text:
Toll-like receptor 3 upregulation in macrophages participates in the initiation and maintenance of pristane-induced arthritis in rats
- Liesu Meng†1, 2,
- Wenhua Zhu†1, 2,
- Congshan Jiang1, 2,
- Xiaojing He1, 2,
- Weikun Hou1, 2,
- Fang Zheng1, 2,
- Rikard Holmdahl3 and
- Shemin Lu1, 2Email author
DOI: 10.1186/ar3034
© Meng et al.; licensee BioMed Central Ltd. 2010
Received: 05 January 2010
Accepted: 25 May 2010
Published: 25 May 2010
Toll-like receptors (TLRs) are involved in both innate and adaptive immune responses and are likely to play a complex role in the pathogenesis of human rheumatoid arthritis (RA) and experimental arthritis. The objective of this study was to identify the key TLR in pristane-induced arthritis (PIA), a rat model for RA, and to clarify its roles in the initiation and maintenance of arthritis.
Arthritis in DA rats was induced by pristane and the severity was evaluated by macroscopic and microscopic score systems. Spleen TLR and cytokine expression was detected at different time points by real-time polymerase chain reaction (PCR) and flow cytometry. Polyinosine-polycytidylic acid (polyI:C, a ligand of TLR3) or TLR3 specific short-hairpin RNA plasmid for RNA interference was administrated to PIA rats in vivo. Serum nitrogen oxide concentration was determined by Griess method, and tumor necrosis factor alpha (TNF-α) was determined by L929 biotest. In splenic macrophages, TLR3 expression was measured by flow cytometry. A rat macrophage cell line (NR8383) was stimulated by pristane, and anti-TLR3 antibody were used to block TLR3 pathway. TLR3 and cytokine expression in NR8383 were detected by real-time PCR.
By screening the TLR expression profile in spleen of DA rats after pristane injection, we found that TLR3 was the most early and prominently upregulated TLR. Both TLR3 mRNA and protein expression of spleen were upregulated at 6 and 26 days after pristane injection. Furthermore, administration of polyI:C exacerbated, whereas RNA interference targeting TLR3 ameliorated, the arthritis. Particularly, TLR3 expression was induced in splenic macrophages of PIA rats, and also in the NR8383 cell line after pristane stimulation in a dose- and time- dependent manner. Upregulation of interferon beta (IFN-β) and TNF-α by pristane stimulation was blocked by anti-TLR3 antibody in NR8383.
TLR3 plays a pivotal role in the initiation and development of PIA which may dependent on macrophage. These findings are useful to understand the pathogenesis of RA and may provide an intriguing therapeutic opportunity for RA.
Rheumatoid arthritis (RA) is an autoimmune chronic inflammatory syndrome affecting 0.5 to 1% of the world population, and is characterized by cellular proliferation in the synovial lining and cartilage and bone destruction of diarthrodial joints [1]. Genetic and serologic evidence in both RA and experimental arthritis favors the involvement of both innate and adaptive autoimmune processes [2, 3]. Toll-like receptors (TLRs) are pattern recognition receptors, which form a bridge between innate and adaptive immune systems, and have been considered to be an important factor in the development of RA [4].
TLRs are involved in activation of antigen-presenting cells (APCs) by influencing the uptake and processing of various exogenous and endogenous antigens [5]. Activation of TLRs in APCs not only leads to the upregulation of the costimulatory molecule expression and cytokine secretion [6, 7], but also promotes the T cell polarization [8, 9]. In addition, TLRs could orchestrate the function of regulatory T cells [10, 11].
TLRs are likely to play a complex role in RA, and certain TLRs exhibit a high expression, such as TLR2, 3, 4, 7 in synovium [12–15], TLR3 in fibroblast-like synoviocytes (FLS) [14], TLR2, 4 in CD14+ macrophages and peripheral blood cells from RA [16]. Both exogenous and endogenous TLR ligands have been detected in synovia, synovial fluids and sera of RA patients [14, 17, 18]. Importantly, these ligands are capable of stimulating FLS and/or immunocytes via triggering TLRs to produce proinflammatory cytokines [12, 19–24], and may also activate the autoreactive T and B cells [25–27]. In particular, injection of TLR2 and TLR9 ligands, peptidoglycan (PGN) and CpG DNA, into articular cavities induces arthritis in mice [28, 29].
Based on the above-mentioned reasons, it seems that TLRs play essential roles in the pathogenesis of RA. However, most studies on RA are descriptive and focus on TLR2 and 4 in synovium. Thus, a systemic study about TLR roles in the initiation of immune response of RA will provide new insight for elucidating the pathogenesis of RA. The TLR genes are highly conserved and using animal models should be reasonable to address the issue of TLR roles in RA development.
Various animal models represent different or partly different disease processes of RA, and a few studies have concerned TLR roles in the pathogenesis of arthritis. TLR2 and 4 were reported to be involved in the chronicity and erosive destruction of streptococcal cell wall induced arthritis and spontaneous arthritis in IL-1 receptor antagonist-knockout mice [8, 30, 31]. TLR4 takes part in the initiation and progression of both autoantibody/lipopolysaccharide (LPS)-triggered arthritis and serum transferred arthritis [32, 33]. It has also been suggested that cathepsin K could ameliorate mycobacteria induced arthritis in rats through suppression of TLR9 signaling in dendritic cells (DCs) [34]. Of particular interest is the pristane-induced arthritis (PIA) model as it is induced with a single subcutaneous injection with a well-defined compound lacking antigenic properties. The induced disease is joint specific and fulfills the clinical criteria of RA [35, 36]. Importantly, the arthritogenic adjuvant stimuli may mimic such a trigger in humans represented by an infectious agent or a non-specific inflammatory response. It is known that pristane induces a strong innate immunity through the triggering of IL-6 production [37], suggesting that TLRs may participate in and mediate the immune response. Altogether, the mechanism of TLRs involved in experimental arthritis is not understood completely.
In the present study, we tried to find the key TLR in arthritis development by screening the TLR1-9 expression profile in spleen from PIA rats. The results indicated TLR3 showed a unique expression pattern compared with other TLRs in spleen of PIA rats. And in PIA rats treated with polyinosine-polycytidylic acid (polyI:C) or RNA interference (RNAi) of TLR3, arthritis manifestations could be modified. Furthermore, TLR3 may exert its influence in a macrophage-dependent manner. These data illustrated the crucial role of TLR3 in the initiation and development of PIA.
Materials and methods
DA rats were bred in a specific pathogen-free animal house. Each group contained 8 to 10 rats at the age of 8 to 12 weeks. The experiments were approved by the Institutional Animal Ethics Committee of the University (permission No. 2006-21).
Induction and evaluation of arthritis
Arthritis was induced by a single subcutaneous injection with 300 μl of pristane (ACROS Organics, Morris Plains, NJ, USA) at the base of the rat's tail. Arthritis development and severity were monitored every two to four days by the perimeters of ankle and mid-paw, and a macroscopic scoring system as described [35].
Histopathological examination of joints
Left ankle joints of rats were sectioned and stained with hematoxylin and eosin. Pathological severities were estimated by extent of 10 items: a. thickness of synovium lining layer; b. pannus; c. synovium inflammatory cells; d. angiogenesis; e. cartilage erosion; f. bone erosion; g. joint ankylosis; h. change of overall articular structure; i. new bone formation; and j. new cartilage formation. Each pathological item was scored on a scale consisting of score 0 (normal) to 3 (most severe). The maximum histopathological scores of arthritis are 30 for each ankle. Synovitis was estimated by addition of item a. to d. scores; joint destruction by e. to h. scores; joint healing by i. and j. scores.
RNA quantitation
Information of primers for conventional PCR and Real-time PCR
Gene name
NCBI Accession No.
Size (bp)
annealing temperature (°C)
Fluorescence-activated cell sorting (FACS) analysis of TLR expression in splenocytes
Three tubes containing 106 splenocytes for cell surface TLR detection were incubated with TLR2, TLR3 and TLR4 antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA) respectively. In the other tube for intracellular TLR3, the cells were fixed and perforated before TLR3 antibody incubation. For detection of macrophage TLR3, two tubes containing 106 splenocytes were incubated with HIS36-PE (anti-rat macrophage, eBioscience, San Diego, CA, USA) firstly. And for cell surface TLR3 detection, the cells were incubated with TLR3 antibody at the same time. For intracellular TLR3 detection, the cells in the other tube after His36-PE staining, were fixed and perforated before TLR3 antibody incubation. The cells from both tubes were incubated with goat anti-rabbit IgG-biotin followed by streptavidin-cychrome. Cells were analyzed by using a FACScanto (Becton Dickinson, Franklin Lakes, NJ, USA), and datum analysis was performed using FlowJo software (Tree Star, Ashland, OR, USA).
Administration of TLR ligands to PIA rats
Thirty-six rats were randomly divided into four groups. The control group was injected subcutaneously with 300 μl phosphate buffered saline (PBS) only. Other groups were injected subcutaneously with 300 μl pristane, then administered subcutaneously with 150 μl saline containing 100 μg polyI:C (Amersham Biosciences, Piscataway, NJ, USA) in the polyI:C group, 50 μg LPS (Sigma-Aldrich, St. Louis, MO, USA) in the LPS group, and none in the saline group, respectively, at two days after pristane injection [38, 39].
Cell transfection and treament with TLR3 RNAi in vivo
The target sequences of TLR3 gene are GAA TCA CAT CTC GAA GAT A, called as shRNA1-TLR3; CTC TCA ATT TAA CGA AGA A, as shRNA2-TLR3; ACC TCG ACC TCA CAG AGA A, as shRNA3-TLR3; and CCT GAC AGA GCT CCA TCT A as shRNA4-TLR3. The sequence of negative control shRNA is TTC TCC GAA CGT GTC ACG T. These different short-hairpin RNA (shRNA) sequences were inserted into pGCsilencer™ U6/Neo/GFP/RNAi plasmid (Genechem, Shanghai, China). The plasmids were extracted with E.Z.N.A.™ Endo-free Plasmid Maxi Kit (Omega Bio-tek, Doraville, GA, USA), and NR8383 cells were transfected with Lipofectamin™ 2000 (Invitrogen). TLR3 expression of cells was detected by RT-PCR after 24 hours and by FACS after 48 hours. DA rats randomly divided into three groups were injected subcutaneously with 300 μl pristane. At one day before pristane injection, and 5, 11 and 17 days after pristane injection, 1 ml of saline was injected peritoneally into the rats in the saline group; 50 μg shRNA3-TLR3 in 1 ml of saline in the shRNA-TLR3 group; and 50 μg negative control shRNA in 1 ml of saline in the shRNA-NC group [40, 41]. At 20 days after pristane injection (d20), GFP mRNA expression was detected by RT-PCR to monitor the plasmid presence in spleen (Primer information was depicted in Table 1). And TLR3 mRNA expression in the spleen was detected by real-time quantitative PCR for determination of the RNAi effectiveness.
Measurement of NO concentration
Serum protein was removed, and NO3- was reduced to NO2- by using cadmium filings. Then NO2- was measured by a microplate assay method based on the Griess reaction.
Measurement of TNF-α concentration
The mouse fibroblast cell line, L929, was cultured in RPMI 1640 (Hyclone, Logan, UT, USA) containing 10% FCS. Ten thousand cells in 100 μl were incubated for 24 hours. Then sera containing 1 mg/ml actinomycin D were added to replace the original medium. After an 18-hour incubation, cell viability was evaluated by the methyl thiazolyl tetrazolium method. Rat recombinant TNF-α (Bender MedSystems Inc., Burlingame, CA, USA) was used as the standard.
Stimulation of macrophages with pristane
The rat macrophage cell line, NR8383, was cultured in F-12 K medium (Sigma-Aldrich) with 20% FBS (Hyclone). Emulsion of pristane was made by repeated aspiration with PBS. Half-million cells per well were put in a six-well plate, then a 50 μl pristane emulsion was added in the culture medium. The theoretical pristane concentration is 0.1, 1, 10, 100 μM, 1 mM and 10 mM, calculated on the basis of original volume of pristane employed. The cells were collected after 8, 24, 48 and 72 hours of stimulation.
TLR3 blockade assay in NR8383
NR8383 were seeded in six-well plates at a density of half-million per well. In the TLR3 Blockade assay, cells were treated with anti-TLR3 or isotype control antibodies at a concentration of 20 μg/ml for 90 minutes before being stimulated with 100 μM pristane or 10 μg/ml polyI:C. The cells were collected after 24 hours' stimulation.
Quantitative data were expressed as means ± SEM. The statistical analysis of differences between experimental groups was performed by Mann-Whitney U test or Student's T test. Correlation analysis was performed by Pearson test. P < 0.05 was considered significant.
TLR3 expression of spleen exhibits a remarkable increase in the PIA rat model
Based on the expression at mRNA level, we determined the protein expression of TLR2, 3 and 4 in the spleens of PIA rats by flow cytometric staining. The results showed that the proportion of cell surface TLR3 positive cells increased significantly at d6 and d26, while that of intracellular TLR3 positive cells and cell surface TLR2 and 4 positive cells increased significantly only at d26 (Figure 1d).
The administration of TLR3 ligand increases arthritis severity
Histopathologically, polyI:C administration aggravated the pathologic changes with abundant inflammatory cell infiltrates and bone destruction (Figure 2c). PolyI:C group showed significant differences with the LPS group and the saline group in the pathological score of synovitis, joint destruction, new bone and cartilage formation (Figure 2d). Meanwhile, the total pathological score of polyI:C group had significantly increased compared with other groups (Figure 2d). All results from the clinical, serological and pathological data indicated that TLR3 ligand administration to the PIA rats could make the disease more severe, and further highlighted the important role of TLR3 in PIA development. Then RNAi of TLR3 in vivo was performed to convince the authors that TLR3 is essential in PIA pathogenesis.
The RNAi of TLR3 significantly ameliorates PIA in rats
TLR3 expression was induced in macrophages with pristane stimulation
To confirm the role of pristane on TLR3 expression upregulation in macrophages, we stimulated NR8383, a rat macrophage cell line, with pristane. We found that TLR3 mRNA expression was increased in a dose-dependent manner after pristane stimulation, and reached the peak at 1 mM pristane (Figure 4b). In contrast, TLR4 mRNA expression did not show a dose-dependent tendency. TLR3 expression was induced already at 8 hours after pristane stimulation, and was highest at 24 hours (Figure 4c). IFN-β mRNA expression also showed a pattern similar to TLR3, and increased significantly in the group stimulated at 24 and 48 hours (Figure 4d). TNF-α mRNA expression increased significantly at 24 and 48 hours (Figure 4d). However, as NR8383 were pretreated with anti-TLR3 antibody, the upregulation of IFN-β and TNF-α expression stimulated by both pristane and polyI:C was inhibited significantly (Figure 4e). The result indicated that in macrophages pristane certainly orchestrated high TLR3 mRNA expression, which activated the downstream cytokine pathway.
A single subcutaneous injection of pristane in DA rats leads to chronic relapsing arthritis similar to RA in humans. We now show that pristane injection also leads to an upregulated expression of TLRs, in particular TLR3. TLR3 was found to play a critical role in the development of arthritis as an injection of TLR3 ligand enhanced arthritis and downregulation of TLR3 through treatment with RNAi alleviated arthritis. And TLR3 expression was induced with pristane stimulation particularly in macrophages.
PIA is a T cell dependent disease, which can be transferred with αβT cells [42], and there is no evidence so far arguing for a pathogenic role of B cells. In the human disease it is clear at present that many patients benefit from depletion of B cells through injection of antibodies to CD20. In contrast, in PIA antibodies to citrullinated proteins are not evoked and there is no evidence for pathogenic antibodies mediating the disease, unlike in the collagen-induced arthritis model. However, the immune response in PIA also shows some striking similarities with RA since rheumatoid factors and antibodies to RA33 are produced in both diseases [43]. Importantly, a strong activation of an innate response is apparent in both RA and PIA, although in RA it has been possible to analyze the response only in already established disease. In RA synovial tissues a strong overexpression of TLR3 and 4 has been reported, whereas isolated macrophages show high expression of TLR2 and 4 [13, 16]. But there has been no study on TLRs in the PIA rat model so far.
A subcutaneous injection of mineral oil or phytol, structurally similar to pristane, leads to an accumulation of oil at the injection site but also a rapid spread of small volumes to draining lymph nodes and subsequently spreads systemically, reaching the spleen [44, 45]. Previous studies have shown that T cells isolated from spleens, less than eight days after pristane injection, transfer severe arthritis, showing that the critical arthritogenic change induced by pristane is present in the spleen [42]. Therefore we selected the spleen to study the innate immune reaction after pristane injection.
Screening TLR mRNA expression at different time points after pristane injection, we found that expression of TLR1, 2, 3, 4 and 8 in the spleen is upregulated in the development stage of arthritis (d26), which is consistent with previous studies from arthritis models and RA patients [20, 22, 23, 46]. A compelling TLR is TLR3 as it was highly expressed uniquely in the early phase, and increased progressively with the development of the disease. It has been reported that TLR3 is overexpressed in synovial tissue from both early and longstanding RA [12, 13]. A possible pathogenic role of the upregulated TLR3 could be that it recognizes RNA released from necrotic synovial fluid cells, and then activates RA synovial fibroblast [14]. TLR3 combined with TLR4, TLR7/8 is also involved in the regulation of DC activation and cytokine production in RA patients [12]. It indicates that TLR3 in the spleen may play a vital role in PIA. To examine whether the observed TLR overexpression has functional consequences, downstream cytokines of TLR pathway, such as TNF-α, IL-6, IFN-γ and IFN-β mRNA expression in the spleen were determined. IFN-β expression was found to increase at both d6 and d26, whereas TNF-α and IL-6 expression only increased at d6, and the IFN-γ expression level did not change. These proinflammatory cytokines are known to be regulated by many TLRs via several intracellular pathways but are mainly associated with NF-κB. TLR3 and possibly TLR4 activate interferon regulatory factor 3 which regulates IFN-β expression [47]. By flow cytometric staining we confirmed high TLR3 protein expression in splenocytes. The expression of TLR3 on splenocyte surfaces increased at d6 and d26, reflecting its mRNA expression pattern.
Based on the experimental data from the gene expression profile and TLR2, 3, 4 protein expression during the disease course, we suggest that TLR3 plays an important role in the pathogenesis of PIA. Such a role could be confirmed through in vivo experiments with TLR3 ligands and RNAi. In a previous study, intra-articular treatment with polyI:C to rats caused a rapid and strong synovitis [48], which suggested the important role of TLR3 signaling in arthritis pathogenesis. And in our research, the systemic treatment with polyI:C synergism with pristane showed that TLR3 ligand could exacerbate arthritis. This dominant role of the TLR3 ligand was also proved by pathology of the PIA rat ankles. The possible mechanism of polyI:C's effect on PIA could be that more TLR3 is activated by polyI:C, which triggers more severe inflammatory reaction in joints. Meanwhile, the results also emphasize the important roles of the ligand in arthritis pathogenesis. But at present, it is still not defined which endogenous ligand of TLR3 participates in PIA. Thus, priority should be given to exploring the probable ligand and investigating its roles in arthritis.
A crucial role of TLR3 in the PIA pathogenesis could also be confirmed by treatment with RNAi in vivo. The shRNA plasmids did not show any immunostimulatory effect in vivo because the shRNA-NC group shows no differences from the saline group in all indexes. The results indicated that the RNAi of TLR3 participated in the initiation of PIA significantly delayed the onset day and decreased arthritis severity. NO concentration of shRNA-TLR3 group was also significantly lower than that of the shRNA-NC and saline groups. The result validates the importance of TLR3 in PIA, however, the ligands and the functional consequences of TLR activation in PIA are not yet fully understood.
TLR3 expresses mainly on DC for humans but on macrophages for mice [49]. It has been found that macrophages are involved in the pathology of K/BxN serum-induced arthritis [50]. Macrophage-derived reactive oxygen species (ROS) participate in T cell selection, maturation and differentiation, and also mediate protection against arthritis by suppressing T cell activation [51]. In addition, macrophages stimulated by excessive interleukin-15 may activate the characteristic autoreactive CD4+ T cells in RA [27]. The increased macrophage activation may be mediated by TLRs in RA or other autoimmune inflammatory diseases [16, 52]. All findings indicate macrophages are a main player in the development of arthritis. We found that TLR3 positive macrophages were increased in the spleen of PIA rats, and cell surface TLR3 showed high expression in the initiation stage of arthritis. It suggests that cell surface TLR3 on macrophage, but not the intracellular expression, exerts a crucial influence on the pathogenesis of PIA.
Using a rat macrophage cell line, NR8383, we could confirm that TLR3, but not TLR4, was upregulated after stimulation with pristane, which excludes LPS contamination in the experiment. Furthermore, the mRNA expression of IFN-β, the main downstream cytokine of TLR3, also increased with the same tendency as that of TLR3. The increase of IFN-β and TNF-α was inhibited by anti-TLR3 antibody, suggesting that this induction was TLR3 dependent. The above mentioned data indicate TLR3 and its downstream molecules in macrophage were activated by pristane, and then took part in the pathogenesis of PIA. More TLR3 expression on macrophages could affect the cell function, such as the capacity of antigen presentation, ROS production, phagocytosis, secretion of cytokines and chemokines and so on. Macrophages with upregulated TLR3 may affect the balance of Th1/Th2 and the activations of autoreactive T cells and B cells [25, 42].
However, it is still unclear how pristane induces TLR3 expression. The interaction between TLR and alkanes like pristane is not completely known. It is reported that oxidized alkane polymers induced activation of the TLR1/2 pathway as its ligands. And pristane could augment the effect of TLR7 ligands but does not directly activate TLR7. In a PIA rat model, further study is needed as to whether pristane or its metabolism products are the ligand of TLR3, or if it induces TLR3 by another pathway. Further studies are also needed to understand underlying mechanisms as to how TLR3 activates joint-specific autoimmune T cells via macrophages.
In summary, we screened TLR and downstream cytokine expression in the spleen of PIA rats, and found that only TLR3 expression was increased at the initiation stage. Regulating TLR3 by using ligand or shRNA in vivo could directly aggravate or relieve symptoms of arthritis. TLR3 may play its role via splenic macrophage, since it had a high expression on macrophages after pristane injection in vivo. And the induction of TLR3 was specifically due to pristane stimulation in macrophage cell lines with a dose- and time- dependent manner. These findings are not only useful in understanding the underlying pathogenesis of RA, but also provide a valuable clue for other immunological disease research.
antigen-presenting cell
dendritic cell
fluorescence-activated cell sorting
fibroblast like synoviocyte
- IFN-β:
interferon beta
mean fluorescence intensity
phosphate buffered saline
pristane-induced arthritis
- polyI:
C: polyinosine-polycytidylic acid
rheumatoid arthritis
- RNAi:
RNA interference
reactive oxygen species
- shRNA:
short-hairpin RNA
toll-like receptor
- TNF-α:
tumor necrosis factor alpha.
We should express gratitude to Prof. Yili Wang and Xu Li for critical reading the manuscript. And we are also grateful to Mr. Fujun Zhang, Ms. Yan Han and Qilan Ning for their expertise and assistance in the experiments. The work was supported by grants from the National Natural Science Foundation of China (30801027 and 30630058) and Shaanxi Province International Cooperation Foundation of China (2007-KW-06).
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- Isnardi I, Ng YS, Srdanovic I, Motaghedi R, Rudchenko S, von Bernuth H, Zhang SY, Puel A, Jouanguy E, Picard C, Garty BZ, Camcioglu Y, Doffinger R, Kumararatne D, Davies G, Gallin JI, Haraguchi S, Day NK, Casanova JL, Meffre E: IRAK-4-and MyD88-Dependent Pathways Are Essential for the Removal of Developing Autoreactive B Cells in Humans. Immunity. 2008, 29: 746-757. 10.1016/j.immuni.2008.09.015.PubMed CentralView ArticlePubMedGoogle Scholar
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- Joosten LA, Koenders MI, Smeets RL, Heuvelmans-Jacobs M, Helsen MM, Takeda K, Akira S, Lubberts E, van de Loo FA, van den Berg WB: Toll-like receptor 2 pathway drives streptococcal cell wall-induced joint inflammation: Critical role of myeloid differentiation factor 88. J Immunol. 2003, 171: 6145-6153.View ArticlePubMedGoogle Scholar
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- Vingsbo C, Sahlstrand P, Brun JG, Jonsson R, Saxne T, Holmdahl R: Pristane-induced arthritis in rats - A new model for rheumatoid arthritis with a chronic disease course influenced by both major histocompatibility complex and non-major histocompatibility complex genes. American Journal of Pathology. 1996, 149: 1675-1683.PubMed CentralPubMedGoogle Scholar
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- Okamoto S, Matsuura M, Akagi T, Akashi M, Tanimoto T, Ishikawa T, Takahashi M, Yamanishi K, Mori Y: Poly(gamma-glutamic acid) nano-particles combined with mucosal influenza virus hemagglutinin vaccine protects against influenza virus infection in mice. Vaccine. 2009, 27: 5896-5905. 10.1016/j.vaccine.2009.07.037.View ArticlePubMedGoogle Scholar
- Filleur S, Courtin A, Ait-Si-Ali S, Guglielmi J, Merle C, Harel-Bellan A, Clezardin P, Cabon F: SiRNA-mediated inhibition of vascular endothelial growth factor severely limits tumor resistance to Antiangiogenic thrombospondin-1 and slows tumor vascularization and growth. Cancer Research. 2003, 63: 3919-3922.PubMedGoogle Scholar
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- Holmberg J, Tuncel J, Yamada H, Lu SM, Olofsson P, Holmdahl R: Pristane, a non-antigenic adjuvant, induces MHC class II-restricted, arthritogenic T cells in the rat. J Immunol. 2006, 176: 1172-1179.View ArticlePubMedGoogle Scholar
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- Barton GM, Medzhitov R: Toll-like receptor signaling pathways. Science. 2003, 300: 1524-1525. 10.1126/science.1085536.View ArticlePubMedGoogle Scholar
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- Heinz S, Haehnel V, Karaghiosoff M, Schwarzfischer L, Muller M, Krause SW, Rehli M: Species-specific regulation of Toll-like receptor 3 genes in men and mice. Journal of Biological Chemistry. 2003, 278: 21502-21509. 10.1074/jbc.M301476200.View ArticlePubMedGoogle Scholar
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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.


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a) Burn down the granary and pretend it was an accident.
b) Punish the cowardly, disloyal workers and replace them with new ones.
c) Tell the lazy workers that the game is up and confiscate the arms.
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Showing Results: Most Recent
This review is from: Scythe GentleTyphoon D1225C12B3AP-13 120mm Case cooler
Pros: Very quiet, moves alot of air
Cons: None
Other Thoughts: I was really impressed. My new build is sitting on the desk right next to me and I can barely hear it running. Great fansREAD FULL REVIEW
This review is from: OCZ XMP Ready Series 6GB (3 x 2GB) 240-Pin DDR3 SDRAM DDR3 1600 (PC3 12800) Desktop Memory Model OCZ3X1600LV6GK
Pros: Works well, had no problems in my overclocking, looks great too.
Cons: NoneREAD FULL REVIEW
This review is from: EVGA E758-A1 3-Way SLI (x16/x16/x8) LGA 1366 Intel X58 ATX Intel Motherboard
Pros: Amazing board so far. Ive been putting it through its passes since receiving it and its really a joy to use. The knowledge and help on the EVGA forums is top notch and I feel like I would have been doing myself a discourtesy by buying any other motherboard.
Cons: None
Other Thoughts: With the level of customer support, and service (US based 24/7 PHONE support), as well as the fact that they are incredible boards, why wouldn't you buy an EVGA product?READ FULL REVIEW


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Several days ahead of Samsung's September 4th debut date for its Galaxy Gear smartwatch, the notorious tech leaks outlet Evleaks has tweeted screenshots that tell us a bit more about how the smartwatch's complementary smartphone app will allegedly work.
The image indicates the Galaxy Gear will be equipped with both NFC and Bluetooth capabilities. It also appears you'll be configuring the watch through an Android app, similar to the way both the Sony SmartWatch and the Pebble watch work. And a "Find my watch" feature suggests the watch will come with built-in GPS.
There's still much we don't know, such as what kinds of apps Samsung will offer for the watch, but stay tuned—we'll have all the details come launch day next week.
[Image: Evleaks]


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Submit Content | Media Center | NCAA.COM
As a former gymnast at Southern Connecticut State and now a maternal-fetal medicine specialist, Dr. Courtney Stephenson knows the value of dedicating herself to a greater goal.
Last year, Florida shared a national title with Oklahoma. This year's championship once again came down to the final event. Rather than sharing the podium, though, Florida stood alone at the top.
National Collegiate NCAA.com Championships Recap All-Time Championships Records and Results (through 2015 Championships)
University of Alabama gymnast Kim Jacob's senior season started with a fractured tibia. It ended with the 2014 NCAA title.


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Read about the talk that stole the show at a recent ME/CFS conference in Simon McGrath's two-part blog.
Discuss the article on the Forums.
Could this be an early XMRV? The JHK virus
Discussion in 'XMRV Research and Replication Studies' started by currer, Aug 15, 2011.
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Page 3 of 5


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You are hereLearn S4L - LTAD › Ten S's of Training and Performance
Ten S's of Training and Performance
In considering the trainability of different athlete skills and qualities, LTAD looks at 10 S's of Training and Performance that describe 10 distinct capacities of the athlete.
The 10 S's of Training and Performance include five basic physical literacy capacities:
Beyond these five physical capacities, there are five general S's that complete the holistic development of the athlete:
Each of these 10 capacities is trainable throughout an athlete's lifetime. However, research shows that there are sensitive periods of accelerated adaptation to training when each of the first five capacities is especially receptive to training.
If these sensitive periods are missed, the athlete may never grow to have as much stamina, strength, speed, skill and suppleness as they could have had. There are also other considerations in trainability.
S4L For
- Coaches
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text: Like, comment, & watch to the end in hd! HD is good for your eyes;) hope you guys enjoyed! i tried a little bit of a different intro this time:P don't know how to feel about it ... xD BUTTT I feel like it's appropriate, because I came up with tTags:


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Adam Serwer points out today that the very name of the planned Islamic community center in Lower Manhattan has led the most paranoid members of society to believe that it is an explicit announcement of the intention to impose Sharia law on the United States. Serwer points out that this is "cartoon-villain stupid." So it makes sense that leading the charge here is Newt Gingrich, who divines the magic forces at work thusly:
The true intentions of [Faisal Abdul] Rauf are also revealed by the name initially proposed for the Ground Zero mosque --"Cordoba House" -- which is named for a city in Spain where a conquering Muslim army replaced a church with a mosque. This name is a very direct historical indication that the Ground Zero mosque is all about conquest and thus an assertion of Islamist triumphalism which we should not tolerate.
Serwer notes that the Cordoba Initiative has announced ideals of its own, chief among them being a restoration of "the atmosphere of interfaith tolerance and respect that we have longed for since Muslims, Christians and Jews lived together in harmony and prosperity eight hundred years ago."
As Serwer goes on to say, this basically boils down to whether you accept that Faisal Abdul Rauf, the imam behind the so-called "Ground Zero mosque" -- after having a long career building interfaith bridges with the stated intention of out-competing Osama bin Laden and his al Qaeda death cult for the hearts and minds of young Muslims -- has now accidentally tipped his hand to the secret plot to destroy America that he's been hiding all along. (If only he'd been smart enough to call the proposed center "The Newt Gingrich House of Pancakes!")
And then, if all that is true, you have to imagine that somehow, it's even possible for Sharia law to be imposed somehow on America. One need only look at how difficult it is for a global superpower to impose Western democracy on Iraq and Afghanistan to see how foolish an idea that is. From my perch inside the Beltway, I find it hard to believe that the "Impose Sharia Law Act of 2011" wins Ben Nelson's vote in the Senate. I suppose it could be done through sorcery, but al Qaeda will really have to step up its game, from crotch-based incendiary discharges, to make that work.
Cordoba: Lost In Translation [Adam Serwer]
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Donald Campbell Dewar
Daily Forecast for February 21st 2017
Donald Campbell Dewar is a male celebrity.
He was born on Saturday August 21st 1937, in Glasgow, Scotland,.
February 21st 2017 means business for Donald Campbell Dewar and requires his full managerial and organizational skills. In return, the day may bring Donald financial gain.
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Loved ones may complain about lack of attention on his part.
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You and Donald
About Donald's Numerological Forecast & Personal Day Cycle
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Read more about our numerological daily forecast for


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Sci Fi Channel, with one of the simplest and most recognizable brands in TV, is going and making things complicated by changing to the ridiculous moniker "Syfy."
What's next? The Phood Network? The Travville Channel? See Enn Enn?
The July 7 name change for the NBC-owned cable outlet is described as "phonetically identical" to Sci Fi. And those are among the dumbest two words you'll see strung together in a press release.
Unsaid in that release is that this is designed to clean up the nerdy image of a cable outlet for guys with pocket protectors who spend too much time at the comic-book store.
"The new brand broadens perceptions and embraces a wider and more diverse range of imagination-based entertainment," according to the formal announcement, which goes on to talk about the money reason behind the silly name:
"It also positions the brand for future growth by creating an ownable trademark that can travel easily with consumers across new media and non-linear digital platforms, new international channels and extend into new business ventures."
Another great press release word: "ownable."
Not all these name changes are silly. Last week, Nickelodeon announced it was renaming its sister channels to fit in to the bigger Nick "brand."
In that change, which kicks in this fall, Noggin becomes Nick Jr. and "The N" will become TEENick, with everybody being tied together in the nick.com Web site.
That name change not only makes good sense; it's not annoying.
CHANNEL SURFING: Channel 58 will air CBS' "March Madness" college basketball coverage starting this week on its main channel and on its "This TV Network" affiliate, available on Time Warner digital Channel 201, Charter Communications Channel 970 and over the air on digital Channel 58.3.
• Tuesday's "American Idol" performance show features Milwaukee's Danny Gokey and the 10 other finalists singing songs from the Grand Ole Opry. Former "Idol" winner Carrie Underwood performs on the two-hour "Idol," which airs at 7 p.m. on Channel 6.
• Time Warner Cable adds WPXE HD - Milwaukee's ION Network affiliate - and MLB HD to its high-definition service at Channels 563 and 583, respectively. HD subscribers with the standard versions of these channels will get the high-def versions.
• MTV's "True Life" is looking for young people between the ages of 14 and 24 who are at odds with their parents over some issue that could be resolved by the end of an episode. If you fit the bill, e-mail your name, age, location, phone number and recent picture to email@example.com.
• President Barack Obama visits Jay Leno on NBC's "Tonight Show" at 10:35 p.m. Thursday on Channel 4. If another sitting president has done one of the late-night talk shows, I can't find any evidence of it.
• Fox News Channel is certainly happy with Glenn Beck's move from CNN's HLN. Friday's show was his best-rated, with just over 3 million viewers. That was second only to Bill O'Reilly, according to Nielsen Media Research numbers.
THE MARCH CHAT: This month's cyberchat starts at noon Wednesday.
Reach Tim Cuprisin at (414) 224-2397 or firstname.lastname@example.com. Follow him at twitter.com/cuprisin.


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Who'll get the BB boot?
BB evictions are set for a nail-biting finale this evening.
Advertisement - Continue Reading Below
Who goes - bitchy Laura or bunny boiler Chanelle?
Despite Ziggy's plea for 'space' in their budding relationship (hardly possible when you're cooped up in a tiny house being watched by millions of people) he urged viewers yesterday: "If anyone cares about my wellbeing, keep [Chanelle] in." We say vote Chanelle, just to see how quickly her devoted boyfriend moves onto someone new.
Who do you think will get evicted. Place your bets on the Cosmo gossip forum.
Click here for more celeb gossip.


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The Indian Space Research Organization (ISRO) achieved a major milestone last night (Feb. 14) with a successful record-setting launch of 104 satellites on a single rocket.
If you missed out on spotting the green comet that zoomed past Earth over the weekend, don't worry — Comet 45P was only the first of three comets to pass near Earth between now and the end of 2018.
Astronomers have spotted more than 100 new potential alien planets, including one in the fourth-closest star system to the sun, a new study reports.
NASA's Juno spacecraft has flown by Jupiter for the fourth time.
Spending a year in space affected former NASA astronaut Scott Kelly's body in subtle but potentially significant ways, new research suggests.
Another long-term space simulation is underway at the Mauna Loa volcano in Hawaii, where six crewmembers entered a small, isolated dome Jan. 19 as a part of the HI-SEAS mission.
A massive iceberg the size of Delaware is now even closer to breaking free from Antarctica, due to a widening crack in the ice shelf, scientists report.
NASA is known for its awe-inspiring missions to explore the far reaches of the cosmos, but this month, the space agency is preparing for very different kinds of rocket launches: ones to explore the mysteries of Earth's auroras.


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Lake Ice
January 4, 2015
I'm a photographer based out of Anchorage, Alaska. I've recently become a wife, a stepmother, and a homeowner. Life is pretty awesome. You can email me anytime. Learn more...
Husband and I went different directions in the grocery store, forgetting his phone is dead. It's like being in the dark ages. 28 hours ago
Me: My skin is super dry and itchy Also me: https://t.co/DBNF7OCFKC 34 hours ago
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#fbf February 2010, Friday work day is over, and it's #twitterafterdark somewhere https://t.co/OcVGWOFwVK 53 hours ago
I AM IN POSSESSION OF GIRL SCOUT COOKIES THIS IS NOT A DRILL 55 hours ago


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Two people were taken into custody today following a raid in the 1300 block of South 116th Street that police say turned up chemicals used in the manufacture of synthetic drugs.
The raid, which took place at 2:30 p.m., involved the West Allis and Milwaukee police departments, the Federal Drug Enforcement Agency, the Wisconsin Division of Criminal Investigations, the Milwaukee High Intensity Drug Trafficking Area, the Milwaukee Metropolitan Drug Enforcement Group and the West Allis Fire Department.
As a precaution, part of 116th Street was initially blocked off as entry teams went in wearing hazardous materials protection suits.
There was no one in the building at the time, but authorities took two suspects in to custody at another location.
Police released no details about what chemicals or how much were confiscated. The investigation is continuing.
More from News and Features
- Suburban Milwaukee Now websites to get new look, realignment
- 1940s radio noir comedy to cap Village Playhouse season in West Allis
- West Allis Police blotter: July 28, 2016
- West Allis rechecking pay to ensure market rate compensation, fairness
- Race for Wisconsin's 6th District Senate seat heats up
- It's 'Curtains' for West Allis Players presenting murder and music
- Nathan Hale High School tuneful summer musical set at high school
- West Allis Police blotter: July 21
- In brief, July 7: Gun control, gourmet store, home for the poor
- West Allis bans e-cigarette smoking where tobacco smoking is banned


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text: The Correct Number Of Exhausts Is As Many As Physically PossibleRaphael Orlove8/10/16 9:04am19611EditPromoteGo to permalinkCar exhausts. Some have one, others have two, more still have more than two. But what is the most optimum number of car exhausts for a car to have? The answer is simple: as many as possible.


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The police re-opened Route 90 from Ein Gedi to the Dead Sea, Thursday afternoon. They also re-opened Highway 204 from the Halukim Junction to Yerocham.
Route 234 in the Tze'elim Bridge area and Highway 222 from Tze'elim to Retamim are blocked.
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Banana Chip Muffins
Total Time
Prep 10 mins
Cook 20 mins
These are wonderful little muffins. I always have to have one just out of the oven warm. These are so good.
Ingredients Nutrition
- 1 egg
- 1⁄3 cup vegetable oil
- 3⁄4 cup sugar
- 3 medium bananas, mashed
- 2 cups flour
- 1 teaspoon baking soda
- 1⁄2 teaspoon salt
- 1⁄2 cup oatmeal
- 3⁄4 cup semisweet mini chocolate chips
- Beat eggs, oil and sugar in a bowl until smooth.
- Stir in bananas.
- Combine dry ingredients and stir into the banana mixture, just until moistened.
- Stir in chocolate chips.
- Fill greased muffin cups (or use cupcake paper liners) 3/4 full.
- Bake at 375ºF.
- for 18 to 20 minutes or until muffins test done.
Most Helpful
The best banana muffin recipe I have found so far. The oatmeal adds a very nice texture to what could be an ordinary banana muffin. And of course you can't go wrong with chocolate chips. Thanks for a delicious recipe.
These were easy, included ingredients I had on hand, were easily adaptlable to family allergies, and tasted great. My children, who don't normaly like muffins, enjoyed these.
WOW!! These are great. The flavor combinations go great together. I had one after I took it out of the oven~it was sooo good. Thanks for another great muffin recipe Miss Annie!! I'll have to make another batch tomorrow.


text:
"The Closer" season finale revolved around a series of brutal crimes, a jurisdictional pissing match, and multiple manipulations. Plus, Daddy returns, and Charlie tugs the heartstrings.
We open on a comely blonde sitting in the pool, talking on the phone, not a care in the world… until someone stabs her multiple times. Her husband said a ninja did it — ok, fine, a guy dressed all in black, like a ninja. The squad doesn't believe it, of course, and set about trying to get him to confess. But guess what? Someone else reported a ninja prowler — and he got away driving a car with Texas dealer plates.
So Brenda calls El Paso for a bit of info, and gets Detective Landry on the phone. He immediately knows what kind of case they're talking about, and says he'll be there in four hours. That's not what Brenda wants to hear.
Thus begins the jurisdictional fight of the evening. The ninja-clad killer murdered two women in Texas, and Landry wants to take him back to Texas for execution — "Try 'em and fry 'em, or, as we may say it these days, collect 'em and inject 'em." Brenda doesn't want that — in all likelihood, this man has killed a lot more than three women, and she wants answers for as many families as possible. They decide that the cop with the most evidence to try the case can have it. Landry is pretty sanguine when they track down their first suspect, the owner of the getaway car. Yes, he owns the car, and he just relocated from Texas, but he just picked that car up from the depot near the airport today.
Brenda tracks down the driver of the truck that delivered Webb's car — Jesse Ray Moore. Brenda and the team use LoJack to find him at a truck stop in California, and Gabriel and Sanchez try to find him. Moore tries to drive away, but Sanchez jumps on the truck. Cue the swerving and the hitting and the out-of-control truck as Sanchez tries to get Moore to stop. It's a brutal fight — Moore breaks Sanchez's gun arm with a baseball bat, but Sanchez eventually gets the upper hand and stops the truck.
Moore has plenty of souvenirs, but there's no blood and no knives in the truck. Then Landry drops a bombshell. Oops! We do happen to have DNA evidence from one of the crime scenes, we just haven't processed it yet. And DNA… well, that's the best evidence there is, right? So I'll just be taking him back to Texas…. Not so fast, Brenda replies. I plan to get him to revoke his right to counsel and confess to everything.
Which, of course, she does — but threatening him with Texas-style execution. You go with Landry, and you're a cinder within a year. California, on the other hand, hasn't executed anybody in ages. You stay here, you live. All you have to do is confess.
He does…. and it's sickening. He remembers details of all his crimes, and he happily shares them with Brenda, who he sees as a a sympathetic ear. She barely keeps her lunch down, but after more than seven hours, she gets him to confess to 16 murders across the Southwest.
Landry congratulates her — confessions trump DNA, so she gets the case. But just for some light reading, her gives her a file on the 16 victims. Take a look at their before-and-after photos, just for kicks. Brenda does… and decides that maybe Moore should be extradited to Texas after all. Landry gloats in the video room — "I played that just right, hmm?" Ick.
The home front
Brenda's Daddy is there to take Charlie back… but Charlie doesn't want to go. She begs Brenda to let her stay, and Brenda says that if Grandpa agrees, they'll talk about it. Fritz overhears, and is not happy. No worries, Brenda says — Daddy will never agree!
Ah, but Daddy refuses to be the bad guy. I told her it was entirely up to you two. Now figure out whether she's getting on the plane with me. I'm staying out of it. So even after her horrific case, Brenda has to tell Charlie that no, she can't stay with them. Yes, she's changed while she's been out here, but she can carry on with that change back in Atlanta. We gave you a second chance, and now your parents deserve the same thing. Besides, I'll come visit, I promise! "You always say that and you never do," Charlie sobs. Ouch. Well, maybe I deserve a second chance too, Brenda says.
Highlights, thoughts and odds and ends
- When the squad realizes that yes, there is someone dressed up like a ninja terrorizing LA. Sanchez turns to Tao: "What do you know about ninjas?" he asks. "What, I'm Asian-American so I'm supposed to know about ninjas?" Tao scoffs. There's a slight pause, then Sanchez says, "Uh, yeah." And Tao, of course, launches into the history and weaponry on the ninja. But that's less because he's Asian-American and more because he's a fervid and unrepentant geek — which is why I adore him.
- Barry Corbin plays he grumpy, long-suffering father to perfection. Still, too many comments like "We both know I'll be gone from this world soon, so you might as well get a jump on it by forgetting about me now…" and I sort of want to smack him.
- Poor Charlie. She's devastated by the whole situation. I don't know if Brenda and Fritz should have let her stay — but I'm surprised, after Brenda used her as a stooge, that Charlie would want to stay. Are things that bad at home?
- Sanchez kicks Moore in the ribs several times after the truck stops — and it's all caught on aerial footage. I can't help but think that's going to come back to bite them someday.
- Oof, that confession scene was brutal. Poor Brenda, having to smile and nod and sit through seven hours of that sort of horror!
OK, folks — That's it for me. I'm stepping down from recapping duties, but it's been a pleasure writing for you. You can check out more of my TV writing at the CW Source — and thanks!
Follow Zap2it on Twitter for all the latest TV, movie and celebrity news.


text: What Car Is Most Likely To Get Pulled Over?Travis Okulski4/22/13 11:46amFiled to: Question of the DayQOTD4172EditPromoteShare to KinjaToggle Conversation toolsGo to permalink Breaking the law on the road will get you pulled over. We know this. But some cars tend to fly under the radar while others are immediate cop magnets. Which ones will almost certainly get pulled over first? Advertisement If you have a lowered Honda Civic with a loud exhaust and it's painted red, I tend to think you're basically asking to be pulled over. Here's my logic:It's red, which people can see for miles. It's loud, which gives cops a clue you're coming. It doesn't matter how well the exhaust is tuned or how much work is done. To a non-car enthusiast, a Civic with a giant pipe and no other work will sound and look the same as one that's fully built and a legit hot rod. Advertisement I see cars like this pulled over a lot in my area, and I don't imagine that all of them around going 90 in a 55 or doing something aggressive. I think that's just the reputation that has been earned in Northern NJ.What say you? What car is a ticket for a ticket?(QOTD is your chance to address the day's most pressing automotive questions and to experience the opinions of the insightful insiders, practicing pundits, and gleeful gearheads that make up the Jalopnik commentariat. If you've got a suggestion for a good Question of the Day, send an email to tips at jalopnik dot com.) Sponsored Photo Credit: WillVision PhotographyReply417 repliesLeave a reply


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Top Definition
The most insulated of suburbs along the Mornington Peninsula in VIC. Its residents are generally old snobs, cashed up asians, painful and wealthy soccer mums and rich teenagers. The pinacle of Mount Eliza's community is 'The Village' a shopping village consisting of 80% hair salons 10% cafes and 10% overpriced retail stores.
Person A: "Dude do you live in Mount Eliza?"
Person B: "Nah dude, I'm not rich or snobby!"
Person B: "Nah dude, I'm not rich or snobby!"
by chris pero November 07, 2011
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text: This Is Your Life, and It's Ending One Minute at a TimeJesus Diaz8/25/09 10:30amFiled to: Datadata visualizationYour life39EditPromoteShare to KinjaToggle Conversation toolsGo to permalink Matt showed you Microsoft's Gordon Bell life in data files since 2001. But how does all that translate to your average real world daily activity? Click image to zoom in. Advertisement Advertisement Shan Carter, Amanda Cox, Kevin Qualy, and Amy Schoenfeld show how most people's typical day goes in this nice data visualization graphic. On average, we spend up two-thirds of your day sleeping, eating, working and watching television. I think I can pass on the last part, to be honest. And I'll sleep when I'm dead in my grave.Check out their cool interactive graphic at the NYT. [NYT via Dark Roasted Blend]Reply39 repliesLeave a reply


text: Wireless connections are a great thing, as you can connect devices to the Internet and one another without the need for cables. Sadly, many of us don't use wireless devices much beyond Wi-Fi. As we continue to use an increasing number of devices, all of which use cables, it can be hard to keep them in order.


text:
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Помогите нам: эти порнозвезды присутствуют в этом видео?
Pornstar Name:
Very sexy.. double fuck
What a classic
Great. Very sensuous and well acted.
Wanna have sex? So cute and sexy Beautiful women
What is the title of the movie
So natural and unplanned, bet she was wet and well lubricated by the time her hubby got in her cunt.
yes ..!! your right .
dirty bitch
what movie is this
Normal (2007)
so nice & sexy
omg this is so hot wow
Lol funny
Not "porn" but sexy girl and sexy film. why don't porn actresses learn to act like film actresses (or film actresses do porn films :))?
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text:
A Promise Is a Promise
€ 10,99
Lieferbar innert 2 Wochen
August 2001
BeschreibungNow in trade paper, popular self-help author Dyer presents an extraordinary biography about a courageous mother and the child she will never abandon. Eduarda Oberra is a Miami woman who has been in a coma for 26 years.
PortraitDr. Wayne W. Dyer, affectionately called the "father of motivation" by his fans, is one of the most widely known and respected people in the field of self-empowerment. He became a well-known author with his bestselling book Your Erroneous Zones, and has gone on to write many other self-help classics, including Meditations for Manifesting, Staying on the Path, Your Sacred Self, Everyday Wisdom, and You'll See It When You Believe It. Despite his childhood spent in orphanages and foster homes, Dr. Dyer, who has a doctorate in counseling psychotherapy, has overcome many obstacles to make his dreams come true. Today he spends much of his time showing others how to do the same. His latest titles include: Getting in the Gap and 10 Secrets for Success and Inner Peace.
Untertitel: Revised. Sprache: Englisch.
Verlag: HAY HOUSE
Erscheinungsdatum: August 2001
Seitenanzahl: 128 Seiten
Format: kartoniert
Alle Preise inkl. MwSt., ggf. zzgl. Versandkosten


text: Despite the last few days near 80 degrees. This overall cool Spring has delayed trees from blooming in far Northwest and Northeast CT. The highest elevations such as Mohawk Mountain are still showing mainly bare branches as you can see in the above picture. Here in Hamden the trees are a few weeks late but are finally becoming more green. So It's hard to believe Memorial Day the unofficial start to summer is just 2 weeks from Today.


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Wilson Women's Star Ruffle Woven Skirt 35,90 €
Mix and match any tennis top with cute athletic piece. The stretch-woven Wilson Star Ruffle Skirt features an elastic waistband, built-in compression short with ball pockets at sides, cute ruffle trim around skirt hem, flat seams for comfort, moisture-wicking performance, UV protection, and a heat transfer Wilson logo at left side. Content: Body: 86% Polyester, 14% Elastane. Inner Short: 92% Polyester, 8% Elastane. Colours: Black w/Silver Dark Plumberry w/Silver
Wilson Women's Star Striated Skirt 19,90 €
Continue to raise the level of fashion with the Wilson Star Striated Skirt. This stylish knit skirt features an elastic waistband, built-in compression short with ball pockets at each side, sublimated graphic print around skirt hem, colourblocking waistband, contrast stitching, flat seams for comfort, nanoWIK moisture-management technology, and a heat transfer Wilson logo at left hem. Content: 88% Polyester, 12% Spandex. Colours: Blue Mirage Striated w/Silver White Striated w/Silver
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text:
Greystones House
No prices found through our partners. Please contact the business directly or check some of our recommended alternatives.
How has this hotel rated in the past?
- Tennis Court
- Non-smoking rooms
- Free High-Speed Internet
- Continental Breakfast
- Non-smoking hotel
- Wireless internet connection in room (free)
Photos of Greystones House
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- Tennis Court
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TripAdvisor Reviews Greystones House Castle Cary
Travel Blogs from Castle Cary
Final flight home
I waited around in the airport for about 5 hours until I boarded my final flight. When I got on, it was extremely quiet, I had a row to myself, as well as the rows surrounding me! The flight was so quiet, it was easy to sleep as everyone had a row to lie down on. I watched a few films, had some naps and are the food and drink that was brought to me. It may have been ...
Glastonbury and Stonehenge
... on this Day of the Dead, Glastonbury was covered in thick fog. Trees were either bare naked or still shedding their brown leaves and made silhouettes in the fog. We hiked to the the tor and began our ascent up the mythical island of Avalon with wet grass and mud under our feet. The hill was steep and deceivingly tall. We could never see the top while climbing. It just kept going up. I followed the dark outlines of my fellow travellers through ...
Halloween in Roman Britain
I had booked a trip through the school to visit Bath, Glastonbury, and Stonehenge on Halloween weekend. The bus left early in the morning before breakfast was being served at the dorms so I had to wait to eat until our first stop. We took a break at an eco-friendly rest stop which was covered by a mound of earth with grass growing all on the top of it. There was also a section on the top that had a solar power grid. They were serving ...
Cooking in Castle Cary
... was good in a teacher who was preparing us for if we ever wanted to cook professionally. She could also get frustrated, but in general she was patient, knowledgable and helpful. Roger is Lou's husband. At work during the day, he would join us for dinner. Full of information, he loved to talk and teach us things and made for very interesting table conversation. He would also help us clean up after dinner, and his super speed would make our lives much easier. I ...
Have I told you about the best Christmas ever?
... Not being entirely mercenary about money, this decision was also swayed by a log fire, use of a car, a TV to watch the Queen's speech, a dog and family members in the locality; not necessarily in that order.
Christmas for us was just for us and it was wonderful. I didn't even mind the nauseating bus journey to avoid the extortionate train fare. On Christmas morning we walked the dog, swapping Christmas greetings and ...


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Getting ready to show at an expo or conference? Corporate swag is a must-have to gift showroom attendees. After all, free stuff is a huge draw for people to visit your table and gets them talking about your brand! At ePromos, you'll find our selection of trade show products at ePromos appeals to every type of customer and business executive. Our mission is outfitting companies with memorable items that fulfill their promotional needs. Make your brand a part of everyone's grab bag with our unique logoed trade show gifts!
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Dylan Phoenix didn't show the proper respect to OT. Sure, she dressed really nice to come and see him, but she should have shaved her pussy clean, knowing he was going to want to play with it. Sure, it looks smooth, but as soon as he runs his hands over her little mound he can feel the stubble starting to form. If she isn't going to groom herself properly for him, then he is going to have to do it in a way that will remind her for next tme.Format: mp4Duration: 43:22Video: 1280x720, AVC(H.264), 7284kbpsAudio: 109kbps
IT Girl (Apr 17, 2015)
IT girl
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Thank you very much, I'll try to burn in my brand new sennheiser cx500
post #31 of 35
11/29/08 at 12:03pm
I have been very sceptical about the benefits of burn-in, especially after my flac debacle where I acidentally my whole collection.
However, I decided to give it an extra try, because I had essentially nothing left to lose. And also, if you cannot trust expert audiophiles who know their stuff, then who can you trust in this world?
So today I have listened to pink noise and frequency sweeps (at about 70 % volume) for 10 hours and then when I switched to some Mozart I was AMAZED by how good it sounded. It was crappy 320 kbit mp3s but I really enjoyed it, and my headache went away too.
Thanks a lot!!!


text:
Love Tiki Totem 12" Stained - Tropical Decor
Price: $39.90
Sku: YDA1101330B
This wooden Love tiki - will bring you eternal love and happiness... This tiki idol measures 12 inches tall. Made of Monkey Pod wood: stained color! Each tiki totem was hand carved with great attention to details. Each piece was hand waxed in Hawaii to give a nice finish!
Great addition to any Tiki collection!


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Hebrews 6:11New King James Version (NKJV)
11 And we desire that each one of you show the same diligence to the full assurance of hope until the end,
New King James Version (NKJV)
Scripture taken from the New King James Version®. Copyright © 1982 by Thomas Nelson. Used by permission. All rights reserved.
Hebrews 6:18-20New King James Version (NKJV)
18 that by two immutable things, in which it is impossible for God to lie, we might[a] have strong consolation, who have fled for refuge to lay hold of the hope set before us.
19 This hope we have as an anchor of the soul, both sure and steadfast, and which enters the Presence behind the veil, 20 where the forerunner has entered for us, even Jesus, having become High Priest forever according to the order of Melchizedek.
New King James Version (NKJV)
Scripture taken from the New King James Version®. Copyright © 1982 by Thomas Nelson. Used by permission. All rights reserved.
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