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Reduced plasma levels of soluble interleukin-7 receptor during graft-versus-host disease (GVHD) in children and adults
BMC Immunology volume 15, Article number: 25 (2014)
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Interleukin 7 (IL-7) signals via the IL-7 receptor (IL-7R) and drives homeostatic T-cell proliferation in patients after allogeneic hematopoietic stem cell transplantation (aHSCT).
We performed a prospective study in adults (n = 33) and children (n = 29) undergoing aHSCT measuring plasma IL-7 and soluble IL-7R (sIL-7R) concentrations between 1 and 12 months after HSCT in order to investigate the link between sIL-7R and clinical events after aHSCT.
sIL-7R, but not IL-7, increased with time after HSCT in plasma from all patients enrolled in the study. sIL-7R values were higher at 2, 3, and 6 months (p < 0.01) if the donor was a sibling as compared to an unrelated donor. Increased sIL-7R levels were also identified in plasma from patients who were not treated with anti-thymocyte globulin (ATG). Low sIL-7R was associated with any grade of acute graft-versus-host disease (GVHD) at 2 and 6 months (p = 0.02) and with a positive CMV PCR at 2 months after HSCT (p < 0.05). Patients with cytomegalovirus (CMV) reactivation had increased IL-7 values at 2 and 3 months (p = 0.02) after HSCT. In multivariate analysis, lower sIL-7R levels were associated with acute GVHD (relative hazard (RH): 0.70, p > 0.01) and sibling donors (RH: 2.23, p = 0.004). Recipients of sibling grafts showed high levels of IL-7 (RH: 1.38, p < 0.05) and bone marrow recipients had low IL-7 levels (RH: 0.73, p = 0.04).
Measurement of the sIL-7R/IL-7 axis will help in guided immune monitoring after HSCT and guided interference with sIL-7R may be explored in GVHD management.
Interleukin-7 (IL-7) is a key cytokine in allogeneic stem cell transplantation (aHSCT) and drives homeostatic, extrathymic T-cell expansion in lymphopenic hosts [1]. IL-7 may also promote expansion of alloreactive T-cells that mediate graft-versus-host disease (GVHD) [2]. Elevated IL-7 levels in serum have been shown to be associated with acute GVHD [3, 4]. Several cell types have been described to contribute to IL-7 production, e.g. stromal cells [5], macrophages [6], B-cells [7] and thymic epithelial cells. Several conditions may impair IL-7 production, including infections associated with tissue damage of stromal cells, the fibroblastic reticular network (FRC) [8] and cytotoxic therapies in the course of anti-cancer treatment or in conditioning regimens for HSCT. Murine studies have shown that radiation, provided in the course of HSCT, can result in reduced thymic stromal IL-7 production [9]. However, it has not been unequivocally proven that thymus-derived IL-7 contributes substantially to systemic IL-7 levels. IL-7 is consumed by the pool of available immune cells expressing the heterodimeric IL-7 receptor (IL-7R, CD127) along with the common gamma chain (CD132) [1, 3]. Expression of the IL-7R on T-cells is associated with T-cell differentiation and maturation, based on CD45RA and CCR7 expression [10]. Precursor T-cells exhibited the highest numbers of IL-7R molecules per cell and terminally differentiated T-cells were found to express the lowest number of IL-7R molecules per cell.
The role of IL-7 in immune reconstitution after HSCT is multifaceted: it promotes thymopoiesis by driving development of immature thymocytes [11, 12]. Some reports have suggested that IL-7 treatment leads to improved—but transient—immune reconstitution [11] without increased alloreactivity [13, 14]. In contrast, other studies have shown that IL-7 aggravates GVHD [2] and that subsequent blockade of the alpha chain of the IL-7R may prevent GVHD [15]. The situation is even more complex, since the IL-7R is not only available in the cell-membrane-bound format, but also as a soluble form (sIL-7R). The soluble IL-7R binds IL-7 with an affinity similar to that of the membrane bound IL-7R [16], leading to sIL-7R-mediated inhibition of IL-7 signaling in T-cells [17]. sIL-7R is not only generated by shedding of membrane-bound receptors, it is also associated with polymorphism in the IL-7R gene (rs6897932), which leads to increased splicing in the transmembrane domain of exon 6 in the 8-exon Il-7R gene [18–20] resulting in increased sIL-7R generation. This SNP has been associated with autoimmune diseases, i.e. type-I diabetes mellitus [21] and rheumatoid arthritis [22]. IL-7R polymorphism has also been studied in adults after HSCT with inconclusive results concerning SNP analysis of donors [23] and recipients [24]. Up to now, the protein concentrations of IL-7 combined with its receptor IL-7R have not been measured in plasma from children and adults during 12 months after HSCT. We therefore designed a longitudinal study to determine IL-7/IL-7R plasma levels in 61 individuals after HSCT and we investigated associations between IL-7/IL-7R and clinical events after HSCT.
Patients and controls
The study involved 29 children and 32 adults (Table 1). Forty-eight patients underwent HSCT for malignant disorders, 16 patients were in first complete remission. 21 patients received grafts from HLA-identical sibling donors and the remaining patients received grafts from matched unrelated donors (n = 33), or HLA-mismatched unrelated donors (n = 8). The sources of stem cells were bone marrow, peripheral blood stem cells, or in a few individuals, cord blood transplants. Genomic HLA typing (MHC class I and class II four-digit typing) was performed as described previously [25]. IRB approval (Stockholm Ethical Committee South 2010/760-31/1) was in place. Peripheral blood mononuclear cells (PBMCs) were from adult participants (n = 32) and in the case of children (n = 29), consent was obtained from their parents or legal guardians (on file at CAST; Center for allogeneic stem cell transplantation). Samples were obtained from 61 patients at 1, 2, 3, 6, and 12 months after HSCT and 26 controls (15 adults and 11 children). Plasma was obtained after centrifugation and stored at -20°C; PBMCs were isolated from heparinized blood over a ficoll hypaque gradient. The cells were preserved in liquid nitrogen using fetal bovine serum (FBS) containing 10% DMSO.
HSCT regimen
Conventional myeloablative conditioning was given to 34 patients and consisted of cyclophosphamide (Cy) at 60 mg/kg for two days in combination with fractionated TBI (FTBI) at 3 Gy/day for four days (n = 15), or busulphan (Bu) at 4 mg/kg/day for four days (n = 17) [26]. Two patients received other protocols. Reduced-intensity conditioning (RIC) was given to 27 patients and consisted of fludarabine (Flu) at 30 mg/m2 for 3–6 days in combination with either Bu at 4 mg/kg/day for two days (n = 7), FTBI at 3 Gy/day for two days and Cy at 60 mg/kg/day for two days (n = 7), Cy at 30 mg/kg/day for two days (n = 7), treosulphan at 12–14 g/m2/day for 3 days (n = 5), or TBI (2 Gy) (n = 1).
GVHD prophylaxis and CMV PCR
Immunosuppressive treatment mainly consisted of cyclosporine (CsA) in combination with a short course of methotrexate (MTX) (n = 41), or tacrolimus and sirolimus (n = 16) [27]. All patients with an unrelated donor or a non-malignant disease received anti-thymocyte globulin (ATG) (Thymoglobulin, Genzyme, Cambridge, MA) (n = 45) or alemzumab (Genzyme) (n = 2) for 2–4 days during conditioning [28]. During the first month, blood CsA levels were kept at 100 ng/mL in patients with malignancies when a sibling donor was used and at 200–300 ng/mL when an unrelated donor was used and also in patients with non-malignant disorders regardless of the donor. In the absence of GVHD, CsA was discontinued after three to six months for patients with malignancies and after 12–24 months for patients with non-malignant disorders. Patients were monitored for CMV viral load with a quantitative PCR on whole blood from the time of engraftment weekly until day 100 after HSCT as published previously. Later than three months after HSCT, weekly monitoring was continued only in those patients who had experienced CMV reactivation or had severe GVHD, while the other patients were monitored at each visit to the transplant center occurring every 2-3 weeks until 6 months after HSCT. Pre-emptive antiviral treatment with either i.v. ganciclovir 5 mg/kg BID or oral valganciclovir 900 mg BID was given at the center's chosen intervention limit > 1000 copies/mL blood. The duration of therapy was a minimum of two weeks and was discontinued when the CMV viral load was < 500 copies/mL [29, 30].
Supportive care
All patients were kept in reversed isolation or they were treated at home, as described in detail previously [31].
Statistical analysis
Differences between patient groups (i.e. children versus adults) or within each group (i.e. comparing different time points) were analyzed using Mann-Whitney U-test or the Wilcoxon test using the Statistical software program (version 10) and GraphPad Prism 4 software. In the multivariate analysis of factors affecting the levels of sIL-7R at different time points, multiple regression were used. To determine whether there was any correlation between CD127 (IL-7R) positive immune cells and soluble IL-7R levels, we used linear regression analysis with the GraphPad software.
Quantification of plasma IL-7 and IL-7R
IL-7 quantification was performed using the ELISA IL-7 Eli-pair (Cat. 851.680.010; Cell Sciences, Inc., Canton, MA) according to the manufacturer's protocol (standard range between 200 and 3,125 pg/mL). Soluble CD127 was measured using an IL-7R ELISA; the anti-IL-7R alpha chain-directed antibody R34.34 (anti-CD127 purified Ab, 1 μg/ml; Beckman Coulter Inc., Brea CA) served as the capture antibody (50 μL/well) by incubation overnight with plasma at 4°C. The recombinant human IL-7 R alpha/CD127 Fc Chimera (306-IR; R&D Systems, Minneapolis, MN) served as the standard (ranging between 0.78125 and 100 ng/mL). Standard and samples were incubated for 4 h, followed by washing steps (PBS, 0.05% Tween) as described earlier [16]. sIL-7R was detected with a biotinylated anti-CD127 antibody (BAF306; R&D Systems). Incubation was for 1 h at RT, followed by washing as described above. Streptavidin-HRP was applied (554066: BD Biosciences, Frankin Lakes, NJ) for 30 min, with subsequent development using Tetramethylbenzidine (TMB). The absorbance was read at 450 nm.
Flow cytometry
PBMCs (0.5 × 106 cells) were first stained with anti-CCR7 for 15 min at 4°C, followed by addition of the 10-color antibody mix as described in detail previously [10]. The PBMC-antibody mixture was incubated for 15 min at 4°C. The anti-CD27 antibody was then added to the cells, which were incubated at 4°C for 15 min, followed by washing with 1 mL of PBS containing 0.1% BSA. The cell pellet was resuspended in 200 μL of PBS (with 0.1% BSA) and the cells were analyzed as described previously [10].
For analysis of PBMCs from children, frozen PBMCs were thawed and 1 × 106 cells were incubated at 4°C for 15 min with the following antibodies: peridinin-chlorophyll-protein complex- (PerCP-) conjugated anti-CD3 (SK7), allophycocyanine 7- (APC-Cy7-) conjugated anti-CD8α chain (SK1), phycoerythrincyanin 7- (PE-Cy7-) conjugated anti-CCR7 (3D12) purchased from BD Biosciences (Stockholm, Sweden), Krome Orange-conjugated anti-CD4 (13B8.2), fluorescein isothiocyanate- (FiTC-) conjugated anti-CD8β chain (2ST8.5H7), APC-Alexa Fluor 700-conjugated anti-CD107a (H4A3), PE-Texas Red-conjugated anti-CD45RA (2H4), APC-conjugated anti-CD127 (R34.34) purchased from Beckman Coulter (Marseille, France), and Brilliant Violet-conjugated anti-CD117 (104D2) purchased from BioLegend (London, UK). After washing, flow cytometric analysis was performed using a Navios flow cytometer (Beckman Coulter, Miami, FL, USA) and data were analyzed by using FlowJo software (Tree Star Inc., Ashland, OR; USA).
Different dynamics of immune reconstitution in children and adults after HSCT
sIL-7R and IL-7 were determined at different time points after HSCT in biological material from thirty-two adults and twenty-nine children (Table 1). The frequency of CD4+ cells (in CD3+ T-cells) between 1 and 12 months after HSCT was 30% for adults after aHSCT (Figure 1), the frequency of CD3+CD4+ T-cells was lower in children in months 1–3 (first month, p = 0.01; second month, p = 0.003; and third month, p = 0.01) after HSCT as compared to adult HSCT recipients. PBMCs from children also showed a not-significant trend of a lower frequency of CD3+CD8+ T-cells in PBMCs, compared to PBMCs from adults, at months 1–3 after HSCT (Figure 1, bottom panel) with a proportionate increase in CD3+CD4-CD8- T-cells in PBMCs (data not shown). PBMCs from children exhibited lower leukocyte counts than adults prior to transplantation (p = 0.02). There were no significant differences in proportion of CD8+ cells between PBMCs from children and before and after HSCT.
PBMCs from children showed a higher frequency of the CD127+ (IL-7R+) subpopulation in CD4+ and CD8+ T-cells as compared to adults (Figure 2). The frequency of CD127+CD3+ cells (total CD3+ T-cells) was lower at 6 months (p = 0.02) in adults as compared to children and the median for the CD127+CD4+ T-cell population at 1 month after HSCT was 76.5% as compared to 23.7% in adults (p < 0.05). A different situation was found to be true for the frequency of the CD127+CD8+ population, which was lower in PBMCs from adults at 2 months (p = 0.03), 3 (p = 0.02), 6 (p = 0.01), and at 12 months (p = 0.03) as compared to children.
Dynamics of sIL-7R and IL-7 in adults and children after HSCT
The median sIL-7R protein content in plasma from adults was 5.17 ng/mL at 1 month after HSCT, it increased to 8.72 ng/mL at 12 months. In plasma from children, the median sIL-7R protein concentration was 5.43 ng/mL at 1 month after HSCT and 7.46 ng/mL at 12 months (Figure 3). No significant differences in plasma levels of sIL-7R or IL-7 were found in samples from adults and children. Significant differences concerning the sIL-7R in plasma from adults were seen at 3 months relative to 1 month (p = 0.03), 3 months relative to 2 months (p = 0.04), 6 months relative to 2 months (p = 0.002) and 12 months relative to 2 months (p = 0.03). In plasma from children, the sIL-7R protein levels increased at 5–7 months (p = 0.02) and at 12–14 months (p = 0.03), relative to the first month after HSCT. The median sIL-7R values in plasma from controls (adults and children) were respectively 9.55 ng/mL and 95.05 ng/mL (see online Additional file 1: Figure S1, left panel).
In addition to the soluble IL-7R, we measured IL-7 protein values in plasma from adults and children after aHSCT. The median IL-7 in plasma from adults was 7.36 pg/mL at 1 month versus 6.50 pg/mL at 12 months after aHSCT. In plasma from children, the corresponding values were 6.85 pg/mL and 31.05 pg/mL, respectively (Figure 3). The median IL-7 plasma level for controls (adults) was 4.70 pg/ml and 5.81 pg/mL (for children; Additional file 1: Figure S1, left panel). Regression analysis showed no significant correlation between sIL-7R and IL-7 values in plasma from patients (Additional file 1: Figure S1, right panel).
Effect of donor age and conditioning on sIL-7R and IL-7 levels
Adults with stem cell donors above 30 years of age tend to have higher sIL-7R values at 2 months than to those receiving grafts from younger donors (p = 0.06). In contrast, adults with donors aged < 30 years had higher IL-7 levels at 6 months than those with older donors (p < 0.05, Additional file 2: Figure S2). Patients conditioned with RIC exhibited lower sIL-7R values 1 month after HSCT as compared to patients conditioned with MAC (p = 0.03, Additional file 2: Figure S2). There were no statistically significant differences between the patients who received RIC and MAC regarding IL-7 levels.
Effect of donor, HLA-match, ATG, and stem cell source on sIL-7R and IL-7 levels
sIL-7R values were higher at 2, 3 (p < 0.01) and 6 months (p < 0.001) if the transplant was from an HLA-identical sibling rather than a MUD (Figure 4). Plasma from recipients of HLA-mismatched grafts exhibited elevated sIL-7R value at 12 months as compared plasma from recipients of grafts from HLA-matched donors (p < 0.001) (Figure 4). sIL-7R values were significantly higher at 2 and 6 months after HSCT if the conditioning regimen did not contain ATG (p < 0.05) (Additional file 3: Figure S3).
In plasma from children, sIL-7R levels were associated with the stem cell source. Plasma from children who received bone marrow grafts rather than peripheral blood stem cell grafts showed increased sIL-7R levels at 3 months after HSCT (p = 0.04) (Additional file 3: Figure S3). Patients treated with donor lymphocyte infusion (DLI) had significantly higher sIL-7R levels 1 month after HSCT than those who were not treated with DLI (Additional file 3: Figure S3).
Association of sIL-7R and IL-7 with post-HSCT complications
We determined whether IL-7 or sIL-7R levels were associated with clinically and biologically relevant events in the course of HSCT in adults and children. Patients with acute GVHD of grades I–IV had lower levels of sIL-7R at 2 and 6 months after HSCT than those without any acute GVHD (Figure 5).
Plasma from patients with CMV infection exhibited lower sIL-7R values at 2 months after HSCT as compared to patients without CMV infection (p < 0.05). Plasma from patients with CMV infection showed higher IL-7 protein values at 2 and 3 months after HSCT than those without CMV infection (p = 0.02) (Figure 5). Plasma from patients with late-stage disease had higher IL-7 levels in the first month after transplantation as compared to patients with early disease (p < 0.05) (Figure 5).
Multivariate analysis
In multivariate analysis of factors affecting the levels of sIL-7R at one month after HSCT, we found that acute GVHD was the only factor that affected sIL-7R levels (Table 2). At 3 months after HSCT, there was a correlation between the absence of acute GVHD and the use of a sibling donor and higher levels of sIL-7R. At 3 and 6 months after transplantation, use of stem cells from sibling donors was the factor that was most strongly correlated to higher levels of sIL-7R. At 12 months after HSCT, there was a correlation between receipt of an HLA-mismatched graft and high levels of sIL-7R.
In multivariate analysis of factors affecting IL-7 levels, we found that late disease (beyond CR1) was associated with higher levels of IL-7 and that the use of bone marrow as the stem cell source was associated with lower levels of IL-7 at 1 month after HSCT. At 6 months after transplantation, the use of stem cells from a sibling donor was associated with higher levels of IL7 (Table 3).
The motivation for examining soluble IL-7R after HSCT was threefold. First, increased sIL-7R levels have been shown to be associated with an increased risk of developing autoimmune responses [18] and we hypothesized that altered levels of sIL7R may drive GVHD. Secondly, increased soluble IL-7R has been shown to bind free IL-7 and to inhibit IL-7 signaling (and therefore immune effector functions) in CD8+ T-cells from patients with infections [33]; inhibition of IL-7, via binding to soluble IL-7R, could potentially impact on GVHD development. Thirdly, increased sIL-7R has been associated with improved immune reconstitution and immune competence in patients with HIV infection [34]. All three biological scenarios, i.e. immune reconstitution, resistance to infection, and higher risk of developing autoimmune responses are clinically relevant after HSCT. Up to now, it has not yet been well defined which cell type or tissue is responsible for generating sIL-7R. The soluble IL-7R could be generated by shedding from cells or by splicing of the IL-7r associated with a polymorphism in the IL-7R gene (rs6897932) [18–20].
Recipients of grafts from HLA-identical siblings showed higher sIL-7R levels than recipients of grafts from URD (unrelated donors) (Figure 4). This difference was not only significant in the univariate analysis but also in the multivariate analysis at 2, 3, and 6 months and with a trend at 12 months after HSCT (Table 2). It is possible that minor histocompatibility antigens (mHags) contributed to immune reconstitution and increased sIL-7R levels after HSCT. Poor immune reconstitution, including low sIL-7R plasma levels, may be associated with increased risks for infection in recipients of grafts from URD as compared to recipients of HLA-identical sibling grafts [35].
Earlier studies showed a close correlation between CMV infection and GVHD [36, 37]. A significant finding in the univariate and the multivariate analysis in the current study was the decreased level of plasma sIL-7R in patients with any grade of acute GVHD (Figure 5, Table 2). Acute GVHD and also chronic GVHD have a profound effect on immune functions after HSCT [38, 39] including increased risk of CMV infection: In the univariate analysis, patients with CMV infection exhibited lower levels of sIL-7R at 2 months after HSCT as compared to patients without CMV infection. This is the timeframe when most patients experience CMV reactivation after HSCT. Immune responses to herpesviruses in general, and CMV infection in particular may trigger acute GVHD. Not mutually exclusive, GVHD, may also pave the way for CMV infection, which delays immune recovery and increases risk of infections [40, 41] after HSCT. sIL-7R and CMV infection was not significant in the multivariate analysis, which may suggest that GVHD is more important than CMV infection leading to decreased sIL-7R plasma levels.
We also found a tendency of lower sIL-7R levels in plasma from patients treated with ATG (Table 2). We treat all recipients of unrelated bone marrow grafts with ATG to prevent GVHD [42] at our center. In addition, patients with non-malignant disorders are also treated with ATG, since they do not benefit from GVHD. ATG has a prolonged effect on T-cell immune reconstitution and may therefore interfere with the generation of (soluble) IL-7R. The use of ATG also affects the rate of infections after HSCT in a dose-dependent fashion [28]. The data from the present study suggest that circulating T-cells (the numbers of which are reduced upon ATG treatment) contribute substantially to generation of soluble IL-7R generation, particularly since IL-7R expression is associated with T-cell maturation and differentiation [10].
Reduced levels of IL-7 protein were also identified in plasma from recipients of bone marrow as compared to patients who received peripheral blood stem cell transplants (Table 3). These two transplant types have different composite of the graft, which may be biologically relevant for IL-7 and soluble IL-7R production as well as IL-7 consumption. Blood cell grafts contain ten times more T-cells and NK-cells than bone marrow grafts [43, 44], supporting the notion that sIL-7R is produced from circulating immune cells. Decreased sIL-7R levels were also identified in plasma from recipients of grafts from unrelated donors as compared to recipients of grafts from HLA-identical siblings. Furthermore, there was a correlation between acute graft-versus-host disease—and to some extent also CMV infection after HSCT—and lower sIL-7R levels.
Three independent studies showed that elevated sIL-7R is associated with an increased risk of to develop autoimmunity, a situation which maybe at first glance counter-intuitive: since soluble IL-7R may bind free IL-7 and neutralizes its effects [16]. Two alternative mechanism, not mutally exlusive, could explain the increased risk of developing autoimmunity due to elevated sIL-7R. The sIL-7R/IL-7 complex may serve as a buffer system and will first neutralize free IL-7. Serum IL-7 levels (which are one tenth of sIl-7R levels) are tightly controlled [45]. Firstly, IL-7, complexed to the sIL-7R, could be released later from its (soluble) receptor and drive autoimmune responses. Secondly, sIL-7R/IL-7 complexes may be more potent in driving expansion of CD8+ T-cell subsets (in murine experiments) [46]. It could very well be that IL-7, complexed to sIL-7R, delivers a more potent signal to the cell-bound IL-7R, an event which would be even more accentuated in 'hypersensitive' autoreactive T-cells [33]. To summarize, the IL-7/sIL-7R complex represents a double-edged sword: free IL-7 supports immune reconstitution and promotes increased immune reactivity in infections [47, 48]; yet free IL-7 may also drive GHVD [3]. 'Neutralized' IL-7, by binding to the sIL-7R may not accessible to IL-7R-positive immune cells; this situation may be associated with increased risk of infections. Subsequently, IL-7, released from the sIL-7R, may be available to antigen-specific T-cells and ensure T-cell survival, which may also include immune cells mediating GVHD. Until now, the detailed dynamic of sIL-7R and IL-7 interaction in ex vivo collected clinical material has not been determined, yet a biologically and clinically relevant time frame to test for sIL-7R would be monthly within the first three month after HSCT; a time frame with a high risk to develop CMV infection and/or GVHD.
Our data suggest that lower sIL-7R may be associated with increased risk of GVHD, i.e. that sIL-7R is not available in suffient amounts to serve as the IL-7 'buffer system'. Measurement of sIL-7R plasma levels, in combination with IL-7, may aid to identify individuals at higher risk to GVHD and potentially CMV infection.
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The authors are grateful to the nurses at CAST for excellent help and advice, paricularly to Karin Fransson; we are indebted to our patients who agreed to participate in the study. This work was supported by Barncancerfonden, Sweden to IE, MM and O.R; KID funding to LR; VR to MM.
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Competing interests
The authors declare that there have no competing interests.
Authors' contributions
TP carried out analyses and performed statistics, processed patient samples and wrote the manuscript, LR carried out analyses and established the sIL-7R ELISA, MR performed statistical analyses, BO was involved in patient recruitment, clinical management and data interpretation; NKV was involved in sample procurement and analyses, RA performed flow cytometric analyses, IE was involved in study design, data analyses and writing the manuscript, JW and AGJ were responsible for children's care management, sample procurement, data analyses and interpretation, IM was responsible for flow cytometry, OR was involved in patient care and management, data analysis, study design and writing the manuscript, MM was responsible for the study design, data analyis and wrote the final version of the manuscript. All authors read and approved the final manuscript.
Electronic supplementary material
Additional file 1: Figure S1: Plasma IL-7 and soluble CD127 levels in healthy children (top) and adults (bottom). B No correlation between plasma IL-7 and IL-7R levels in children (top) and adults (bottom) after aHSCT. R2, goodness of fit. (TIFF 131 KB)
Additional file 2: Figure S2: Analysis of clinical endpoints and IL-7/soluble IL-7R after HSCT: increased IL-7 is associated with younger donor (<30y, 6months) and RIC (12 months after HSCT), there is a trend for increased sIL-7R with the age of the donor (>30y) and MAC. * = p < 0.05. (TIFF 504 KB)
Additional file 3: Figure S3: Plasma sIL-7R levels after aHSCT and analysis of clinical endpoints, effect of DLI (donor lymphocyte infusion) compared to no DLI; effects of sources: PBSC (peripheral blood stem cell) vs. BM (bone marrow) vs. CB (cord blood) and immunosuppression with ATG (antithymocyte globulin) or not. * = p < 0.05, ** = p < 0.01. (TIFF 416 KB)
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Poiret, T., Rane, L., Remberger, M. et al. Reduced plasma levels of soluble interleukin-7 receptor during graft-versus-host disease (GVHD) in children and adults. BMC Immunol 15, 25 (2014). https://doi.org/10.1186/1471-2172-15-25
DOI: https://doi.org/10.1186/1471-2172-15-25
- Interleukin-7
- Interleukin-7 receptor
- Immune reconstitution


text:
Volkswagen L1 Concept - Click above for high-res image gallery
There is no way that Volkswagen can prove their claim that the new 1-liter concept vehicle, the L1, is truly "the most fuel-efficient automobile in the world," but that isn't stopping them from using the phrase to describe the second version of a car that has been making the rounds since 2002. The new 1-liter car – so named because it burns 1.38 liters of diesel per 100 km (rounding down from 1.38 to 1 is acceptable, right?) – that will be on display in Frankfurt features a carbon fiber/plastic body and weighs just 380 kilograms.
The L1's CO 2 emissions are crazy low – just 36 grams per kilometer – from a hybrid TDI and 10 kW electric motor powerplant. The 0.8-liter TDI unit was derived from the 1.6 TDI that is making its debut in Frankfurt in the new Golf BlueMotion and Passat BlueMotion. Top speed is 100 mph. The tandem seats are accessible from a hinged roof cover. The L1 has two driving modes – eco and sport – and, of course, features Auto Start-Stop. What does all this get you? An astonishing 170 mpg (U.S.).
If the car looks a bit slippery to you, you're not alone. The L1 has a Cd 0.195. While this concept retains some of the fanciful design and joy of the 2002 model, VW is calling the L1 "revolutionary yet close to production readiness" and is talking about 2013 as a possible production date. Want one?
[Source: Volkswagen]
To the Point: Breakthrough for the 1-Litre Car
Volkswagen presents the most fuel-efficient automobile in the world
- L1 Concept is driven by new high-tech TDI and E-motor
- Carbon Fibre Reinforced Plastic body of the L1 weighs just 124 kilograms
- Maximum speed 160 km/h; L1 consumes just 1.38 litres diesel per 100 km
- CO2 emissions reduced to a new best value with just 36 g/km
Wolfsburg / Frankfurt, 14 September 2009 - Technological breakthrough for the 1-litre fuel consumption car: International Motor Show (IAA), Volkswagen is presenting the concept of the future L1 in a world premiere that points the way toward the production version. The full hybrid vehicle weighs just 380 kilograms thanks to its carbon-fibre reinforced body (CFRB). With a combined fuel consumption figure of 1.38 litres of diesel per 100 kilometres, this extremely aerodynamic (Cd 0.195!) Volkswagen suitable for everyday use is intended to become the most fuel-efficient automobile in the world. CO2 emissions of the 160 km/h L1 are similarly low at 36 g/km.
Retrospective: Seven years ago, Dr. Ferdinand Piëch, at that time Chairman of the Board of Management and today Chairman of the Supervisory Board of the Volkswagen Group, drove a prototype from Wolfsburg to Hamburg that was unlike any other car before it: the Volkswagen 1-Litre car – the world's first car with fuel consumption of one litre fuel per 100 kilometres. The man and machine wrote automotive history. In April 2002, however, it was also clear that the time for a production version of the 1-Litre car lay far in the future. Production of the body itself – from carbon-fibre reinforced plastic (CFRP) – was not realistic due to cost considerations. Yet, in 2002 Dr. Ferdinand Piëch already prophesised that the time would soon come for the 1-Litre car and CFRP as a material for industrial applications. By 2009 the time had come: Volkswagen is making a clear statement at the IAA in Frankfurt (September 17 to 27) with the second generation of this ingenious car: The L1 represents a step forward into the future with completely new technology and a new design; revolutionary yet close to production readiness.
"It is an enormous challenge to control costs in producing the monocoque out of CFRP," says Dr. Ulrich Hackenberg, member of the Board of Management for the Volkswagen Brand with responsibility for development. Both technically and visually, the CFRP body is already considered a significant achievement in car design. Unique on this car are the proportions of its dimensions. While the length of the L1 at 3,813 millimetres is still similar to that of a Volkswagen Fox, and its height of 1,143 millimetres nearly matches that of a Lamborghini Murciélago, the car's aerodynamically optimised width (1,200 millimetres) has no comparisons in the world of today's production cars.
L1 philosophy – defining a new type of automobile
In developing both prototype generations of the L1, Volkswagen simply questioned everything that typically characterised an automobile. The key starting point was body construction, and a core question was raised here: How would a car have to look and be built to consume as little energy as possible? The logical answer: extremely aerodynamic and lightweight. Yet these objectives had to be achieved under a non-negotiable precondition: a maximum of safety. The approach taken: a narrow two-seater with a CFRP body!
The seat layout fitting this design goal was dictated by the uncompromising aerodynamic form of a glider: One seat behind the other. Entry to the concept car is also similar to that of a glider; through a roof cover hinged at the side. On this second generation of the L1, the concept has been further honed; each component has been redesigned, a special chassis with aluminium components was developed, and above all the crucial CFRP technology from Formula-1 racing and airplane construction was transferred to automotive manufacturing. This has been combined with a unique form of hybrid drive to create a near-production vehicle. 2013 is the target year for this future dream to become a reality.
Note: TDI, TSI, DSG and Twincharger are registered trademarks of Volkswagen AG or other companies of the Volkswagen Group in Germany and other countries.
World Premiere of the L1 - Drivetrain
The Future Needs the TDI:
Small TDI leverages minimal fuel consumption and maximum range
Downsizing – 0.8 TDI is smallest diesel engine intended for production applications ever built by Volkswagen
Electrifying idea – E-motor plus TDI pushes CO2 emissions down to 36 g/km
Wolfsburg / Frankfurt, 14 September 2009
The TDI, E-motor and 7-speed DSG are located at the rear, and they combine to create the most fuel efficient road-legal car hybrid drive in the world. Proof of this are its 1.38 litre per 100 kilometres fuel consumption and 36 g/km CO2 emissions. Serving as the primary drive source is a completely redeveloped two-cylinder turbo-diesel with common rail direct injection (TDI). It is operated in two different modes depending on the load conditions. In the standard "ECO" mode, the 800 cm3 TDI develops a power of 20 kW / 27 PS (at 4,000 rpm); in "Sport" mode – used to reach top speed, for example – the car's power rises to 29 kW / 39 PS (at 4,000 rpm). The TDI's maximum torque is 100 Newton-meter (at 1,900 rpm). Naturally, the L1 also has a Stop-Start system that automatically shuts down the engine when vehicle has stopped and restarts when the accelerator or E-pedal is pressed.
The hybrid module has been integrated into the housing of the 7-speed DSG (Direct Shift Gearbox). It is located between the TDI engine and the DSG gearbox and consists of a 10 kW / 14 PS electric motor and a clutch. The E-motor is supplied with energy from a lithium-ion battery located at the front of the car. An electronic power control module, operating at around. 130 Volts manages the flow of high voltage energy the battery and to the E-motor. In parallel, the vehicle's low voltage electrical system is supplied with the necessary 12 Volts through a DC/DC converter.
Electric motor – details of the E-motor
In normal operation the electric motor can support the TDI engine in conditions such as by electronic load point shifting and in acceleration. If necessary – generally during acceleration – the E-motor can supply 40 percent additional torque over the entire speed engine speed range. Moreover, the E-motor can propel the L1 over short distances by itself. In this case, an auxiliary clutch decouples the TDI from the drivetrain. Restarting the TDI is a very easy process. In so-called "pulse starting" of the TDI, the electric motor is sped up and is then coupled to the TDI unit to provide almost instant starting. The entire process takes place automatically and without jolts, so the driver hardly notices the restarting of the TDI engine.
In braking phases, the E-motor operates as a generator to charge the lithium-ion battery by recovering braking energy. The gears of the automatically shifting DSG are always selected with the aim of achieving the best possible fuel economy. The engine controller regulates all energy flow and drive management tasks taking into account the moment by moment demands for power made by the driver. Some of the parameters used to calculate the optimum propulsion mode for the given conditions are: accelerator pedal position, engine load, momentary fuel demand, energy supply and the mix of kinetic and electrical energy at any given time.
Diesel engine – details of the 0.8 TDI
The TDI engine in the L1 is a new development. Yet, even here Volkswagen has been able to exploit synergies to design an engine that is both innovative and cost-effective. Hence, this 0.8 litre TDI unit has been derived from the 1.6 TDI just introduced a few months ago. The 1.6 TDI is making its debut at the IAA in cars such as the new version of the Golf BlueMotion (3.8 l/100 km) and the Passat BlueMotion (4.4 l/100 km) – which are currently the world's most fuel-efficient production cars in their respective classes.
Based on their common origins, the 0.8 TDI and 1.6 TDI have identical cylinder spacing (88 millimetres), bore (79.5 millimetres) and stroke (80.5 millimetres). These high-tech TDI engines also share key internal engine features for reducing emissions. They include special piston crowns, multi-injection and individual orientations of the specific injection jets. On both drivetrains there is exhaust gas recirculation, an oxidation catalytic converter and a diesel particulate filter. Equipped this way, the TDIs in each Volkswagen fulfil the limits of the Euro-5 emissions standard with ease.
The 1.6 TDI, thanks to its common rail injection, is also an exceptionally quiet and low-vibration diesel engine. These positive properties have been successfully transferred to the two-cylinder unit. The TDI's aluminium crankcase was also constructed with high precision to achieve very low friction losses. The oil pump, designed to operate at a maximum oil pressure of 4.0 bar, also contributes to engine efficiency.
Another example of how the entire drive system is configured for high efficiency is the L1's cooling system. Its external water pump is controlled by engine management so that cooling is only activated while engine operating conditions require it. This thermal management also contributes to reduced fuel consumption. A second electric water pump, also activated only when needed, provides cooling required for the starter generator and the power electronics in a separate water circulation loop operating at a lower temperature level.
Automatic transmission – details of the 7-speed DSG
Gear shifting work aboard the L1 is handled by the 7-speed DSG, which is one of the most innovative automatic transmissions in production. Compared to the version equipping the new Polo, for example, the design of the Direct Shift Gearbox has been developed to include clutch control for the hybrid module. Furthermore, individual gear ratios have been optimised to attain responsive driving performance despite the car's extremely low fuel consumption. The hybrid module is integrated into the DSG housing as previously mentioned. It is located where the flywheel is usually to be found.
Driving performance – economical and yet responsive
The L1, equipped with ABS and ESP, has a top speed of up to 160 km/h – this is remarkable considering its fuel efficiency. With maximum acceleration from a standstill, the two-seater reaches 100 km/h after just 14.3 seconds. The fuel tank holds just ten litres yet, this is sufficient for a theoretical driving range of about 670 kilometres, given the car's 1.38 litre average fuel consumption.
World Premiere of the L1 - Interior
The World's Most Fuel-Efficient Car with a Jet's Cockpit:
180-degree cockpit offers ideal ergonomics
Tailored to the driver – Steering wheel is the centre point of the interior
Comfortable for two – Seats offer fully-fledged long-distance comfort
Wolfsburg / Frankfurt, 14 September 2009
Talking about car driving as 'piloting' might sound out of place, but in the case of the L1 it is wholly appropriate. The driver (in the CFRP tube frame seat) and passenger (in the fixed CFRP seat that is part of the monocoque) sit one behind the other. At both locations, the seat position is ergonomical and very comfortable. All instruments and controls are arranged over a 180 degree radius for the driver, which places them perfectly within view and reach. The instrument panel itself has been integrated into the monocoque and is made of CFRP. The interior applications are produced from glass fibre reinforced plastic (GFRP). One of the materials Volkswagen is introducing for interior side trim is the new "Sport Esteem" material that is as robust as it is touch-friendly.
To start the L1, the driver pushes a button on the right side of the steering wheel. When it is rotated, the round start button simultaneously serves as a gear selector switch and is used to activate the electronic handbrake (Drive, Neutral, Reverse and Park). The entry canopy and rear hatch are opened electrically by touch controls to the left and right of the driver. Also designed as touch sensors are controls for the entire air conditioning control system. Via multifunctional keys in the steering wheel, the driver controls the on-board computer, navigation and entertainment systems.
The classic door mirror and rear-view mirror have been completely eliminated on the L1. In their place, cameras display images on OLED-active (OLED = organic light emitting diode) displays located on the left and right sides of the instrument panel. A Park Distance Control (PDC) system makes parking easier as well.
In case of a crash, not only are the driver and passenger protected by the monocoque, which is designed as a highly rigid CFRP safety cell, as well as aluminium crash elements in the front of the car, but also by a steering wheel airbag and head/side curtain airbags to the left and right inside the entry canopy.
World Premiere of the L1 - Body
Car Design of the Future:
The L1's exceedingly safe CFRP safety cell weighs just 124 kilograms
Lightweight – L1 is no heavier than a good touring motorcycle
Revolution – Never before in carmaking history has CFRP been applied so comprehensively to improve fuel efficiency
Wolfsburg / Frankfurt, 14 September 2009
Both technically and visually, the CFRP body is already a significant achievement in car design. Unique on this car: the proportions of its dimensions. While the length of the L1 at 3,813 millimetres is still similar to that of a Volkswagen Fox, and its height of 1,143 millimetres nearly matches that of a Lamborghini Murciélago, the car's aerodynamically optimised width (1,200 millimetres) has no comparisons in the world of today's production cars.
CFRP body – Monocoque and exterior skin
The two-seat monocoque, including the tubular frame driver's seat and passenger seat as well as the exterior body skin, all consist of CFRP. There are no doors. Instead, the driver and passenger climb into the L1 from the top. An electrically actuated entry canopy above the seats is opened and closed for this purpose. Headlights and taillights all utilize LED technology, which consume a lot less energy. The rear wheels are completely covered; their wheel covers can be removed to change the Michelin low resistance tires ("Energy Saver": front 95/60 R16, rear 115/70 R16). The underbody is also completely enclosed. The 0.8 TDI is cooled via adaptive air channels integrated in the sides of the car body. These automatically open and close based on the hybrid unit's operating state and vehicle speed. The tailgate is opened in the usual, manual way. It too consists of CFRP. Inside is a stowage space of 50 litres.
CFRP advantages – composition and weight
Carbon fibre reinforced plastic, as the name implies, consists of multiple layers of high-strength carbon fibres, which are integrated in a very tough matrix. This mix results in an extremely strong and lightweight composite material. Until now, producing a body like that of the L1 from CFRP, while conforming to industrial standards, was an insurmountable task. Up to now CFRP was only practical for very small production runs, as in aircraft manufacturing or motorsport. Now Volkswagen has succeeded in finding a production-viable and cost-effective way to produce CFRP parts in suitable volumes.
The reason that CFRP is the ideal material for the L1 body is demonstrated by considering its weight and strength. The L1 weighs just 380 kilograms, which is equivalent to the weight of a very high-end, fully equipped touring motorcycle of the 1200-cc class. The L1, on the other hand, is an automobile through and through. Of the 380 kilograms curb weight, 122 kilograms are taken by the drivetrain, 79 kilograms by the chassis, 35 kilograms by interior furnishings and 20 kilograms by the electrical system. 124 kilograms remain, and this is precisely the weight of the body.
These 124 kilograms can be further broken down: 64 kilograms are accounted for by the CFRP monocoque including integrated passenger seat, 28 kilograms is the weight of the entire CFRP exterior skin, 19 kilograms for the CFRP entry canopy, 9 kilograms for the CFRP driver's seat and 4 kilograms for the LED lights. By way of comparison: The body of the legendary Lupo 3L – until today the smallest Volkswagen production car ever built – weighed 306 kilograms, and the entire car weighed a lightweight 813 kilograms. That is 433 kilograms more than the L1.
And there are other advantages: the material's extremely high stress limits and its ideal forming properties for even the most challenging of design features.
Design – anything but typical, yet a typical Volkswagen
The design and styling of the L1 – function and form – combine to form one uncompromising unit. Walter de Silva, Head of Design for the Volkswagen Group has this to say: "The design of the L1 redefines classic and aesthetic vehicle traits. Especially significant, of course, is how the nearly rocket-shaped lines catch one's attention. All of its moving parts are integrated so accurately that the body resembles a rocket or jet. It is a body that cuts through the air with minimal aerodynamic resistance." A top Cd value of 0.195 and 1.02 m2 frontal area (Cd x A = 0.199 m2) is a statement that is sculpted into the CFRP.
And this is how it looks, the most aerodynamic front end in the world: "The typical layout of conventional headlights with a radiator grille in the middle would be entirely inappropriate here," says de Silva. "That is why we chose a more minimalistic layout and integrated the headlights into a neutral horizontal stripe that conveys a far-sighted and contemporary feeling." The Head of Group Design continues: "In a sense, we applied the same principle to the front end of the Scirocco, where the headlights are joined by a glossy black stripe, and the brand logo is also placed on the engine bonnet. This underscores the dynamic character of this automobile."
The extremely aerodynamic design also shapes the rear with its diffuser and wheels that are completely enclosed. The most distinctive feature of the overall appearance in the rear are the LED taillights that were worked into the TDI's air outlet ports. De Silva: "The same stylistic features as in front are repeated in the rear, in the taillights – which we have integrated in the grilles of the two air outlet ports; they have an even more aerodynamic appearance. Another identifying feature is the air outlet directly behind the cockpit – a nearly abstract, graphic element that underscores the purposeful aesthetics of this vehicle." Due to the driver's low seating position, there is an additional window in the roof that is ideal, for example, to view traffic lights.

subset: commoncrawl


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Resolving Shortfall Debt
We are seeing an increasing number of borrowers being contacted by UK Solicitors collecting outstanding debts for Overseas lenders. Using various treaty's once a judgement is gained in the country the property is located & the lender can collect in the borrower's home state. You will be contacted confirming the balance owed.
At this stage a lump sum settlement will be required, the lender will have paid legal costs for which you are liable. This will not go away.
EU Property Solutions can act using the backdrop of our in depth knowledge of processes in the home country to counter act arguments put forward by UK solicitors. We can protect your Assets, Credit Files and ensure legal action is ceased. Our settlements typically range from 10%-20% of the outstanding shortfall balance.
We read many online forums and see individuals commenting that they have handed the keys back to the Bank and walked away. This is false, do not bury your head in the sand. There is always a solution.
If ignored we have seen Overseas Lenders appoint UK Legal Representation to submit County Court Claims, Issue Writs, Place Charges on UK assets and Place Defaults on UK Credit Files.


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In conjunction with the SoAR Foundation, The National Academies has officially announced the Breakthroughs 2030 study to produce a 10-year agenda for food and agriculture research. Nominations for the study's lead committee are due by March 22. Please contact Tim Fink (email@example.com or 571-257-7625) with any questions.
For more info, see: http://bit.ly/2n3MijK


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Review: The Best Minds of My Generation
There are so many books about the Beat Generation that focus on the writers' roles as rebels and...Read More
by David S. Wills | Dec 30, 2016 | Reviews | 1 |
There are so many books about the Beat Generation that focus on the writers' roles as rebels and...Read More
by Richard Kostelanetz | Oct 14, 2015 | Beatdom #16, Beatdom Content, Essays | 1 |
Not many prose writers alive (Céline, Genet, a few others) would dare the freedom and intelligence to trust their own minds, remember they made that jump, not censor it but write it down and discover its beauty. That's what I...Read More
by GK Stritch | Dec 6, 2013 | Memoirs, Fiction & Poetry | 0 |
Hot chocolate . . . delicious Blustery deep freeze winds Cut through cruel canyons of Manhattan...Read More
by Nick Meador | Oct 24, 2011 | Beatdom #9, Beatdom Content, Essays | 2 |
At the turn of the 1960s, Jack Kerouac found himself in a profound state of limbo, the climax of...Read More
by David S. Wills | Jul 10, 2009 | Beatdom #4, Beatdom Content, Essays | 0 |
by David S. Wills In Issue Two of Beatdom, we ran a story about the women of the Beat Generation, and we obviously talked a little about Joan Vollmer. However, we didn't say enough to do her justice, for she was a fascinating...Read More
by David S. Wills | Feb 8, 2009 | Beatdom #3, Beatdom Content, Essays | 0 |
by Adi Rajkovic There have been many pivotal experiences and events that have influenced my vision as an artist, but the most arresting event (historically speaking) has been the Beat Generation. Although short lived and long...Read More


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Join Vancouver magazine on November 9th, 2017 to experience a Hawaiian culinary journey by exploring the culture, adventure, and cuisine of Maui.
On Thursday, November 9th, guests will sample delectable bites from Kā'anapali chefs but that's not all! Each guest at our pop-up party will be entered to win round trip tickets for two to the Kā'anapali Beach Resort in Maui (with a unique stay of 2 nights at Sheraton Maui Resort & Spa plus dinner at Black Rock Kitchen and 2 nights at The Westin Nanea Ocean Villas plus dinner at Mauka Makai).
How to Enter:
For your chance to win tickets to this invite-only event, do one of the following:
- Complete the entry form below.
- Post to Facebook, Twitter or Instagram with #CraveKaanapali telling us why you'd love to experience Maui


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Pieces of Boston's colonial history are now being uncovered right in the heart of downtown. On Sept. 7, the National Park Service and the City of Boston opened an archaeological excavation site next to historical Faneuil Hall, digging up artifacts from as long ago as the 17th century.
The dig is making way for the new visitors center expansion that will include enhanced exhibits and information for National Park and Freedom Trail visitors, as well as new restrooms, vendor stalls and handicap access, according to Sean Hennessey, a National Park Service spokesman.
Hennessey said the National Park Service wants to give visitors a great welcoming experience so they can better learn about Boston's vibrant past, he said.
Since Faneuil Hall is part of the Boston National Historical Park, the National Historic Preservation Act requires that the excavation be completed before an expansion of the visitors center can be constructed, he said.
"I think it's a great law because we can uncover untold stories," he said.
"[Diggers are] continually uncovering artifacts that contribute to an evolving understanding of history," he said.
Hennessey said that the excavation requires "digging, sifting, sorting and documenting in a very, very careful way."
The digging process in the 15-by-15 foot test pit involves both mechanical and manual digging, he said.
"You excavate to dig up the top layers literally with a shovel," he said. "The deeper you go, you start to hit a water table, and then the hole fills with water that you need to pump out."
The dig is expected to conclude within the next few weeks, but the archaeologists, contractors and University of Massachusetts-Boston graduate students working on the project have already uncovered at least 2,000 objects, he said.
Items found so far include clay stems from smoking tobacco pipes, chicken bones in a trash pit, bits of pottery and wig curlers, he said. Excavators send all artifacts to the Boston Archaeology Lab for further study and preservation, he said.
In addition to searching for historical objects, archaeologists and students are also analyzing the soil in the contexts of entomology and palynology, according to City Archaeologist Ellen Berkland. Excavators will take geochemical and botanical samples to acquire information about how the site changed overtime, she said.
"It's different in an urban area because you're dealing with multiple layers of fill and types of soil," she said. "You have to maintain complete control of a site in order to recreate it."
Faneuil Hall's colonial history is rich with revolutionary events that make up the story of the founding of the U.S., according to Hennessey.
The Hall was built on top of a landfill that covers what used to be the old Town Dock, which served as Boston's port, center of commerce and city meeting place in the 17th and 18th centuries, he said.
The original Faneuil Hall was built in 1742, and was expanded in 1806 by renowned Boston architect Charles Bulfinch, who also designed the statehouse, according to the City of Boston website. It is one of 16 sites on the famed Freedom Trail.
Some of the country's most celebrated orators found their platform at Faneuil Hall, including Samuel Adams, who organized efforts to protect colonial rights in what would later become the Revolutionary War, the website said.
Today, the first floor still serves as a marketplace, as it has for hundreds of years, and the second floor meeting room is still used for debates, the website said.
Although archaeologists are not looking for any specific artifact and don't expect to find anything especially groundbreaking in the current dig, they hope that the items they do find will provide clues for a fuller picture of what life was really like in colonial Boston, Hennessey said.
In a similar excavation during the restoration of Revolutionary War stronghold Dorchester Heights in the mid-1990s, archaeologists uncovered significant evidence of an old fort from the 1770s, he said.
Because the Town Dock area, and later Faneuil Hall, has been a central meeting place since the very birth of Boston, archaeologists hope that a dig in this location could prove to be particularly fruitful, he said.
Hennessey said researchers are also hoping that the dig might shed some light on some of the lesser known, more shameful parts of Boston's history that escape wider public attention, such as Boston's burgeoning slave trade in the early 1700s.
Colonists held slave auctions right next to where Faneuil Hall was built, he said.
Ironically, Peter Faneuil, the wealthy merchant and benefactor of Faneuil Hall, was involved in the slave trade, he said. The "Cradle of Liberty," as the Hall is often called due to the revolutionary activities that took place there, was actually built from funds derived from the slave trade, he said.
Hennessey said it is important for people to understand the relevance of the Faneuil Hall excavation.
"I hope [the dig] increases awareness of the important science of archaeology," he said. "It allows us to preserve and interpret what lies beneath our feet, and to understand who we are and where we came from."
"This is a story that needs to be told, and archaeology can make a vital contribution," he said. "It humanizes Boston's history."
College of Communication sophomore Mike Barry said he agrees with archaeologists that the dig has great cultural significance.
"Part of why Boston is so great is its history, it's unique among American cities," he said.
College of General Studies sophomore Kimberly Hammett said she believes the dig is especially relevant to BU students because of the close proximity of Faneuil Hall.
After hearing about the excavation, many BU students said they were intrigued by the historical possibilities right outside their door.
"It's really interesting," said Katherine Keegan, a College of Arts and Sciences sophomore.


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Their musical reunion at the Olympics was only temporary, every Spice Girl in search of an interview is telling us. So where next for the peppery pentahedron? Here are some ideas…
The concept: Ambition-themed gameshow in which the Girls audition potential replacements from a crowd of needy schoolchildren
Emma: And now it's Kayleigh from Swindon's turn! Kayleigh, which Spice Girl do you… "wannabe"?
Small child: I wannabe Geri!
Geri: Kayleigh, you're aware that to become the perfect simulacrum, your established interiority is forfeit? And that you may pass from physical existence into a theoretical state of hyperreality?
Small child: I like the dress with the flag on it! Girl Power!
The concept: Mel B oversees proceedings as the remaining girls pair up – quite literally! – to live as Siamese twins for a week. In the grand finale the winning pair undergo a full and irreversible surgical joining procedure.
Mel B: (bursting into room in which Emma and Victoria are sat glumly in a giant three-legged jumpsuit) Hi girls! How are you getting on?
Emma: (wriggling) I'm worried about what'll happen if we win the challenge and have to live like this permanently.
Victoria: David says he's alright with it. And I could get a new fashion line out of it.
Emma: (viciously) Stop fooling yourself Victoria!
Mel B: Chin up, ladies! It's nearly time for the breakdancing challenge!
The concept: Undeterred that a programme of the same name already exists, the Girls present a fun exploration into the concepts of identity and consciousness.
Mel C: So you see Mel, the self is a narrative we create for ourselves, but an intangible one. Look at this hula hoop! Its centre of gravity is a point in the middle of thin air!
Mel B: Are we going to do hula hooping now?
Mel C: Well YOU'RE not, Mel! Being sporty is MY narrative!
Mel B: You're really starting to episteme off.
The concept: Each Spice Girl tries out a different method of attaining immortality. Hosted by Esther Rantzen.
Esther: Well I think we can all agree Geri's quietened down A LOT since we put her into cryogenic suspension. We'll be checking back in ten years to see how she's getting on.
Emma: How much more of this disgusting black fungus water do I have to drink, Esther? It tastes like the bottom of a carpet. And I've got terrible shits.
Esther: It's Ling-Zhi, the Chinese mushroom of immortality, Emma – keep going and don't be a baby! Victoria, how are you getting on uploading your intelligence onto that floppy disk?
Victoria: I've written my name on the label already!
The concept: The Spice Girls call for help repeatedly from the centre of an existential void. But no-one ever comes.


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Our Purpose
To form successful work relationships.
Our Mission Statement
Work is an essential necessity for our survival, but it can also be an experience that is fulfilling and meaningful. Our jobs also define us as individuals and impact every aspect of our daily lives.
For many of us, the tasks required to find the job we want can be daunting and unattainable, and we often end up settling for a work experience that is unsuccessful for ourselves and our employers. We believe that the right recruitment process can play an integral role in attaining (1) the jobs we really deserve, and (2) the positive impact we want to make with our employer. To make sure this happens, Matura Farrington is committed to truly understand the needs, values, and goals of those we serve and matching employers with employees to form successful work relationships.
Our Ethics, Values and Philosophy
Matura Farrington Staffing Services consists of ethical professionals who take pride in their work. As a Company, we achieve professional satisfaction from maintaining a set of standards that have been defined by us, and the leading association in our industry, The American Staffing Association. By maintaining these standards, we earn our clients' trust.
Our clients are comprised of Los Angeles' top law firms and corporations who demand of their staffing firms a set of ethics, standards, and values that are similar to their own. These include; (1) delivering the services promised in the initial stages, (2) providing accurate and reliable information requested by a client, (3) rectifying problems to the satisfaction of the client, and (4) standing by a client to meet their overall objectives in staffing – from the most basic of direct hire placements to the most demanding long-term, high volume temporary projects.
Our clients consistently tell us that we understand their needs and deliver the service they expect. We do not put our emphasis on marketing hype and promises. Our job is to match employers with employees to form successful work relationships. This can only be accomplished through accurately assessing and considering the needs of the parties involved and finding the balance. This sensitivity to the placement process is key to our success. It is an uncompromising commitment to the people and companies we represent.


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03 June 2013
"muted" by Jessica Bell
Long story short, "muted" is, as the cover says, a short story in verse. Set in a dystopian future in which self-expression through art and dress and (as concerns the main character) music is strictly controlled, the story traces the crisis of Concetta, a singer who has ended up on the wrong side of the law.
"muted" is richly imagined. Jessica doesn't hold back on gory detail - something I always admire. As a science fiction piece, the story has a lot going for it, opening a world's worth of weird and terrible in its short length. The main thing here is the language, though. This is poetry, written by a musician, about a musician. As a whole, "muted" is about art and sacrifice. If you are looking for something that takes sci fi lyricism to its logical extreme, you could do worse. I liked it.
8 comments:
- Michael Offutt, Phantom Reader said...
Damn. That cover reaches out, grabs me, slams me against the computer, and says buy me. I shall follow my instincts. Besides, I like to support Jessica who was kind enough to ask me to contribute an essay to her upcoming book.
- June 3, 2013 at 12:23 PM
- Elizabeth Twist said...
It's awesome, right? I mean to ask her who designed it. If she did, well, holy heck. Some people are just multi-talented.
- June 3, 2013 at 12:25 PM
- Mina Lobo said...
I agree that it's a stunning cover. Have checked out the blurb and it kinda twisted my gut a bit, as a currently non-singing singer. But I'm trying to stretch myself as a reader and so, with your recommendation, will give it a shot. :-)
Some Dark Romantic
- June 3, 2013 at 7:07 PM
- Emily Genther said...
Sounds interesting.
- June 3, 2013 at 9:45 PM
- Jessica Bell said...
Thank you so much, elizabeth!!! Wow, so glad you liked it. :-) Yes, I did design the cover myself. I absolutely love to do it. And surprisingly, it's very calming.
Michael, you're awesome. Thank you!
And thanks for your kind comments, Mina, and Emily!
- June 4, 2013 at 4:02 AM
- Catherine Stine said...
I was involved in the conversation when Jessica was figuring out the colors, so this is especially cool. Who can resist a good short piece involving music and suspense?!
- June 4, 2013 at 11:14 AM
- Elizabeth Twist said...
For those who picked it up, I hope you like it! Jessica, I am impressed by that cover. Beautiful work.
- June 4, 2013 at 10:48 PM
- Amelia Davis said...
Really inspiring stuff!!! This post is fantastic, and equally fantastic is this conversation thread. This information regarding printers and different category of printers & its machinary is so inspiring! Awesome Work. Keep it Up!
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- August 6, 2013 at 5:49 AM


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This is somewhat embarrassing, isn't it? It looks like nothing was found at this location. Maybe try a search?
Dear Valued Customers,
Greetings and Welcome!
As our country enters a period of economic uncertainty, we at district25 remain committed to serving our customers with the highest standards.
With this in mind, we are experiencing order fulfillment delays, but are working with our delivery partners in order to ensure your orders are delivered as soon as possible.
Our team continues to work effortlessly to provide you the highest level of service.
We thank you for your loyalty and continued support.
Happy shopping,
Team D25 "


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County OKs Stagecoach Community Center lease agreement
Although there was an email submitted to county officials expressing concerns, the Lyon County Board of Commissioners voted unanimously May 15 to lease the Stagecoach Community Center to Stagecoach Bingo, a Nevada nonprofit.
The lease agreement is similar to leases to nonprofit groups for the Dayton Community Center and Mason Town Hall, with a lease term of $1 per year.
Lyon County Public Works director Mike Workman said the lease arrangements had been pursued as part of budget and staff cuts the county has faced in recent years, to where the county's facilities division had trouble properly maintaining and overseeing various county facilities.
Stagecoach resident Margot Easley expressed several concerns about the proposed lease agreement in an email to the county but during the meeting she only spoke about concerns whether a 4-H group in Stagecoach would still be able to use the building as it has in the past.
Commissioner Bob Hastings said that was outlined in the lease agreement, which takes effect June 1, and they would still be able to continue that use.
Easley noted the stove isn't now used for fire reasons but said Stagecoach 4-H was working with Silver Stage CoOp about obtaining a grant to upgrade the community center's kitchen.
Commission Vida Keller said she had received a number of calls and emails about the kitchen situation.
Workman said in a separate interview, "It's a really, really good opportunity for the community to come together and make that a much more useable facility."
He said the county's goal is to increase usage of the facility and the lessees also have that goal to expand the use. He said the group that leased the Dayton Community Center reported it had increase usage of that facility.
Workman said Stagecoach Bingo has been operating under an agreement for several years to provide custodial service in exchange for rent for the facility located at 8105 U.S. 50 West in the eastern portion of Stagecoach.
Penny Slattery, Stagecoach Advisory Board chairman, also favored this action.
"I want to let the commission know that as far as I'm concerned, this is really a hallmark moment" for the community, Slattery said.
She said the situation isn't perfect but felt it was a good idea and thanked those involved with reaching that moment, especially Shawn Keating, who was in charge of facilities in that area of the county.
The lease notes Lyon County only uses a small portion of the building for minor storage, yearly voting and monthly meetings of the Stagecoach Advisory Board.
It continues later that Stagecoach Bingo "has agreed to operate and provide the routine maintenance" for the center, which includes the former fire station bays, "and also to expand the uses of the facility for the community of Stagecoach."
Workman said initially Lyon County had asked the Stagecoach General Improvement District, which operates the community water system in Stagecoach, if it wanted to take over ownership of the building, but the reply was no due to the fact the GID is an enterprise fund.
The lease is for one-year but it includes a provision that the lease would automatically renew for three additional one-year periods, on the same terms, unless terminated or modified by written agreement of both parties.
If the lessee changes its status from a nonprofit corporation, the lessee would be responsible for rent at fair market value as determined by the Lyon County assessor.
Workman said the county and lessee would meet yearly to see if any modifications were needed in the agreement.
Regarding use, the lease states, "It is Lessor's intention that the Stagecoach Advisory Board or other county hosted function, 4H, Central Lyon County Youth Connection and Healthy Communities Coalition continue to provide services from that location and that those organizations pay no rent but will be required to schedule events through the Lessee except" for the Stagecoach Advisory Board, "which holds their meetings on the first Wednesday of each month."
Otherwise it says organizations would be subject to lessee's rental and cleanup agreement and cleaning deposits and/or fees, and that Lyon County would have priority for use of the building for voting but it would give advanced notice of at least 30 days.
Lyon County would be responsible for maintaining the property in "good, safe and substantial condition," including heating, electrical, air-conditioning, etc., while Stagecoach Bingo would be responsible for cleaning inside and outside the building and "normal day-to-day maintenance and upkeep."
The lessee could make alterations and improvements to the premises as long as they didn't weaken or impair the structural strength or lessen the value of the building, the agreement states, but it must inform the county of the plans and gain written consent beforehand.
Lyon County, per the agreement, would be responsible for all utility costs related to water, basic garbage and electric and for propane costs up to $6,000 per fiscal year, with Stagecoach Bingo responsible for all other utility costs.
Lyon County would maintain property and fire insurance on the property, although the lessor would also maintain commercial general liability insurance in the amount not less than $1 million and fire insurance on the contest owned by the lessee at the premises.


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- Decade (Neil Young album)
Decade Greatest hits album by Neil Young Released October 28, 1977 Recorded 1966-1976 Genre Country rock, folk rock, rock Length 143:40 Label Reprise Producer Neil Young, Elliot Mazer, Tim Mulligan, David Briggs Neil Young chronology American Stars 'N Bars
Comes a Time
Decade is a triple album compilation by Neil Young, released in 1977, now available on two compact discs. It contains 35 of Young's songs recorded between 1966 and 1976, among them five tracks that had been unreleased up to that point. It peaked at #43 on the Billboard Top Pop Albums chart.
Compiled by Young himself, with his hand-written notes about each track, Decade represents almost every album from his career and various affiliations through 1977 with the exception of Four Way Street and Time Fades Away. Of the previously unreleased songs, "Down to the Wire" features the New Orleans pianist Dr. John with Buffalo Springfield on an item from their shelved Stampede album; "Love Is a Rose" was a minor hit for Linda Ronstadt in 1975; "Winterlong" received a cover by Pixies on the Neil Young tribute album from 1989, The Bridge; and "Campaigner" is a Young song critical of Richard Nixon. The track "Long May You Run" is a different mix to that found on the album of the same name, featuring the harmonies of the full Crosby Stills & Nash before David Crosby and Graham Nash left the recording sessions.
For many years, Decade was the only Neil Young compilation album available. A 1993 compilation called Lucky Thirteen was released, but it only covered Young's 1982–1988 output. It was not until 2004 that Reprise Records released a single-disc retrospective of his best-known tracks, titled Greatest Hits. Throughout the 1980s and '90s, Young promised fans a follow-up to the original Decade collection, provisionally titled Decade II; eventually, this idea was scrapped in favor of a much more comprehensive anthology to be titled Archives, spanning his entire career and ranging in size from a box set to an entire series of audio and/or video releases. The first release of archival material since Decade and Lucky Thirteen would appear in 2006, Live at the Fillmore East, a recording from a 1970 concert featuring Crazy Horse with Danny Whitten. Several other archival live releases followed, and in 2009 the first of several planned multi-disc box sets, The Archives Vol. 1 1963–1972, was issued.
Alternate early version
Initially, Decade was to be released in 1976, but was pulled at the last minute by Young. It was shelved until the following year, where it appeared with two songs removed from the original tracklist (a live version of "Don't Cry No Tears" recorded in Japan in 1976, and a live version of "Pushed it Over The End" recorded in 1974). Also removed were the following comments on those two songs and Time Fades Away, from Young's handwritten liner notes[1]:
Time Fades Away. No songs from this album are included here. It was recorded on my biggest tour ever, 65 shows in 90 days. Money hassles among everyone concerned ruined this tour and record for me but I released it anyway so you folks could see what could happen if you lose it for a while. I was becoming more interested in an audio verite approach than satistfying [sic] the public demands for a repetition of Harvest.
Don't Cry No Tears. Initially titled 'I Wonder,' this song was written in 1964. One of my first songs. This is a live recording from Japan with Crazy Horse.
Pushed It over the End. Recorded live on the road in Chicago, 1974. Thanks to Crosby & Nash's help on the overdubbed chorus, I was able to complete this work. I wrote it for Patty Hearst and her countless brothers and sisters. Also, I wrote it for myself and the increasing distance between me and you.
Professional ratings Review scores Source Rating Allmusic [2] Robert Christgau (A)[3]
The album has been lauded in many quarters as one of the best examples of a career retrospective for a rock artist, and as a template for the box set collections that would follow in the 1980s and beyond. However, in the original article on Young from the first edition of the Rolling Stone Illustrated History of Rock and Roll and a subsequent article in the 1983 Rolling Stone Record Guide, critic Dave Marsh used this album to accuse Young of deliberately manufacturing a self-mythology, arguing that while his highlights could be seen to place him on a level with other artists from his generation like Bob Dylan or The Beatles, the particulars of his catalogue did not bear this out. The magazine has since excised the article from subsequent editions of the Illustrated History book; a transcription of it can be found at the link below (despite his scathing view of Young's career, Marsh gave the album the highest possible rating).
Track listing
All songs written by Neil Young.
Side one
- "Down to the Wire" – 2:25
- previously unreleased; performed with Buffalo Springfield members Stephen Stills and Richie Furay along with Dr. John
- "Burned" – 2:14
- performed by Buffalo Springfield; appears on the album Buffalo Springfield
- "Mr. Soul" – 2:41
- performed by Buffalo Springfield; recorded live in the studio in New York City, with guitar overdubs added subsequently; appears on the album Buffalo Springfield Again
- "Broken Arrow" – 6:13
- performed by Buffalo Springfield; appears on the album Buffalo Springfield Again
- "Expecting to Fly" – 3:44
- appears on the album Buffalo Springfield Again but no band member other than Neil Young appears on the track
- "Sugar Mountain" – 5:43
- recorded live in concert on November 10, 1968 at the Canterbury House, Ann Arbor, Michigan; released as the b-side to "The Loner," 1968
Side two
- "I Am a Child" – 2:17
- appears on the Buffalo Springfield album Last Time Around but features no members of the band other than Neil Young and drummer Dewey Martin
- "The Loner" – 3:50
- appears on the album Neil Young
- "The Old Laughing Lady" – 5:59
- appears on the album Neil Young
- "Cinnamon Girl" – 2:59
- performed by Neil Young & Crazy Horse; appears on the album Everybody Knows This Is Nowhere
- "Down by the River" – 9:16
- performed by Neil Young & Crazy Horse; appears on the album Everybody Knows This Is Nowhere
Side three
- "Cowgirl in the Sand" – 10:01
- performed by Neil Young & Crazy Horse; appears on the album Everybody Knows This Is Nowhere
- "I Believe in You" – 3:27
- performed by Neil Young & Crazy Horse; appears on the album After the Gold Rush
- "After the Gold Rush" – 3:45
- appears on the album After the Gold Rush
- "Southern Man" – 5:31
- appears on the album After the Gold Rush
- "Helpless" – 3:34
- performed by Crosby, Stills, Nash & Young; appears on the album Déjà Vu
Side four
- "Ohio" – 2:56
- performed by Crosby, Stills, Nash & Young; released as a single, 1970
- "Soldier" – 2:28
- edited from the album Journey Through the Past
- "Old Man" – 3:21
- appears on the album Harvest
- "A Man Needs a Maid" – 3:58
- appears on the album Harvest
- "Harvest" – 3:08
- appears on the album Harvest
- "Heart of Gold" – 3:06
- appears on the album Harvest
- "Star of Bethlehem" – 2:46
- appears on the album American Stars 'n Bars
Side five
- "The Needle and the Damage Done" – 2:02
- recorded live in concert on January 30, 1971 at Royce Hall, Westwood, California; appears on the album Harvest
- "Tonight's the Night" [Part I] – 4:41
- appears on the album Tonight's the Night
- "Tired Eyes" – 4:33
- appears on the album Tonight's the Night
- "Walk On" – 2:40
- appears on the album On the Beach
- "For the Turnstiles" – 3:01
- appears on the album On the Beach
- "Winterlong" – 3:05
- previously unreleased; appeared on certain acetate pressings of Tonight's The Night
- "Deep Forbidden Lake" – 3:39
- previously unreleased
Side six
- "Like a Hurricane" — 8:16
- performed by Neil Young & Crazy Horse; previously unreleased; different lead vocal dub than version on American Stars 'n Bars
- "Love is a Rose" — 2:16
- previously unreleased
- "Cortez the Killer" — 7:29
- performed by Neil Young & Crazy Horse; appears on the album Zuma
- "Campaigner" — 3:30
- previously unreleased
- "Long May You Run" — 3:48
- performed by Crosby, Stills, Nash & Young; previously unreleased; original mix (without Crosby and Nash) appears on the Stills-Young Band album Long May You Run
The CD release combined sides 1-3 onto disc one, and sides 4-6 on disc two.
- Elliot Mazer – Producer
- Neil Young – Guitar, Harmonica, Piano, Vibes, Vocals
Year Chart Peak
1977 Billboard Pop Albums 43
- ^ Williams, Paul. Neil Young: Love To Burn. p. 115. ISBN 0-934558-19-1.
- ^ "Allmusic review". http://www.allmusic.com/album/r22512.
- ^ "CG: Neil Young". Robert Christgau. http://www.robertchristgau.com/get_artist.php?name=Neil+Young. Retrieved 2011-08-31.
External linksCategories:
- 1977 greatest hits albums
- Neil Young compilation albums
- Triple compilation albums
- Reprise Records compilation albums
- Albums produced by Neil Young
- Albums produced by David Briggs
- Albums produced by Elliot Mazer
- "Down to the Wire" – 2:25
Wikimedia Foundation. 2010.


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Seeing what cannot be seen, and we do not even see the observer. What is over the horizon? What can the observer see that we cannot?
The endless task, an imaginary means of providing energy, always trying but never getting there.
From the early days of the internet, when we "surfed" from site to sit, full of adventure, and not even thinking about falling off the board
Joy? childhood? The spirit of play? in front of a shadowy darkness?
A different era, the 1970s, family?
Old and new energy forms? or just competing?
Contrasting the old and the new, putting the familiar in an unexpected place. Or the loss of old ways?
A found broken ornament, it pretty much can be a symbol of sadly what is common in the world these days
The road as a path, a symbol of life, having big goals.
Clicking forward or backwards.