IFN-γ protects from lethal IL-17 mediated viral encephalomyelitis independent of neutrophils 9:104 DOI: 10.1186/1742-2094-9-104
© Savarin et al.; licensee BioMed Central Ltd. 2012
Received: 7 March 2012 Accepted: 10 April 2012 Published: 29 May 2012 Abstract Background
The interplay between IFN-γ, IL-17 and neutrophils during CNS inflammatory disease is complex due to cross-regulatory factors affecting both positive and negative feedback loops. These interactions have hindered the ability to distinguish the relative contributions of neutrophils, Th1 and Th17 cell-derived effector molecules from secondary mediators to tissue damage and morbidity.
Methods
Encephalitis induced by a gliatropic murine coronavirus was used as a model to assess the direct contributions of neutrophils, IFN-γ and IL-17 to virus-induced mortality. CNS inflammatory conditions were selectively manipulated by adoptive transfer of virus-primed wild-type (WT) or IFN-γ deficient (GKO) memory CD4
+ T cells into infected SCID mice, coupled with antibody-mediated neutrophil depletion and cytokine blockade. Results
Transfer of GKO memory CD4
+ T cells into infected SCID mice induced rapid mortality compared to recipients of WT memory CD4 + T cells, despite similar virus control and demyelination. In contrast to recipients of WT CD4 + T cells, extensive neutrophil infiltration and IL-17 expression within the CNS in recipients of GKO CD4 + T cells provided a model to directly assess their contribution(s) to disease. Recipients of WT CD4 + T cells depleted of IFN-γ did not express IL-17 and were spared from mortality despite abundant CNS neutrophil infiltration, indicating that mortality was not mediated by excessive CNS neutrophil accumulation. By contrast, IL-17 depletion rescued recipients of GKO CD4 + T cells from rapid mortality without diminishing neutrophils or reducing GM-CSF, associated with pathogenic Th17 cells in CNS autoimmune models. Furthermore, co-transfer of WT and GKO CD4 + T cells prolonged survival in an IFN-γ dependent manner, although IL-17 transcription was not reduced. Conclusions
These data demonstrate that IL-17 mediates detrimental clinical consequences in an IFN-γ-deprived environment, independent of extensive neutrophil accumulation or GM-CSF upregulation. The results also suggest that IFN-γ overrides the detrimental IL-17 effector responses via a mechanism downstream of transcriptional regulation.
KeywordsCentral nervous system Encephalomyelitis CD4 +T cells IFN-γ IL-17 Neutrophils Neurotropic coronavirus Background
IL-17 and IFN-γ play diverse and often opposing functions during microbial infections, as well as autoimmune diseases. These interactions are partially attributed to their distinct regulation of the neutrophil response. Both IL-17A and IL-17 F signal through the IL-17R to induce granulocyte colony-stimulating factor and stem cell factor, thereby expanding neutrophil progenitors in the bone marrow and spleen as well as increasing mature neutrophils in the blood [1–3]. IL-17 also induces ELR
+ CXC chemokines, which attract neutrophils [2, 3]. By contrast, IFN-γ opposes neutrophil recruitment by downregulating expression of neutrophil chemoattractants [4]. Analysis of polarized T cell subsets and genetically deficient mice has provided insight into the distinct effector functions of IL-17 and IFN-γ; however, the interplay between IL-17 and IFN-γ in vivo remains complex [5, 6]. Moreover, downstream effector mechanisms mediating pathological consequences may be tissue- and pathogen-specific and are largely unresolved. For example, Th17 cell-mediated protection is critical during bacterial pneumonia [2]. IL-17-mediated neutrophil recruitment to the infection site also indicates a protective role for Th17 cells during oropharyngeal candidiasis [7]. By contrast, Th17-mediated inhibition of both protective Th1 responses and antimicrobial neutrophil functions increased tissue destruction following gastric candidiasis and pulmonary aspergillosis [8]. These differences may reflect distinct infection sites, as indicated by the distinct immune responses to Candida albicans, which are dependent upon the anatomical site of infection [7].
Viral infections are often dominated by Th1 responses. However, the coemergence of Th17 and Th1 cells has recently been documented in several infections, including human immunodeficiency virus [9], simian immunodeficiency virus [10] and cytomegalovirus [11]. A deleterious role of IL-17 is implied by acute lung injury associated with IL-17-mediated neutrophil recruitment during influenza virus infection [12]. By contrast, Th17 responses are protective against lethal influenza virus infection in IL-10-deficient mice [13]. Similarly, IFN-γ-mediated protection during herpes simplex virus-1 corneal infection correlated with reduced IL-17 production and subsequent neutrophil infiltration [14]. However, the function of IL-17 during central nervous system (CNS) viral infections, including human immunodeficiency virus encephalitis, is unclear, although Th17 cells promote Theiler’s murine encephalomyelitis virus persistence and chronic demyelination by limiting the antiviral cytotoxic T-lymphocyte response [15].
In contrast to the limited information on IL-17 function during viral encephalitis, analysis of experimental autoimmune encephalitis (EAE) has revealed numerous insights into effector mechanisms as well as crosstalk between Th1 and Th17 cells [16]. Although the inflammatory CNS disease multiple sclerosis and its animal model EAE were historically associated with a Th1 immune response [17, 18], a pro-inflammatory role of IFN-γ was contradicted by substantially increased disease severity and mortality in mice deficient in IFN-γ (GKO) or the IFN-γR [19, 20]. The correlation between increased EAE severity, enhanced Th17 responses and neutrophil infiltration into the CNS of GKO mice suggested that IFN-γ might be protective by inhibiting the Th17 response [21]. Although IL-17
−/− mice are susceptible to EAE [22], adoptive transfer of polarized encephalitogenic CD4 + T cells support Th17 cells as detrimental participants in EAE [23, 24]. However, the pathogenic mechanisms associated with Th17 cells remain an ongoing challenge and may involve multiple pathways. These include excessive CNS neutrophil infiltration and release of degrading enzymes, free radicals and pro-inflammatory cytokines, direct IL-17-mediated neuronal toxicity [25], and/or secretion of granulocyte macrophage colony-stimulating factor (GM-CSF) as the pathogenic effector molecule [26–28]. These data suggest that the balance between IFN-γ and IL-17 effector functions, as well as their regulation of neutrophils may dictate the outcome of non autoimmune-driven CNS inflammation, such as viral encephalitis.
During encephalomyelitis induced by the strain designated JHMV, CD4
+ T cells not only contribute to antiviral effects by enhancing CD8 + T cell function within the CNS [29] but also mediate viral control in absence of CD8 + T cells [30]. Nevertheless, they also contribute to both clinical disease and demyelination [30]. To define the role of CD4 + relative to CD8 + T cells in viral encephalitis, memory CD4 + T cells from immunized donors were transferred into infected severe combined immunodeficiency (SCID) mice [31]. This study revealed an early morbidity and mortality in infected recipients of CD4 + T cells lacking the ability to secrete IFN-γ compared to recipients of IFN-γ-sufficient CD4 + T cells or infected unreconstituted control mice [31]. Notably, both memory populations were equally effective in controlling virus replication [31]. The lethal outcome was specific for CD4 + T cells lacking IFN-γ [31], but not for a similar memory CD8 + T cell population deficient in IFN-γ [32]. These data suggest that mortality was related to immune effector functions specific to CD4 + T cells and controlled by IFN-γ.
In this study, SCID recipients of GKO CD4
+ T cells infected with JHMV were characterized by extensive neutrophil accumulation and IL-17 expression within the CNS. Neutrophil infiltration in the absence of IFN-γ correlated with significantly elevated levels of CXCL1, independent of IL-17. Moreover, comparison of infected recipients of wild-type (WT) CD4 + T cells depleted of IFN-γ and recipients of GKO CD4 + T cells depleted of IL-17 revealed mortality was due to IL-17, irrespective of abundant neutrophil accumulation. IFN-γ introduced by co-transfer of WT CD4 + T cells with IL-17-producing GKO CD4 + T cells abrogated the detrimental effects of IL-17 without affecting IL-17 transcription within the CNS. These data thus segregate the effects of toxic neutrophil components from IL-17-mediated pathogenesis. Material and Methods Mice
Homozygous BALB/c Thy1.1 mice, provided by Dr. J. Harty (University of Iowa, Iowa City, IA, USA) and GKO BALB/c mice, provided by Dr. R. Coffman (DNAX Research, Palo Alto, CA, USA), were bred locally at the Cleveland Clinic. SCID mice were obtained from the National Cancer Institute (Frederick, MD, USA). Recipients and donors were maintained under sterile conditions and all procedures were performed in compliance with Cleveland Clinic Institutional Animal Care and Use Committee-approved protocols.
Virus
The gliatropic JHM strain of mouse hepatitis virus (JHMV)-neutralizing mAb variant designated 2.2v-1 was used for intracerebral infection [33]. JHMV was propagated and plaque assayed on monolayers of DBT cells, a continuous murine astrocytoma cell line [32]. SCID mice were injected in the left hemisphere with 30 μl volume containing 500 PFU of JHMV diluted in endotoxin-free Dulbecco’s modified PBS. The severity of the JHMV-induced clinical disease was graded as follows: 0, healthy; 1, ruffled fur and hunched back; 2, partial hind limb paralysis or inability to turn to the upright position; 3, complete hind limb paralysis; 4, moribund or dead. Virus titers were determined on plaque assay on monolayers of DBT cells as previously described [32, 33]. Briefly, brains were homogenized in ice-cold Dulbecco’s PBS using Ten Broeck tissue homogenizers (Kimble Chase, Vineland, NJ, USA). After clarification by centrifugation at 400 x g for 7 minutes at 4°C, supernatants were stored at −70°C whereas pellets containing CNS-derived cells were suspended in Percoll (GE Healthcare Bio-Sciences AB, Uppsala, Sweden) and used for flow cytometry analysis (see below).
T cell purification and adoptive transfer
BALB/c Thy1.1 and GKO donors were immunized by intraperitoneal (i.p.) injection with 2 × 10
6 PFU of JHMV. Donor splenocytes were prepared four to sixteen weeks post immunization. CD4 + T cells were purified by positive selection using anti-CD4-coated magnetic beads (Miltenyi Biotec Inc., Auburn, CA, USA). Purity of the purified population was assessed by flow cytometry using fluorescein isothiocyanate- (FITC) labeled anti-CD4 (clone GK1.5), phycoerythrin- (PE) labeled anti-CD8 (clone 53-6.7) and peridinin chlorophyll protein- (PerCP) labeled anti-CD19 (clone 1D3) mAbs (BD Pharmingen, San Diego, CA, USA). Recipients received 5 × 10 6 donor CD4 + T cells composed of 100% Thy1.1 (WT), 100% GKO or a 50/50% mixture of Thy1.1/GKO (WT/GKO) CD4 + T cells by intravenous (i.v.) injection coupled with a single i.p. injection of 250 μg of anti-CD8 mAb (clone TIB.210). Mice were challenged with virus two to three hours after adoptive transfer. For neutrophil depletion, mice received i.p. injections of either 500 μg of anti-Ly-6G (clone 1A8) or anti-Gr1 (clone RB6-8C5) mAb every other day until sacrifice, starting two days before infection. Depletion was confirmed in both cases by flow cytometric analysis using anti-Ly-6G (clone 1A8) mAb in addition to examination of hematoxylin and eosin- (H&E) stained sections of brain. Only data for the anti-Ly6G experiments are shown. No differences in survival relative to control-treated mice were observed following treatment with either neutrophil-depleting mAb. Similarly, for anti-IFN-γ treatment, mice received i.p. injections of 500 μg of anti-IFN-γ (clone XMG1.2) mAb every other day, starting two days before infection. For anti-IL17 treatment, mice received i.p. injections of 1 mg of anti-IL-17A (clone 1D10) mAb at day zero and six post infection (p.i.). Isolation of central nervous system-derived cells
After brain homogenization and centrifugation to obtain supernatants for virus determination as described above, cell pellets were resuspended in RPMI containing 25 mM HEPES, pH 7.2 and adjusted to 30% Percoll (GE Healthcare Bio-Sciences BA). A 1 ml underlay of 70% Percoll was added prior to centrifugation at 800 x g for 30 minutes at 4°C. Cells were recovered from the 30% to 70% interface and washed with RPMI medium prior to analysis.
Flow cytometry
CNS mononuclear cell suspensions were blocked with anti-mouse CD16/CD32 (clone 2.4G2, BD Pharmingen) mAb on ice for 15 minutes prior to staining. Cells were then stained with FITC-, PE-, PerCP- or allophycocyanin-conjugated mAb for 30 minutes on ice in PBS containing 0.1% BSA. Expression of surface molecules was characterized using the following mAbs (all obtained from BD Pharmingen except when indicated): anti-CD45 (Clone Ly-5), anti-CD4 (clone GK1.5), anti-Thy1.1 (clone OX-7), anti-CD8 (clone 53-6.7), anti-CD11b (clone M1/70), anti-F4/80 (Serotec, Oxford, UK), anti-Ly6G (clone 1A8) and anti I-A/I-E (clone 2G9). Samples were analyzed on a FACSCalibur flow cytometer using CellQuest software (Becton Dickinson, Mountain View, CA, USA).
Gene expression analysis
RNA was isolated from three or more individual brains per group using TRIzoL reagent (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions. cDNAs were prepared using SuperScript II Reverse Transcriptase (Invitrogen) and oligo (dT)
12–18 primers (Invitrogen). Semi-quantitative RNA expression was assessed using LightCycler and SYBR Green kit (Roche, Basel, Switzerland) and the following primers; ubiquitin: F: 5’- TGGCTATTAATTATTCGGTCTGCAT-3’, R: 5’- GCAAGTGGCTAGAGTGCAGAGTAA -3’; IFN-γ: F: 5’- TGATGGCCTGATTGTCTTTCAA-3’, R: 5’- GGATATCTGGAGGAACTGGCAA-3’; IL-17: F: 5’-CTTCATCTGTGTCTCTGATGCTGTT-3’, R: 5’- TCGCTGCTGCCTTCACTGT-3’; IL-22: F: 5’- CATGCAGGAGGTGGTACCTT-3’, R: 5’- CAGACGCAAGCATTTCTCAG-3’; IL-21: F: 5’- GGACAGTATAGACGCTCACGAATG-3’, R: 5’- CGTATCGTACTTCTCCACTTGCA-3’; MHC class II: F: 5’- TCAACATCACATGGCTCAGAAATA-3’, R: 5’- AGACAGCTTGTGGAAGGAATGG-3’; GM-CSF: F: 5’- TTTCCTGGGCATTGTGGTCTA -3’, R: 5’- AAGGCCGGGTGACAGTGAT -3’; IL-6: F: 5’- ACACATGTTCTCTGGGAAATCGT -3’, R: 5’- AAGTGCATCATCGTTGTTCATACA-3’; IL-1β: F: 5’- GACGGCACACCCACCCT-3’, R: 5’- AAACCGTTTTTCCATCTTCTTCTTT-3’; CCL7: F: 5’-GGGAAGCTGTTATCTTCAAGACAAA-3’, R:5’-CTCCTCGACCCACTTCTGATG-3’; CCL20: F: 5’-GGTGGCAAGCGTCTGCTC-3’, R: 5’-GCCTGGCTGCAGAGGTGA-3’; CXCL2: F: 5’-CCTGCCAAGGGTTGACTTCA-3’, R: 5’-TTCTGTCTGGGCGCAGTG-3’; MMP9: F: 5’- CCATGCACTGGGCTTAGATCAT-3’, R: 5’- CAGATACTGGATGCCGTCTATGTC-3’; MMP3: F: 5’- TTTAAAGGAAATCAGTTCTGGGCTATA-3’, R: 5’-CGATCTTCTTCACGGTTGCA-3’; MMP12: F: 5’- GGAGCTCACGGAGACTTCAACT-3’, R: 5’-CCTTGAATACCAGGTCCAGGATA -3’. TaqMan primers and 2X TaqMan fast master mix (Applied Biosystems, Carlsbad, CA, USA) were used to assessed CXCL1 and CCL2 mRNA levels. Levels of mRNA expression were normalized to ubiquitin mRNA using ΔCt method as previously described [31]. Immunofluorescence
After ice-cold PBS perfusion, brains in OCT were frozen in liquid nitrogen and stored at −80°C until 10 μm sections were prepared. Sections were fixed with methanol/acetone (1:1 ratio) for 15 minutes and then treated with blocking solution for 30 minutes at room temperature. Rat anti-mouse IL-17 (R&D systems, Minneapolis, MN, USA) and hamster anti-mouse CD3 primary mAbs (Serotec) were incubated overnight at 4°C. Alexa Fluor 488 goat anti-rat (Invitrogen) and Alexa Fluor 546 goat anti-hamster (Molecular Probes, Eugene, OR, USA) were added for 1 hour at room temperature. Sections were mounted with Vectashield mounting medium with 4’-6-Diamidino-2-phenylindole (DAPI) (Vector Laboratories, Burlingame, CA, USA) and analyzed using a Leica DM4000B fluorescent microscope (Leica, Wetzlar, Germany).
In vitro T cell stimulation
Cytokine expression by CD4
+ T cells derived from cervical lymph nodes of SCID recipients were analyzed directly at day eight p.i. without stimulation with viral antigen. For analysis of cytokine production by cells prior to transfer, JHMV was adsorbed to donor splenocytes for 60 minutes at 4°C and cells cultured for six days in RPMI complete, 10% FCS at 2.5 × 10 6 cells/ml. Cytokine production from both splenic cultures or ex vivo lymph node cells was measured following four hours stimulation with PMA (10 ng/ml) (Acros Organics, Geel, Belgium) and ionomycin (1 μM) (Calbiotech, Spring Valley, CA, USA). Monensin (2 μM) (Calbiotech) was added to the cultures for the last two hours. After stimulation, cells were harvested and stained for surface expression of CD4. Cells were then permeabilized using the cytofix/cytoperm kit (BD Pharmingen) according to the manufacturer’s instructions and stained for intracellular FITC-IFN-γ and PE-IL-17. Statistical analyses
Statistical differences were calculated using the two-tailed unpaired Student’s
t-test. P values <0.05 were considered significant. *p < 0.05, **p < 0.01, ***p < 0.001 Results IFN-γ mediated control of central nervous system neutrophil infiltration is not the sole factor regulating survival +T cell recipients infected with JHMV was the large CNS infiltrating neutrophil population (72.3% compared to 17.5% in WT CD4 +T cells recipients) (Figure 1A) [31]. Increased neutrophil accumulation in GKO recipients is consistent with IFN-γ-mediated downregulation of ELR +neutrophil chemokines [4]. Indeed, analysis of cytokine and chemokine mRNA expression in infected T cell recipients demonstrated that high IFN-γ mRNA correlated inversely with mRNA expression of the neutrophil chemoattractant CXCL1 (Figure 1B). Thus, IFN-γ mRNA in WT CD4 +T cell recipients was associated with sparse CXCL1 expression and neutrophil recruitment, while low IFN-γ mRNA expression in both GKO CD4 +T cell recipients and infected SCID controls correlated with high CXCL1 expression and extensive neutrophil recruitment. Infected mice were depleted of neutrophils to explore a possible correlation between neutrophil-derived proteases, free radicals and proinflammatory cytokines with virus-induced mortality. Depletion was confirmed by the absence of Ly6G +CD11b +neutrophils within the CNS-derived inflammatory cells (Figure 1C). However, the absence of neutrophils did not prevent early mortality of GKO CD4 +T cell SCID recipients (Figure 1C), implicating alternate mechanisms inducing mortality in GKO recipients. +T cells derived from JHMV-immunized donors, memory GKO CD8 +T cells did not trigger early mortality in infected SCID recipients [32]. These data suggest that IFN-γ deficiency was not the sole factor controlling early death. WT CD4 +T cell recipients were depleted of IFN-γ to confirm that a CD4 +T cell factor distinct from IFN-γ controls disease outcome. The modestly reduced survival rate of IFN-γ-depleted WT CD4 +T cell recipients (Figure 2A) demonstrated IFN-γ blockade did not reproduce the mortality of GKO CD4 +T cell recipients. The efficiency of IFN-γ blockade within the CNS was confirmed by analyzing IFN-γ-dependent MHC class II expression on microglia [34]. In contrast to class II expression on the vast majority of microglia in recipients of WT CD4 +T cells, class II remained undetectable in anti-IFN-γ-treated WT recipients (Figure 2B), confirming inhibition of local IFN-γ signaling within the CNS. IFN-γ depletion also had minimal effects on T cell recruitment into the CNS, reducing the CD4 +T cells within the inflammatory population from 15.4% to 12.3% (data not shown). In support of the role of IFN-γ in regulating neutrophils, IFN-γ-depleted WT recipients exhibited vastly increased CNS neutrophil infiltration, approaching the numbers found in GKO CD4 +T cell recipients (Figure 2C). In addition to confirming IFN-γ-mediated control of CNS neutrophil recruitment [4], these data reassert that abundant CNS neutrophils are insufficient to account for early mortality. IL-17 mediates mortality, independent of neutrophils +T cell recipients (Figure 3A). Previous data demonstrated that TNF and inducible nitric oxide synthase were also not associated with early mortality of GKO recipients [31]. Indeed, passive transfer of neutralizing anti-TNF mAb was unable to alter the mortality of the GKO CD4 +T cell recipients (data not shown). These results suggested additional factor(s) intrinsic to GKO CD4 +T cells in mediating disease outcome. Inhibition of IL-17 production by IFN-γ [37], suggested IL-17 as a potential candidate. Consistent with this concept, IL-17 mRNA expression was increased in the CNS of GKO CD4 +T cell recipients (Figure 3B), although IL-17 is not expressed in the CNS of infected WT mice [38]. Importantly, IL-17 mRNA remained below detection not only in SCID-infected control mice lacking T cells, but also in recipients of WT CD4 +T cells depleted of IFN-γ (Figure 3B), both of which are characterized by vast CNS neutrophil infiltration (Figure 1A). Although neutrophil-derived IL-17 has been implicated in enhancing tissue damage during reperfusion injury [39], these data suggest that neutrophils recruited into the CNS do not secrete IL-17 during acute viral encephalitis. Expression of IL-17 mRNA only in GKO CD4 +T cell recipients also ruled out a potential contribution of resident CNS cells. IL-17 expression exclusively in the CNS of GKO recipients thus implied that the source of IL-17 was the GKO-derived CD4 +T cell population itself. In support of this concept, transcript levels encoding IL-22, another cytokine produced by Th17 cells [40], were also significantly increased in infected GKO recipients compared to WT recipients and infected SCID control mice (Figure 3C). By contrast, IL-21, a CD4 +T cell-derived cytokine known to provide helper functions to CD8 +T cells and B cells [41] was expressed at similar levels in both the GKO and WT CD4 +T cell recipient groups (Figure 3C). IL-17 production by CD4 +T cells in the CNS of GKO recipients was confirmed by immunofluorescence histochemistry. A substantial fraction of T cells within the CNS of GKO recipients expressed IL-17. Moreover, all IL-17 positive cells co-expressed CD3 (Figure 3D), indicating that T cells are the predominant source of IL-17 within the CNS of SCID recipients. In contrast to the CNS, only ~8% of T cells in the cervical lymph nodes of GKO recipients secreted IL-17 at day eight p.i. (Figure 3E), suggesting enrichment of IL-17-expressing T cells within the CNS. To determine if IL-17 expression is imprinted during the primary response following immunization of GKO donor mice, cytokine expression was analyzed in the memory WT and GKO T cell populations prior to transfer (Figure 3F). WT memory CD4 +T cells prominently expressed IFN-γ and very little, if any, IL-17 following in vitrostimulation. By contrast, immunization of GKO mice primed a small fraction of memory CD4 +T cells capable of producing IL-17. These results were consistent with IFN-γ-mediated inhibition of Th17 cells [42] and suggested that IL-17 expression was imprinted prior to transfer and re-expressed in the infected recipients. To confirm a role of IL-17 in the early mortality of GKO recipients, WT and GKO recipients were treated with anti-IL-17 mAb. Consistent with the absence of IL-17 mRNA in the CNS of the WT recipients, anti-IL-17 treatment had no effect on the survival of WT recipients (Figure 4A). By contrast, inhibition of IL-17 in GKO recipients lead to a significant decrease in mortality, with 73% of mice surviving to day 18 p.i. (Figure 4A). In support of the concept that mortality was not influenced by neutrophils, the increased neutrophil infiltration in the CNS of GKO recipients was not altered by anti-IL-17 treatment (Figure 4B), confirming their primary regulation by IFN-γ [4]. IFN-γ overcomes IL-17-derived CD4 + T cell mediating mortality +T cell subsets is well established during primary T cell activation and expansion [43, 44]; however, less is known about cross-regulation during antigen-induced restimulation of memory T cells. IL-23 has recently been shown to promote GM-CSF expression by Th17 cells, which in turn enhances detrimental disease outcomes [27, 28]. Based on the observation that GM-CSF is downregulated by IFN-γ [27], we tested whether IFN-γ-producing CD4 +T cells can override the detrimental function of GKO CD4 +T cell-mediated IL-17 expression. Memory CD4 +T cells from WT Thy1.1 immunized mice were co-transferred with Thy1.2 GKO CD4 +T donor cells (WT/GKO recipients) prior to infection. MHC class II expression was measured in the CNS of all recipient groups to confirm functional IFN-γ expression [34]. Class II mRNA was maximal in the CNS of WT recipients, reaching 22-fold higher levels than in GKO recipients. MHC class II mRNA was increased 10-fold following the co-transfer of WT and GKO CD4 +T cells compared to mice receiving GKO CD4 +T cells alone (Figure 5A). Increased mRNA expression correlated with MHC class II protein expression by the majority of microglia in both WT and co-transfer groups (Figure 5A). Although the proportion of microglia expressing MHC class II was decreased in the co-transfer compared to the WT groups, differences were not statistically significant (Figure 5A). Co-transfer of WT and GKO CD4 +T cells protected recipients from early mortality (Figure 5B). Disease severity was similar in WT and WT/GKO recipients and significantly reduced compared to GKO-only recipients, especially after day six p.i. (data not shown). In addition, control of CNS virus replication was identical at day eight p.i. in recipients of WT, WT/GKO or GKO CD4 +T cells (Figure 5C), confirming previous results that accelerated mortality does not correlate with uncontrolled virus replication [31]. CNS leukocyte infiltration in WT/GKO CD4 +T cell recipients was similar to WT recipients, and lower than GKO recipients (Figure 5D). Specifically, neutrophils within CD45 hibone-marrow-derived inflammatory cells were reduced to 16% in WT/GKO recipients, resembling the proportion in WT recipients (Figure 5D and data not shown). +T cells migrated to the CNS in the presence of WT CD4 +T cells, the relative proportions of each donor population was examined by co-transfer of Thy1.1 WT CD4 +T cells and Thy1.2 GKO CD4 +T cells. Surprisingly, T cells recruited into the CNS of infected co-transferred recipients were essentially derived from the GKO memory CD4 +T cells, as less than 20% of CD4 +T cells expressed Thy1.1 +(Figure 6A). Given the large population of infiltrating GKO CD4 +T cells, we next determined if IFN-γ-mediated protection correlated with reduced IL-17 mRNA expression. Although protective, the minor population of WT CD4 +T that infiltrated the CNS did not reduce expression of IL-17 mRNA in the CNS (Figure 6B). Protection mediated by IFN-γ, despite elevated IL-17, suggested that IFN-γ interferes with IL-17-mediated signaling events, rather than directly influencing Th17 expression. This notion was tested by in vitrostimulation of memory CD4 +T cells derived from GKO donors in the presence of recombinant IFN-γ. Exogenous IFN-γ was indeed unable to downregulate IL-17 production (Figure 6C), supporting the in vivoobservation that IFN-γ-expressing WT CD4 +T cells did not alter CNS expression of IL-17 mRNA in WT/GKO recipients (Figure 6B). The maintenance of IL-17 in the presence of IFN-γ in vitroand in vivoindicates that the phenotypes acquired during in vivoprimary responses are retained in the transferred memory cells following reactivation in recipient mice. To confirm this assumption, IFN-γ was depleted in WT/GKO recipients. WT/GKO recipients treated with anti-IFN-γ succumbed to infection by day nine p.i. similar to infected recipients of GKO CD4 +T cell (Figure 6D). These data actually suggest that IFN-γ diminishes the detrimental effects of IL-17, despite the apparent expansion/survival advantage of GKO relative to WT CD4 +T cells in the infected recipients. +T cells. Similar expression of CCL2, CCL7 and CCL20 was detected comparing infected SCID controls and GKO recipients; by contrast CCL2 and CCL7 were upregulated and CCL20 downregulated in recipients of WT CD4 +T cells (Figure 7A). These data suggest that in contrast to EAE, CCL2, CCL7 and CCL20 chemokine expression is regulated by IFN-γ rather that IL-17 during JHMV infection. Moreover, no significant difference in CXCL2 mRNA was found comparing SCID-infected controls and recipients of either WT or GKO CD4 +T cells (Figure 7A), supporting CXCL1 as the major neutrophil chemoattractant during JHMV infection. CNS infection with JHMV induces a limited number of MMPs, that is, MMP9, MMP3 and MMP12 [46]. As MMP9 is specifically expressed by neutrophils [47], abundant neutrophil recruitment in the CNS of GKO T cell recipients (whether or not treated with anti-IL17) correlated with MMP9 expression (Figure 7B). MMP3 and MMP12 mRNA expression were also upregulated in GKO recipients compared to infected SCID controls and WT recipients, suggesting a potential role of these MMPs in GKO mortality by mediating tissue destruction (Figure 7B). However, survival of GKO recipients treated with anti-IL17 also expressed increased MMP3 and MMP12 mRNA (Figure 7B), suggesting that MMP3 and MMP12 play no role in the early mortality of GKO recipients. Finally, to investigate a potential contribution of GM-CSF to the rapid disease progression, relative levels of GM-CSF were measured in the CNS of SCID-infected controls, and recipients of WT and GKO CD4 +T cells. GM-CSF mRNA expression was increased in GKO recipients relative to controls and WT CD4 +T cell recipients. These data were reminiscent of enhanced GM-CSF expression by Th17 compared to Th1 cells in EAE [27] and suggested a potentially detrimental role during JHMV encephalomyelitis. However, the increased survival of GKO recipients treated with anti-IL17 mAb did not correlate with a decrease in GM-CSF expression. These results indicate that GM-CSF expression correlated with IFN-γ deficiency, but not with an IL-17 mediated feedback loop. Nevertheless, these data suggest that IFN-γ directly affords protection from mortality by interfering with detrimental IL-17-mediated events, distinct from those mediating EAE. Discussion
IFN-γ and IL-17 are major effector molecules of tissue inflammation that play opposing roles in neutrophil recruitment/accumulation [4, 48, 49]. While their distinct influence on disease has been demonstrated during autoimmune-mediated neuroinflammatory responses, the interplay between IL-17 and IFN-γ, specifically the effects on downstream targets remain controversial. Furthermore, during microbial infections, protective and detrimental effects of IFN-γ and IL-17 depend on the pathogen and prominent cell types affecting microbial control [50–52]. The present study evaluated how the absence of IFN-γ secretion by CD4
+ T cells contributes to a rapid lethal outcome during viral encephalomyelitis, without altering viral control. Early virus-induced mortality in SCID recipients of GKO virus-specific memory CD4 + T cells correlated with both IL-17 production and extensive neutrophil accumulation in the CNS. Selective blockade of either neutrophils or IL-17 demonstrated that early mortality did not correlate with CNS neutrophil recruitment, but rather with IL-17. This was confirmed by the prolonged survival of recipients of anti-IFN-γ mAb-treated WT recipients, which were characterized by extensive neutrophil inflammation, but an absence of IL-17.
Neutrophil-independent pathogenic effects of IL-17 in the JHMV model contrast with non-CNS viral infectious models including the influenza virus and herpes simplex virus-1 infections, which attribute Th17 cell-mediated pathogenesis to neutrophil attraction [12, 14]. However, neutrophil depletion following severe influenza virus infection also suggests that neutrophils play a protective, rather than a deleterious role [53]. Our data also contrast with the deleterious role of neutrophils during EAE [49, 54]. Adoptive transfer of Th17 cells leads to excessive CNS neutrophil migration after EAE induction, while impaired neutrophil recruitment restrains leukocyte access into the CNS [49], indicating a prominent role of neutrophils in disrupting the blood-brain barrier. However, in contrast to EAE, neutrophils are not essential for the loss of blood-brain barrier integrity following sublethal JHMV infection [55]. By contrast, JHMV-induced encephalomyelitis demonstrates that IFN-γ plays a more prominent role than IL-17 in regulating CNS neutrophil recruitment and/or retention by downregulating ELR
+ neutrophil chemokine expression. Increased neutrophils correlated with high CXCL1 expression in the CNS of both IFN-γ-depleted WT recipients lacking IL-17, as well as in GKO recipients treated with anti-IL-17 Ab. Moreover, neutrophil infiltration was reduced by co-transfer of WT and GKO CD4 + T cells, despite sustained IL-17 expression in the CNS. These results are consistent with early studies identifying IFN-γ as a critical factor regulating CNS neutrophil infiltration [4], as well as recent observations implicating IFN-γ as a dominant molecule controlling CNS inflammation [26].
Despite evidence implicating IL-17 as a pathogenic mediator, independent of neutrophils, the mechanism(s) involved in IL-17-induced mortality of JHMV-infected mice remain unclear. Identical viral burden at day eight p.i. in all recipients [31] indicated that IL-17 does not alter control of virus replication, in contrast to its role in facilitating viral persistence following Theiler’s murine encephalomyelitis virus infection [15]. Sustained Ag independent interaction between Th17 and neuronal cells during EAE correlated with increased neuronal damage due to IL-17-mediated neurotoxicity [25]. Increased gray matter infection, especially in neuronal cells, is associated with premature death following JHMV infection of mice deficient in innate immune components [56]. In addition, there is a preferential distribution of CD4
+ T cells in the gray matter of GKO recipients compared to WT recipients [31], suggesting the possibility that in absence of IFN-γ, IL-17-secreting CD4 + T cells localize proximal to uninfected neurons, contributing to neuronal dysfunction and premature death. However, few neurons are infected early during JHMV pathogenesis in SCID mice and the types of infected cells were similar in all groups, suggesting no alteration in viral tropism [31]. In addition, no differential neuronal loss was found comparing GKO and WT recipients [31]. Similarly, increased expression of GM-CSF in GKO recipients compared to the WT counterparts suggested that GM-CSF might also contribute to disease outcome following JHMV infection. GM-CSF was implicated as a pathogenic effector molecule secreted by Th17 cells during EAE [27, 28]. However, the survival of GKO recipients treated with anti-IL17 did not correlate with a decrease in GM-CSF expression. Although GM-CSF expression is reduced by IFN-γ [27], the data do not support a pathogenic role of GM-CSF in early mortality of JHMV-infected GKO recipients.
IL-17 mRNA expression in GKO CD4
+ T cell recipients suggested Th17 cells as the primary mediators of disease. Nevertheless, IL-17 can also be produced by neutrophils, γδ T cells, NK and CD8 + T cells [57, 58]. A deleterious contribution of neutrophil-derived IL-17, suggested during kidney ischemia-reperfusion [39], was ruled out by the inability of neutrophil-depletion to rescue mice from early death, as well as the absence of IL-17 mRNA in WT recipients treated with anti-IFN-γ, despite high CNS neutrophil infiltration. IL-17 production by CD4 + T cells derived from immunized GKO donors prior to transfer supports GKO CD4 + T cells as the primary source of IL-17. Moreover, stimulation of WT donor CD4 + T cells strongly induced IFN-γ, but not IL-17, indicating that virus-specific Th17 cells only differentiate in the absence of IFN-γ. These results support previous observations of a minor, if any, role of Th17 cells in the pathogenesis of JHMV-infected immunocompetent WT mice [38] and corroborate the inhibitory function of IFN-γ on Th17 differentiation during T cell priming [59]. However, our data are novel in demonstrating that memory GKO CD4 + T cells are committed in their ability to produce IL-17 when restimulated in the recipient host, even in the presence of IFN-γ. Although unanticipated, this finding was confirmed by the inability of IFN-γ to downregulate IL-17 production in GKO donor cells in vitro, as well as on in vitro-differentiated mature Th17 cells [15]. Similarly, the IL-27 suppressive function on Th17 differentiation from naïve CD4 + T cells could not be reproduced on memory Th17 cells [60], supporting the stability of committed Th17 cells. Importantly, the prolonged survival of co-transferred recipients, despite sustained CNS IL-17 expression, suggests that IFN-γ overcomes the deleterious effects of IL-17. However, the mechanisms by which IFN-γ overrides IL-17 function remain unclear. In EAE, IL-17 exerts detrimental effects via signaling in resident CNS cells, with astrocytes implicated as major targets [45]. However, Th17 cell localization proximal to neurons also implicates potential dysregulation of neuronal function [25]. Responsiveness of both cell types to IFN-γ [61, 62] suggests IFN-γ may counteract signaling molecules downstream of the IL-17R. Conclusions
This study demonstrates that IL-17, in the absence of IFN-γ, can accelerate mortality during viral encephalomyelitis by a mechanism independent of the magnitude of CNS neutrophil infiltration and reversible by IFN-γ.
Abbreviations BSA:
bovine serum albumin
CNS:
central nervous system
DAPI:
4’-6-diamidino-2-phenylindole
EAE:
experimental autoimmune encephalitis
FITC:
fluorescein isothiocyanate
GKO:
IFN-γ deficient
GM-CSF:
granulocyte monocyte colony-stimulating factor
H&E:
hematoxylin and eosin
IFN-γ:
interferon-gamma
IL:
interleukin
i.p.:
intraperitoneal
i.v.:
intravenous
JHMV:
gliatropic JHM strain of mouse hepatitis virus
mAb:
monoclonal antibody
MHC:
major histocompatibility complex
MMP:
matrix metalloproteinase
PCR:
polymerase chain reaction
p.i.:
post infection
PE:
phycoerythrin
PerCP:
peridinin chlorophyll protein
SCID:
severe combined immunodeficiency
SD:
standard definition
SEM:
standard error of the mean
TNF:
tumor necrosis factor
WT:
wild-type.
Declarations Acknowledgments
The authors thank Wenqiang Wei, Kate Stenson and Shabbir Hussain for technical assistance. This work was supported by National Institutes of Health grant NS18146 and National Multiple Sclerosis Society fellowship grant FG-1791-A-1 to CS.
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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.


Elmo is sending a message to military families on the Sesame Street/ USO Experience tour: Moving is OK. Visiting 65 military installations over 235 dates, Elmo is accompanied by a new friend named Katie, who is worried about a military move. The USO and Sesame Street hope that this message will be reassuring to children and families experiencing military relocation.
A recent report by the Handson Network shows that military families are four times more likely to move than average Americans. This data mirrors the results of a study published in Military Medicine in 2005 that suggests a link between frequent relocation and behavioral problems in a sample of 179 military children.
Because moving can be stressful, this show opens a dialogue and allows families to enter into conversations with their children regarding the subject. The Military Relocation Network suggests that parents discuss the new location, school and potential friends with their children to ease anxiety.
Further, stress can be reduced for the entire family by staying organized and preparing for the arrival of the moving company. With Elmo’s help, Katie’s story guides these children in coping with both the fear of moving and the worry of settling into a new life.


CALGARY, Dec. 21, 2015 /CNW/ - The Alberta Court of Appeal has dismissed two appeals from defendants seeking a right to a trial by jury for
Securities Act offences. The appeals – one from Calgary and one from Edmonton – claimed that the maximum penalty under Alberta securities laws of five years less a day in prison, or a $5 million fine, or both, triggered the right to a jury trial under section 11(f) of the Canadian Charter of Rights and Freedoms. That section provides a right to a trial by jury for any person charged with an offence where the maximum sentence is imprisonment for five years or a more severe punishment.
In dismissing the appeals, the majority of the Court concluded "…the maximum penalty of five years less a day found in the
Securities Act was deliberately chosen to avoid jury trials of complex securities prosecutions. The Provincial Legislature cannot be criticized for attempting to enact Charter-compliant legislation by deliberately setting a maximum penalty which reflects the de facto cut-off in the Charter, and the procedural limitations of the Provincial Court."
An appearance to set a date for trial in the Calgary proceeding against Ronald Aitkens will be held in mid-January 2016. The Edmonton proceeding, involving defendants Jeremy and Robert Peers, is scheduled to begin in February 2016.
A copy of the decision is available on the ASC website at www.albertasecurities.com.
The ASC is the regulatory agency responsible for administering the province's securities laws. It is entrusted with fostering a fair and efficient capital market in Alberta and with protecting investors. As a member of the Canadian Securities Administrators, the ASC works to improve, coordinate and harmonize the regulation of Canada's capital markets.
SOURCE Alberta Securities Commission
For further information: For Media Inquiries: Mark Dickey, Senior Communications Advisor, 403.297.4481; For Investor Inquiries: ASC Public Inquiries, Toll Free 1.877.355.4488
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The National Pork Producers Council (NPCC) expressed strong disappointment in a U.N. commission's failure to approve an international standard for a widely used feed additive.
The Codex Alimentarius Commission, which was established by the U.N.’s Food and Agriculture Organization and its World Health Organization to promote food safety and fair practices in trade, refused to adopt a science-based standard for ractopamine, a feed additive used to promote leanness in pork and beef. Establishment of a standard will be held at the final stage before approval for the fourth consecutive year.
Ractopamine, like all feed additives, was evaluated and approved by the U.S. Food and Drug Administration and has been approved for use in 26 countries, including Australia, Brazil, Canada, Indonesia, Mexico, the Philippines and South Korea.
A Codex panel of international scientists, including scientists from the European Union, three times has confirmed the safety of ractopamine and reaffirmed the safety of the product at this week’s commission meeting in Geneva, Switzerland.
Despite those findings and the support of the United States, Canada and countries in Africa, Asia, Latin America, and the Pacific Islands for adoption of the standard, opposition from the European Union, China, Thailand and Russia blocked it for non-scientific reasons outside the scope of the Codex. Except for Russia, those countries ban imports of pork from pigs fed
ractopamine.
“U.S. pork producers are very disappointed that the Codex commission succumbed to the bullying of countries that had no scientific reasons for opposing adoption of a standard for ractopamine,” said NPPC President Doug Wolf, a producer from Lancaster, Wis. “This is a scientifically proven safe product, and the commission failed again to act on its mandate to base standards and guidelines on science. This lack of action calls into question Codex’s legitimacy as the international reference body for scientific standards for food.”


Abstract
The reduction and removal of user fees is currently one of the top policy priorities in Africa today and several countries have reduced or abolished user fees. However, there is concern that without due attention to the process of fee removal, the expected utilization and affordability benefits may not be achieved. This policy brief examines the challenges of implementing fee reduction in the Kenyan context, and assesses an innovative approach to alleviate these challenges through direct funding of health facilities.
Citation
CREHS policy brief, October 2009, 2 pp.


Performance review scoring sheets include questions addressing the employee performance that are answered on a scale of one to five. A score of one indicates an employee is performing unsuccessfully. Two shows partial success, while a three is awarded to employees who are fully successful. A score of four is given to those with superior performance, and a score of five is reserved for those employees exhibiting distinguished performances.Continue Reading
Unsuccessful performers do not meet job expectations, and their jobs may be lost. If the performance review is the employee's first, supervisors may demand that goals be met in a timely manner; otherwise, their jobs may be lost. Unsuccessful performers typically are denied raises.
Partially successful performers may do well in some areas of their jobs, but they do not meet expectations in other areas. These employees show signs of burnout or low motivation. They may need more training to grasp the technical skills related to the jobs. These employees may also be successful in their workplaces but exhibit undesirable conduct. A partial raise is sometimes awarded.
Fully successful performers are dedicated to their job and always try to improve their skills. They are considered essential members of their teams.
Superior performers are ambitious and work well with other employees. While they may not be ready for promotions at that time, they should be encouraged to continue honing their skills for possible future promotions.
Distinguished performers are ready to be promoted. They are typically the company's most valued employees.Learn more about HR


Being comfortable reinforces what you already know, which can be very beneficial. Feeling uncomfortable creates the opportunity to learn something new.
I’ve had a stammer all my life. A stammer, like Parkinson’s, is a chronic condition; it has to be dealt with every day. Again, like Parkinson’s, there is no cure for a stammer. It has been a long, uncomfortable road but through my stammer I learnt Heidegger’s idea of thrownness (the state in which I exist is blameless, it just
is); that I can’t choose my thrownness but I can choose my reaction to it. I also learnt resilience dealing with my stammer. It turns out that all these valuable lessons are applicable to Parkinson’s. A stammer was the perfect dress rehearsal for coping with the challenge of a Parkinson’s diagnosis. Therefore, quite unexpectedly, my stammer had a positive influence on my life and in the forging of who I am. My stammer can be seen as the classroom wherein I learnt the clarity to cope with my Parkinson’s. I’m glad I concentrated in that lesson!
We all face adversity during our lives. Sometimes things happen outside of our control. This can be incredibly disempowering. However, real control and real empowerment comes from how we react to a situation. The adversity of my stammer, because I reacted to it in the right way (I
learntfrom it), shines a light on the adversity of my Parkinson’s. I am grateful to my stammer because without it my Parkinson’s would be shrouded in darkness.


Epoxy flooring, often used in garages and industrial settings, consists of epoxy resins that coat and protect the floor. Durable epoxy flooring provides a waterproof barrier, resists chemicals, mold and oil, and requires little maintenance to keep it clean. Pulling in or backing out of the garage too quickly, however, can leave tire marks on your epoxy floor. These marks need a stronger method of cleaning than mopping alone to remove them completely, but don't always require the use of harsh chemicals.
1
Mix 1 gallon of warm water with about 2 tablespoons of dish detergent and apply it to the tire marks. Allow it to sit for at three to five minutes. Alternatively, put on rubber gloves and pour or spray enough citrus solvent on the tire marks to cover them completely.
2
Scrub the marks with a melamine sponge to remove them from the epoxy floor. If you used citrus solvent, scrub the floor with a firm-bristled brush.
3
Mop the floor with clean water in a bucket to remove any cleaning residue and allow it to dry.
Things You Will Need Dish detergent Rubber gloves Citrus solvent Melamine sponge Firm-bristled brush Mop Bucket Warnings Don't use solvents near open flames or while smoking. Avoid touching solvents. They can cause skin and eye irritation.


Nonprofit corporations differ from profit-making organizations in that nonprofits are not operated to personally benefit an individual nor can their money be distributed to stockholders. A nonprofit organization must devote all of its money to the organization’s designated purpose. This does not mean that nonprofits cannot pay salaries. However, salaries are reviewed regularly and cannot not be excessive. In addition, nonprofits differ from profit-making organizations in their purposes, tax status, political involvement and governance.
Purposes
Whereas the profit-making organization is driven to make money for the company’s owners, the nonprofit organization’s sole purpose for existence is to fulfill its mission. Nonprofit organizations apply to the IRS and show bylaws that demonstrate that they fit into one of the categories that the government deems worth of receiving 501(c)( 3) tax-exempt status. Most 501(c)(3) tax-exempt organizations fall into the categories of educational, charitable, scientific, literary or religious. Examples include soup kitchens and charitable foundations.
Tax Status
Organizations that are granted tax-exempt status are not required to pay taxes at all with the exception of payroll tax for employees. However, not all nonprofits are tax-exempt. The Maine Association of Non-Profits notes that donations to some social service organizations such as the National Rifle Association or the National Organization of Women, which are categorized by the IRS as 501(c)(4), are not tax deductible.
Political Activities
Nonprofits classified as 501(c)(3) are very restricted in their political activities. They cannot participate at all in campaigns for or against candidates for elective office. However, they may offer education programs and information about issues and may otherwise engage in nonpartisan activities such as urging people to register and vote. They may lobby on issues, but lobbying cannot be a substantial part of the organization’s activities. A 501(c)(4) organization has more leeway to participate in lobbying and political campaigns.
Governance
A board of directors governs nonprofits. This board has the responsibility of making sure that the organization remains true to its mission as stated in the bylaws. Nonprofits that stray from their stated purpose or violate other relevant regulations can have their tax-exempt status withdrawn. In profit-making organizations, the owners are free to decide to run the corporation in any way they deem desirable. The owner may be a single individual or a few founders in a privately held company. With large publicly held companies, the stockholders vote at annual meetings to determine the direction the company should take.


Empathy: "Hath not a Jew eyes? ..."
A recent article in ScienceDaily, "Less Empathy Toward Outsiders: Brain Differences Reinforce Preferences For Those In Same Social Group" (July 1, 2009) reports that
... perceiving others in pain activates a part of the brain associated with empathy and emotion more if the observer and the observed are the same race. The findings may show that unconscious prejudices against outside groups exist at a basic level.Published in the Journal of Neuroscience, The study confirms an in-group bias in empathic feelings, something that has long been known but never before confirmed by neuroimaging technology ... the finding raises as many questions as it answers, Farah said. "For example, is it racial identity per se that determines the brain's empathic response, or some more general measure of similarity between self and other?" she said. "What personal characteristics or life experiences influence the disparity in empathic response toward in-group and out-group members?"One can think of a few, including slightly dissimilar facial expressions. The researchers chose race as the characteristic to study, but race is rarely a category by itself. It is bound up with class, status, lifestyle expectations, history, and culture. Some other, more tightly specified, category may have been more informative.
Remember Shylock's wrenching plea in Shakespeare's Merchant of Venice?
I am a Jew. Hath not a Jew eyes? Hath not a Jew hands, organs, dimensions, senses, affections, passions, fed with the same food, hurt with the same weapons, subject to the same diseases, healed by the same means, warmed and cooled by the same winter and summer, as a Christian is? If you prick us, do we not bleed? If you tickle us, do we not laugh? If you poison us, do we not die? And if you wrong us, shall we not revenge? If we are like you in the rest, we will resemble you in that. - From The Merchant of Venice (III, i, 60-63) Hat tip:Stephanie West Allen at Brains on Purpose
Labels: empathy


Reducing the Risk of a Shark Encounter: Advice to Swimmers
The relative risk of a shark attack is very small (The Relative Risk of Shark Attacks to Humans Compared To ...) but, risks should always be minimized whenever possible in any activity. The chances of having an interaction with a shark can be reduced if one heeds the following advice:
Always stay in groups since sharks are more likely to attack a solitary individual. Do not wander too far from shore --- this isolates an individual and additionally places one far away from assistance. Avoid being in the water during darkness or twilight hours when sharks are most active and have a competitive sensory advantage. Do not enter the water if bleeding from an open wound, and enter with caution if menstruating --- a shark's olfactory ability is acute. Wearing shiny jewelry is discouraged because the reflected light resembles the sheen of fish scales. Avoid waters with known effluents or sewage and those being used by sport or commercial fisherman, especially if there are signs of bait fishes or feeding activity. Diving seabirds are good indicators of such action. Sightings of porpoises do not indicate the absence of sharks --- both often eat the same food items. Use extra caution when waters are murky and avoid uneven tanning and bright colored clothing --- sharks see contrast particularly well. Refrain from excess splashing and do not allow pets in the water because of their erratic movements. Exercise caution when occupying the area between sandbars or near steep dropoffs --- these are favorite hangouts for sharks. Do not enter the water if sharks are known to be present and evacuate the water if sharks are seen while there. And, of course, do not harass a shark if you see one!


Being a horse jockey involves much more than sitting on a horse. To be a successful jockey, you must be exceptionally physically fit. The exercises involved with being a jockey focus on building muscle strength, stamina and endurance, which include exercises such as yoga and running.
Balance and Flexibility
Yoga, Pilates, situps, planks, lunges and squats are effective at working on a jockey's balance and flexibility. Some exercises replicate the stances of a jockey on a horse, like squats. Yoga moves such as the Cat and Cow work core muscles of the abdomen and lower back. To do these moves, get on all fours. Exhale and round your spine up while keeping your shoulders down. This is known as the Cat. Hold this position for a few seconds then return to a neutral position. Exhale then inhale and stick your head and tailbone up while your belly sinks toward the floor. This is known as the Cow.
Aerobic Exercises
The goal of performing aerobic exercises is to get your heart pumping. This increases your heart and lung capacity. Aerobic exercises also work key muscle groups, such as the legs and core, which are crucial for jockeys to control their horse. Jockeys partake in daily aerobic exercises such as swimming, running, jogging and biking. During racing season, professional jockey Martin Lane rides his bikes 150 miles per week. Aerobic exercises should be done for 30 minutes, three to five times per week.


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The federal Form I-9, Employment Eligibility Verification, is used by employers as a record of their basis for determining eligibility of an employee to work in the United States. The form is kept by the employer and made available for inspection by officials of the Department of Homeland Security, the Department of Labor, and the Office of Special Counsel for Immigration Related Unfair Employment Practices.
Submission of the information required in this form is voluntary. However, an individual may not begin employment unless this form is completed, since employers are subject to civil or criminal penalties if they do not comply with the Immigration Reform and Control Act of 1986.
All employees, citizens and noncitizens, hired after November 6, 1986, must complete Section 1 of this form at the time of hire, which is the actual beginning of employment. The employer is responsible for ensuring that Section 1 is timely and properly completed.
Employers must complete Section 2 by examining evidence of identity and employment eligibility within three (3) business days of the date employment begins. If employees are authorized to work, but are unable to present the required document(s) within three business days, they must present a receipt for the application of the document(s) within three business days and the actual document(s) within ninety (90) days. However, if employers hire individuals for a duration of less than three business days, Section 2 must be completed at the time employment begins.
Employers must record:
document title issuing authority document number expiration date, if any the date employment begins
Employers must sign and date the certification. Employees must present original documents. Employers may, but are not required to, photocopy the document(s) presented. These photocopies may only be used for the verification process and must be retained with the I-9. However, employers are still responsible for completing the I-9.
Employers must complete Section 3 when updating and/or reverifying the I-9. Employers must reverify employment eligibility of their employees on or before the expiration date recorded in Section 1. Employers CANNOT specify which document(s) they will accept from an employee.
File Descriptions:
These files are in Adobe Portable Document Format (.pdf), and you will need the free Acrobat Reader to view and print the files. More information:
The I-9 Form


Organisations have begun to reflect on the culture and sub-cultures of their organisations. They are intent on assessing whether issues exist, their underlying causes and what actions could be taken to either change or enrich their organisational culture.
Organisations need to tackle the issue of culture in its broadest sense to effect lasting change. While conduct risk and risk management are critical factors – and gain significant attention from regulators, boards and management – they are not enough in themselves.
This article,
Organisations’ cultural assessment and transformation, explores taking a more holistic approach, understanding and addressing an organisation’s core strengths and weaknesses as regards leadership, individual beliefs and other organisational mechanisms, and associated operationalisation, so a genuine transformation can occur.
While cultural change is a challenging undertaking, it is an essential element in an organisation’s long-term sustainability.


"One of the most important functions that any marketing organization should be tasked with concerns strategy. Strategy, being quite distinct from tactics, concerns the choices and tradeoffs that an institution makes… The strategy is responsible for defining and communicating the institution’s core brand and relevant attributes to external constituents."
Michael E. Porter, 1998
As the number and quality of higher education institutions continue to increase throughout the broader Middle East, it remains vital for AUB to uniquely define its core brand and communicate it with a greater degree of pervasiveness and consistency across various marketing mediums. In this respect, the Office of Communications is actively involved in monitoring external trends impacting higher education in the broader region. The ability for AUB to attract academically gifted students beyond Lebanon requires the institution to position itself in advantageous ways vis-à-vis regional alternatives. To that effect, the Office of Communications works to identify the institution’s strategic differentiators and actively promote them. The following illustration depicts AUB’s strategic marketing process:
For assistance with strategic marketing and communications, please contact:
Martin AsserAssociate Vice President for CommunicationsExtension: 2674 (Building 42, Room 208)Email address: ma389@aub.edu.lb


Systemic Risk: Regulatory Oversight and Recent Initiatives to Address Risk Posed by Credit Default Swaps GAO-09-397T: Published: Mar 5, 2009. Publicly Released: Mar 5, 2009. Additional Materials: Contact:
The U.S. financial system is more prone to systemic risk today because (1) the current U.S. financial regulatory system is not designed to adequately oversee today's large and interconnected financial institutions, (2) not all financial activities and institutions fall under the direct purview of financial regulators, and (3) market innovations have led to the creation of new and sometimes complex products that were not envisioned as the current regulatory system developed. Credit default swaps (CDS) are one of the products that have assumed a key role in financial markets. My statement will discuss (1) the extent to which U.S. financial regulators and the UK regulator oversee CDS, (2) risks and challenges that CDS present to the stability of financial markets and institutions and similar concerns that other products may pose, and (3) the recent steps that financial regulators and the industry have taken to address risks pose by CDS and similar efforts that may be warranted for other financial products. GAO reviewed research studies and congressional testimonies. We interviewed financial regulators and a variety of financial market participants. In January 2009, GAO designated the financial regulatory system as a high-risk area in need of congressional attention. Issues involving systemic risk regulation in general and CDS in particular should be considered as part of that effort.
The current regulatory structure for CDS does not provide any one regulator with authority over all participants in the CDS market, making it difficult to monitor and manage potential systemic risk. Federal oversight of CDS trading and monitoring of the CDS market are largely conducted through the banking regulators' safety and soundness oversight of supervised banks that act as CDS dealers. The Securities and Exchange Commission and the Commodity Futures Trading Commission lack the authority to regulate CDS broadly as financial products. Regulators have sought to address potential systemic risks arising from CDS activities mainly through collaborative efforts with other supervisors and key market participants. However, the extent to which regulators routinely monitor the CDS activity of unregulated market participants is unclear. The Financial Services Authority in the United Kingdom has authority over most CDS products and can collect information about the CDS market, but it has pursued most of its regulatory efforts in collaboration with U.S. regulators. CDS pose a number of risks to institutions and markets, many of which are not unique. These include counterparty credit, operational, concentration, and jump-to-default risks. Market participants and observers noted that CDS referencing asset-backed securities (ABS) and collateralized debt obligations (CDOs), particularly those related to mortgages, currently pose greater risks to institutions and markets than other types of CDS. Other risks and challenges from CDS relate to the lack of transparency in CDS markets, the potential for manipulation related to the use of CDS as a price discovery mechanism, and the use of CDS for speculative purposes. Regulators and market participants noted that over-the-counter (OTC) derivatives, to varying degrees, may pose some similar risks and a few identified equity derivatives as the OTC derivatives that were most similar to CDS. Financial regulators and market participants have initiated several efforts to mitigate these risks. These efforts target primarily operational and counterparty credit risks and include improving the operational infrastructure of CDS markets, creating a clearinghouse or central counterparty process to clear CDS trades, and establishing a central trade registry for CDS. If effectively implemented and sustained, these initiatives couldbegin to address some of the risks noted. But the effectiveness of these recent initiatives could be limited because participation is voluntary and regulators lack the authority to require all market participants to report their trades to a repository. Moreover, customized and highly structured CDS, which can include CDS with complex reference entities that may present additional risks, generally lack the standardization necessary for centralized clearing. Other ideas to reform CDS markets, such as mandatory clearing or limiting some types of trades, have important limitations that would need to be addressed. Finally, many participants and observers agreed that OTC derivatives other than CDS generally share some of the same risks and could benefit from similar efforts to mitigate their impact.


Introduction: Accurate outcomes data and predictors ofoutcomes are fundamental to the effective care of patients with COPDand in guiding them and their families through end-of-lifedecisions. Design: We conducted a retrospective cohortstudy of 166 patients using prospectively gathered data in patientswith COPD who required mechanical ventilation for acute respiratoryfailure of diverse etiologies. Results: Thein-hospital mortality rate for the entire cohort was 28% but fell to12% for patients with a COPD exacerbation and without a comorbidillness. Univariate analysis showed a higher mortality rate among thosepatients who required > 72 h of mechanical ventilation (37% vs 16%;p < 0.01), those without previous episodes of mechanical ventilation(33% vs 11%; p < 0.01), and those with a failed extubation attempt(36% vs 7%; p = 0.0001). With multiple logistical regression,higher acute physiology score measured 6 h after the onset ofmechanical ventilation, presence of malignancy, presence of APACHE(acute physiology and chronic health evaluation) II-associatedcomorbidity, and the need for mechanical ventilation ≥ 72 h wereindependent predictors of poor outcome.
Conclusions:We conclude that among variables available within the first 6 h ofmechanical ventilation, the presence of comorbidity and a measure ofthe severity of the acute illness are predictors of in-hospitalmortality among patients with COPD and acute respiratory failure. Theoccurrence of extubation failure or the need for mechanical ventilationbeyond 72 h also portends a worse prognosis.


In its renewable portfolio standard (RPS) report filed with the Hawaii Public Utilities Commission (PUC), Hawaiian Electric Co. Inc. is reporting record-high renewable energy use for the state in 2015.
The consolidated RPS for Hawaiian Electric, Maui Electric and Hawaii Electric Light combined was 23.2%, an increase from 21.3% for 2014.
According to Hawaiian Electric, the achievement, which far exceeded Hawaii’s 2015 RPS goal of 15%, came from a diverse mix of renewable sources, including wind power, waste-to-energy, biomass, geothermal, hydro, biofuels and solar (both utility-scale and customer-sited rooftop systems).
Hawaii Island, which reported that 48.7% of customer electricity use came from renewable resources, led the state. Maui County, including Molokai and Lanai, recorded 35.4%, and Oahu achieved 17.2%.
The RPS reported to the PUC represents the calculation for the entire year. However, for monthly records, Hawaii Island hit a renewable high of 68% in December, Maui County hit 63% in April and Oahu reached 42% in May.
The utility says electricity generated using renewable energy sources increased by 39,153 MWh in 2015. The total amount of electrical energy generated from customer-sited, grid-connected sources increased by an estimated 128,060 MWh, a 24.9% jump from the previous year.
The Hawaiian Electric companies are preparing updated power supply improvement plans for achieving the state’s future RPS goals of 30% by 2020, 40% by 2030, 70% by 2040 and 100% by 2045.
“A balanced portfolio of renewable resources is critical to our successful transition from imported oil,” says Alan Oshima, Hawaiian Electric’s president and CEO. “Energy conservation and storage, rooftop and grid solar, wind power, geothermal, and demand response programs will all continue to help us reach that goal while providing value to our customers.”


Article Title Recommended Citation
Aya Gruber,
A Provocative Defense,103 Cal. L. Rev. 273 (2015). Available at: http://scholarship.law.berkeley.edu/californialawreview/vol103/iss2/2 Abstract
It is common wisdom that the provocation defense is, quite simply, sexist. For decades, there has been a trenchant feminist critique that the doctrine reflects and reinforces masculine norms of violence and shelters brutal domestic killers. The critique is so prominent that it appears alongside the doctrine itself in leading criminal law casebooks. The feminist critique of provocation embodies several claims about provocation’s problematically gendered nature, including that the defense is steeped in chauvinist history, treats culpable sexist killers too leniently, discriminates against women, and expresses bad messages. This Article offers a (likely provocative) defense of the provocation doctrine. While fully acknowledging widespread gender inequity in society, the Article argues that the feminist critique may overestimate the provocation doctrine’s contribution to such inequality and underestimate its value to marginalized defendants. Provocation, like many legal doctrines, has a complex history. Further, the limited empirical evidence available appears to undermine rather than confirm assertions that the defense disproportionately burdens women and proves strategically vital to murderous men. Moreover, efforts to utilize criminal punishment to express an anti-masculinity, anti-violence message may, in the end, reinforce destructive masculine norms, exacerbate racial hierarchies, justify extant unequal power distributions, and, ironically, increase violence and suffering. In the end, the Article cautions that the feminist critique of provocation and similar progressive critiques of doctrinal leniency may unintentionally instantiate and entrench the punitive impulses that create and sustain mass incarceration.


UNICRI’s Investigating Crimes against the Environment course aims to provide media and public relations professionals, as well as those planning a career in public information and journalism, with an in-depth understanding of current environmental threats to the natural environment, sustainable development and human rights.
Climate change, together with wildlife crime, illegal exploitation of natural resources and illicit trafficking and dumping of hazardous material, is putting at danger the economic, environmental and cultural lives of communities worldwide. Clear, objective and reliable journalism and information sharing are crucial to guarantee a public understanding regarding the potential global catastrophic impacts of ecological crimes and climate change on the environment and humankind.
Over the two-day course participants will deepen their knowledge of transnational organized environmental crime, corruption, natural and human factors of climate change and the role of green governance to pursue sustainable development. Journalists and communication professionals will acquire key tools to communicate environmental threats, report compelling news stories, disseminate clear information out of complex scientific issues, detect inaccurate reporting, and verify reliable sources.
Participants will also be engaged in a series of brainstorming sessions and practical exercises with eminent scientists, high-level officers and experienced journalists, expanding their professional network in a select, international setting.


1. Hong, Sae-Yong: 20 articles (01/2014 - 03/2002) 2. Gil, Hyo-Wook: 17 articles (01/2014 - 09/2005) 3. Lee, Eun-Young: 15 articles (06/2011 - 03/2002) 4. Lin, Ja-Liang: 14 articles (01/2014 - 11/2002) 5. Yang, Jong-Oh: 13 articles (06/2011 - 05/2002) 6. Jian, Xiang-dong: 10 articles (11/2013 - 04/2007) 7. Liu, Zhi: 9 articles (05/2015 - 05/2012) 8. Cory-Slechta, Deborah A: 9 articles (03/2014 - 07/2002) 9. Yen, Tzung-Hai: 9 articles (01/2014 - 07/2010) 10. Singh, Mahendra Pratap: 9 articles (12/2013 - 04/2006)
1. Poisoning
10/01/1991 - "The most effective treatment of severe paraquat poisoning in man is uncertain. "
01/01/1987 - "There is a striking discrepancy between the efficacy of the kidneys, haemodialysis and haemoperfusion in removing paraquat from the body and the poor prognosis of paraquat poisoning even when the blood and urine concentrations (which are good indices of concentrations in lung and other tissues) are very low. "
01/01/1986 - "There is a striking discrepancy between: the efficacy of the kidneys, haemodialysis (HD) and Haemoperfusion (HP) in removing paraquat from the body and the poor prognosis of paraquat poisoning even when the blood and urine levels (which are good indices of concentrations in lung and other tissues) are very low. "
12/01/1997 - "It is also suggested that early haemoperfusion is effective in treating paraquat poisoning even in very severe cases due to its possible detoxicating effect in addition to toxin removal."
12/07/1974 - "Effective treatment for paraquat poisoning in rats and its relevance to treatment of paraquat poisoning in man."
2. Lung Injury
01/01/2008 - "Anti-TGF-beta1 antibody: an effective treatment for lung injury caused by paraquat in the future."
06/01/2014 - "[The efficacy of traditional Chinese medicin in animal model of lung injury induced by paraquat: a meta-analysis]."
06/01/2009 - "The purpose of this study was to report the long-term evolution of lung lesions in eight survivors with significant paraquat-induced lung injuries who could be followed-up for longer than 6 months. "
10/12/2004 - "The aim of this study was to evaluate the time course of in vivo and in vitro respiratory mechanics and examine whether these parameters could reflect the temporal changes in lung parenchyma remodelling in paraquat (PQ)-induced lung injury. "
09/01/1992 - "The purpose of this study was to establish a chronic model of paraquat-induced lung injury. "
3. Acute Lung Injury
01/01/2015 - "We hypothesized that SF and OMT combination treatment might have protective effects on paraquat-induced acute lung injury. "
06/01/2012 - "This study investigated the effect of the bark extract of Bathysa cuspidata on paraquat (PQ)-induced extra-pulmonary acute lung injury (ALI) and mortality in rats. "
02/01/2015 - "In conclusion, paraquat can induce IL-1β/IL-18 secretion via NLRP3-ASC-caspase-1 pathway, and the NLRP3 inflammasome is essential for paraquat-induced acute lung injury. "
02/01/2015 - "Acute lung injury was induced by administering paraquat (PQ) intraperitoneally. "
02/01/2015 - "The innate immune response is important in paraquat-induced acute lung injury, but the exact pathways involved are not elucidated. "
4. Fibrosis (Cirrhosis)
04/01/1987 - "[An ultrastructural study of paraquat-induced intra-alveolar fibrosis of the lung]."
01/01/1987 - "Studies in experimental animals have demonstrated that paraquat produces an acute damaging phase in the lung, followed by a reparative phase dominated by an extensive fibrosis. "
01/01/2014 - " immune system plays an important role in exacerbating paraquat-induced lung fibrosis. "
06/01/2013 - "[The relationship of endoplasmic reticulum stress with paraquat induced lung fibrosis in rats]."
07/01/2012 - "Paraquat (PQ) is the second most widely used herbicide worldwide, but it selectively accumulates in human lungs to cause oxidative injury and fibrosis with high mortality. "
5. Pulmonary Fibrosis (Hamman Rich Syndrome)
10/01/2008 - "TET has antagonistic effect against acute toxicity of paraquat through significant reduction of pulmonary fibrosis."
07/01/2013 - "Docosahexaenoic acid (DHA) ameliorates paraquat-induced pulmonary fibrosis in rats possibly through up-regulation of Smad 7 and SnoN."
11/15/2013 - "This study was carried out to highlight the role of PPARγ in the paraquat (PQ)-induced pulmonary fibrosis. "
07/01/2002 - "The next target of study will be to verify the mechanism of pulmonary fibrosis by paraquat on the basis of the outcome of studies such as this review."
06/01/1999 - "Paraquat is a commonly used herbicide world wide and is a well documented cause of pulmonary fibrosis in studies of laboratory animals and in humans after exposure to a high dose (usually accidental or as parasuicide). "
1. Cyclophosphamide (Cytoxan) 2. Methylprednisolone 3. Ascorbic Acid (Vitamin C) 4. Xuebijing 5. Antioxidants 6. Charcoal (Charbon) 7. Dexamethasone (Maxidex) 8. Sodium 9. 1- Methyl- 4- phenyl- 1,2,3,6- tetrahydropyridine (MPTP) 10. Vitamin E
1. Hemoperfusion 2. Renal Dialysis (Hemodialysis) 3. Transplantation (Transplant Recipients) 4. Continuous Positive Airway Pressure 5. Emergency Treatment


On May 11, 2016, the federal Defend Trade Secrets Act (DTSA) became law. The DTSA provides trade-secret protections on the federal level that are similar to those available through the Uniform Trade Secrets Act (UTSA) adopted (with variations) in 48 States. The DTSA will have at least three effects upon private funds, particularly those with public investors.
First, the DTSA has a broader definition of “trade secret” than the UTSA, which means that more fund materials (e.g., formation and governing documents) may now qualify for protection from disclosure under State open-records and open-meetings laws. For example, where a State law provides that “other law” can exempt certain information from disclosure, the DTSA may now qualify as that separate basis to protect fund documents possessed by public investors.
Second, the DTSA independently may provide further protections for fund documents as “trade secrets” under its broader definition. The DTSA (through the Economic Espionage Act of 1996) defines a “trade secret” to include “all forms and types of financial, business, scientific, technical, economic, or engineering information,” which is broader than the “information” without elaboration protected under the UTSA.
Further, while the UTSA only specifically includes “formula, pattern, compilation, program, device, method, technique, or process” as examples of protected “information,” the DTSA also lists plans, compilations, procedures, and codes, regardless of how they are memorialized and whether they are tangible or intangible. These broader categories of specifically protected information may assist private funds not only in protecting information from disclosure under open-records laws, but may provide additional protection and enforcement options for unauthorized disclosure of fund documents or information.
Third, the DTSA may affect a private fund’s disclosure strategy and litigation options when unauthorized disclosure occurs. The DTSA provides for
ex parte civil seizure of trade-secret information in extraordinary circumstances, a remedy not previously available under the UTSA. The DTSA, however, also grants civil and criminal immunity to whistleblowers who disclose trade secrets to the government in confidence, and requires employers to notify employees of whistleblower immunity. Private funds should therefore consider reviewing employment manuals and contracts to account for these new whistleblower protections.
With the passage of the DTSA, private funds should be cognizant of their obligations regarding whistleblower immunity, and of potential new grounds to assert trade-secret protection and seek additional enforcement options where misappropriation occurs.


The Governor's Office of Planning & Budget is committed to helping Utah grow in an intelligent and responsible manner by creating tools and managing programs that assist our communities avoid the costs and inefficiencies associated with the lack of planning and coordination.
The mission of the Planning Section is to:
- promote quality growth -
- plan for critical land conservation -
- improve planning practice -
-facilitate intergovernmental relations -
Providing technical assistance to local governments is one of the critical aspects of the mission of GOPB. Unfortunately, GOBP staff is limited in its ability to reach every community desiring assistance. To address this issue, GOPB occassionally brokers arrangements between local governments and planing students from Utah's universities.
Each project is unique. Issues such as scope, schedule, and budget will vary. However, in every case, GOPB staff members can serve in the capacity of a mentor.


Gingival Recession Around Implants: A 1-Year Longitudinal Prospective Study
Paula N. Small, DDS, MPH, Dennis P. Tarnow, DDS
PMID: 10960986
A longitudinal study was performed, which measured the soft tissue around implants following surgery, to determine if a predictable pattern of soft tissue changes could be identified. This study evaluated 63 implants in 11 patients. Baseline measurements were recorded at stage 2 surgery in 2-stage implant systems, and at stage 1 surgery in the 1-stage system. Subsequent measurements were recorded at 1 week, 1 month, 3 months, 6 months, 9 months, and 1 year after baseline measurements. The majority of the recession occurred within the first 3 months, and 80% of all sites exhibited recession on the buccal. It is therefore recommended that one wait 3 months for the tissue to stabilize before either selecting a final abutment or making a final impression. As a general rule, one can anticipate approximately 1 mm of recession from the time of abutment connection surgery. A comparison of this studys results with data recorded in previously reported studies reveals clinically significant trends in the nature of soft tissue healing. (INT J ORAL MAXILLOFAC IMPLANTS 2000;15:527–532) Key words: dental abutment, dental implantation, endosseous dental implants, gingival recession, mouth mucosa