text:
Effects of inflammatory cytokine IL-27 on the activation of fibroblast-like synoviocytes in rheumatoid arthritis
DOI: 10.1186/ar3067
© Wong et al.; licensee BioMed Central Ltd. 2010
Received: 4 February 2010
Accepted: 6 July 2010
Published: 6 July 2010
Interleukin (IL)-27 is a novel member of the IL-6/IL-12 family cytokines that are produced early by antigen-presenting cells in T helper (Th)1-mediated inflammation. Elevated expression of IL-27 has been detected in the synovial membranes and fluid of rheumatoid arthritis (RA).
We investigated the in vitro effects of IL-27, alone or in combination with inflammatory cytokine tumor necrosis factor (TNF)-α or IL-1 β on the pro-inflammatory activation of human primary fibroblast-like synoviocytes (FLS) from RA patients and normal control subjects, and the underlying intracellular signaling molecules were determined by intracellular staining using flow cytometry.
Significantly higher plasma concentration of IL-27 was found in RA patients (n = 112) than control subjects (n = 46). Both control and RA-FLS constitutively express functional IL-27 receptor heterodimer, gp130 and WSX-1, with more potent IL-27-mediated activation of signal transducers and activators of transcription (STAT)1 in RA-FLS. IL-27 was found to induce significantly higher cell surface expression of intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 and release of inflammatory chemokine IL-6, CCL2, CXCL9, CXCL10 and matrix metalloproteinase-1 of RA-FLS than that of control FLS (all P < 0.05). Moreover, an additive or synergistic effect was observed in the combined treatment of IL-27 and TNF-α or IL-1 β on the surface expression of ICAM-1 and VCAM-1 and the release of CXCL9 and CXCL10 of RA-FLS. Further investigations showed that the expression of ICAM-1, VCAM-1 and chemokines stimulated by IL-27 was differentially regulated by intracellular activation of phosphatidylinositol 3-OH kinase-AKT, c-Jun amino-terminal kinase and Janus kinase pathways.
Our results therefore provide a new insight into the IL-27-activated immunopathological mechanisms mediated by distinct intracellular signal transductions in joint inflammation of RA.
Rheumatoid arthritis (RA) is a chronic autoimmune disease with 1% prevalence in the industrialized countries, characterized by cytokine-mediated inflammation of the synovial lining of the diarthrodial joints and the destruction of cartilage and bone [1]. Together with T and B lymphocytes and macrophages, fibroblast-like synoviocytes (FLS) play crucial roles in both joint damage and the propagation of inflammation in RA [2]. Normal synovial tissue consists of two anatomically distinct layers: a surface layer (intima or synovial lining) and an underlying layer (subintima). The predominant cell types in the normal intima and subintima are macrophage-like (type A) synoviocytes and FLS (type B synoviocytes). FLS are bipolar, spindle-shaped cells with prominent secretory machinery, including extensive endoplasmic reticulum, regular ribosomal arrays and well-developed Golgi apparatus [2]. FLS can mediate cartilage and bone destruction in RA mainly via the elucidation of matrix metalloproteinases (MMPs) such as MMP-1 [3]. Additionally, FLS can secrete receptor activators of nuclear factor-κB ligand (RANKL) that attract macrophages from the vasculature; stimulate the differentiation of vascular- and tissue-derived macrophages into osteoclasts; and activate osteoclasts at the bone surface, leading to bone erosion [4].
FLS mediates inflammation and autoimmunity through a wide range and complex mechanisms during the development of RA [2]. In RA, FLS respond to inflammatory cytokines including interleukin (IL)-1β, 6, 8, 12, 17, 18, 21, tumor necrosis factor (TNF)-α and interferon (IFN)-γ through the activation of multiple intracellular signaling pathways including extracellular signal-regulated protein kinase (ERK), c-Jun amino-terminal kinase (JNK) and p38 mitogen activated protein kinase (MAPK), leading to the expression of multiple cytokines such as IL-1β, IL-6, and TNF-α, as well as the secretion of MMPs that contribute to tissue destruction [3, 5–8]. Adhesion molecules present on the FLS surface, including CD44, vascular cell adhesion molecule (VCAM), and intercellular adhesion molecule (ICAM), regulate the trafficking of leukocytes into and/or through the synovial tissue [9]. Direct contact between FLS and T cells can induce T cell activation [10]. Secretion of IL-15 from FLS, along with the other cytokines, activates T cells, neutrophils and macrophages [10].
IL-27 is a heterodimeric cytokine composed of EBV-induced protein 3 (EBI3) and p28 subunit that signals through a receptor complex composed of the unique IL-27Rα (IL-27R) (WSX-1/T cell cytokine receptor (TCCR)) and gp130 signaling subunit. IL-27 belongs to IL-6/IL-12 family of structurally related ligands that include IL-12, IL-23 and IL-6 [11, 12]. IL-27 receptor has been found to be expressed on naïve T cells, NK cells, monocytes, mast cells, activated B cells, Langerhan's cells and activated dendritic cells, whereas IL-27 is produced mainly by activated macrophages and dendritic cells [11, 12]. IL-27 can induce a T helper (Th) type 1 response and the expression of T-bet and interferon (IFN)-γ by naïve T cells [11, 13]. In innate immunity, IL-27 can provoke the production of IL-1β, TNF-α, IL-18 and IL-12 in monocytes [14]. However, more recent studies revealed that IL-27 can play a regulatory role by suppressing the acquired immunity, inducing the development of Th cells, and expansion of inducible regulatory T cells to produce IL-10 [11, 15–17]. However, the molecular basis for pleiotropic actions of IL-27 in various immune responses has not been well elucidated yet. In animal studies of RA, administration of IL-27 can attenuate collagen-induced arthritis at the disease onset stage in mice [17]. However, in clinical studies, elevated protein and gene expression IL-27 has been detected in the established RA synovial membranes and fluid [17]. Moreover, IL-27 can induce a Th1 immune response and susceptibility to proteoglycan-induced arthritis in murine model [18]. Consistent with these findings, adjuvant-induced arthritis in rats and experimental autoimmune encephalomyelitis in mice can be abolished by anti-IL-27 antibodies [19]. Therefore, the immunopathological roles of IL-27 on the activation of FLS in RA remained unsettled, particular at the local inflammatory joints. In an attempt to further elucidate the immunopathological roles of IL-27 in the joint inflammation of RA, the in vitro activating effect of IL-27 in combination with TNF-α or IL-1β on FLS from RA and control subjects and its underlying intracellular signal mechanism was investigated in this study.
Materials and methods
RA patients, control subjects and blood samples
Patients with active RA (n = 112), who fulfilled the American College of Rheumatology 1987 criteria for RA [20], were enrolled in this study. Patients eligible for the study included those with active RA despite treatment with at least 12.5 mg of methotrexate per week. Active RA was defined by the presence of at least four swollen and tender joints and in addition at least two of the following: morning stiffness lasting at least 45 minutes, erythrocyte sedimentation rate (ESR) Westegren of at least 28 mm/h, and a serum C-reactive protein concentration of at least 20 mg/L [21]. Forty-six sex- and age-matched healthy Chinese volunteers were recruited as control subjects. Six mL of EDTA venous peripheral blood were collected from each patient and control subject. The above protocol was approved by the Clinical Research Ethics Committee of The Chinese University of Hong Kong-New Territories East Cluster Hospitals, and informed consent was obtained from all participants according to the Declaration of Helsinki.
Recombinant human IL-27 and TNF-α were purchased from R & D Systems, Minneapolis, MN, USA. ERK inhibitor PD98059, JNK inhibitor SP600125, p38 MAPK inhibitor SB203580, phosphatidylinositol 3-OH kinase (PI3K) inhibitor LY294002 and Janus kinase (JAK) inhibitor AG490 were purchased from Calbiochem Corp, San Diego, CA, USA. SB203580 was dissolved in water while PD98059, LY294002, SP600125 and AG490 were dissolved in dimethyl sulfoxide (DMSO). In all studies, the concentration of DMSO was 0.1% (vol/vol).
Endotoxin-free solutions
Cell culture medium was purchased from Cell Applications Inc., San Diego, CA, USA, free of detectable lipopolysaccharide (< 0.1 EU/ml). All other solutions were prepared using pyrogen-free water and sterile polypropylene plastic ware. No solution contained detectable LPS, as determined by the Limulus amoebocyte lyase assay (sensitivity limit 12 pg/ml; Biowhittaker Inc, Walkersville, MD, USA).
Cell culture of FLS
Human FLS isolated from synovial tissues obtained from normal healthy subjects and patients with RA were purchased from Cell Applications. FLS was cultured in synoviocyte growth medium including 10% synoviocyte growth supplement (Cell Applications) in 5% CO2 - 95% humidified air at 37 C [22].
Quantitative PCR of GP-130 and WSX-1 gene expression
Total RNA of FLS was extracted using RNeasy Mini Kit (Qiagen Inc., Valencia, CA, USA). All RNA samples were pre-treated with deoxyribonuclease I (Invitrogen Corp., Grand Island, CA, USA) and then stored at -70°C. For each treatment, approximately 1 μ g of total RNA was reversely transcribed to complementary DNA (cDNA) with TaqMan Reverse Transcription Reagents (Applied Biosystems Inc., Foster City, CA, USA). The mRNA expression of GP-130, WSX-1 and GAPDH (endogenous control) was quantified by real-time PCR using SYBR Green probe (Roche Diagnostics Corp., Indianapolis, IN, USA) with the use of Applied Biosystems 48-well StepOne™ Real Time PCR System. The primers human GP-130, WSX-1 and GAPDH were as follows, human hgp130 forward: 5'-TCTGGGAGTGCTGTTCTGCTT-3', human gp130 reverse: 5'-TGTGCCTTGGAGGAGTGTGA-3'; human wsx-1 forward: 5'-GCTCCTGCCTCTATGTTGGC-3', human wsx-1 reverse: 5'-CCTTCATGTTCTTGGACCAGC-3'; GAPDH forward: 5'-ATGGGGAAGGTGAAGGTCG-3', GAPDH reverse: 5'-GGGGTCATTGATGGCAACAATA-3'. Real-time PCR was performed in a 25 L reaction mixture containing primers, FastStart Universal SYBR Green master (ROX) reagent (Roche) and cDNA sample in duplicate. A negative PCR control without template and a positive PCR control (GAPDH) with a template of known amplification cycle were included. The real-time PCR reaction was performed with 95°C for 15 sec to denature cDNA, 60°C for 60 sec to allow the SYBR green probe and primers to anneal to the denatured cDNA. The cycles were repeated 40 times after an initial 10-minute denaturation at 95°C. The threshold cycle (Ct) is the PCR cycle at which an increase in reporter signal above the baseline signal can first be detected. The relative GP-130 and WSX-1 mRNA expression (n = 3) was obtained by comparing with the relative expression of GAPDH using 2-ΔCt (Ct,gp130/wsx-1 - Ct,GAPDH).
Western blot analysis
Cells were washed, lysed, and an equal amount of proteins to ensure equal protein loading was subjected to SDS-PAGE and then blotted onto PVDF membrane (GE Healthcare Corp, Piscataway, NJ, USA). The membrane was blocked with 5% skimmed milk and probed with primary antibody against WSX-1 at 4°C overnight. After washing, the membrane was incubated with secondary antibody coupled to horseradish peroxidase (GE Healthcare) for one hour at room temperature. Antibody-antigen complexes were then detected using an ECL chemiluminescent detection system (GE Healthcare).
Assay of human cytokines, chemokines and MMP-1
Plasma concentration of IL-27 was quantitated by ELISA reagent from R & D Systems. The concentration of chemokine CXCL9, CXCL10, CCL2 and CCL5 in culture supernatant with equal cell number loading was measured simultaneously by bead-based multiplex cytokine assay with BD cytometric bead array (CBA) (BD Biosciences Pharmingen, San Diego, CA, USA) using a four-color FACSCalibur flow cytometer (BD Biosciences Corp, San Jose, CA) [23]. Human MMP-1 in culture supernatant was assayed by ELISA reagent (RayBiotech Inc. Norcross, GA, USA).
Flow cytometry of adhesion molecules and gp130 on cell surface
Cells were resuspended in cold PBS and blocked with 2% human pooled serum at room temperature for 15 minutes, followed by washing with PBS, cells were incubated with the FITC-conjugated mouse anti-human ICAM-1 or VCAM-1 monoclonal antibodies, or FITC-conjugated mouse IgG1 isotypic control (BD Pharmingen) at 4°C in dark for 45 minutes. For the detection of surface expression of gp130, unconjugated mouse-anti-human gp130 (R & D Systems) followed by secondary FITC-conjugated anti-mouse IgG antibodies (BD Pharmingen) were used for the staining. After washing, cells were fixed with 1% paraformaldehyde in PBS. Expression of cell surface adhesion molecules and gp130 on 5,000 viable cells was then quantitatively analyzed by flow cytometry (FACSCalibur flow cytometer, BD Biosciences) in terms of mean fluorescence intensity (MFI).
Intracellular staining of activated (phosphorylated) signaling molecules
The intracellular expression of phosphorylated signaling molecules was determined using previously established intracellular staining assay [23]. Briefly, cells were fixed with pre-warmed BD Cytofix Buffer (4% paraformaldehyde) for 10 minutes at 37°C after stimulation by IL-27. After centrifugation, cells were permeabilized in ice-cold methanol for 30 minutes and then stained with mouse anti-human phosphorylated AKT, phosphorylated JAK2, phosphorylated JNK, phosphorylated signal transducers and activators of transcription (STAT)1(pY701) (BD Pharmingen) and phosphorylated PI3K (Cell Signaling Technology, Beverly, MA, USA) monoclonal antibodies or mouse IgG1 isotypic control (BD Pharmingen) for 60 minutes followed by FITC conjugated goat anti-mouse secondary antibody (BD Pharmingen) for another 45 minutes at 4°C in the dark. Cells were then washed, resuspended and subjected to analysis. Expression of intracellular phosphorylated signaling molecules of 5,000 viable cells was analyzed by flow cytometry (FACSCalibur, BD Biosciences) as MFI.
Statistical analysis
Plasma concentrations of IL-27 were expressed as median (interquartile range, IQR) as they were not in Gaussian distribution and its difference between RA and control groups was assessed by Mann-Whitney rank-sum test. The statistical significance of differences of other parameters was determined by one-way ANOVA. The values were expressed as mean ± SD from three independent experiments. Any difference with a P-value less than 0.05 was considered significant. When ANOVA indicated a significant difference, the Bonferroni post hoc test was then used to assess the difference between groups. All analyses were performed using the Statistical Package for the Social Sciences (SPSS) statistical software for Windows, version 16.0 (SPSS Inc, Chicago, IL, USA).
Plasma concentration of IL-27 in RA
The recruited RA patients (female:male = 97:15) cohort was found to have mean age of 53 ± 9 (range 31 to 83 years) while control subjects (female:male = 40:6) have a mean age of 47 ± 7 (range 25 to 62 years). Plasma concentration of IL-27 was found to be significant higher in RA patients than that in control subjects (11.7 (7.2 to 19.2) vs 7.1 (4.9 to 10.0) ng/ml, P < 0.001).
FLS express functional IL-27 receptor
IL-27 up-regulated ICAM-1 and VCAM-1 expression on the cell surface of FLS
IL-27 could enhance CCL2, CXCL9, CXCL10 and MMP-1 production from FLS
Furthermore, IL-27 (50 ng/ml) could induce MMP-1 (13.8 ng/ml) release comparing with medium control (9.4 ng/ml) from RA-FLS but IL-27 (10 to 100 ng/ml) cannot induce any MMP-1 release from control FLS.
Enhanced up-regulation of adhesion molecules and the release of chemokines upon combined treatment of IL-27 and TNF-α or IL-1β
Effects of IL-27 on the activation of STAT1, JAK-2, AKT, PI3K and JNK signaling pathways in FLS
Effects of signaling molecule inhibitors on the expression of adhesion molecules and release of chemokines from FLS activated by IL-27
To investigate the immunopathological roles of inflammatory cytokine IL-27 on joint inflammation, we first confirm the significantly elevated plasma level of IL-27 in RA patients than normal controls. Apart from immune effector cells, recent study showed that IL-27 can also activate non-immune effector cells such as keratinocytes [25]. Our present expression analysis of IL-27 receptor complex has suggested that IL-27 could activate other non-immune cell targets such as joint FLS. Human FLS constitutively expressed functional IL-27 receptor complex WSX-1 and gp130, and STAT1 was phosphorylated upon IL-27 stimulation (Figures 1 and 6). The present results are therefore in concordance with the expression of gp130 and WSX-1 on synovial biospsies [26–28]. IL-27-mediated activation effects have been shown to be mainly through the regulation of STAT1 phosphorylation [13, 25, 29]. For example, IL-27 could activate STAT1 to induce IL-12Rβ2 expression in naive CD4+T cells for Th1 differentiation [13]. IL-27-induced STAT1 activation was associated with proinflammatory effects in human monocytes and keratinocytes [25, 29]. To further explore the difference of the activating effects of IL-27 on RA and control FLS, we performed functional and underlying mechanistic studies.
In the present study, we found that IL-27 was able to induce significantly higher expression of ICAM-1 and VCAM-1 and to augment TNF-α- and IL-1β-induced ICAM-1 and VCAM-1 on RA-FLS than that of control FLS (Figures 4 and 5), thus providing a novel immunopathological mechanism of IL-27 mediated joint inflammation in RA. ICAM-1 and VCAM-1 are transmembrane glycoprotein of the immunoglobulin supergene family expressed on many cell types. ICAM-1 and VCAM-1 participate in leukocyte-leukocyte, leukocyte-endothelium, and leukocyte-epithelium interactions and transendothelial migration [30]. Expression of VCAM-1 on synovial fibroblasts is also a clinical hallmark of RA [31]. Expression of these adhesion molecules on FLS has been implicated in the pathogenesis inflammation in RA by the induction of the infiltration of leucocytes into local joint [32]. Because of the pivotal role of ICAM-1 and VCAM-1 in inflammation, antisense oligonucleotides, monoclonal antibodies or blockage compound against ICAM-1 and VCAM-1 have been demonstrated with therapeutic beneficial effects in inflammatory diseases [33]. Expression of ICAM-1 and VCAM-1 on epithelial cells can be up-regulated by cytokines such as TNF-α, IFN-γ or IL-1δ [34, 35]. We herein provided evidence that IL-27 is a potent inducer of ICAM-1 and VCAM-1, especially in combination with TNF-α or IL-1δ, on RA-FLS but not control FLS, thereby elucidating the selective pathological roles of IL-27 in RA.
A panel of inflammatory cytokines such as TNF-α, IL-1, IL-6, IL-23, and IL-2 families are active in the joints of the patients with RA by causing inflammation, articular destruction, and the comorbidities [24]. We also found that IL-27 could induce a significantly higher amount of IL-6 from RA-FLS than control FLS (P < 0.05, data not shown). To examine whether IL-27 also modulates the expression of chemokines in FLS, we found that IL-27 can induce significantly higher release of inflammatory chemokines CCL2, CXCL9 and CXCL10 from RA-FLS than control FLS. Combined treatment of IL-27 and TNF-α or IL-1β also resulted in the synergistic production of CCL5, CXCL9 and CXCL10 of RA-FLS (Figures 4 and 5). CCL2, CCL5, CXCL9 and CXCL10 are crucial inflammatory chemokines for the chemoattraction of Th1 cells and macrophages [36]. Those chemokines may also stimulate FLS to release other cytokines and MMP for the cartilage degradation and pannus formation [36]. In fact, we observed that IL-27 could induce the release of MMP-1 from RA-FLS but not control FLS. MMP-1 has been suggested to correlate with the invasive growth of FLS in RA [3]. Furthermore, CCL2, CCL5, CXCL9 and CXCL10 have been shown to be highly expressed in synovial fluid and synovial tissue [37–39]. Stimulation of RA-FLS with CCL2 and CCL5 can enhance the production of IL-6 and CXCL8 [40], while CCL2, CXCL10, CCL5 and CXCL9 induce the gelatinase and collagenase activities in the supernatants of cultured FLS [41]. CCL2 and CCL5 can stimulate MMP production by chondrocytes, inhibit proteoglycan synthesis and enhance proteoglycan release from the chondrocytes [42]. In addition, CCL5 can induce the expression of inducible nitric oxide synthase, IL-6 and MMP in chondrocytes [43]. FLS cells are a kind of primary cell without cell proliferation and division. The FLS cell numbers were constant during culture and therefore the constant cell number did not have any effect on the chemokines and MMP concentrations being measured in the cell culture supernatant.
Although IL-27 alone could not activate CCL5 release from RA and control FLS, IL-27 exhibited synergistic effect with TNF-α to induce CCL5 release in RA-FLS. Actually, TNF-α can induce the expression of CCL5 in cultured FLS [44]. Our results show that RA-FLS exhibits a lower level of TNF-induced ICAM-1 and VCAM-1 expression as compared to control FLS. It may be due to the lower cell surface expression of TNFRp55 (data not shown), the crucial TNF receptor component for the expression of adhesion molecule ICAM-1 and VCAM-1, on RA-FLS compared with control FLS [45, 46]. However, our previous study showed that the synergistic activating effects of IL-27 and TNF-α on bronchial epithelial cells were partially due to the IL-27 up-regulated expression of TNF-α receptor p55TNFR [47]. IL-27 stimulation could also lead to the increased expression of TNF-α and IL-1β in primary human monocytes [14, 48]. The above may account for the synergistic effect of IL-27, TNF-α and IL-1β for chemokine release. Since FLS, monocytes and Th1 cells play important roles in the pathogenesis of RA, the enhanced production of ICAM-1 and VCAM-1, and chemokines for macrophages and Th1 cells of RA-FLS induced by IL-27, TNF-α and IL-1β should contribute to the development of inflammation.
IL-27 has been reported to activate the JAK/STAT signaling pathway in T lymphocytes [49], but the signaling pathways of IL-27 in FLS have not been reported. We have previously reported the involvement of ERK, JNK, JAK and p38 MAPK pathways in the expression of adhesion molecules and release of cytokines and chemokines from activated bronchial epithelial cells upon exposure to diverse stimuli such as TNF-α, IL-4, IL-13 and IL-31 [50, 51]. IL-27 stimulation could lead to receptor-mediated tyrosine phosphorylation of the STAT family [52]. Our study showed that incubation with IL-27 resulted in STAT1 tyrosine phosphorylation within five minutes and maintained it for 30 minutes in both control and RA-FLS, with a more potent activation in RA-FLS (Figure 6A). This discrepancy may account for the higher induction of adhesion molecules and release of chemokine upon IL-27 stimulation in RA-FLS. We also found that PI3K-AKT and JNK pathways were activated in both RA and control FLS in response to IL-27, whereas JAK-2 in RA-FLS was activated upon IL-27 activation (Figure 6). Actually, previous studies have revealed that JNK and AKT involve the inflammatory mechanisms of FLS in RA [53, 54].
Using selective inhibitors that could suppress the activation of their corresponding signaling pathways, we further elucidated the involvement of different signaling pathways in regulating expression of adhesion molecules and chemokines of FLS. These inhibition experiments demonstrated that the IL-27-induced up-regulation of ICAM-1 on control FLS, and ICAM and VCAM-1 on RA-FLS was regulated by the activation of intracellular PI3K-AKT and JNK pathways (Figure 7). Regarding the chemokine release, IL-27 induced CXCL9 and CXCL10 from control FLS, CCL2 and CXCL10 from RA-FLS and CXCL9 from RA-FLS was found to be mediated by PI3K and JNK pathways, JAK, PI3K and JNK pathways, and JAK and PI3K pathways, respectively (Figure 7). The above discrepancy of the intracellular signaling mechanisms between RA and control FLS for the induction of adhesion molecules and chemokines indicates the distinct signaling pathways in inflammatory responses. Such induction of chemokines by IL-27 in RA-FLS was not completely inhibited by those inhibitors, other unidentified signaling pathways might therefore contribute to the chemokine expression. Further experiments may be required to investigate the regulatory mechanisms of different signaling molecules and transcription factors for the induction of chemokines in human FLS upon cytokine treatment.
Using a murine model, IL-27 can exhibit a suppressive effect on inflammation and can attenuate collagen-induced arthritis when administered at the onset of the disease via the blocking of Th17 differentiation from naïve T helper cells [17, 55, 56]. On the other hand, the late onset of IL-27 was unable to downregulate Th17 in established RA, thereby inducing the inflammation of the ongoing adjuvant-induced arthritis in vivo [57]. In view of that, our study using FLS derived from normal subjects and established RA patients, therefore, only exhibits the activating effects rather than the suppressive effects of IL-27. Since IL-27 is detected at significant concentrations in the joints of patients with ongoing RA, neutralization of IL-27p28 has been suggested as a novel therapy [57]. Previous therapeutic approaches relied on disease-modifying antirheumatic drugs (DMARDs) such as methotrexate and sulfasalazine that had only partial clinical beneficial effects and were associated with toxicity [24]. In view of the application of antibodies for the blockade of TNF-alpha, IL-1β and IL-6 receptor as treatments for RA, other cytokines such as IL-27 may therefore offer alternative targets for therapeutic intervention or may be useful as predictive biomarkers of established RA [57–59]. Because of the recent advances in the application of MAPK and NF-κB inhibitors as potential anti-inflammatory agents [60], our present study should also provide new clues for the development of novel treatment for IL-27-mediated RA inflammation.
Our results indicate the differential inflammatory IL-27-mediated intracellular signaling mechanisms including STAT1, PI3K-AKT, JNK and JAK pathways for the activation of RA and control FLS. They suggest the distinct signaling pathways during inflammatory responses in RA. As a result, IL-27 plays crucial immunopathological roles in joint inflammation in RA by the induction of adhesion molecules, cytokines and chemokines, especially combined with TNF-α and IL-1β. Our study also provides a biochemical basis for the development of a new modality for RA.
cytometric bead array
- DMARDs:
disease-modifying antirheumatic drugs
dimethyl sulfoxide
EBV-induced protein 3
extracellular signal-regulated protein kinase
erythrocyte sedimentation rate
fibroblast-like synoviocytes
intercellular adhesion molecule
Janus kinase
c-Jun amino-terminal kinase
mitogen activated protein kinases
mean fluorescence intensity
- MMPs:
matrix metalloproteinases
phosphatidylinositol 3-OH kinase
rheumatoid arthritis
receptor activator of nuclear factor-κB ligand
signal transducers and activators of transcription
T cell cytokine receptor
- Th:
T helper
tumor necrosis factor
vascular cell adhesion molecule
This work was supported by a Direct Grant from The Chinese University of Hong Kong (Reference No.: 2008.1.018).
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This article is published under license to BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


text: Food Open for breakfast, lunch and dinner every day, The Princess Royal is one of Farnham's best places to eat. Our chefs make use of locally sourced ingredients to create mouth-watering home-cooked pub-food classics, which can be enjoyed in the contemporary Waverley Room, the spacious bar or even al fresco in summertime. Right now, we're putting the final finishing touches on our exciting Spring menu to make the most of fantastic seasonal produce - so watch this space for a sample menu coming soon. We take taste personally "Every dish is a signature dish" In our Fuller's Kitchen, we take taste personally. That means our talented chef team have seasonal, local produce delivered every day. It also means they have the freedom to put their stamp on every dish, and it is why you will see Head Chef, Paul Deakin's signature on every menu. Learn more about the passion behind Fuller's Kitchen.

subset: commoncrawl


text: I have an old electric Youngstown Kitchens wall oven. Recently the oven door would not stay shut. The problem is with a broken safety cable (about 10inches) attached to a spring within the door. I was originally thinking it was made by Tappan. Looking at the tag in the oven, it reads Youngstown Kitchens and Stiglitz Corporation, Model ED024. I'm speculating it was assembled by the Stiglitz Corporation for Youngstown Kitchens. It looks like a simple piece but I cannot seem to locate a replacement part. Anyone know if this part exists? I'm thinking I may have to just have the part made up.


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Fasten your Seatbelts. Turbulence Ahead.
As a child, yearly visits back to India meant weeks of frenzied shopping and the launch of a certain degree of verbal warfare at home. Words like "Flash visit", "shopping" and "luggage" flew through the air dodging furniture and landing on the opponent. When Mummy flung 'Shopping' and 'One more?' at Daddy, he retaliated with "Money" and "Luggage!!!"
Suitcase after suitcase were crammed to bursting levels. Torches, emergency lamps, jars of Tang powder, packets of chocolates and enough Nido milk powder to meet the calcium requirements of a small village, all jostled for space in there.
Passenger Comfort like never before.
Onboard the flight, we smiled smugly as the perky air-stewardess dutifully performed the emergency drill. We had our own private stash of food and supplies in the luggage bin. All we needed was to cling onto our bulky suitcase and allow it to drift us ashore to some uninhabited island in the midst of the Indian Ocean.
We would spend the rest of our days sipping thick, milky morning cuppas and snacking on pistachios with tall glasses of Tang juice later in the evening. An island lifestyle which would have forced Robinson Crusoe to go hide behind the coconut trees in shame.
God's Own Country's very own Car.
The arrival at Cochin airport was always followed with the same ritual. The airport doors would whoosh open so that we would be rudely awakened from our breezy island dreams and instead be hit by a balmy gust of tropical air. Year after year, our trusty driver Jose (Jose-Uncle for us kids), greeted us at the entrance grinning beside his gleaming white Ambassador car.
Now for those of you who have not had the pleasure of travelling in an Ambassador, I need to dedicate a few lines to this man-made marvel. An Ambassador is not just a car. It is THE Car. Move over, Benz and BMW.
Want to fit an entire football team in a single car? Ambassador is the answer.
Want to stuff the luggage of an NRI with several 'conditionally' grateful relatives? Again, Ambassador fits the bill.
Mummy masters the Magician Hat trick!
Once home, eager relatives would steadily trickle in. Helping hands would break open stubborn clasps on some of the suitcases. Others would crowd around trying to perfect that correct expression of surprise and gratitude when Mummy distributed the 'A Little Somethings'.
Unpacking the suitcases was like pulling things out of a Magician's Hat. One surprise after another. It still amazes me how many torches, emergency lamps and flasks could be stuffed inside a suitcase. On top of that, 'Gulf' quality handkerchiefs would be stuffed inside these flasks. And it didn't stop there. Gold rings and chains (again 'Gulf' quality) were wrapped inside these handkerchiefs. It was a bit like a game of 'Pass the Parcel' gone haywire.
Family Scripture saying: "And not a single leaf of the Family Tree shall be unvisited".
Then we proceeded to the nightmarish part of the trip - visiting relatives - which was like Halloween 'Trick o' Treat' ing, except the candy was replaced by cups of tea and plates of 'mixture'. Almost every house in a 5 km radius was visited because everybody was related and everybody was on the 'Cannot Avoid' list. Of course, Jose-Uncle and his trusty Ambassador ferried us across to each house and back.
Back to the Future.
And 20 years down the line, Jose-Uncle still greets me each time I arrive at Cochin airport laden with my own set of stuffed suitcases. Some things haven't changed since then. I am still terribly motion-sick all the way home. Jose-Uncle still has a very strong opinion about Kerala politics, which I don't understand at all. The Ambassador still rocks (literally and figuratively).
But the difference is that nobody wants emergency lamps anymore. The zillion inverter companies around the state saw to that. And candies that come in packets? No way! Nothing less of a Ferrero Rocher or Toblerone is acceptable.
Journey back to Dubai and Deja Vu.
Predictably enough, keeping up with the superhuman packing and hoarding skills of Indians, we took back as much luggage as we arrived with. Watermelon-sized mangoes, bunches of bananas, innumerable spices, and pickles of every possible vegetable and meat known to the Malayali. Enough supplies to get us through any impending crash scenarios and reach that magical island.
That is if the weight of Mummy's newly purchased Kanjeevaram sarees don't cause a buoyancy problem.


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Toll Free Number
12:00 Noon CST
Please enter the dates of your stay, number of rooms, and guests to see a list of available rooms.
This Zapata hotel offers great service and amenities for business or leisure travelers with warm, Texas style hospitality!
Its proximity to Falcon Lake on the Rio Grande River makes the Best Western Inn by the Lake an ideal location for any outdoor enthusiast or tourist looking for adventure. Falcon Lake is especially known for its fishing, boating and bird watching. Also close by lies the historic Spanish village of San Ygnacio. The hotel's location is convenient to several major Texas cities as well as to the Mexican border.
Each comfortable room features king or queen-size beds, a microwave, refrigerator and high-speed Internet. Guests can choose to relax in the outdoor pool or have a picnic in the pavilion located on-site. The hotel also serves a complimentary full breakfast every morning.
Whether for a business trip or just for a little relaxation, come and experience the unique flavor of southern Texas. Make an online hotel reservation with today at this Zapata, Texas hotel today!
Latest Reviews
Stopped here on travel and the hotel is great. Service was wonderful. And very convenient to everything in this area. I would definitely consider staying here again in the future. Best Western is so...READ FULL REVIEW
had to move to another room after first night a/c failure ice machine broke day 2 parking limited for my boat trailer breakfast was good lots of road construction at entrance tolerable convenient to...READ FULL REVIEW
This is the first time I have been to Zapata and trip advisor said this was the #1 place to stay. Definitely must have been an off night for them. Room was spacious but when I walked in it had only...READ FULL REVIEW
I really liked the way it looked outside and inside the lobby smelled and looked very clean. Need to have that person working up front clean the rooms, or manage people cleaning....looked clean but...READ FULL REVIEW
Very nice clean hotel, very roomy. It's has plenty of all the little extra stuff that I look for. The decor is very modern. Although it's right by 83, the highway, you'd couldn't hear the traffic...READ FULL REVIEW


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The deal to extend the payroll tax cut includes increases to new federal employees' retirement contributions.
Under the Middle Class Tax Relief and Job Creation Act of 2012, feds hired after Dec. 30, 2012 and those with less than five years of previous federal service, will have to pay 2.3 percent more of their salaries toward their defined benefit pensions.
The Office of Management and Budget refers to these future feds as revised annuity employees (RAE). The chart below details the RAE and agency contributions. OMB released the chart as part of the OMB's A-11 budget guidance for agencies' fiscal 2014 budget requests.
Source: A-11 Budget Guidance
Hiring freeze result of Trump team's review of federal agencies
Hiring freeze result of Trump team's review of federal agencies
New feds' pension contributions increase under payroll tax cut deal
Federal pay and benefits tracker
Photos of the amazing, moving, important and amusing things happening in the federal community.
A daily update of important moments in the history of the U.S. government.


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Riad Ghita
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TravelPod Member ReviewsRiad Ghita Fes
What a beautiful introduction to Moroccan style! Lovely rooms in a refurbished 16th (??) century riad, just inside the gates of the fantastic Fes medina. We had excellent dinner and breakfast here, too. Watch the stairs, and thank heavens for the lovely staff who carried our luggage up and down!!
This review is the subjective opinion of a TravelPod member and not of TravelPod.com.
TripAdvisor Reviews Riad Ghita Fes
Travel Blogs from Fes
Arrival in Fez, Morocco
... vegetables: fried eggplant, zucchini with a light tomato sauce, spicy olives, caramelised carrots, and some other mix of vegetables and luscious flavours that we couldn't identify - this was my favourite. Next, chicken tagine with olives and lemon, and that doesn't begin to describe its deliciousness. Finally, a light sponge-type cake with a streak of apricot (I think) through it and a light yoghurty topping. It made a late night but made a perfect start to our stay in ...
Big day. Sun up (You don't see that everyday!).
... think he is fully awake at the moment as although he sounds American he has only muttered a single 'mornin'. I think he too is here for the sunrise although his timing is probably a bit better than mine. I have that dilemma about continuing to type or saving battery power again. As there's no power up here I'm using battery and then I suffer a bit through the day although as the wifi is so poor (did I mention that?) I may not need the iPad so much. I ...
For our journey to Fez we decided to save some money by getting a bus instead of a train. We couldn't find too much info about the mystical bus so decided to just walk around the bus stops and try and figure it out with our handful of French words and traveller spirit. The spirit disappeared pretty quickly thanks to the pouring rain, the strain of walking with our very heavy bags and the frustration of having no idea where we were going. Eventually someone pointed us in a ...
Adventure in Morocco...F
... a wonderful fountain that graces the center of the courtyard. The walls are covered with intricate mosaics or exquisitely carved stucco, that is typical of the interior decorations of royal buildings in Fez. We enjoy an excellent Moroccan lunch here, with the highlight being the opportunity to take in all of the beauty that surrounds us here. It is definitely like something out of a movie. in beautiful courtyard of an old home.
After lunch, we spent ...
The Medina
... can easily imagine that the Medina has not actually changed that much over the centuries and we can only hope it stays that way. While we did not do lots here, just the things we wanted, there are numerous other activities offered through accommodation providers and also Fez can be a useful base to visit other nearby towns if they spark your fancy. OK post is long enough time to leave it at that and head off to the next stop: Chefchouen. ...


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Kid-friendly B. Spinaroos offers fun and excitement with soft intertwining bristles that even a small hand can handle. Kids can create characters, creatures and vehicles for custom fun.
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|Spinaroos (75 pcs)
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Comments about Spinaroos (75 pcs):
We bought these for my son at the age of one and he is now 4 and still plays with them. We have purchased an extra bucket (of simple peices) so he can create more things. He is forever building them into animals and buildings-they are a wonderful way for him to use his imagination and they are durable and safe.


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I believe all of the people in the legion team, in this match, cheated on purpose. They clearly made parasite a neutral and thus they could kill him constantly and get farmed without leaving the base.
This is an annoying feat and I do hope that you can ban them, would be much abliged.
Match ID: 81135473
Obviously it's a bug with parasite. You can infest one of your own in lvl 3 (Now need lvl 2 in infest) then you click on yourself (while infested) and spam on the infest button. Thus you die and become a neutral enemy.
1. Go out and get lvl 3.
2. Infest one of your own creeps.
3. Click on yourself and click the "Infest" button multiple times.
4. You are now dead and is seen as a neutral.
How big of a bug isn't this? You can even do it in a MM game.
It's a known bug, pretty big.
Rob Cherry | Former Technical Producer / Programmer


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15 Parks near Dorchester, South Carolina
Ridgeville, South Carolina 5.01 miles SE
Saint George, South Carolina 11.78 miles NW
St George, South Carolina 12.07 miles NW
Holly Hill, South Carolina 12.54 miles N
Summerville, South Carolina 14.59 miles SE
269 Clark Ln Dorchester, South Carolina 29437 (map) | (843) 462-2255
Sites: 12 | Latest Rate: $23.00 | 3 Reviews
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Sep 2012 - $23.00
We found this park in the Passport America Directory, and it is no more than a field in the backwoods. The grass and weeds are 8-10 inches high. Upon our arrival we were met by the owner's wife in a car and the owner on his ATV with two loose dogs running beside them. It was late in the day and too far back to the freeway to leave. Two sad rundown RV's were in the middle of the field with pitbulls running loose. We will NEVER return. - Arizona Traveler
Nov 2010 - $20.00
Did not honor Passport America rate. We camped in RV site and set up our tent. Intended to pay full PA rate but on checking out, we were charged $20, we complained and were told that they do not allow tent campers. Why did they take our money, why didn't they say anything before checkout time? I suggest avoiding this rundown campground. - Steve_M
Jun 2010 - $26.00
We spent three days here only because it was conveniently located next to Francis Beidler Forest, an Audubon sanctuary and well worth a visit, but many a camper will simply turn around and leave upon first viewing this campground-with zero "curb appeal" as it is simply a treeless/shadeless open field with twelve pull through sites. We were the only guests other than two men who lived there. In a nutshell, the place reeks of neglect; grass does not appear to have been mowed in months, possibly years. True, they offer full hook ups, but no restrooms. We had to remove ticks from ourselves and our dogs after walking the premises. There is NOTHING to do in park itself, and this is worth no more than an overnight stay while in transit to some place else. Owners need to visit a few other RV parks to have a point of reference for how a campground CAN look! - GLCDP
746 Givhans Ferry RD Ridgeville, South Carolina 29472 (map) | (843) 873-0692
Sites: 25 | Latest Rate: $21.00 | 9 Reviews
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Oct 2016 - $21.00
Small campground with some large sites, only 5 with sewer. Other sites with 30 amp electric and water. 1 bathroom for all on side of campground so we saw lots of people walking thru the middle sites. Trees all over so satellite TV hard to get. Internet is very poor. With all the rain, we had many bugs, even during the day (after hurricane Matthew). Dusty road but in good condition. Dumpster and recycle away from campground. Host checked us in (very nice and helpful); rangers never waved or said Hi. Gate locked at 6 pm this time of year but you get a code for the lock. Rating down due to position of bathroom and dumpster. Dump station is short for larger motorhomes and there are many trees next to the road. - pegdiver
Jun 2016 - $15.00
Small campground. 25 spacious sites. Seven sites have full hookups. Remainder have water and electricity. Dump station is easily accessible as you leave the campground. Numerous tent sites in the woods. RV sites are packed gravel/sand. Roads are packed sand. There were heavy rains prior to our arrival. Workers were spreading sand on the roads to smooth them out. Brand-new electrical utility connections worked perfectly. Water and sewer connections worked fine. Excellent television reception. Virtually no AT & T cell coverage. No wi-fi. Bathrooms were ok, but in need of updating. No trash receptacles in the campground. Dumpster facility accessible as you leave the campground. NOTE: bring lots of mosquito repellent! Mosquitos were aggressive and abundant, especially along the trails through the woods. - tomnorman457
Jan 2016 - $15.00
Stayed 3 days. Campground host, was very nice & accommodating. All dirt roads with potholes. Sites are packed dirt which on rainy days would be very messy. Very overgrown trees & shrubs at sites. Only E&W (30 amp). Walked through tent & cabin areas which looked very nice. Bathroom & showers were kept clean. Area to dispose of trash is a long distance away from sites. Personally would not camp here again. - rvgirl54
289 Horne Taylor Road St George, South Carolina 29477 (map) | (843) 563-8303
Sites: 32 | Latest Rate: $36.00 | 90 Reviews
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Aug 2016 - $36.00
The rate reflects $3/day upcharge for 50amp service. Though the park is about 5 miles off I-95, it is an easy drive over good roads. Our site was a pull thru, with the site and roads throughout the park crushed stone/gravel all in good condition. We were given a wifi password, but did not have any wifi service at our site. AT&T service is very weak, but Verizon is quite strong. There is no park cable, but our site had good satellite access as do many of the sites in the park. There is a small playground, but no pool. One of the park rules stipulated all dogs were to be leashed, max 6 ft. However we saw several instances of dogs off-leash, something that makes us feel a bit uncomfortable, particularly when the dog is fairly large as was the case here. It's hard to understand why some people don't seem to think common RV park rules like this don't apply to them, or that the park doesn't enforce their own rules. Other than this, the park was acceptable. Given the roaming dogs and the fact that the park is a 10 mile round trip off I-95, we would probably not stay here again. - Johnson's
Jun 2016 - $30.00
We stayed here for three nights while visiting friends. This is a well-maintained park. The owner is friendly and the park is quiet. Roads are gravel; majority of sites have paved pads with patios. The owner is very friendly. Close (45 miles) to Charleston and about 30 miles to Summerville. Being out in the country, phone service was weak (AT&T). There is no cable. WiFi available in the laundry room only. Be sure to remember that check-in is at 2:00. We arrived at 1:00 (forgot about check-in time) and the office was closed. We would stay here again. The majority of those staying here are permanent residents; but looks as if they're leaving the next day. - oldbluejay
Jun 2016 - $28.00
We stayed here 1 month and a week. Once again, had a great time. This park is in the country so phone service is weak. The park is about 45 minutes from downtown Charleston and 30 minutes from Summerville. Roads are rocked and the park is well maintained. - Grammy8
139 Motel Drive St George, South Carolina 29477 (map) | (843) 563-4180
Sites: 30 | Latest Rate: $17.00 | 13 Reviews
|Clean Restrooms:
|Clean Showers:
Apr 2016 - $17.00
We saw this campground in Passport America so we pulled in and this place should not be listed in Passport America. High grass, electrical hookups were knocked over, sites could not be more than 30' back-in and mud if you get off the pavement. Please don't waste your time. - sportscoach 1
Oct 2013 - $27.00
We drove by and decided not to stay here. It is close in and fuel and food handy. As said in a previous report, it looks 'seedy'. The RV park is behind the motel and down in a low area prone to standing water. The road through is narrow, the sites narrow and all back-ins. Vehicle parking could be a problem for your tow/toad. Virtually next door to Flying J and other services as well as right off the Interstate could make this an OK overnight. - Lew
Feb 2013 - $16.00
Stayed one night with Passport America rate. Site was level, and we used the 30 amp and sewer. Limited TV channels over the air; got acceptable 3G with AT&T. Place had a seedy feel. Several permanent residents and only a couple of us overnighters. Standing marshy water in the grass (this is the low country) but also puddles in the open walkway to bathrooms. Train at 9 pm vibrated the RV like magic fingers but didn't bother us. Would not stay here again. - NC Writer
147 Wayside Ln. Canadys, South Carolina 29488 (map) | (843) 538-8206
Sites: 25 | Latest Rate: $20.00 | 13 Reviews
|Clean Restrooms:
|Clean Showers:
Jul 2016 - $20.00
This quiet state park is just a few miles from I-95 and a good place to stay overnight. Restrooms/showers are dated but clean. The interior road is paved, but sites are not and are very sandy. There are lots of mature trees and plenty of shade. We received several OTA TV stations and the free WIFI worked. Our water/electric hookups were OK. We called in to reserve for one night, then when we arrived the site was not level enough for our unit. Phoned the office from the site and they happily changed us. We liked it here and found it to be good value. We would stay again. - R&S
Apr 2016 - $20.00
$20 per night was for a "little tippy" packed sand pull-though site with 30-amp electric and water. Our site, #17, was shaded by large trees. The site had a wooden picnic table and fire ring. It was pleasantly quiet during our stay. The small facility, at only 35 acres, is well maintained and very attractive. The campground host was very friendly and helpful. There are many activities available, which you can find listed on their website. The campground is near I95 but we heard no traffic noise at our site. AT&T and Verizon cell service was acceptable. There is free Wi-Fi available at the office. We received 17 digital over-the-air TV stations and many radio stations. - DD Rolling Ranch
Nov 2015 - $15.00
This is a very nice state park with electric and water hookups. A dump site is available. The heavy rains this past fall caused some damage to the gravel base of the sites. We were in Site 14 which was wide and long enough for our 40' Class A and tow, but the drop from the paved road into the site was deep and rutted. Other sites are in better shape. Repairs are on-going. Park is very clean and well managed. Camp host on site. Note: if you make reservations online, there is a two night minimum purchase. If you call the park, a ranger can reserve a site for one night. - DiplomatTraveler
11592 Jefferies Hwy Walterboro, South Carolina 29488 (map) | (843) 538-8731
Sites: 33 | Latest Rate: $27.00 | 1 Reviews
|Clean Restrooms:
|Clean Showers:
Nov 2016 - $27.00
Just an overnight stay. Easy in and out. Electric and water was good.
Approx, 4 - 5 miles off I-95. Lots of permanent campers, but all was quite in the evening. - JJ & CC
11592 Jeffries Hwy Canadys, South Carolina 29433 (map) | (843) 538-8206
Sites: 20 | Latest Rate: $27.00 | 15 Reviews
|Clean Restrooms:
|Clean Showers:
Sep 2015 - $27.00
Good CG for an overnight, several options for entering. This is essentially a house owner who took extra land and made an rv park. Electricity is so-so. There is a new owner and the name has changed to Eagles(town has also changed to Walterboro) and the rate has gone up to $30. The office is not clearly marked, walked up to house(run down) and "office" not open. No notes as to what to do.
The host walked up and gave us a site. No one was expecting us, they had messed up our reservation by a day, luckily there was space.
OK for an overnight but no longer. - Wandering
Dec 2012 - $15.00
This is a wonderful park, but if you require restrooms, laundry, swimming pool, etc. it is not the park for you, as there are none. The park is clean and spots are clean underfoot. There is nothing extra but the spots. The Wi-Fi here is good, a little slow, but stable and adequate. Management is a wonderfully nice woman and really goes out of her way to be helpful. There is plenty of shade. Road noise could be a problem if you are close to the highway. Further back from the highway, the noise is mitigated well by the trees and structures. Voltage was only slightly low, but never became a problem. In the two weeks we were there, only one morning was low voltage a problem. That was the only day the park was full, and in the morning, voltage dipped a little. But the rest of the time we were there we had no problems with low voltage. We would definitely stay at this park again. Price reflects a daily rate for our two week stay. - John&Sandy
Jul 2012 - $17.00
The other reviewer (July 2012) wrote an accurate review of this campground. It's only 3 miles The other reviewer (July 2012) wrote an accurate review of this campground. It's only 3 miles from I95 exit 68. It's easy in and easy out. It is well maintained and a great spot to get off the road for a night or two. The price is fantastic. We've stayed at New Green Acres 10 miles south on I95 and it was double the price and not near as nice. The comment about the lady owner being very nice and helpful was right on the money. An older gentleman who was working on a tractor when we got here got me parked and was very friendly. I suspect most travelers don't know about this place, try it you'll like it - floridaguy
Cross, South Carolina (map) |
Sites: 20 | 0 Reviews
2060 Ranger Dr Cross, South Carolina 29436 (map) | (843) 753-2818
Sites: 62 | Latest Rate: $38.00 | 1 Reviews
|Clean Restrooms:
|Clean Showers:
Mar 2012 - $38.00
We are full-time RV'ers and travel for work. The main reason for this rating is we arrived around 8:30 pm, after dark, to find the entrance impossible to see due to lack of lights and the addition of crushed asphalt. This campground is in the country and turning around is not an option. Once we got in the campground, we found no maps or instructions for late arrivals. The phone numbers posted on the door were useless as there was no cell coverage. No campground personnel were to be found. We stumbled around the park in the car to try and find a site we could fit in, not easy again due to the black crushed asphalt and poor lighting. When we checked in the next morning, the staff was rather gruff and unhelpful. Did not use the bath facilities, but they appear to be new. It is a quiet and peaceful setting with docks that you can fish from or enjoy the water. The crushed asphalt tracks terribly and they have a resident Canadian Goose who has injured wings that will drive you a little nuts. Would not stay here again. - full-time jane
2020 Ranger Dr Cross, South Carolina 29436 (map) | (843) 753-2818
Sites: 50 | Latest Rate: $25.00 | 1 Reviews
|Clean Restrooms:
|Clean Showers:
Sep 2006 - $25.00
Outstanding park with three boat landings, several docks for fishing, lots of trees, family oriented area great for fishing and relaxing. Staff and personnel at the site are always willing to help. Highly recommended for a stay on Lake Moultrie. Cable, phone, modem etc. available for more permanent stays only. There are no bath facilities or designated swimming areas. All campers in the site are larger with their own facilities. - luvour5er


text:
From bug 499654 comment 8. There's code in http://mxr.mozilla.org/mozilla-central/source/browser/base/content/sanitize.js#171 with no UI for clearing offline cache and DOM storage data:
We should either kill the code or add back the UI.
(BTW bug 436248 claims the code is also broken)
other issues with the code that could be re-enabled here are in dependencies.
Another thing to remember is that currently Clear Recent History -> Cookies clears the DOM storage (although not always - bug 527667). The currently disabled code also clears DOM storage, so if it's enabled as is, it will be confusing. On the other hand, if it's changed, the logic in the Preferences window (Options -> Advanced -> Network -> Offline data -> Remove) should be tweaked too.
Created attachment 572929 [details] [diff] [review]
Comment on attachment 572929 [details] [diff] [review]
Fine by me.
(In reply to Nickolay_Ponomarev from comment #0)
> We should either kill the code or add back the UI.
> (BTW bug 436248 claims the code is also broken)
Are there any automated tests for this functionality?
Comment on attachment 572929 [details] [diff] [review]
Needs also browser peer review, Ehsan, feel free to forward to anyone else.
I will write test for the app cache/user data clearing.
Looks like this gets closer related to bug 538588. Making the code I want to test here shared between modules and also the test seems quit reasonable now. See bug 397719 as mentioned in bug 538588 comment 13.
However, bug 397719 seems to be quit old and this bug and bug 538588 are Q4 goals that need to get in ASAP. So I would rather go a followup way for sharing.
This should land after bug 538588. We also want to add the removal of offline privileges to the shared module (to fix bug 383014). The clear recent history dialog is not doing it right now.
Created attachment 578614 [details] [diff] [review]
v1.1 + test
Includes test. Deps on patch from bug 538588.
Comment on attachment 578614 [details] [diff] [review]
v1.1 + test
>diff --git a/browser/base/content/offlineCache.jsm b/browser/base/content/offlineCache.jsm
> let OfflineCacheHelper = {
> clear: function() {
>+ // Remove all privileges for offline-apps
Why are we including this here? For the sanitize dialog, this is handled separately by the "site specific settings" option (siteSettings), so I don't think having this option also clear permissions makes sense.
Created attachment 578850 [details] [diff] [review]
v1.2 + test
Aha, wasn't aware. I wanted to be consistent with the Advanced/Clear button, but there is now a discussion to change its behavior too.
Comment on attachment 578850 [details] [diff] [review]
v1.2 + test
>diff --git a/browser/base/content/sanitizeDialog.css b/browser/base/content/sanitizeDialog.css
>+#itemList {
>+ padding-bottom: 0.25em;
Why is this needed? Assuming it is needed, this should probably live in the platform-specific theme CSS file rather than this content CSS file.
>diff --git a/browser/base/content/test/browser_sanitizeDialog.js b/browser/base/content/test/browser_sanitizeDialog.js
>+ function () {
>+ // Give www.example.com privileges to store offline data
>+ var pm = Cc["@mozilla.org/permissionmanager;1"]
>+ .getService(Ci.nsIPermissionManager);
>+ pm.add(URI, "offline-app", Ci.nsIPermissionManager.ALLOW_ACTION);
>+ pm.add(URI, "offline-app", Ci.nsIOfflineCacheUpdateService.ALLOW_NO_WARN);
This should probably go in the next test, where it actually gets cleared? Or is it needed for setItem/openCacheEntry to work?
(In reply to Gavin Sharp (use firstname.lastname@example.com for email) from comment #10)
> >+#itemList {
> >+ padding-bottom: 0.25em;
> >+}
> Why is this needed? Assuming it is needed, this should probably live in the
> platform-specific theme CSS file rather than this content CSS file.
I can see a scroll bar that disappears after clicking one of the items. This avoids that scroll bar to appear and has actually no affect on the dialog's layout.
I'll try to find the proper CSS and move it there. This is AFAIR specific to Windows.
> >diff --git a/browser/base/content/test/browser_sanitizeDialog.js b/browser/base/content/test/browser_sanitizeDialog.js
> >+ function () {
> >+ // Give www.example.com privileges to store offline data
> >+ var pm = Cc["@mozilla.org/permissionmanager;1"]
> >+ .getService(Ci.nsIPermissionManager);
> >+ pm.add(URI, "offline-app", Ci.nsIPermissionManager.ALLOW_ACTION);
> >+ pm.add(URI, "offline-app", Ci.nsIOfflineCacheUpdateService.ALLOW_NO_WARN);
> This should probably go in the next test, where it actually gets cleared? Or
> is it needed for setItem/openCacheEntry to work?
It's obviously needed for the first test.
Created attachment 578858 [details] [diff] [review]
v1.3 + test
only thing changed: removed the bottom padding, it seems like the scroll bar is not any more appearing with the newly added item
Adding dev-doc-needed, since https://developer.mozilla.org/en/Offline_resources_in_Firefox explicitly calls out this bug as being an existing issue.
(Once Firefox 11 is released, I assume we'll want to update that MDN article.)
Doc updated:
(the one mentioned in comment 42 has been moved to that location)


text:
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text:
Regulations that prohibit the disposal of mercury-added products in any manner other than recycling, disposing as hazardous waste or alternative method approved by MassDEP.
310 CMR 76.00: Mercury-Added Product Disposal Ban
These regulations were last amended in September 2009.
310 CMR 76.00: Proposed Amendments & Public Comment
- There are no proposed regulations out for public comment at this time.
310 CMR 76.00: Recently Promulgated Amendments
- There have been no recent amendments to these regulations.
Mercury-Added Lamps
- Fluorescent & Mercury-Added Light Bulbs
Guidance for homeowners & businesses, including proper management, broken bulb cleanup and recycling drop-off locations.
- Changes in Mercury-Added Lamp Manufacturer Requirements
Please Note: The mercury lamp provisions of the Massachusetts Mercury Management Act were amended in July 2014. As a result, mercury lamp manufacturers are no longer required to submit annual certifications to MassDEP. Follow this link for additional information.
- Product Labeling Plan
Don't see what you're looking for? View all Mercury-Added Product Management Reporting Forms categories.
[ Permitting Assistance ] • [ Fees and Payments ] • [ Transmittal Number and Form ]


text:
Q: Do I qualify?
A: Our table (below) lists which accounts and customers qualify, and which do not. The key was to be a customer by the end of last year.
Q: How much will the windfall actually be worth?
A: The pounds 800 figure is only an average, and an estimated one at that. It comes from dividing the Alliance & Leicester's possible stock market value - pounds 2.5bn - by the number of qualifying customers - 2.5 million savers and 500,000 mortgage borrowers. If you are both a saver and borrower you stand to get two lots of free shares.
The minimum handout for anyone who qualifies is likely to be for shares worth at least pounds 500. The other pounds 300 a head might be doled out according to savings (or mortgage) balance, or according to how long people have been customers. But the possibility of such a variable bonus is only a guess. The Alliance & Leicester has not yet said whether the share handout will be at a flat rate, with everyone getting the same amount, or whether there will be a basic and account-related entitlement.
In addition, because the windfall is in the form of shares, there is no fixed value. As with all shares, their value will depend on what price the stock market puts on them when they start trading. And once on the stock market, dividends will probably be paid and the shares will rise or fall like other shares. Anyone who hung on to their free Abbey National shares, for example, has seen the value of their windfall enhanced by the shares' strong outperformance of the stock market since 1989.
Q: When will I get the shares?
A: Early next year is the plan, when the society joins the Stock Market as a medium-sized bank in the FT-SE 100 index of blue chip shares.
Q: Can I touch my savings?
A: Difficult to say - the society is not being very helpful on this one. It will not say to what degree savers might reduce their entitlement if they take money out of their accounts now. Until the society announces the details of any bonus share distribution and any further qualifying dates for that bonus - and the details of the scheme will not be released until later this year - savers are in the dark. Cynics might say the society is delaying coming clean so that it can keep savers locked into poor interest rates.
What the society has said so far is simply that to qualify, your total savings balance, 56 days before the voting date, must be more than pounds 100, or your mortgage debt must have been pounds 100 last 31 December and still be so on the voting date.
Q: What about "carpetbaggers"?
A: When the Woolwich announced its switch to bank status last month, its chief executive, Peter Robinson, almost seemed to relish excluding people who had opened accounts this year from the windfall, dubbing them "carpetbaggers".
Alliance & Leicester has also excluded people who opened accounts this year. It claims the number of people who might have hoped for windfalls but have been excluded is "minimal", but this may still run into thousands. In December the society stemmed the speculative tide by closing its instant access account and requiring new savers to put in pounds 5,000 (pounds 1,000 for a Tessa) to open an account. It followed this, after the Woolwich windfall announcement last month, by barring the opening of new accounts carrying "share" status - those that carry voting and membership rights and therefore are most likely to qualify for windfalls.
Q: How does this deal compare with others?
A: It's quicker, inasmuch as you should get your shares sooner than those from the Halifax and Woolwich building societies - the Halifax is pencilled in for summer 1997 and the Woolwich for autumn 1997.
The Alliance & Leicester deal is not as lucrative as when the Cheltenham & Gloucester building society was taken over by Lloyds Bank - when savers were paid an average of pounds 2,200 in cash.
Since we do not yet know the details of the share allocation, it is difficult to make a full comparison with the other share deals - but on average they are all likely to be worth pounds 800 or so. For windfall watchers, shares worth at least pounds 500 would make it a fairly standard deal. And for those who have qualified with balances of just pounds 100, that is a fabulous investment return.
However, the Alliance & Leicester deal excludes a greater proportion of its customers - 2 million out of 5 million - than any other of the big deals announced to date. This is because it owns Girobank, whose customers are not members of the society and do not qualify. Girobank operates through post offices and issues giro cheques to millions of social security claimants.
Q: What happens next?
A: Letters went out last week from the Alliance & Leicester to customers explaining its proposed changes and enclosing a leaflet called Alliance & Leicester conversion: some questions answered. There is also a helpline on 0345 221144. You do not have to do anything for now, although it might be advisable for savers to ensure their savings total pounds 100 with the society.
The Alliance & Leicester expects to publish details of the share distribution later this year. This will be sent to members, along with an invitation to vote on the proposals. The vote is planned for later this year, to be followed by the share handout early next year.
Q: Who is next?
A: Some analysts think this may be the last windfall announcement for some time. Five of the biggest building societies have now declared in recent times - Halifax and Leeds (which merged), Woolwich, Alliance & Leicester, National & Provincial, and Cheltenham & Gloucester. The Nationwide is the only remaining society that analysts consider big enough to become a bank on its own. But it seems more committed to remaining a building society and will announce rate improvements for savers and borrowers in the spring as part of a loyalty bonus.
A number of societies remain takeover targets, however. The A&L might even take over a society before its own share handout, allowing customers of the swallowed society effectively to join in the A&L share deal.
One potential takeover target, Bristol & West, said last week it continued to review its position and does not rule out coming up with a windfall deal, while the Woolwich is reported to be interested in buying the Britannia. In each case the savers and borrowers of any society that was bought would stand to receive a windfall.
Looking forward over the next few years it is almost certain there will be more rationalisation among societies.
The only real question is which will go. So instead of trying to spot the next, savers with spare cash should simply try to open accounts with as many of the remaining societies as possible. This, however, is becoming more difficult, as societies try to deter speculators by raising minimum opening balances and closing qualifying accounts.
Who gets the shares?
2.5 million savers with the following accounts open on 31 December 1995:
Bonus 180; Bonus 90; Bonus Share; Capital Choice; CashPlus; Extra Interest Share; Fixed Rate Bond (1 and 2 Year); Gold; Gold Plus; Gross Interest; Hambros; High Income Term Share; High Yield Term Share; Instant Access (Card); Instant Access (Passbook); Investment Bond; Investment Plus; Keysaver; Leicestercard Gold; Leicestercard 3 Year Bond; Midas; MoneyMonthly Term Share; MoneyBuilder Plus; MoneyReady; Monthly Income Bond; Monthly Income Investment Bond (3 Year); Monthly Income Option Bond (3/5 Year); Ninety Day; Option Bond (3/5 Year); Premier; Premium Plus; Prime Plus; ReadyMoney Plus; Regular Savings; Save-As-You-Earn (SAYE); Seven/7 Day; Share; Sharesave; Special Edition; Special Return; Step-Up Bond; Step-Up Bond Issue 2; Stock Market Bond; Tempus; TESSA.
To qualify, savers must be the first-named accountholder and must have at least pounds 100 in total in any number of accounts at the qualifying date, 56 days before the voting date (probably late this year, but yet to be finalised).
500,000 mortgage borrowers (those who are savers as well will get two sets of shares) with at least pounds 100 of mortgage debt owing on 31 December 1995 and on the voting date.
The plan is that the entitlement of people who die before getting their shares will pass to their beneficiaries. Alliance & Leicester employees and pensioners will also qualify.
Who doesn't get the shares?
2 million holders of other accounts and financial products including: Giro Current; Keyway; Alliance; Flexiplan Plus; Flexiplan; Tessa Deposit; Prime 90 Deposit; Prime Deposit; Offshore accounts (90 Day, Instant Access, Island); Visa credit cards; unsecured personal loans.
Also excluded are the thousands of people who opened accounts or took out mortgages this year; people who open accounts from now; those under the age of 18 at the voting date who hold accounts directly; and second- named accountholders.
- More about:
- Alliance And Leicester
- Bonds
- Bonds And Funds
- Mortgages


text:
The people who run Harrods are happy this year. Punters are rolling in through the doors, tills are bleeping and spending at the checkouts is up to the limits one would expect for so famous a store. A Harrods spokesperson says: "We are out of the recession and people feel confident."
But for the rest of us, whose spending is likely to take place in far more prosaic surroundings, the festivities can lead to liquidity problems, and not just the hangover type.
Credit cards are one way of easing cashflow worries. Most of us have one and, with more than 21 million Visa cards in the UK, the indications are that our plastic friend will bear the brunt of the Christmas bills.
Royal Bank of Scotland has researched consumer behaviour over the Christmas period. At RBS, Jayne Goodwin thinks we have learned to use credit cards as a temporary source of borrowing. "We see a lot of card spending in the four weeks leading to Christmas. Last year this totalled pounds 125m, with a 30 per cent increase expected this year. In January, personal overdrafts and loans take over as customers refinance their borrowing."
The basic principle of card spending is simple enough. Most offer up to 57 days' interest-free credit on spending up to their limit, usually set at a minimum balance of pounds 500.
But there are some snags, quite apart from whether you can afford to clear the balance. Securing the maximum period of interest- free credit depends on spending at just the right time. This in turn depends on when your monthly card statement is prepared. The art lies in shopping just after this date. The amount will be added to your next statement, with payment due 21 days later.
So if your statements are prepared on 10 December, you will have to pay by the end of January. If your statements are dated 20 December, you can delay paying until near the middle of February. Check your last statement for the date it was prepared and shop accordingly.
But if you can't clear the whole balance by the due date, most cards from high street banks offer no interest-free credit. Instead, they charge daily on the whole amount of the balance. Late payment usually costs pounds 10 to pounds 12, with NatWest top at pounds 20.
The exact basis on which interest is charged may also vary. Some cards, like Bradford & Bingley's, charge from date of purchase. Others, like TSB's, from the date at which a purchase is charged to your account. A gap of two days between spending and when the charge is added is average. If you use the card just once each month and pay it off in full, then charging from date of purchase will add 24 days of interest to the annual cost of your card.
In exchange for notional periods of free credit, you do pay more expensive annual percentage rates (APRs), mostly between 18 and 21 per cent. Worse still, lenders add between 1 and 2 per cent to this on credit balances arising from cash withdrawals. Gold cards, open to those earning pounds 20,000 or more, charge on average 1 per cent less than standard versions. There is often a cost attached to having a card in the first place, but if you have one already, this won't affect your spending either way.
Many cards offer extra "free" benefits; loyalty points, travel insurance and purchase protection. For instance, Alliance & Leicester offers cover against loss, theft or accidental damage for 100 days on purchase values between pounds 50 and pounds 5,000. Cover is all risk, but includes a clause stating "unless the item is otherwise insured", in which case you can't claim.
Loyalty points are also on offer from card providers. Spend pounds 250 with Sainsbury's Reward Card and you accumulate points enough for a store voucher worth pounds 2.50.
Using credit cards allows the "double-dipping" option, where spending entitles you to loyalty points, cash off your gas bills, Air Miles and a plethora of special offers.
Store cards are more expensive. Aside from John Lewis, charging 18 per cent APR, most levy rates between 29 and 30 per cent. Timecard, valid at Comet, Woolworths and B&Q, comes top with a rate of 31.90 per cent. There are alternatives. New bank cards such as Royal Bank of Scotland's Advanta, are now being launched with discounted rates. Advanta's APR is just 9.9 per cent, but only until July next year, when it goes up to 18.55 per cent, giving you six months to pay for Christmas presents at an attractive rate.
Elsewhere, the Co-operative Bank offers its Advantage Visa card with a discounted APR of 7.9 per cent until April, when rates go up to 10.9 per cent. This is cheap borrowing, but there is no interest-free period.
According to the Co-op's Dave Smith: "We've segmented the market and aim for those who want to use a card as a continuous source of credit." The card is open to non-account holders at the Co-op.
Personal overdrafts charge equivalent annual rates (EARs) on outstanding balances. The best deal comes from Alliance & Leicester at 9.5 per cent, but most cost 15 per cent or more, with Clydesdale and RBS coming top at 19.56 per cent. Monthly arrangement fees of around pounds 10 will usually be added. To get the best deal you have to switch banks, and it may be too late in the day to do it for this Christmas.
Unsecured loans are another option, but care is needed. As a general rule, the shorter the term and the smaller the amount borrowed, the higher the APR. This will be fixed at outset, with monthly repayments combining interest and capital.
If early redemption of the loan is planned, expect to be penalised with an excess charge of two months' interest.
On loans of pounds 500 over six months, expect to pay not less than 16.9 per cent from Midland and as much as 25.8 per cent from Yorkshire Bank.
Some lenders also offer flexible loans with no minimum repayment period and lump sum repayment facilities.
Clydesdale Bank is competitive with an EAR of 19.2 per cent on loans between pounds 500 and pounds 5,000, repayments on a weekly, fortnightly or monthly basis and no early redemption penalty. Drawbacks include a pounds 35 arrangement fee and minimum pounds 100 drawdown, but if you want the early redemption option, this is good value.
If in doubt about whether you can afford Christmas, apply the Scrooge test: which is worse, being in debt, or not spending at all? And remember, he only had to deal with the Ghost of Christmas Past.
Best borrowing deals - a Christmas selection
Co-Operative Bank Advantage Visa
offers 7.9% APR until 31 March 1998, then 10.9%. Phone 0800 109000
RBS Advanta
offers 9.90% APR until 1 July 1998, then 17.9%. Phone 0800 077770
Loans (pounds 500 over six months)
Midland Bank (unsecured) Clydesdale Bank (flexible)
19.9% APR. Phone 0800 180180 19.2% EAR. Phone 0800 240024Reuse content


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Do You Have Closet Fear?
How often do you look at some of the clothes in your closet and wonder why they are there? Maybe, you just don't feel like they represent the real... Read More
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How often do you look at some of the clothes in your closet and wonder why they are there? Maybe, you just don't feel like they represent the real... Read More
Boost Your Confidence by Embracing Your Personal Style We all need to update our look from time to time because wearing the same hairstyle, wearing the same clothes and... Read More
Do you take way more than you need on a business trip? The plain truth is that we all do it at some time or another. Why? Is it... Read More
Everyone once in a while, I want to share with you a man Caught in Good Style so you can share with your spouse, significant other or friends. Hey,... Read More
I Caught another very talented and successful business women in Good Style this week at the local National Speakers Association Meeting. Mary Kausch is the founder of HRetc!! and... Read More
The more I know this woman, the more I am amazed at her accomplishments and as you can see from this photo, she is truly rocking her personal style.... Read More
This woman knows how to rock her look. Lacha Hughes, owner of This is It Home Care, was Caught in Good Style at the Sisters of Hope conference in... Read More
I was recently invited to a semi-formal dinner and it got me thinking about what that really means. Since it is the season for all kinds of parties, I... Read More
Last night I attended one of the most fun fundraisers ever. The Jane Doe Advocacy Center hosted a murder mystery mock trial dinner at The Bevo Mill. What a... Read More
Last week I had the opportunity to speak at the Prosper on Purpose conference hosted by Rainie Howard. It was a great day filled with new connections and amazing... Read More


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17 Charlemont Street East Ranfurly Otago
1 Bute Street Ranfurly Otago 9332
11 Pery Street Ranfurly Otago 9332
26 Charlemont Street East Ranfurly Otago
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AQuality Distribution
Tropical Fish, Aquariums
L-No catfish fom our own Tanks
Specialist importers, Breeders , associate breeders or well-selected Wildcaught specimens.
We quarantine for at least 1 month
Contact us with your request!
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BRIGHT, Vibrant, Healthy imports of exciting Malawi Cichlids for the keen collector, breeder, or hobbyist.
The best Alround Source of Discus. Tankbred, Wild, Hybrid AND Strain Discus from premium collectors, traders, and Breeders. Sourced Globally.
Essential Foundations Begins with BEST source ALL_CURED. DOSED SEEDED BIODIVERSITY ALL under ONE ROOF Caribbean, Atlantic, Pacific, Indian, Indo-Pacific
Fish health and quality comes FIRST! We tailor-make imports of these and other items. Specialist fishkeeper or trade professional, amateur or hobbyist, spa owner or personal user ? We can advise and supply these and other speciea with best , longterm success. We KNOW how to KEEP them alive, and make them THRIVE. Not just suppliers for business purposes.
Livestock and equipment.-Direct source Aquarium problems?
SOLVED! Science and long-term experience based Guaranteed results!
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