3% acetone for 10 min at −20°C. Cells were then washed thrice with PBS and incubated with 0.2% Triton/PBS for 10 min, followed by washing and 30 min blocking in 3% nonfat milk before the addition of antibodies overnight to detect Hsp27 (1:500) and AR (1:250). Antigens were visualized using anti-rabbit or anti-mouse antibodies coupled to FITC or rhodamine (1:500; 30 min). Photomicrographs were taken at 20× magnification using Zeiss Axioplan II fluorescence microscope, followed by analysis with imaging software (Northern Eclipse, Empix Imaging, Inc.).
Transfection and luciferase assay. LNCaP cells (2.5 × 105) were plated on six-well plates and transfected using lipofectin (6 μL per well; Invitrogen Life Technologies, Inc.). The total amount of plasmid DNA used was normalized to 2 μg per well by the addition of a control plasmid. Media were replaced by CSS ± R1881 24 h after transfection for another 24 h. Luciferase activities measured using the Dual-Luciferase Reporter Assay System (Promega) with the aid of a microplate luminometer (EG&G Berthold). All experiments were carried out in triplicate wells and repeated five times using different preparations of plasmids.
Northern blot analysis. Total RNA was isolated from LNCaP cells treated with OGX-427 or mismatch control, transfected with empty vector, Hsp27 wt, or Hsp27 TM, and treated with SB203580 before R1881 (10 nmol/L) treatment for 24 h using TRIzol/chloroform extraction (Invitrogen Life Technologies, Inc.). A 10-μg aliquot from each sample was subjected to horizontal electrophoresis and followed by hybridization with Hsp27 (700 pb) or prostate-specific antigen (PSA; 2 kb) cDNA probe as described ( 23) and human glyceraldehyde-3-phosphate dehydrogenase (GAPDH) cDNA probe for normalization of loading levels.
Electromobility shift assay. LNCaP cells were treated with indicated concentrations of OGX-427 or mismatch control. Forty-eight hours after the second treatment, nucleoplasmic proteins were extracted using CeLytic NuCLEAR Extraction Kit (Sigma). The nuclear extract (10 μg) was incubated in a final volume of 20 μL containing DNA-binding buffer [10 mmol/L Tris-HCl (pH, 7.5), 150 mmol/L NaCl, 1 mmol/L EDTA, 1 mmol/L DTT, 1 mmol/L phenylmethylsulfonyl fluoride, 20% v/v glycerol] and 1.5 μg of poly(deoxyiosinic-deoxycytidic acid) (Roche) for 10 min at room temperature and then incubated for 20 min at room temperature with double-stranded 32P-labeled (with specific activity of about 300,000 cpm) PSA-ARE oligonucleotide ( 27).
Chromatin immunoprecipitation. LNCaP cells were treated with 10 nmol/L of R1881 for 4 h and parafolmaldehyde cross-linked and sonicated. Chromatin immunoprecipitation (ChIP) assay was done using EZ ChIP kit according to the manufacturer (Upstate) on the PSA gene regions ARE I and ARE III as described in ref. 28.
Reverse transcription-PCR. About 4 μg of total RNA were reverse transcribed in cDNA using 100 pmol of random hexamer primers (Pharmacia) and Moloney murine leukemia virus reverse transcriptase (Life Technologies). The cDNA was successively amplified with two pairs of AR-specific primers. About 5 μL of cDNA were used as the starting DNA template in the PCR assay. To verify RNA quality, each sample was amplified with a set of primers specific to β-actin (5′-TGATCCACATCTGCTGGAAGGTGG-3′ sense and 5′-GGACCTGACTGACTACCTCATGAA-3′ antisense). Analysis of PCR products was done by electrophoresis in 2% agarose gel and visualized by ethidium bromide staining.
Protein stability. LNCaP cells were plated and treated with OGX-427 or mismatch control or transfected with Hsp27 wt or empty vector as described above. Media were changed 48 h later to RPMI + 5% serum containing 10 μmol/L of cyclohexamide incubated at 37°C for 2 to 6 or 16 h. Western blot was done using AR, Hsp27, and vinculin antibodies.
In vivo imaging of luciferase activity. Bioluminescence imaging was done with a highly sensitive, cooled CCD camera mounted in a light-tight specimen box (Xenogen Corporation) using the protocol as previously described ( 29). Before imaging, mice were injected i.p. with 100 mg/kg of the substrate d-luciferin and anesthetized with 1% of isoflurane. At 15 min postinjection, mice were imaged by IVIS 200, and bioluminescence was monitored by Living Image software.
Animal treatment. Male athymic nude mice (Harlan Sprague-Dawley, Inc.) were injected s.c. with 1 × 106 LNCaP-Probasin–driven luciferase cells. Once tumors were palpable with serum PSA levels ∼25 ng/mL and bioluminescence detectable using the IVIS-Imaging System, mice were treated with 20 mg/kg of OGX-427 or mismatch control i.p. once daily for 7 days. Tumor volume, serum PSA, and bioluminescence measurements were done at baseline and on days 4 and 8.
Statistical analysis. All data were analyzed by Student's t test. Levels of statistical significance were set at P < 0.05.
Androgens and Hsp27 protect LNCaP cells from apoptotic stress. Androgen is assumed to have an important role as a survival factor in prostate epithelial cells. The synthetic androgen R1881 enhances LNCaP cell survival in the presence of cytotoxic stress, including etoposide ( 30), cyclohexamide ( 10), and PI3K inhibitors ( 31). We recently reported that Hsp27 conferred resistance to androgen ablation in LNCaP ( 23) and next set out to explore whether androgen- and Hsp27-induced prosurvival activities were interrelated. Before beginning to study relationships between androgen and Hsp27, we first confirmed that R1881 added to androgen-depleted CSS increased LNCaP cell growth ( Fig. 1A, left ) and decreased apoptotic rates ( Fig. 1A, right) compared with CSS alone. R1881 protected LNCaP cells to paclitaxel treatment, lowering apoptotic rates and increasing cell growth (Supplementary Data S1). R1881 also protected LNCaP cells from apoptosis induced by OGX-427, a second-generation ASO that potently decreases Hsp27 levels ( 23). OGX-427–induced knockdown of Hsp27 was associated with decreased apoptotic rates in R1881-treated cells, as measured by fluorescence-activated cell sorting (FACS) and PARP cleavage ( Fig. 1B and C). Collectively, these results indicate that androgens suppress apoptosis induced by Hsp27 knockdown or paclitaxel.
Androgens lead to rapid Hsp27 phosphorylation via p38 MAPK pathway. Androgens stimulate growth and survival via both genomic and nongenomic pathways. Recently identified nongenomic effects include activation of Src, PI3K, and Akt ( 3, 10). Because Akt has been reported to phosphorylate Hsp27 in intact neutrophils ( 32), we tested whether androgen leads to Hsp27 phosphorylation. Interestingly, androgen induced rapid phosphorylation of Hsp27 on both Ser78 and Ser82 residues in a dose- and time-dependent manner ( Fig. 2A ). Within 15 min after incubation with 10 nmol/L R1881, Ser78 and Ser82 phosphorylation levels increased 3.2- and 5.3-fold, respectively ( Fig. 2A, right).
To identify upstream effectors of androgen-induced Hsp27 phosphorylation, we analyzed R1881-induced changes in p38 kinase and Akt phosphorylation levels, both previously associated with Hsp27 activation ( 32, 33). R1881 enhances p38 kinase and Akt phosphorylation in a time-dependent manner, with maximum stimulation at 15 and 5 min, respectively ( Fig. 2B, left and right). These results suggest that Hsp27 may be phosphorylated via p38 and/or Akt pathways. Preincubation of LNCaP cells with the p38 kinase inhibitor, SB 203580, abolished R1881-induced Hsp27 phosphorylation ( Fig. 2C, left), whereas the Akt inhibitor, LY 294002, did not alter androgen-induced Hsp27 phosphorylation ( Fig. 2C, right), indicating that androgen-induced Hsp27 phosphorylation occurs via the p38 kinase pathway.
Androgen-induced Hsp27 phosphorylation is AR dependent. To determine whether AR is required for androgen-induced phosphorylation of Hsp27, changes in Ser78 and Ser82 phosphorylation levels were analyzed after treatment with R1881 ± the anti-androgen bicalutimide. Bicalutimide inhibited R1881-induced Hsp27 phosphorylation at both sites ( Fig. 3A ), suggesting that ligand binding to AR is required for Hsp27 phosphorylation by R1881. Next, PC3 cells, which do not express endogenous AR, were transfected with increasing amounts of AR or empty vector with or without R1881. As shown in Fig. 3A, Hsp27 phosphorylation levels at both Ser78 and Ser82 sites increased with increasing AR levels after R1881 treatment. In AR-negative PC3 cells, Hsp27 phosphorylation is insensitive to androgens, but is enhanced when AR is transiently overexpressed in PC3 cells. These results confirm that AR is required for androgen-induced Hsp27 phosphorylation.
To further define mechanisms of androgen-induced Hsp27 phosphorylation, we determined whether Hsp27 and AR interact using coimmunoprecipitation for Hsp27 and AR in LNCaP cells. Hsp27 is detected in AR immunoprecipitated complexes, and conversely, AR is detected in Hsp27 immunoprecipitated complexes ( Fig. 3C, left). The domain of AR required for the interaction with Hsp27 was next analyzed using glutathione S-transferase (GST)–pulldown assays. GST pulldown after initial equimolar normalization of GST-AR domain fusion as previously described ( 34) indicates that Hsp27 binds with NH2-terminal and ligand binding (COOH-terminal) domains, but not the DNA-binding domain of AR (Supplementary Data S2). Furthermore, immunofluorescence illustrates that Hsp27 and AR colocalize in the cytoplasm of LNCaP cells cultured in the absence of androgens. Importantly, and of potential functional relevance, both proteins translocate and colocalize in the nucleus after R1881 treatment ( Fig. 3C, right).
Before ligand binding, AR exists in a complex with Hsp90 and other co-chaperones. The AR-Hsp90 interaction maintains AR in a ligand-binding conformation necessary for efficient response to hormone ( 35). Upon ligand binding, AR is released from Hsp90 and is translocated into the nucleus. Interestingly, AR immunoprecipitation blots show that shortly after androgen treatment, Hsp27 levels increase in complex with AR as Hsp90 levels decrease ( Fig. 3D), suggesting that upon ligand binding, phospho-activated Hsp27 replaces Hsp90 to chaperone AR into the nucleus.
Effect of Hsp27 on genomic activity of AR. Many proteins participate in the activation of AR, either by direct binding or as part of a tertiary complex regulating the activity of other transactivators. Because Hsp27 colocalizes with and shuttles ligand-activated AR during nuclear translocation, we next investigated the effects of phospho-Hsp27 levels on AR transcriptional activity. This was done by gain-of-function strategies using overexpressing wt Hsp27, as well as loss-of-function strategies using an Hsp27 ASO (OGX-427), dominant-negative Hsp27 phosphorylation TM, or an Hsp27 phosphorylation inhibitor (p38 kinase inhibitor, SB 203580). PSA transactivation assays were done using LNCaP cells transiently transfected with the luciferase reporter plasmid regulated by the PSA enhancer-promoter region (6.1 kb) in the presence or absence of an increasing amount of exogenous wt Hsp27 (0, 0.25, 0.5, and 1 μg). R1881 treatment increased AR reporter gene expression 34-fold, whereas wt Hsp27 overexpression increases androgen-stimulated transcriptional activity of PSA a further 3-fold ( Fig. 4A, left ). In contrast, Hsp27 knockdown using OGX-427 decreased the transactivation of the androgen-regulated PSA reporter in a dose-dependent manner, further supporting a role for Hsp27 in AR transactivation ( Fig. 4A, right).
To determine whether Hsp27 phosphorylation is required for R1881-induced AR-mediated gene activation, we overexpressed an Hsp27 TM (Ser15, Ser78, and Ser82 substituted by alanine) in a dose-dependent manner and tested its ability to affect R1881-stimulated PSA transactivation. As shown in Fig. 4B (left), Hsp27 TM transfection inhibited luciferase transactivation from the PSA enhancer in response to R1881. Similarly, inhibition of androgen-induced, p38 kinase–mediated Hsp27 phosphorylation using the p38 kinase inhibitor, SB 203580, suppressed R1881-mediated transactivation of PSA enhancer–driven luciferase activity ( Fig. 4B, right). Collectively, these results indicate that androgen-induced p38 kinase-mediated phosphorylation of Hsp27 is important for AR-mediated PSA expression.
The preceding data identified a role for phospho-Hsp27 in the transactivation of PSA. To provide further evidence to substantiate this hypothesis, Northern blot analyses were done ( Fig. 4C) and indicated that levels of AR-regulated endogenous PSA mRNA decreases significantly after Hsp27 knockdown (OGX-427; Fig. 4C, left), p38 kinase inhibition (SB 203580; Fig. 4C, middle), or Hsp27 phosphorylation inhibition (Hsp27 TM; Fig. 4C, right). These results confirm that Hsp27 levels and phosphorylation status enhance AR-mediated PSA mRNA expression.
As a transcription factor, AR translocates to the nucleus after androgen binding, where it interacts with ARE to transactivate its target genes. Suppression of Hsp27 levels or phosphorylation negatively regulates androgen-stimulated transcriptional activity of PSA. To determine whether AR interaction with its response elements is affected by Hsp27 levels, we next analyzed the effects of Hsp27 knockdown on AR binding to PSA-ARE using electrophoretic mobility shift assay (EMSA) with nuclear lysates of LNCaP cells treated with OGX-427. As shown in Fig. 4D, 10 nmol/L R1881 increased AR binding to ARE, and this effect was specifically blocked by excess cold PSA oligonucleotide. Hsp27 knockdown using OGX-427 decreased AR binding to its ARE, confirming that Hsp27 knockdown decreases AR binding to its ARE. Collectively, the preceding data indicate that phospho-activated Hsp27 plays a critical role in androgen-induced nuclear translocation and transcriptional activity of the AR.
We next explored whether Hsp27 forms a complex with AR on the ARE of the PSA promoter using ChIP assay in LNCaP cells ±R1881. β-Actin fragment was amplified as control. As shown in Fig. 4E, Hsp27 is present, along with AR (which serves as a positive control) on the AREs. In the presence of R1881, higher levels of AR and Hsp27 were recruited to the promoter proximal region (ARE I) and also to the enhancer region (ARE III) of the PSA promoter. These data indicate that Hsp27 continues to interact with the AR transcriptional complex at the level of the ARE and seems to be regulated by androgen.
Hsp27 knockdown induces AR degradation via the proteasome-mediated pathway. Hsp27 knockdown or inhibition of phosphorylation inhibits androgen-stimulated nuclear translocation of the AR with subsequent suppression of AR-regulated gene expression. To investigate the fate of AR after Hsp27 knockdown, changes in AR mRNA and protein levels after treatment with OGX-427 were evaluated. AR mRNA levels did not change after Hsp27 knockdown ( Fig. 5A, left ). In contrast, Hsp27 knockdown decreased AR ( Fig. 5A, middle) and Hsp90 ( Fig. 5A, right) protein levels in a dose-dependent manner. The effect of Hsp27 knockdown on AR protein stability was next evaluated using cyclohexamide, which inhibits protein synthesis. As shown in Fig. 5C, AR protein levels decrease significantly with rapid degradation after OGX-427–induced knockdown of Hsp27. In contrast, Hsp27 overexpression prolonged AR half-life compared with empty vector-transfected controls ( Fig. 5C). AR degradation after Hsp27 knockdown occurs via the proteasome pathway because treatment with the proteasome inhibitor MG-132 suppressed Hsp27 knockdown-induced AR degradation ( Fig. 5C). Taken together, these findings indicate that Hsp27 knockdown induces AR degradation via a proteasome-mediated pathway.
Hsp27 knockdown disrupts AR-Hsp90 association and increases AR-MDM2 association and ubiquitination. AR forms a heterodimer complex with Hsp90 to provide stability for ligand-unbound AR. Indeed, without Hsp90 binding, the unfolded protein will be recognized and degraded by the ubiquitin-proteasome system ( 12, 35). Hsp27 knockdown by OGX-427 significantly disrupts the association between AR and Hsp90 ( Fig. 5D, left). This indicates that Hsp27 knockdown–induced dissociation between AR and Hsp90 may render the AR-Hsp90 heterocomplex vulnerable to degradation by the proteasome-mediated pathway. Because AR has been reported to be ubiquitinated by the E3 ligase, MDM2 ( 36, 37), we next set out to determine whether interaction between MDM2 and AR increased after OGX-427–induced Hsp27 knockdown. As predicted, Hsp27 knockdown increased the association between endogenous MDM2 and AR as shown by co-immunoprecipitation experiments in Fig. 5D (middle). Furthermore, Hsp27 knockdown increased levels of ubiquitinated AR ( Fig. 5D, right). These results indicate that OGX-427–induced Hsp27 knockdown dissociates the Hsp90-AR heterocomplex and increases MDM2-mediated AR ubiquitination and degradation.
In vivo Hsp27 knockdown by OGX-427 decreases LNCaP proliferation rates, serum PSA levels, and AR client protein expression levels. Our results establish a novel mechanism whereby ligand-activated AR phospho-activates Hsp27 to cooperatively enhance AR nuclear translocation and transcriptional activity. Hsp27 knockdown leads to AR degradation and reduced PSA transcription and LNCaP cell growth in vitro. We sought to determine whether in vivo Hsp27 knockdown with OGX-427 decreased AR activity in LNCaP xenografts. Male nude mice were injected s.c. with 1 × 106 LNCaP-Probasin–driven luciferase-transfected cells (Supplementary Data S3), and once tumors were palpable with serum PSA levels ∼25 ng/mL and bioluminescence was detectable using the IVIS Xenogen monitor, mice were treated with 20 mg/kg of OGX-427 or mismatch control. A bioluminescent signal first became detectable when serum PSA levels reached ∼5 ng/mL and was easily detected at serum PSA levels above 20 ng/mL. Mice were subjected to three different types of measurements: bioluminescence of probasin-promoter–driven luciferase activity (a measure of in vivo AR activity in LNCaP xenografts), serum PSA levels, and tumor volume. Measurements were done at baseline before treatment (day 0), during treatment (day 4), and at the end of treatment (day 8). Beginning day 4, OGX-427 decreased bioluminescence and reduced circulating PSA levels by 60% (P < 0.01), with a slight 15% decrease in tumor volume ( Fig. 6A and B ); individual data are presented in Supplementary Data S4. In contrast, mice treated with mismatch control ODN showed an anticipated increasing trend on bioluminescence and PSA levels, coincident with increasing tumor volume during the 8-day treatment period. Western blot of snap-frozen xenograft samples ( Fig. 6C) and immunostaining data (Supplementary Data S5) indicate that OGX-427 decreased LNCaP xenograft levels of AR, Hsp27, and Hsp90. In addition, OGX-427 also decreased Ki67 staining (Supplementary Data). Collectively, these data suggest that the in vivo anticancer activity of OGX-427 results, in part, from AR disruption, and that serum PSA may prove a useful surrogate of pharmacodynamic activity as OGX-427 moves into human trials.
Hsp27 is an ATP-independent chaperone that is phospho-activated by cell stress to prevent aggregation and/or regulate activity/degradation of certain client proteins. The chaperone activity of Hsp27 is regulated by stress-induced changes in phosphorylation and oligomerization ( 38). As a cytoprotective chaperone situated as a Hub at the center of many pathways regulating cellular response to therapeutic stress, targeted inhibition of Hsp27 would inhibit many pathways implicated in cancer progression and resistance. The cytoprotective effects of Hsp27 result from its ubiquitin binding and degradation of IκB ( 20), direct interference of caspase activation, modulation of oxidative stress, and regulation of the cytoskeleton ( 19, 39). Higher levels of Hsp27 are commonly detected in many cancers including prostate ( 22, 23, 40) and is associated with metastasis, poor prognosis, and resistance to chemotherapy or radiation ( 41, 42). We recently reported that overexpression of Hsp27 in LNCaP cells suppressed castration-induced apoptosis and confers androgen resistance ( 23), whereas Hsp27 knockdown using ASO potently decreases Hsp27 levels, increases caspase-3 cleavage and apoptosis, enhances paclitaxel chemosensitivity, and delays tumor progression in vivo ( 22, 23).
Human Hsp27 is phosphorylated on three serine residues, Ser15, Ser78, and Ser82. p38 kinase and Akt were reported to be the Hsp27 kinases ( 32, 33), although protein kinase C (PKC) α and δ and cyclic AMP–dependent kinase can also phosphorylate Hsp27 ( 43). Hsp27 becomes phosphorylated when exposed to angiotensin ( 43), interleukin-6 (IL-6), IL-1, heat shock and tumor necrosis factor-α ( 44). Using LNCaP cells, which express AR and are sensitive to androgens, we show that androgens increase phospho-Hsp27 levels within minutes in a dose-, time-, and p38 kinase pathway–dependent manner. This rapid androgen/AR-mediated Hsp27 phosphorylation identifies a nongenomic mechanism for AR in LNCaP cells. These findings are in agreement with previous studies indicating that steroids can act via classic steroid receptors or through atypical membrane receptors, and that AR mediates nongenomic activation in response to androgens ( 6, 10). Ligand binding to AR induces its association with Src via Src SH3 domain and AR proline-rich domain, triggering Src-dependent pathway activation ( 6, 10). In AR-negative COS-1 and PC3 cells, transfection of AR is necessary for R1881 induction of c-Src/Raf/extracellular signal-regulated kinase and Akt, respectively. In AR-positive LNCaP cells, the anti-androgen bicalutamide suppresses R1881-stimulated Hsp27 phosphorylation. In AR-negative PC3 cells, Hsp27 phosphorylation is insensitive to androgens, but is enhanced when AR is transiently overexpressed in PC3 cells. This nongenomic stimulation is supported further by the interaction and colocalization of Hsp27 with AR. Hsp27 binds with AR via its NH2-terminal domain and may directly or indirectly involve other AR coregulators, including signal transducers and activators of transcription 3 (STAT3) and ARA55, both of which have been reported to associate with Hsp27 ( 45, 46).
In the classic model of androgen action, in response to androgens, AR dissociates from Hsp90 ( 11). Interestingly, we found that following androgen treatment, Hsp27 becomes more abundantly associated with AR as AR dissociates from Hsp90, suggesting a dynamic role for molecular chaperones in AR shuttling. This nongenomic action of AR ultimately influences its classic genomic effects. Once in the nucleus, ligand-bound nuclear receptors like AR are recruited to target gene promoters either through direct binding to hormone response elements or association with other promoter-bound transcription factors ( 11). Many proteins participate in the activation of AR, some directly binding to AR, and others via a tertiary complex with other transactivators. Hsp27 has now been identified as a chaperone interacting with the AR transcriptional complex at the level of its ARE. Recent reports indicate that the AR coactivators STAT3 and ARA55 also interact with Hsp27 ( 23, 45). Hence, Hsp27 and AR may complex with either ARA55 or STAT3 to cooperatively promote AR translocation and transactivation. Interestingly, PSA transactivation in LNCaP cells is enhanced by Hsp27 overexpression and suppressed by Hsp27 knockdown or inhibition of Hsp27 phosphorylation. Similarly, in ER+ MCF-7 breast cancer cells, inhibition of p38 kinase signaling also blocks ER-mediated transcription by inhibiting nuclear translocation of ERα ( 47). Furthermore, Hsp27 knockdown decreases AR nuclear translocation and binding to its ARE.
Previous studies emphasize the importance of Hsps in steroid receptor stability. For example, Hsp90 inhibitors such as geldanamycin induce steroid receptor degradation by directly binding to the ATP-binding pocket of Hsp90 and thereby inhibiting its function ( 12, 35). Results shown in Figs. 5 and 6 indicate that Hsp27 knockdown destabilizes AR by inducing the dissociation of the AR/Hsp90 heterocomplex and increasing AR association with the E3 ligase MDM2, with subsequent ubiquitin-proteasome–mediated AR degradation. Although both OGX-427 and geldanamycin share the ability to abrogate the interaction between AR and Hsp90, they do so by different mechanisms. Interestingly, OGX-427 induces the degradation of Hsp27, AR, and Hsp90, whereas geldanamycin binding to the ATP-binding pocket of Hsp90 inhibits its chaperone activity ( 48), induces degradation of client proteins ( 49), and is accompanied by stress-activated increases in Hsp70 and Hsp27 ( 50).
The effects of OGX-427 on AR expression and activity in vitro were recapitulated in vivo. OGX-427 induced rapid decreases in probasin-luciferase reporter–driven bioluminescence that correlated with early decreases in serum PSA and decreased LNCaP xenograft levels of AR and its chaperones, Hsp90 and Hsp27. In prostate cancer where the AR is critically important, ligand-activated AR leads to rapid p38 kinase–mediated phosphorylation of Hsp27, which, in turn, complexes with and chaperones the AR to enhance its stability, shuttling, and transcriptional activity. OGX-427–induced knockdown of Hsp27 destabilizes the AR and enhances its ubiquitination and degradation. Collectively, these data identify a novel mechanism for Hsp27 as an AR chaperone (as summarized in model illustrated in Fig. 6D) and justifies further investigation targeting Hsp27 as therapeutic for prostate cancer.
Grant support: Terry Fox Foundation of the National Cancer Institute of Canada and the NIH Pacific Northwest Prostate Specialized Programs of Research Excellence.
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
The University of British Columbia has submitted patent applications, listing Dr. Gleave as inventor, on the antisense sequence described in this paper. This IP has been licensed to OncoGenex Technologies, a Vancouver-based biotechnology company that Dr. Gleave has founding shares in.
We thank Virginia Yago and Mary Bowden for their technical assistance in animal experimentation.
Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/).
- Received June 4, 2007.
- Revision received July 25, 2007.
- Accepted August 22, 2007.
- ©2007 American Association for Cancer Research.


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There it is, my second authentically French recipe, just for you! Those "Biscuits à la Cuillère" are among the most typical and old cookies here in France. The story indeed tells that they were once created by a cook named Antonin Carême whose master, a prince (Prince de Talleyrand) had some problems with his teeth… The cook then decided to create soft cookies his "master" could appreciate without a problem… And you will see that these cookies indeed are incredibly soft in the inside. But what makes them so irresistible is that they also are a bit crispy on the outside ! Powdered with icing sugar, these cookies are the perfect choice with a warm cup of good coffee, and young children do appreciate them as a snack, I give you my "mummy's word" !
Ingredients (for about 15 to 20 cookies) :
2 eggs
1/2 cup (60 g) of plain flour or all purpose flour
1/4 cup (50 g) of powdered sugar
Icing sugar
In a bowl, beat the two egg yolks (keep the whites, you'll use them later in the recipe) with a little bit less than half of the powdered sugar, until the yolks turn into a lighter yellow color (for about 3 minutes).
In another bowl or in the bowl of your stand mixer, beat the egg whites. As soon as the whites turn frothy, add the remaining powdered sugar little by little, beating the whites all along until stiff.
Softly add the yolks to the stiff whites with a spatula, mixing really gently.
Now is the step which explains why we call these cookies "Biscuits à la cuillère" which basically means "spoon cookies" in English.
Cover your oven tray with baking paper.
With two tablespoons, drop the dough on the tray in the shape of "sausages" about one inch wide by three inches long (3 cm wide by 7 cm long). Be careful to drop your sausages a bit distanced apart as they will rise when they cook.
Sprinkle some icing sugar on the "sausages" and let them stand for about 10 minutes. During this time, preheat your oven to 340 F (180°C).
When the 10 minutes are over, sprinkle again each sausage of dough with some icing sugar and place the tray in the oven. Let the cookies bake for about 12 minutes or until the edges are golden (not more than 15 minutes).
Let the cookies cool on the tray for some minutes and, with a spatula, gently transfer them on a wire rack to cool completely (a few minutes more). Do not worry if the cookies do stick a bit to the baking paper, it is absolutely normal : you will just need to pull them off delicately.
These cookies can perfectly be kept in an airtight container for a few days. Enjoy !

subset: commoncrawl


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These days it can be difficult to organise staff social events and gifts which are suitable for everyone and which really motivate staff and make them feel valued.
Have you considered cookery parties? These events offer something more than just a staff meal out and can also make things more comfortable for staff. It's a fact that staff events can sometimes be awkward and employees feel compelled to talk about work or feel awkward about socialising with people they only know through work.
Cookery parties provide staff with a common goal and, with a relaxing activity in place, people feel less awkward about making conversation and things flow more naturally.
Cooking events also take the focus off drinking, which can sometimes be a problem at staff dos. Some people may drink too much, whilst others who prefer not to drink may feel excluded. At a cooking party, there's the chance to sip wines and try prosecco for those that want to, but equally there is plenty to do for non-drinkers.
Staff still have the chance to enjoy a sit-down meal together at the end of the session, as they would at the usual Christmas party. However, many find it more fun to have cooked the food themselves and to have learned something new about food and cooking alongside colleagues.
Visit www.venturis-table.com for more information.


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The Red de Comunicadores Populares del Sur has denounced threatens to kill former guerrilla Ramiro Durán, spokesman for the political party People's Alternative Revolutionary Force in Huila.
The army was allegedly responsible for the threats made when Ramiro participated in a meeting with the community of Piedra Pintada, Vegalarga, rural area of eastern Neiva.
According to the first reports, men of the National Army broke into the meeting, attended by over 70 people from the region, many of them accompanied by their minor children.
According to the complaint of the ex-combatants, the meeting began around 9:00 a.m. today, on Monday, November 7, when the National Army troops entered pointing their guns at all the attendees.
"The order is to kill him [Ramiro] because he is a dissent" was the justification that a corporal of the National Army gave to the delegation of former FARC guerrillas.
The documents of Ramiro Durán and his entire security scheme were withheld. The community surrounded the former guerrilla and took him to a local house.
People were expecting the arrival of the institutions is expected that would guarantee the return of the former guerrilla to Neiva.
It is to underline, says former guerrillas that the delegation that carries out the tasks of pedagogy for peace had indeed informed the National Police about their displacement to the area and the development of the meeting.


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Good morning from Augusta. Last week, the Maine Legislature unveiled a list of 272 proposed bill titles that will be screened by a panel of legislative leaders that controls whether or not they will be considered during the 2018 legislative session.
Five constitutional amendments are being floated, including one targeting the governor's power. Sen. Justin Chenette, D-Saco, wants to strip the chief executive's constitutional power to oust elected sheriffs, which Gov. Paul LePage has recently threatened to exercise in a dispute on jails' interactions with immigration officials. Two Republican proposals look as if they would tighten Maine's referendum process, while Chenette wants to bar legislators from changing citizen-initiated laws for a year. Sen. Lisa Keim, R-Dixfield, wants to increase senators' terms from two years to four years.
What are some of the other interesting bills? House minority leader and gubernatorial candidate Ken Fredette, R-Newport, wants a bond issue aimed at student debt consolidation and repayment — something he broke with LePage to block earlier this year. Chenette wants a lifetime ban on legislators becoming lobbyists. Rep. Lydia Blume, D-York, wants all-electronic toll collection on the Maine Turnpike and Rep. Patrick Corey, R-Windham, wants a tax credit on gun safes to promote safety.
But many of these bills will be difficult to pass and most won't be considered. The session will be highly charged, with many top legislators running for higher office. The constitutional amendment will needs two-thirds support in both chambers and a statewide vote. And an equally divided Legislative Council — the panel of legislative leaders that is only supposed to allow "emergency" bills in even-year sessions — let through roughly a quarter of these bills in 2015. They're expected to meet later this month to discuss the bill requests.
Hayes brings on well-known independent operative
State Treasurer Terry Hayes of Buckfield, an independent gubernatorial candidate, has hired Kyle Bailey as her campaign manager. Bailey of Gorham ran the 2016 referendum campaign to implement ranked-choice voting in Maine and was finance director for the 2012 same-sex marriage campaign and independent gubernatorial candidate Eliot Cutler in 2014. He also runs Maine Independents, a new home base for unenrolled legislative hopefuls. Hayes of Buckfield is one of two independents running so far, alongside Alan Caron of Freeport.
Reading list
- One of the most horrible things about heroin addiction is the broken lives of the children it leaves behind. Hailey LeClair, 14, of Bradley, is one of those people. The young woman, who is a writer, and the BDN's Erin Rhoda teamed up for a touching story about her struggles.
- First lady Ann LePage is being courted from on high to launch a run for the U.S. Senate. Stephen Bannon, a former senior adviser for President Donald Trump, is trying to recruit candidates from across the country for the U.S. Senate to topple Senate Majority Leader Mitch McConnell. Ann LePage is among the candidates Bannon has his sights on but she has made no public comments about the possibility and it's unclear if she has political ambitions.
- A Maine man convicted of a 1989 murder is back in court today trying to clear himself months after the state's key witness recanted her testimony. The proceedings in the case of Anthony Sanborn are expected to last 10 days and could exonerate him, get him a new trial or send him back to prison. He was released on bail in April after the person who the state said was the only eyewitness to the killing of 16-year-old Jessica Briggs said she didn't witness the killing and was coerced into testifying by police.
- Environmental groups are rallying against Trump's climate plan. President Trump and his environment chief, Scott Pruitt, are moving toward issuing a new set or rules that would override the Obama-era Clean Power Plan. As the administration decides to do it with or without seeking congressional approval, left-leaning states and environmentalists are squawking. New York Attorney General Eric Schneiderman, for example, has vowed to sue.
Happy [belated] birthday, Mr. Governor
With the Daily Brief on holiday hiatus on Monday, we missed our opportunity to wish the state's top elected official a happy birthday. LePage turned 69 on Monday.
We don't want to show up on any of campaign finance reports so we offer the following gifts: A free subscription to the Daily Brief (it isn't technically a newspaper, a medium that we know he hates) and a soundtrack fit for a chief executive.
Today's Daily Brief was written by Christopher Cousins and Michael Shepherd and edited by Robert Long. If you're reading it on the BDN's website or were forwarded it, click here to get Maine's only newsletter on state politics via email on weekday mornings.


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An easy-to-carry hobo tote in durable waxed canvas will be your go-to bag all year long. This canvas has been hand waxed and sun-dried with pure filtered beeswax, it will soften and become more creased and distressed with use; giving it that sought after well-worn, lived-in look.Our bucket tote has an easy-carry-shape and includes 5 slip pockets (2 outside and 3 inside) as well as a large interior zippered pocket. A double-wide, natural leather strap and nickel plated hardware complete the look.FREE Shipping WorldwideDetails:-Approximately 14"H x 15.5"W x 3"D-Lining is natural twill.-Exterior is waxed cotton canvas -Hand-cut, 100% leather strap in Dark Brown, Medium Brown (pictured), Weathered Tan or Natural. Leather is finished on top and left natural on the underside.
**Extra Straps and Cross-Body Straps are available for purchase here:
http://www.independentreign.com/products/leather-crossbody-strap-leather-straps-Stress points are all reinforced-Estimated processing time prior to shipping is currently 2 to 7 days. Please feel free to contact me for "rush" orders, gift messages, specific delivery deadlines or review my Shop Policies for more info.
Quality:With an ever-growing marketplace, buyers now have an enormous selection of goods to choose from. My items are all handmade, by me, in my studio in New York. It is a labor of love and it insures that each item is crafted with the same exact care and quality every single time. Attention to detail, style, and durability are vastly important to create a finished product that not only looks good, but will last. Have a question? A suggestion or comment? Need more info? Please feel free to contact me. I'm likely sitting in front of my computer or at my sewing machine and will reply as soon as possible ;)Thanks for visiting my shop and have a great day!


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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
Be the first OregonLive.com user to add photos or videos of Popp Roger W Dc
Popp Roger W DC can be found at Young St 979. The following is offered: Doctors & Clinics. The entry is present with us since Sep 10, 2010 and was last updated on Nov 14, 2013. In Woodburn there are 22 other Doctors & Clinics. An overview can be found here.
Popp Roger W DC is located at 979 Young St Ste H, Woodburn, OR. This business specializes in Chiropractic Offices.
Mr Popp has not yet shared his bio with us.


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A truck hauling liquid asphalt caught fire along State Highway 195 near the intersection of Farm-to-Market 2670 Wednesday morning, Killeen police said.
No one was injured, and the fire was contained to the area of the truck, said Carroll Smith, Killeen Police Department spokeswoman. The fire was reported around 6:30 a.m.
"Nothing spilled on the road," she said; however, a hazardous materials crew was sent to the scene as a precautionary measure.
Smith said the driver is working as a contractor for the Texas Department of Transportation and noticed smoke coming from the truck when he pulled over.
She said quick actions by the driver kept the fire from spreading. Smith did not say what caused the fire.
Killeen firefighters extinguished the blaze. Traffic on State Highway 195 was hampered at the time of the fire, but was back to normal early Wednesday afternoon.


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- +6Photos
Tidak Apa'Pa Liveaboard
- Overview
- Schedule & Pricing
- Cabins
- Amenities
- Reviews
- NitroxNo
- Wi-FiFree
- Equipment RentalSurcharge
- Tech Diving FriendlyYes
- Fleet nameTidak Apa'Pa Liveaboard
- Year built2001
- Length20
- Width5.5
- Capacity8
- MaterialWood
Tidak Apa'Pa a traditional Indonesian wooden yacht. Comfortable and friendly, the Tidak Apa'Pa is a great choice for trips among friends or dive buddies. Small capacity guarantees you will get personal attention. The boat is rebreather friendly.
Boat interior and exterior
There are 4 cabins on board that can accommodate 8 divers. Cabins have shared a bathroom. Common areas are great to spend time after diving day and enjoy the gorgeous view. Passengers can rest at the open air salon or relax on soft bean bags.
Meals served on board are various and meets dietary restrictions.
Tidak Apa'Pa equipped with a compressor, she has tenders to get divers to the remote dive sites. There are special facilities also for tech divers. Divers can be divided into groups of four, so personal care is guaranteed. Safety and navigation
Safety is one of the most important things during the trip. Boat's crew pays special attention to that. Tidak Apa'Pa supplied with life rafts, jackets, firefighting equipment, emergency oxygen and first aid kit.
Photo Pro
The boat provides with photo facilities, also cruise director is an underwater photographer so divers can rely on professional advice.
Diving features
Itineraries of Tidak Apa'Pa include diving on Ambon dive sites, Raja Ampat, Banda Sea and Komodo. Amazing variety of corals and fish species, extraordinary underwater landscapes attractions divers from all over the world. There are several options of trip duration. You can choose either 7 nights trip or extended trip with 10 and 12 night duration to explore more dive sites. Komodo 7 nights trips include diving at Central and North dive sites like Crystal Rock and Castle Rock. North dive sites will include diving with sharks on Golden Passage and night dive at Shot Gun. 10 nights trip will be extended with more dives sites of the North and South of Komodo. Also, don't miss a chance to explore volcanic landscaped of Sangean. Another destination represented by volcanic formation is the Banda Sea. Here you can dive at Manuk and Banda Neira. This trip also gives a chance to meet hammerheads.
When to go
Although most of the destinations Tidak Apa'Pa offer can be considered as a year round diving destination, the best time to visit Komodo is between April and August and for Raja Ampat its months from October until April.
Tidak Apa'Pa is the best choice for those who want to spend time in the friendly cozy atmosphere, for those who are looking for something new and exciting.
Schedule & Pricing
Booking process
Select the cabin of your choice and enter your personal information. Submit the booking request. Once the booking is processed, you will receive an email notification with all booking details conveniently laid out in PDF.
Our team will contact the liveaboard on your behalf to confirm availability and other booking details. We put your place(s) on hold and get back to you within 24 hours. If there are any issues, we come back to you with alternative options.
Once your booking is confirmed, you will receive an email notification with payment instructions. You then have up to 72 hours to take care of other travel arrangements and process the transaction to secure your reservation.
Payment Terms & Cancellation Policy
Book with Divebooker and get extra benefits
- No Booking Fees
- Loyalty Club Benefits
- FREE DAN Dive Insurance
- Book Now Pay Later
- Best Price Guarantee
- Special Offers & Deals
- 24/7 Expert Support
Double Cabin
- Deck
- Lower Deck
- Bedding
- Double Bed
- Max occupancy
- 2 Guests
- Number of Cabins
- 2 Cabins
Twin Cabin
- Deck
- Lower Deck
- Bedding
- Twin Beds (Bunk-Style)
- Max occupancy
- 2 Guests
- Number of Cabins
- 2 Cabins
Reviews about Tidak Apa'Pa
- Cabins & Facilities
- Food & drinks
- Guiding & Safety
- Service & Infrastructure
Share your experience with othersWrite a review
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We are glad to share that Retreat venue have been finalized – Fujairah Rotana Resort & Spa, Al Aqha Beach, Fujairah. Participants can check in from 2:00 pm onwards on 27th Feb 2017 and Check out is on 2nd March 2017 morning as per plan.
For those who have missed to register for the retreat, now they can. As we have received some cancellations, we can accommodate new participant again.
Please send your registration to email@example.com with Full name, age, gender, contact number, email address and city you will be coming from.
If you have already registered and due to change in plan, wish to cancel, please advise us urgently. We commit booking to Resort, based on registered numbers and liable to pay.
Retreat payment can be handed over to me at Manav Bhawan during Friday Satsang. For alternate arrangements, you can contact me (phone & whatsapp) on my mobile number 055 44 53 455. Last date for payment is 5th February 2017.
We are making special arrangements to allow family participation, with children. If you are looking for such option, please contact me. Ensuring that main event does not get disturbed, hopefully we can find a way.
Since the retreat is during working days and you may arrive on 27th February 17 by evening and can leave before the retreat ends (on 1st March 17 night). In this manner, 2 days leave from work (28th & 1st).
For those planning to fly into UAE for Retreat and need Visit Visa Assistance or Local Hotel Booking before and after retreat, or for tourism purpose, can contact M/s Monsoon Whispers Travels & Tours LLC on Office +91 4 255 2931 Mobile +971 56 5266189 or email@example.com with subject mentioning "Yoga and Holistic Living Retreat".
If you will need assistance for Transport, we will coordinate this aspect closer to the date for retreat.
Sri Gurubhyo Namah:


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According to a recent study conducted by Pew Research center, 59% percent of US adults between the ages of 18-29 are on Instagram. In addition to this, Facebook also recently announced that the platform currently boasts over 700 million monthly users, far exceeding those on Twitter. These results are staggering and reveal to publishers and private users alike that Instagram is an essential platform to promote content.
In this blog post, we'll reveal some tips on how to promote your digital publications on Instagram and how to potentially grow a strong following of subscribers!
Create an official Business account
In 2016, Instagram revealed new business tools enabling companies of any size to have access to additional features such as a contact button, account insights and adverts to attract potential customers. Creating a business account is a simple way to gain credibility on the platform and has huge brand building value for magazine publishers!
Vary your content
Instagram is all about variety, so make sure that your Instagram profile is full of different kinds of content to keep your audience engaged! Such content can include behind the scenes footage and photos, teaser clips about upcoming issues and inspirational quotes! In addition, Instagram's new photo album feature allows you to share multiple images within a single post to tell a coherent visual story.
Check out an example of this album format below from our Instagram page:
Be Consistent
Producing regular and coherent content on your Instagram page is extremely important as it demonstrates professionalism and reliability to your audience. Many Instagram experts suggest posting between 1 to 2 times a day for optimal results.
You can also schedule your digital publications to be published on Calaméo in just a few clicks with a Premium or Platinum plan. To further enhance your professionalism, we suggest synchronizing the release dates of your Calaméo digital publications and your Instagram posts to create unified image online.
Include a link to boost your traffic
On Instagram you only have one opportunity to share a clickable link, so use it wisely! We suggest optimizing your bio (your page description) with a link to a targeted landing page. For example, a link to page of your digital publication displayed in your post so that your audience can click to browse through the magazine and potentially the rest of your website.
Here's a great example of traffic generation to a specific page by Calaméo user Basic Magazine:
Avoid overly promotional content
Keep in mind that Instagram is about the soft-sell approach, even if your digital publications are catered more to a B2B market. If you post the right images, customers retain your marketing message without the need for a hard sell. Sales pitches can be left to other platforms such as Facebook and Twitter.
Use hashtags!
Instagram organizes popular photos by hashtag, so be sure to include as many pertinent hashtags as possible in your captions to make your content discoverable and to build your audience.
Check out Kobo Books excellent use of pertinent hashtags below:
Interact with your Community
Share your online audience's content (with permission) on your page to diversify your Instagram feed. The implementation of user generated content will help you immensely on Instagram as it is one of the most effective ways to build an engaged community. Indeed, it is far more captivating for your audience to see "real" content posted by real people as opposed to brand curated content.
💡Don't forget that if you'd like to be featured on our Instagram page, tag your favorite Calaméo publications with #CalameoCreatives!
Instagram is a fantastic tool for marketing your digital publications and your business if used effectively. Equipped with our top tips, you'll be well on your way to achieving Instagram success in no time at all.
As fellow Instagrammers, we just love interacting with our Calaméo community on this special platform. Come and follow us @Calameo_publishing!


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(Not) All About Dmitry
Dmitry Medevdev will be sworn in as Russian president today. Instead of trying to psychoanalyze the man at the helm, Washington should judge Moscow by its actions over the coming years.
As Dmitry Medvedev is sworn in as Russia's third post-independence president today many will doubtless succumb to the undying attraction of the Kremlin tea leaves and ask, "Who is Medvedev?" It won't be time well spent.
The interest in Medvedev is understandable; he will be assuming what is at least nominally a very powerful position in an increasingly assertive country that remains mysterious to many Americans almost seventeen years after the collapse of the Soviet Union. And Medvedev's predecessor and soon-to-be prime minister, Vladimir Putin, has unquestionably become a major figure on the international stage.
But if anything should make clear the fruitlessness of the "who is [insert Kremlin leader here]" game, it should be Putin's own presidency. Eight years ago, many American backers of Boris Yeltsin who also supported heavy-handed involvement in Russia's economic policies welcomed Putin. They saw him as a reliable successor to Yeltsin who would be better able to use the powers of the presidency to force further policy changes. When Putin made it his first priority to undermine and effectively exile the so-called "oligarchs" who had supported other candidates for the presidency, the then- president's KGB background became a focus of excited but often oversimplified attention (Putin was a foreign-intelligence officer, after all, not a domestic security informer or enforcer).
Since then, "who is Putin?" has gone on for eight years and has yet to produce much that is useful-mainly because the people who ask the question normally want simple answers and Putin, like most others on this earth, is not a simple person. The same seems likely to be true of Medvedev as well.
Much more informative is the slower, more complex, and considerably-more-nuanced business of actually watching what Putin, Medvedev and other leaders do over time. And there are certain things to watch out for. In Mr. Medvedev's case, we will soon see who is appointed to senior positions and who is not. Understanding the appointments will not be easy-Medvedev is unlikely to have a completely free hand and Mr. Putin will clearly have significant influence; though probably more in shaping the government that he will head than in selecting the Kremlin staff that will work for his protégé and successor. Key roles for an already-strong economic team, including long-serving Finance Minister Alexei Kudrin, could be an important plus. And an extended fight over personnel issues could augur poorly for historically unprecedented power-sharing between Medvedev and Putin, possibly leading to paralysis or worse.
In addition, will Medvedev try seriously to take on corruption after saying that Russia suffers from "legal nihilism"? Mr. Putin said that he wanted to establish a "dictatorship of law" and that the government should be "equidistant" from the oligarchs. Neither happened in practice because Russia's powerful business leaders remain a key political constituency supporting the current political system (and Putin took on only those who did not). Medvedev is unlikely to confront oligarchs who support the regime, but might be able to impose new rules on their behavior-something that would be a major accomplishment.
Medvedev's approach to the media will also be revealing. Under Putin, the Russian media had noticeably more leeway in criticizing the prime minister and the government than in assailing the president and the Kremlin. If the same arrangement continues, unwelcome attention could well be focused on Prime Minister Putin, especially if Putin's government is not fully successful in implementing its ambitious national projects in education, health, housing and agriculture. Yet tighter control over media discussion of the government (as distinct from the Kremlin) could eliminate one of the few remaining "safety valves" that prevents dissent from becoming something more-and provoke Western criticism, though this will probably be secondary in Medvedev's calculations.
In foreign policy, the U.S.-Russian agenda is over-full and underachieving: differences on missile defense, NATO enlargement, Kosovo and a host of other issues have undermined progress in areas of shared interest like combating terrorism and nuclear proliferation. Mr. Medvedev is not likely to spend much effort cultivating an outgoing American president-especially when consolidating his own position may take some time-but a different approach to the new American president in January could help to turn the relationship around. (Some degree of reciprocity would obviously be required for it to do more.) Medvedev's interaction with key U.S. partners in Europe, especially the United Kingdom, which has even-rockier ties with Moscow at present, could be an early indicator of what is to come.
Understanding Russia is difficult at best and predicting Russian behavior doubly so, especially in times of transition. But we could do ourselves a big favor by focusing less on unanswerable questions instead looking at actions over the coming year. It's the only way to develop sound and effective policies for the United States in dealing with another major power.
Paul J. Saunders is the executive director of The Nixon Center and the associate publisher of The National Interest.


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2020 St John's New Year's Day Fell Run
Photographs are now online from the 2020 St Johns Fell Run held on New Year's Day..
Since 2006, my passion for both photography and sport has enabled me to cover many events, with official media accreditation, including the Isle of Man TT, Classic TT, Manx Grand Prix, Southern 100, Pre/Post TT meetings at Billown, also the North West 200 Road Races, British Superbike Championship, FIM World Indoor Enduro Championship and British Sprint Enduro Championship.
I have worked with, and have been commissioned by, various worldwide motorsport organisations/media outlets including:
Isle of Man Government – Pacemaker Press International – The Official Isle of Man TT Programme – Manx Motor Cycle Club – Southern 100 Club – The Official Manx Grand Prix/Classic TT Programmes – Classic Racer – Duke Video – Motor Cycle News – Bike Sport News – Fast Bikes Magazine – Super Bike Magazine – Isle of Man Newspapers
I regularly cover various events here on the Isle of Man and all the images on the website are available to purchase by the general public (for personal use) in addition to the media. (Please get in touch regarding commercial usage of any images or commissioned work).
Photographs are now online from the 2020 St Johns Fell Run held on New Year's Day..
Photographs are now online from the 3rd round of the Isle of Man Athletics Cross Country League held at the Nunnery on 1st December 2019
Photographs are now online from the 2019 Manx Grand Prix and Classic TT…
Whilst I have covered many events on the Isle of Man, the website underwent a major revamp in January 2020 and many galleries were removed to allow space for more.
If you are looking for a particular image from any of the events that are listed below, please contact me here giving as much detail as possible.
Isle of Man TT (2007 – 2019)
MGP & Classic TT (2008 – 2019)
North West 200 (2012 – 2019)
Southern 100 (2008 – 2019)
To get in touch, please fill in the form below, If you are searching for a particular image, please give as many details as possible, (Date, Event, Name, Number, etc)
Necessary cookies are absolutely essential for the website to function properly. This category only includes cookies that ensures basic functionalities and security features of the website. These cookies do not store any personal information.
Any cookies that may not be particularly necessary for the website to function and is used specifically to collect user personal data via analytics, ads, other embedded contents are termed as non-necessary cookies. It is mandatory to procure user consent prior to running these cookies on your website.
Click here to view current promotions/offers. Dismiss


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As you know, we recently moved and the new place has a club which conducts story time every Wednesday. I took my kid there today and the reason for writing this post came to me. The session started with the bubbles session. The organizer blew bubbles and all the babies who were scattered in the room gathered around her. The rest was super fun for the kids but something struck me.
Are we not like kids running after material bubbles? Oooooooh such a deep thought! No? Think about it!
We (me included) are so lost and busy after our man-made set goals that we probably are unaware of others around us. We sometimes bump into others, sometimes we fall but are only busy with the bubbles. I don't have much to say about the topic and my baby is playing and might call me anytime but …. are you after some bubble? The reason why I am calling our material goals a bubble is because once we get it, the happiness quickly bursts and we completely forget it and run after another.
Let me know your views.


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"Dumb Charades?", we were dumbfounded! That's exactly what our colleague told us when he had gone outing with his team to Ooty some years ago. About 30 of them had booked the vacation almost three months prior to taking the trip. After reaching Mettupalayam the team got into a minivan to take them to the resort. A few minutes into the ride and someone decided to play "Dumb Charades". The high energy in the group only elicited good participation. Soon two teams were formed and they started competing. This went on till the van reached the resort. After a quick refresher and breakfast the team is out again on the road visiting places and guess what's going on inside? You guessed right, "Dumb Charades" again. Like a bad habit this continued everytime the team got into the van. Best part, our colleague confesses, there was only one topic, "Tamil Movie Names" and the teams couldn't exhaust the list of names for the entire journey, i.e till they reached Coimbatore to board the train back to Chennai.
Certainly, There are better ways to enjoy boating
Another colleague who we thought was in a slightly better vacation was lucky to get on to a boat to visit an island off the coast of Singapore. For hours the team sat on the open boat in the blistering sun, looking at the waves and the coastline. An MC was having a trying time as he couldn't engage the people on the boat to a game of "Bingo", we repeat "Bingo" and that too with adults. On reaching the island just one couple got into their swimwear to go for a swim. Others just languished on the land sitting under trees and moving about. They had hardly an hour to spend on the island when they were back on that boring two hour boat ride back again for the return trip.
Vacations on the sea will never be the same again!
Why play silly games or spend hours travelling and tiring yourself when you can have the "Team Outing" of your life this Summer at Bay of Life's Summer Surf Camp where you'll be indulging in activities you only watched on National Geographic. For Nature and Wildlife lovers we have Tours, Treks and Camps, for adventurers, Snorkeling, Kayaking, Surfing, Paddleboarding and Swimming in the Sea. Choose from a wide range of accommodation facilities that includes rejuvenating spas, yoga retreats, great food and Internationally certified coaches and instructors who'll ensure your complete safety in or outside the water. Believe Us! You are going to back again next Summer or even earlier, for more fun! We have proof!
For the best team outing, you have had in a very long time, contact Bay of Life Click here


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On our second day in Montana, Alex and I decided to tackle a slightly more ambitious climb. When conference ended around noon, we drove a short distance from Big Sky to the Dudley Creek trailhead and spent most of the rest of the day climbing Wilson Peak (10,705′).
The peak itself is one of the more prominent high points on the skyline, seen from Big Sky resort:
The first 5 miles is a gradual uphill climb – on a well maintained trail – to Dudley Lake.
This 5 mile stretch gains ~2800 vertical feet, leaving another 1.25 miles and ~1500 vertical feet of off-trail climbing. The climb itself is mostly easy class 2, with the exception of a 50-60 foot summit block requiring a few class 3 moves.
We reached the summit in a little under 4 hours. We spent 20 minutes or so taking pictures and eating some snacks, before we decided to start heading for the car.
There were several ominous storms on the horizon, along with the occasional rumble of distant thunder, but fortunately we never had to deal with more than a few light sprinkles or rain on the return trip.
The entire round trip took 6:41, including rest stops. Total vertical gain was 4578′. Total distance was 12.5 miles.


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That's how young people have described their involvement in previous World Youth Days (WYD).
These events - held about every three years - are opportunities for young Catholics to gather with the Holy Father to pray, worship, learn, serve, and mingle.
Hundreds of thousands of young Catholics from around the world will be traveling "down under" for the celebration of World Youth Day in Australia in the summer of 2008.
Diocesan pilgrimage
Joining them will be a delegation from the Diocese of Madison. "We're all excited. The diocesan plan is going well," said Fr. Paul Arinze, pastor of St. Joseph Parish in Dodgeville and chairman of the diocesan pilgrimage planning committee.
He explained that 63 people are registered for the diocesan group. These include almost all of the diocesan seminarians as well as some high school and college-age students and young adults.
Father Arinze said there is still an opportunity for young people from the Diocese of Madison to attend World Youth Day. "We have about seven openings for young people aged 16 to 35," he said. Full story ...
Celebrating feast
By Susanna Herro
|Most recent issue Past issues


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Touch-n-Go Thermostat by Carrier
- Details
- Category: Thermostats
COVID-19 Info: We're here to help. We're listed by NYS as essential, so we can serve you promptly and safely - see our FAQs.
Click here for Air Purification and Virus Protection options.
There are a variety of units available to accompany your heating and cooling system. Superior control over the heat, coolness and moisture are signature features of thermostats offered by Hawn.
Comfort. Savings. Your life – in tune.
Lyric was designed for crazy schedules and spur-of-the-moment activities. You can control Lyric from anywhere on your smartphone, or let it adapt to your life as you go.
Lyric can save energy when you're away, and get the house warm when you're heading home. No more wasted energy when you're out for the night, and no more getting in from work to a cold house. It'll even let you know when the filter needs changing.
Voice Activated Recognition and Control (on-device only) allows the temperature setting to be adjusted in the home by talking to the thermostat
Total Connect Comfort App - Allows user to remotely monitor and manage home's comfort and energy usage anytime, anywhere
On-screen Comfort Information - See indoor and outdoor temperature and humidity information right on the home screen
Customizable Color Touchscreen - Easily change the high-definition display to any color you choose
Auto Alerts - Extreme temperature alerts and filter change reminders are sent to connected network devices and displayed on thermostat
Advance Fan/Control Comfort - Improves air circulation and helps regulate the temperature throughout your home
Energy Savings - Choose programmable mode to save on annual heating and cooling costs
Internet Time Sync - Keeps time during power failure and automatically updates for daylight savings
Simple Programming - Tool-free wiring, manual-free install and setup
5 Year Limited Warranty
View your system performance at the touch of a button, 24 hours a day, seven days a week with the industry's largest, easy-to-read LCD screen.
TrueSense dirty filter detection reminds you when it's time to change your media filter. And when you add a humidifier, germicidal UV lamps or ventilators, the Infinity Control also reminds you for their routine maintenance.
One control adjusts a furnace or fan coil, air conditioner or heat pump as well as controls optional accessories such as zoning, humidifiers, ultraviolet lights and filtration products.Up to 8 zones are supported with the Infinity Zone Control model.
Patented independent fan control by zone for ultimate air flow control.
Five-year limited parts warranty
The revolutionary Infinity® System is the only system in the industry that allows you to control temperature, humidity, air quality, fan speed, ventilation; and even detects when your air filter needs replacing. It's so advanced it automatically adjusts to your system's needs for peak-efficiency and performance.
Carrier's revolutionary Infinity Control is the smart control of the future. Intuitive prompts walk you through every program from humidity levels to fan speeds, giving you the ultimate control over your home comfort.
Automatic humidifier control.
Fan speed control for dehumidifying.
A "cool to dehumidify" mode.
One-button vacation mode.
Entire system control from one location.
Unique field configurable to either a non-programmable or programmable model, depending upon your need.
One control for indoor temperature and humidity.
Smart recovery.
Large push buttons.
Five-year limited warranty
IdealHumidity, controls your homes humidity level even when the system isn't actively heating or cooling. By keeping the proper balance of moisture in your home, you can remain more comfortable at higher temperatures in the winter . The unit offers better efficiency which means lower utility costs.
When setting up your Thermidistat Control, you'll appreciate the time-saving "copy previous day" function. Once you have set your comfort schedule, Smart Recovery makes sure your system delivers the temperature you want at the time you want it.


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Veterinary hospital in Genesee County is seeking a ft/pt licensed veterinarian for a fast-paced, multi-doctor, small animal practice. We are looking for an energetic doctor who enjoys building long-term relationships with their clients. This is a modern, independently owned, walk-in facility that has a busy surgery (by appointment) schedule. We offer Idexx in-house laboratory, Antech reference lab, digital radiograph, K-Laser, and surgical laser. New graduates welcome, we love to mentor! We also can be flexible with our schedules. No emergencies or Sundays, half day on Saturdays. A competitive wage and benefit package is offered with buy-in potential. If you have the drive to serve our great community, please forward cover letter and resume to email@example.com
Attn: Dr. Theresa Driscoll or Kristy
Michigan licensed veterinarian or new graduate.


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Value Proposition Design
15 October 2018
Ashford, Co Wicklow
To run any successful business you need a clearly defined VALUE PROPOSITION that explains how your product/service solves your customers' problem, need or improves their situation. A Value Proposition allows you to define who your customers are and to offer them what they really want. It also helps to secure grant aid or private investment.
In this 1-day workshop we will scope out your Value Proposition to help you to run an efficient and profitable, creative business.
Following a creative profession means spending a lot of time thinking from the perspective of "I" – what is my topic or my specific style, what do I want to stand for etc. However, without an engaged audience and buying customers the necessary revenue cannot be generated.
This is the moment when "they" perspective needs to be included into our thinking and planning. It does not mean changing the way we work, it means being able to clearly articulate why our work meets our customers' needs.
We help you to work out a clear Value Proposition, to understand "they" perspective. We will lead the whole process and provide you with the tools and feedback you need. You will get a chance to apply the newest entrepreneurial techniques (i.e. Creative Business Model Canvas), and to discuss your ideas and challenges with us and a small group of other photographers developing their own endeavours and who are always happy to share their experiences.
Where appropriate we will look at your Value Propositions not only from a business but also from the social and cultural perspective.
General Information
- Duration: 1 Day
- Price: Full – €125
- Early Bird – €85
The programme is 100% attendance, the number of participants is limited to 15
What Will You Learn:
- What is Value Proposition and how do you test and refine your specific Value Proposition?
- Who are the right customers you need to reach and engage to grow your business?
- What are the needs and problems your customers need solving the most?
- How do you effectively communicate your Value Proposition to your customers?
- Is your business scalable?
What You Will Need
- Materials will be provided.
Programme & Timetable
|9.00 – 9.30am
|Arrival & Registration
|9.30 – 11.00am
|Value Proposition Canvas
|Visualise, design & test
|11.15am – 12.45pm
|Your customers profile
|Their needs/wishes/problem you will be solving/meeting
|12.45 – 1.45pm
|Lunch Break
|1.45 – 3.15pm
|Value map of your products/services
|3.30 – 5.00pm
|Price points, go to Market Strategy
|5.00 – 6.00pm
|Presentations , Feedback, Remaining Questions
The Tutors & Speakers
The Tutors & Speakers
Monika Sapielak
Co-Founder of the Creative and Artistic Entrepreneurs Academy
Entrepreneur and Innovator with 25 years of work experience as CEO, Manager, Cultural Leader, Mentor & Educator.
Regular guest speaker and lecturer at conferences, in higher education institutions and for the media in Europe and in Ireland. Monika's recent talks include University Business Forum, in Oct'16 in San Sebastian; Creative Business Cup in Copenhagen, University of Pecs, Hungary, Trinity College Dublin, University College Dublin and Dublin Institute of Technology.
Monika's mentoring and coaching expertise includes Start-up support, Business Strategy, Value Proposition and USP, Stakeholders Relationships, Fundraising, EU Funding – grant writing and grant applications' review, Audience & Customers' base development, Market research.
Recipient of the Merit to Polish Culture awarded by the Ministry of Culture and National Heritage of the Republic of Poland as well as Main Awardee of the Arthur Guinness Fund.
Ian Oliver
Head of Creative Entrepreneurship, Creative and Artistic Entrepreneurs Academy
Ian has over 15 years' senior management and entrepreneurial experience in the Arts, creative and technology Sectors and 4 years' experience working with Local Government, Local Enterprise Offices, Local Arts Offices, Arts Organisations and Venues, Universities, Businesses and Community Groups throughout Ireland, auditing and engaging the artistic and creative community within specific geographic regions.
Currently Ian is working on advising and designing sustainable projects that utilise art and creativity as the main driver to effect sustainable economic and social change. He also teaches and mentors artistic and creative entrepreneurship training to adult learners and undergraduate students.
Your Questions & Feedback form
We would love to hear from you so please feel free to send us your questions or your feedback!
Find Us
The Old Milking Parlour
Ballymurrin Lower, Kilbride
Wicklow, Ireland, A67TK64
Monday–Friday: 10:00AM–6:00PM
tel: +353(0)866084020
email: firstname.lastname@example.com
Your Questions and Feedback Form
We'd love to hear from you so please send us your feedback!
Copyright © 2017 Creative Entrepreneurs Academy


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Monday, July 29, 2013
La Cappella di Sol LeWitt e David Tremlett
The Sol LeWitt and David Tremlett Chapel, Brunate, La Morra, 1999 here
Posted by ahoi! at Monday, July 29, 2013
Labels: architecture, colour, david tremlett, google image search, sol lewitt
1 comment:
- versus said...
Exact points de couleur.
- July 31, 2013 at 2:25 AM
Subscribe to: Post Comments (Atom)


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Our sister publication, BOARDS. has just announced that the latest edition of their printed magazine will be hitting shelves very soon, on the 15th of April to be exact.
This issue is all about ruling on the flat and includes contributions from the likes of Antoine Albeau, Robby Swift, Josh Angulo, Andy Chambers, Max Rowe, Kevin Pritchard, Karin Jaggi, Adam Sims, Steven van Broeckhoven and many more.
Check out all the info about the upcoming issue and how to subscribe right here.


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Meghan has been a successful massage therapist since 2007 starting in Florida at a health clinic and eventually moving to Virginia to help in several fitness & wellness centers. Working with chronic issues such as sports injuries and soft tissue strains, she has learned a diverse array of treatment modalities proven effective in helping clients regain increased mobility, reduce pain and achieve balance in their body structures.
In addition to therapeutic massage, she offers deep tissue, pre-natal, and sports massage. Meghan is especially fond of including an active – passive stretching technique (PNF) into her routines when it's appropriate. Another not-so-common modality, Lymphatic Drainage (MLD), can be amazingly helpful when inflammation is an issue or joint pain is present. Currently, Meghan is one of the few therapists in the area to have been also trained in Thai Yoga Massage – an incredible experience when you need someone to help stretch your muscles and regain your range of motion. Massage and Bodywork has been very rewarding for her as she is able to aid in her clients' fitness and health goals, while making personal connections with those passionate about the healing arts.
Services Offered
Services Offered:
Deep Tissue Massage
Quite different from "a Therapeutic Massage with Extra Pressure," our Deep Tissue Massage focuses on releasing muscle restrictions (adhesions) and re-balancing structural tension (holding patterns) running through the several layers of soft tissue (muscle). Deep Tissue Massage requires a much slower pace than our Therapeutic Massage and more than one session is usually necessary for better results and/or resolution. Very effective if you have strained a muscle resulting in limited (and possibly painful) movement.
IMPORTANT: While it is not unusual for some people to experience bruising and pain for several days following treatment it does not happen to everyone. Soon the pain will decrease and you will begin to experience a range of movement you haven't had for a very long while. Eventually, that every-day pain thing will be just a distant memory.
PLEASE NOTE: 30-MINUTES WILL ONLY ALLOW ENOUGH TIME TO WORK ON ONE REGION OF THE BODY (muscles of your thigh, calf, forearms or hands, etc.). One hour is highly recommended for your first session.
- 30-minutes: $80
- 45-minutes: $95
- 1-hour: $110
- 90-minutes: $150
- 2-hours: $195
Value Packs
- Value Package of 3 – 45-minute Sessions: $270
- Value Package of 3 – 1-hour Sessions: $315
- Value Package of 3 – 90-minute Sessions: $429
Integrative Bodywork
Integrative Bodywork combines several healing bodywork modalities with therapeutic massage. Some of those modalities might include: CranioSacral therapy, Trigger Point therapy, Myofascial Release, Deep Tissue, Positional Release, Neuromuscular Facilitation, Reiki, and acupressure. Very effective in treating some chronic pain issues such as sciatica, acute injuries such as whiplash, sports-related injuries, carpal tunnel and other repetitive stress injuries (RSIs), plus physical traumas such as accidents involving falling-down, bicycling, and automobiles.
1-hour usually gives the therapist enough time to focus on those areas of the body associated with the pain or injury. Several treatments are often necessary to understand and resolve the issue(s).
- 30-minutes: $75
- 45-minutes: $90
- 1-hour: $105
- 90-minutes: $145
- 2-hours: $185
Value Packs
- Value Package of 3 – 45-minute Sessions: $255
- Value Package of 3 – 1-hour Sessions: $300
- Value Package of 3 – 90-minute Sessions: $414
Manual Lymph Drainage
Therapist uses very gentle pressure. Each session lasts from 75 to 90 minutes, and therapy usually is performed once a day several times a week for two to four weeks. The lymphatic system is a vital bodily defense against infection & disease, and lymphatic massage is one of the best things you can do to help your body help itself. Our Manual Lymph Drainage specialists have the experience & skill to help reduce the debilitating effects of stagnant or excessive lymph fluid in the body. Studies have found lymphatic massage to be a medically beneficial form of physical therapy for lymphedema-related issues. Lipedema, an often misdiagnosed and misunderstood disease, currently affects millions of women who don't even realize it.
Please Note: If you have been diagnosed with lymphedema, please make sure to receive clearance from your doctor (or lymphedema specialist) to receive our manual lymph drainage treatment. Please bring a letter from your provider indicating that you are able to receive this service. Thank you.
For additional information visit: http://diseasetheycallfat.tv
- 1-hour: $110
- 90 minutes: $150
Value Packs
- Value Package of 3 – 1-hour Sessions: $315
- Value Package of 3 – 90-minute Sessions: $429
Sports Massage
If you're a runner, we are going to concentrate on your legs; if you're a swimmer, we will spend more time on your upper body and hips; a cyclist, your hip flexors, quads, ITB, calves and upper trapezius will likely be targeted.
A sports massage therapist may use their forearm or the palm of their hand to bear down on the muscles in your legs and back to separate and relax them and their fingers to put direct pressure on a muscle in spasm, which can be painful but will reduce your pain later. Along with massage we'll combine deep tissue massage and stretching when necessary to help maximize your muscles' potential.
PLEASE NOTE: A 1-hour session is recommended for your initial session/evaluation.
- 30-minutes: $75
- 45-minutes: $90
- 1-hour: $105
- 90-minutes: $145
- 2-hours: $185
Value Packs
- Value Package of 3 – 1-hour Sessions: $300
- Value Package of 3 – 90-minute Sessions: $414
Stress-reduction Massage
This is the kind you're most likely to encounter at a spa. Stress-reduction massage can also induce a state of mind called the Relaxation Response, which has been shown to minimize the effects of stress and boost your immune system. Everyone knows that a stressful day will cause the muscles in your shoulders and neck to tighten. For some, symptoms can even include a tense jaw, back ache, fatigue and headaches. When you're feeling overwhelmed and even simple movement is stiff, our Stress-reduction Massage is likely to bring you immediate relief.
For more severe or chronic pain you should consider our Therapeutic Massage.
- 1-hour: $79
- 90-minutes: $120
Value Packs
- Value Package of 3 – 90-minute Sessions: $327
Thai Yoga Massage (Traditional Thai)
Thai Yoga, also known as Traditional Thai, is done on a mat on the floor with the client wearing loose, comfortable clothing. This method uses the whole body to treat the client, blending gentle rocking, acupressure, rhythmic compression, and assisted stretches to relax and revitalized body and mind. The therapist will use their hands, knees, legs and feet to move the client into a series of yoga-like stretches.
PLEASE NOTE: Please wear loose, comfortable clothing to these sessions.
- 90-minutes: $150
Therapeutic Massage
OUR SIGNATURE MASSAGE. Blends a general Swedish-based massage with muscular therapy techniques to reduce significant tension due to either muscle strain or excessive stress. Some discomfort may result during treatment but a renewed sense of ease and feeling relaxed should last for several days. All sessions are customized for individual needs.
With that in mind, we ask that you please communicate with your therapist about the technique and pressure to be used. You may need to upgrade to one of our Advanced Bodywork modalities (see Integrative Bodywork or Deep Tissue Massage) if your pain involves several muscles or fascial lines. But if your wish is to feel relaxed and still have some of your pain & tension reduced our Therapeutic Massage is probably right for you.
PLEASE NOTE: 1-hour is recommended when it's your initial session.
At MTW we strongly feel that because you have a tension pattern that is unique to you, your session should be customized to meet your individual needs/issues. Our therapists are skilled in many healing modalities which they combine to provide you with a truly therapeutic experience for your mind, body and spirit. Our hope is to educate and encourage you to be an active participant in your own health and wellness.
With that in mind, we ask that you please communicate with your therapist about the technique and pressure they use. You may need to upgrade to one of our Advanced Bodywork modalities (see Integrative Bodywork or Deep Tissue Massage) if your pain involves several muscles or fascial lines. But if your wish is to feel relaxed and still have some of your pain & tension reduced our therapeutic massage is probably right for you.
- 30-minutes: $60
- 45-minutes: $75
- 1-hour: $90
- 75-minutes: $110
- 90-minutes: $130
- 2-hours: $175
Value Packs
- Value Package of 3 – 45-minute Sessions: $213
- Value Package of 3 – 1-hour Sessions: $255
- Value Package of 3 – 90-minute Sessions: $372


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"Only let us live up to what we have already attained."
So, I plan on talking more about my faith (though I will not ignore the posting on homeschooling, organizing, and homemaking!) I will try to post with a minimum of apologies for my possible ignorance on a topic, and use as much scripture as I can. This way, where I am wrong you will have the benefit of God's infallible Word.
Thank you for walking this new path with me.
I am happy to share this at WFMW.


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Besides not wasting all of it at the bar, there's a lot booze can teach us about money.
Drinking isn't cheap, especially if you aren't aware of these nine hidden costs.
A story in the New York Times says that exercise helps human brains build more neurons (a process called neurogenesis). It also reports on some mouse studies that show that exe
After quitting smoking, having one or two drinks a day is the best single thing you can do for your cardiovascular health. [more]
The struggle of the homemade gift — what's sure to please, and what's bound for the bin? We've got something that's sure to please; DIY infused vodka!
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Burkett Needle, Smash and Grab, Pipsqueak Peak, East Ridge
Alaska, Coast Mountains
Three years ago Fred Beckey pulled out his little black book (actually a tattered FedEx envelope) and showed me a blown-out photocopy of the unclimbed west side of Burkett Needle. Of course, no tactical information could be gained from scrutinizing the fuzzy blobs and lines on the page, but it showed that half of an amazing peak on the Stikine Ice Cap remained overlooked. That summer Fred, Micah Lambeth, and I flew to Petersburg and then helicoptered in to the Burkett Boulder. During that trip Micah and I attempted the west ridge of the Needle and, on July 4, 2006, climbed the East Ridge (5.7) of a 6,700' pyramidal-shaped rock peak. This peak, which we dubbed Pipsqueak Peak, is situated on the left (west) side of the major col west of the Needle.
It took several summers for everything to align, but on July 3 John Frieh and I caught a helicopter flight with Wally from Temsco Air in to the Burkett Boulder. The weather was set to be absolutely gorgeous for at least the next two days. We quickly packed in the afternoon sun and hiked east a quarter-mile to the base of a ridge that allows access to the upper glacier under the Needle. We followed the scrambly ridge (3rd and 4th class) for 1,800' to a small, flat campsite on a knife-edge ridge. On the morning of the 4th we left camp and climbed the glacier to the base of the South Pillar (Cauthorn-Collum-Foweraker, 1995). From here we traversed on steep snow into a couloir that led to the col at the base of the unclimbed west ridge. One long mixed pitch (M4) got us onto the rock. From here we simul-climbed perfect granite along the crest for many enjoyable pitches (up to 5.8). Higher, we climbed two tricky mixed pitches (M4) to the top of the false summit, then made a short rappel to a col below the imposing summit pinnacle. A short knife-edge traverse with great exposure led to two more fantastic rock pitches, following cracks up and right toward the summit (5.6). A final pitch up an icy fist crack and the last dollop of snow led us to the summit. We rappelled the beautiful South Pillar route, touching down in the hot-pink glow of the setting sun.
We named our route Smash and Grab (IV 5.8 M4), after Dieter Klose's expression for climbers who smash into town and grab a summit before the Stikine's clouds realize what's happened.
Dave Burdick, AAC


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Reliable Technology
Solutions Provider
You Dream It
We Build It
We develop your dreams/ideas
Into a real working thing
What can we < code > or design for you?
Starlabel is the only Creative Agency you will ever need. We provide your company or institution with complete modern technologies that thrive businesses. We are expert in these areas Networking, System Vulnerability Analysis, Server Configuration and Management, Smart Security Systems, Web Development, Mobile Application Development, User Experience (UI) / User Interface design, Computer Hardware Maintenance & Installation, Search Engine Optimization, Graphic Design, Corporate Branding and Consulting.
Computer Networking
CMS Development
Cross-Platform App Dev
UI/UX Design
Cross-platform App Dev
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Graphic Design
Infographic Design
Website Design/Development
Web Development


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Halfling Shire is the home of the Halflings and reside in the Great Plains. They generally are a peaceful race, but do not like people who do no enjoy a variety of food or who are too ambitious.
Finding their hidden home requires walking in the Great Plains until you randomly see the "To the west you see…" message appear.
So basically to avoid being attacked you want to have
- As few Quest Points as possible or
- As many different foods as possible.
The amount in 2 generally goes up with 1… so eventually you'll want as many as 30. Though if you have a seriously high Magic Shield, you can tend to shrug their attacks off like nothing.
When you walk in, its pretty uneventful until you get at the 6 way in the middle.
- If you go Northeast, you will run into a Food Shop, and if your a member of the Thieves Guild, you can join the Daggermaster guild.
- Going Northwest takes you to the Bank and Armor Shop where you can get a Halfling Necklace.
- Going Southwest takes you towards the Trinket Exchange, where you can send and receive amulets and rings.
- Going Southeast takes you to the Amulet Shop, where you can buy an Invisibility Ring and/or a Shining Pendant.
- Going West takes you to another Food Shop, and the Courier Corp.
A — Armor Shop
D — Dagger Guild (Thieves Only)
B — Bank
1 — Tenderleaf's Bar/Food Shop
2 — Rivertoe's Bakery/Food Shop
T — Trinket (Amulet and Ring Exchange/Storage)
3 — Amulet Shop
C — Courier Corp


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People have now a-days, (said he,) got a strange opinion that every thing should be taught by lectures. Now, I cannot see that lectures can do so much good as reading the books from which the lectures are taken. I know nothing that can be best taught...
Southey's Common-place Book - Pagina 516
di Robert Southey - 1850
Visualizzazione completa -