Inducible Protein-10 Research Articles

Association of plasma and synovial fluid interferon-γ inducible protein-10 with radiographic severity in knee osteoarthritis

Objectives The objective of this study was to investigate interferon-γ inducible protein-10 (IP-10) concentrations in plasma and synovial fluid of patients with knee osteoarthritis (OA) and to analyze their relationship with disease severity. Design and methods Forty OA patients and 15 healthy controls were enrolled in this study. OA grading was performed according to the Kellgren–Lawrence criteria. IP-10 levels in plasma and synovial fluid were assessed using enzyme-linked immunosorbent assay. Results Plasma IP-10 levels in the knee OA patients were significantly lower than those of controls ( P = 0.006). IP-10 levels in plasma were markedly higher with regard to paired synovial fluid ( P < 0.001). Furthermore, IP-10 concentrations in plasma and synovial fluid displayed significant inverse correlation with radiographic severity ( r = − 0.713, P < 0.001 and r = − 0.561, P < 0.001, respectively). Subsequent analysis revealed that plasma IP-10 levels were positively correlated with synovial fluid IP-10 levels ( r = 0.424, P = 0.006). Conclusions IP-10 levels in both plasma and synovial fluid were inversely associated with the severity of knee OA. Accordingly, IP-10 could serve as a biomarker for determining disease severity and might play a possible role in the pathophysiology of osteoarthritis.

Green Tea Epigallo-Catechin-Galleate Ameliorates the Development of Obliterative Airway Disease

ABSTRACTLung transplantation has the worst outcome compared to all solid organ transplants due to chronic rejection known as obliterative bronchiolitis (OB). Pathogenesis of OB is a complex interplay of alloimmune-dependent and -independent factors, which leads to the development of inflammation, fibrosis, and airway obliteration that have been resistant to therapy. The alloimmune-independent inflammatory pathway has been the recent focus in the pathogenesis of rejection, suggesting that targeting this may offer therapeutic benefits. As a potent anti-inflammatory agent, epigallo-catechin-galleate (EGCG), a green tea catechin, has been very effective in ameliorating inflammation in a variety of diseases, providing the rationale for its use in this study in a murine heterotopic tracheal allograft model of OB. Mice treated with EGCG had reduced inflammation, with significantly less neutrophil and macrophage infiltration and significantly reduced fibrosis. On further investigation into the mechanisms, inflammatory cytokines keratinocyte (KC), interleukin-17 (IL-17), and tumor necrosis factor-α (TNF-α), involved in neutrophil recruitment, were reduced in the EGCG-treated mice. In addition, monocyte chemokine monocyte chemoattractant protein-1 (MCP-1) was significantly reduced by EGCG treatment. Antifibrotic cytokine interferon-γ–inducible protein-10 (IP-10) was increased and profibrotic cytokine transforming growth factor-β (TGF-β) was reduced, further characterizing the antifibrotic effects of EGCG. These findings suggest that EGCG has great potential in ameliorating the development of obliterative airway disease.

New and Evolving Management Paradigms for Hepatitis C after Liver Transplantation

Hepatitis-C induced liver failure is the leading indication for liver transplantation in the United States, and the burden of hepatitis C virus (HCV)-induced liver disease is not expected to peak for at least another decade. 2011 will usher in a new era of directly acting antiviral therapies and personalized medicine that will assist patients and clinicians in choosing the best drug regimen. Specific markers to predict sustained virologic response (SVR) in the posttransplant setting are under development, and the role of graft genetic markers like interleukin-28B and interferon-γ inducible protein-10 have yet to be fully defined. Lessons and experiences from treating the pretransplant population will be applied to patients with recurrent posttransplant HCV while studies specific to this population proceed. New paradigms for HCV treatment give promise to reducing the pretransplant burden of disease and improving SVR rates in the posttransplant population.

The genome of self-complementary adeno-associated viral vectors increases Toll-like receptor 9–dependent innate immune responses in the liver

AbstractAlthough adeno-associated viral (AAV) vectors have been successfully used in hepatic gene transfer for treatment of hemophilia and other diseases in animals, adaptive immune responses blocked long-term transgene expression in patients on administration of single-stranded AAV serotype-2 vector. More efficient vectors have been developed using alternate capsids and self-complimentary (sc) genomes. This study investigated their effects on the innate immune profile on hepatic gene transfer to mice. A mild and transient up-regulation of myeloid differentiation primary response gene (88), TLR9, TNF-α, monocyte chemotactic protein-1, IFN-γ inducible protein-10, and IFN-α/β expression in the liver was found after single-stranded AAV vector administration, regardless of the capsid sequence. In contrast, scAAV vectors induced higher increases of these transcripts, upregulated additional proinflammatory genes, and increased circulating IL-6. Neutrophil, macrophage, and natural killer cell liver infiltrates were substantially higher on injection of scAAV. Some but not all of these responses were Kupffer cell dependent. Independent of the capsid or expression cassette, scAAV vectors induced dose-dependent innate responses by signaling through TLR9. Increased innate responses to scAAV correlated with stronger adaptive immune responses against capsid (but not against the transgene product). However, these could be blunted by transient inhibition of TLR9.

Reduced surface toll-like receptor-4 expression and absent interferon-γ–inducible protein-10 induction in cystic fibrosis airway cells

ABSTRACTAs part of the innate and adaptive immune system, airway epithelial cells secrete proinflammatory cytokines after activation of Toll-like receptors (TLRs) by pathogens. Nevertheless, cystic fibrosis (CF) airways are chronically infected with Pseudomonas aeruginosa, suggesting a modified immune response in CF. The authors have shown that in CF bronchial epithelial cells, a reduced surface expression of TLR-4 causes a diminished interleukin (IL)-8 and IL-6 response upon lipopolysaccharide (LPS) stimulation. However, there is no information regarding activation of the MyD88 (myeloid differentiation primary response gene 88)-independent TLR-4 signaling pathway by LPS, which results in the activation of adaptive immune responses by secretion of the T cell–recruiting chemokine interferon-γ–inducible protein (IP)-10. Therefore, the authors investigated the induction of IP-10 in CF bronchial epithelial cell line CFBE41o− and its CFTR-corrected isotype under well-differentiating conditions. TLR-4 surface expression was significantly reduced in CFBE41o− by a factor of 2, compared to the CFTR-corrected cells. In CFTR-corrected cells, stimulation with LPS increased IP-10 secretion. Incubating cells with siRNA directed against TLR-4 inhibited the LPS stimulated increase of IP-10 in CFTR-corrected cells. The reduced TLR-4 surface expression in CF cells causes the loss of induction of IP-10 by LPS. This could compromise adaptive immune responses in CF due to a reduced T-cell recruitment.

Systemic Effects of White Adipose Tissue Dysregulation and Obesity‐Related Inflammation

Systemic Effects of White Adipose Tissue Dysregulation and Obesity‐Related Inflammation

IL28B polymorphisms, IP-10 and viral load predict virological response to therapy in chronic hepatitis C

Hepatitis C virus (HCV) is a major cause of chronic liver disease, cirrhosis and hepatocellular carcinoma and the identification of the predictors of response to antiviral therapy is an important clinical issue. To determine the independent contribution of factors including IL28B polymorphisms, IFN-gamma inducible protein-10 (IP-10) levels and the homeostasis model assessment of insulin resistance (HOMA-IR) score in predicting response to therapy in chronic hepatitis C (CHC). Multivariate analysis of factors predicting rapid (RVR) and sustained (SVR) virological response in 280 consecutive, treatment-naive CHC patients treated with peginterferon alpha and ribavirin in a prospective multicentre study. Independent predictors of RVR were HCV RNA <400 000 IU/mL (OR 11.37; 95% CI 3.03-42.6), rs12980275 AA (OR 7.09; 1.97-25.56) and IP-10 (OR 0.04; 0.003-0.56) in HCV genotype 1 patients and lower baseline γ-glutamyl-transferase levels (OR=0.02; 0.0009-0.31) in HCV genotype 3 patients. Independent predictors of SVR were rs12980275 AA (OR 9.68; 3.44-27.18), age <40 years (OR=4.79; 1.50-15.34) and HCV RNA <400 000 IU/mL (OR 2.74; 1.03-7.27) in HCV genotype 1 patients and rs12980275 AA (OR=6.26; 1.98-19.74) and age <40 years (OR 5.37; 1.54-18.75) in the 88 HCV genotype 1 patients without a RVR. RVR was by itself predictive of SVR in HCV genotype 1 patients (OR 33.0; 4.06-268.32) and the only independent predictor of SVR in HCV genotype 2 (OR 9.0, 1.72-46.99) or genotype 3 patients (OR 7.8, 1.43-42.67). In HCV genotype 1 patients, IL28B polymorphisms, HCV RNA load and IP-10 independently predict RVR. The combination of IL28B polymorphisms, HCV RNA level and age may yield more accurate pre-treatment prediction of SVR. HOMA-IR score is not associated with viral response.

The 'interferon paradox' as a predictor of non-response to anti-viral therapy: novel association with apolipoprotein E and hepatitis C virus (HCV) lipoviralparticles (LVP).

Introduction Identification of factors that influence response to treatment are important for individualised care in the era of direct acting antivirals. Interferon-γ inducible protein-10 (IP10) is one of hundreds of interferon stimulated genes (ISGs). Paradoxically, high ISGs, notably IP10 are associated with poor treatment response. Although HCV hijacks the VLDL pathway in its life cycle, another paradox is that high LDL cholesterol is a marker of favourable outcome. We have shown that infectious HCV LVP correlate with insulin resistance and that high LVP ratio (LVP ratio = LVP/LVP + non LVP) is associated with lower EVR.1 Aim To evaluate the interactions between LVP, IP10 and lipid profiles. Methods Fasting samples were collected from 71 consecutive HCV G1 patients. LVP and LVP ratio were determined as previously described.1 Fasting lipids, apoB and apoE were measured using automated enzymatic methods. IP10 was measured using a commercially available ELISA (R&D Systems, Abingdon, UK). Results Univariate analysis demonstrated a relationship between IP10 and LVP ratio (r=0.31, p=0.028) but not between total viral load and IP10 (r=0.005, p=0.973). IP10 levels inversely correlated with LDL cholesterol (r=-0.302, p=0.017) and apoB (r=-0.295, p=0.015). IP10 levels also strongly positively correlated with apoE (r=0.444 p Conclusion This data indicates there is a close link between ISGs and lipoprotein metabolism. High IP10 correlates with apoE which is an exchangeable lipoprotein that transfers from HDL to triglyceride-rich lipoprotein remnants where it mediates clearance via LDLR, a candidate HCV receptor. ApoE is important in HCV replication/assembly and is a constituent of infectious LVP. Hence the effect of the endogenous interferon response on lipids is favourable to the virus by promoting LVP. This provides a possible explanation for the interferon paradox.

Changes in helper lymphocyte chemokine receptor expression and elevation of IP‐10 during acute respiratory syncytial virus infection in infants

It is known that lymphopenia caused by apoptosis may occur during severe respiratory syncytial virus (RSV) infection. However, further evidence about how T-cell subsets may be affected in infants during severe RSV bronchiolitis is needed to understand the mechanisms through which immunological memory may be altered. There is increasingly convincing evidence that RSV may be associated with the development of atopy and asthma. Surrogates of Th1, Th2 and regulatory T-lymphocyte populations were measured in blood from children with acute RSV bronchiolitis and in convalescence using the cell surface receptors CXCR3, CCR4 and CD25, respectively. Samples were also obtained from healthy age-matched controls. Plasma levels of the chemokines interferon-γ inducible protein-10 (IP-10) and thymus and activation-regulated chemokine (TARC), which are known ligands for CXCR3 and CCR4, were also measured. Free plasma DNA was measured using quantitative PCR. CXCR3-positive cells were significantly decreased during acute infection (p = 0.013), while CCR4 and CD25 T-cell populations were unchanged. Plasma levels of IP-10 were markedly elevated in acute infection (p = 0.001). Convalescent samples were not significantly different to control samples for lymphocyte phenotypes or plasma chemokines. Elevated free plasma DNA was detected during acute infection compared with convalescence and controls. A profound reduction in the Th1, but not Th2, and CD25-positive lymphocyte populations associated with exaggerated IP-10 production occurs in severe RSV bronchiolitis. Free DNA is detectable in plasma. This may allow significant alterations in the generation of T-cell memory.

Circulating Interleukin (IL)-8, IL-2R, IL-12, and IL-15 Levels Are Independently Prognostic in Primary Myelofibrosis: A Comprehensive Cytokine Profiling Study

Abnormal cytokine expression accompanies myelofibrosis and might be a therapeutic target for Janus-associated kinase (JAK) inhibitor drugs. This study describes the spectrum of plasma cytokine abnormalities in primary myelofibrosis (PMF) and examines their phenotypic correlates and prognostic significance. Patients included in this study were required to have archived plasma, bone marrow biopsy, and cytogenetic information available at the time of first referral to the Mayo Clinic. Multiplex biometric sandwich immunoassay was used to measure plasma levels of 30 cytokines. In total, 127 PMF patients were studied; comparison with normal controls (n = 35) revealed significantly increased interleukin-1β (IL-1β), IL-1RA, IL-2R, IL-6, IL-8, IL-10, IL-12, IL-13, IL-15, tumor necrosis factor α (TNF-α), granulocyte colony-stimulating factor (G-CSF), interferon alfa (IFN-α), macrophage inflammatory protein 1α (MIP-1α), MIP-1β, hepatocyte growth factor (HGF), IFN-γ-inducible protein 10 (IP-10), monokine induced by IFN-γ (MIG), monocyte chemotactic protein 1 (MCP-1), and vascular endothelial growth factor (VEGF) levels and decreased IFN-γ levels. In treatment-naive patients (n = 90), increased levels of IL-8 (P < .001), IL-2R (P < .001), IL-12 (P < .001), IL-15 (P = .001), and IP-10 (P = .003) were independently predictive of inferior survival. A similar multivariable analysis that included all 127 study patients confirmed the prognostic value of these five cytokines, and IL-8, IL-2R, IL-12, and IL-15 remained significant when risk stratification, according to the recently revised Dynamic International Prognostic Scoring System (DIPSS plus), was added to the multivariable model. Leukemia-free survival was predicted by IL-8, which was also the only cytokine associated with ≥ 1% circulating blasts. Other cytokine-phenotype associations included increased IL-8 and constitutional symptoms; IL-2R, IL-12, and transfusion need; IL-2R, IL-8, and leukocytosis; IP-10 and thrombocytopenia; HGF, MIG, IL-1RA, and marked splenomegaly; and IL-1RA, IL-2R, IP-10, MIP-1β, and JAK2V617F. A two-cytokine (IL-8/IL-2R) -based risk categorization delineated prognostically different groups within specific DIPSS plus risk categories. This study signifies the presence of specific cytokine-phenotype associations in PMF and a prognostically relevant plasma cytokine signature that might prove useful as a laboratory tool for predicting and monitoring treatment response.

인간 호중구의 세포사멸과 시토카인 분비에 대한 아포지방단백 A-I의 영향

Apolipoprotein A-I (apoA-I) has anti-inflammatory and anti-oxidative properties. This study was designed to investigate whether apoA-I affects apoptosis and cytokine production of human blood neutrophils in an in vitro culture system. Spontaneous apoptosis of neutrophils was significantly delayed by apoA-I. In addition, high density lipoprotein containing apoA-I also delayed apoptosis of neutrophils. Apoptosis of neutrophils was inhibited by anti-scavenger receptor type B-I antibodies. The amounts of interleukin-8, interferon (IFN)-inducible protein 10 (IP-10), and tumor necrosis factor-α (TNF-α) in the supernatants of cultured neutrophils treated with apoA-I were significantly increased. Combined treatment of neutrophils with IFN-γ and apoA-I produced higher amounts of IP-10 and TNF-α than did treatment with IFN-γ or apoA-I alone. The present study reveals that apoA-I activates neutrophils to produce cytokines and delays spontaneous apoptosis of neutrophils. These findings suggest that apoA-I, although a well-known negative acute-phase protein, has a pro-inflammatory effect in neutrophils.

Impact of a probiotic Enterococcus faecalis in a gnotobiotic mouse model of experimental colitis

IL-10-deficient (IL-10(-/-) ) mice are susceptible to the development of chronic intestinal inflammation in response to the colonization with commensal Enterococcus faecalis isolates. The aim of this study was to characterize the impact of a probiotic E. faecalis strain in germ-free, wild-type (WT), and disease-susceptible IL-10(-/-) mice. The probiotic E. faecalis and the colitogenic control strain OG1RF induced IL-6 and IFN-γ inducible protein-10 secretion in the murine intestinal epithelial cell line Mode K. Epithelial cell activation involved nuclear factor κ B, p38 and extracellular signal-regulated kinase 1/2-dependent pathways. Mouse embryonic fibroblasts from WT and toll-like receptor-2-deficient (TLR-2(-/-) ) mice confirmed that both E. faecalis strains trigger pro-inflammatory responses via the pattern recognition receptor TLR-2. Monoassociation of germ-free IL-10(-/-) mice with the probiotic E. faecalis strain revealed pro-inflammatory epithelial cell activation and colonic tissue pathology. The non-pathogenic nature of E. faecalis was confirmed in monoassociated WT mice. 2-DE and MALDI-TOF MS identified the ER stress chaperone Hspa5 (glucose-regulated protein 78) and 3-mercaptopyruvate sulfurtransferase as key targets in the epithelium from IL-10(-/-) and TLR-2(-/-) mice. This study shows the potential of probiotic bacteria to initiate pro-inflammatory responses in the disease-susceptible but not the normal host.

Parthenolide inhibits TRIF-dependent signaling pathway of Toll-like receptors in RAW264.7 macrophages.

Toll-like receptors (TLRs) play an important role in induction of innate immune responses for host defense against invading microbial pathogens. Microbial component engagement of TLRs can trigger the activation of myeloid differential factor 88 (MyD88)- and toll-interleukin-1 receptor domain-containing adapter inducing interferon-β (TRIF)-dependent downstream signaling pathways. Parthenolide, an active ingredient of feverfew (Tanacetum parthenium), has been used for centuries to treat many chronic diseases. Parthenolide inhibits the MyD88-dependent pathway by inhibiting the activity of inhibitor-κB kinase. However, it is not known whether parthenolide inhibits the TRIF-dependent pathway. To evaluate the therapeutic potential of parthenolide, its effect on signal transduction via the TRIF-dependent pathway of TLRs induced by lipopolysaccharide (LPS) or polyinosinic-polycytidylic acid (poly [I:C]) was examined. Parthenolide inhibited nuclear factor-κB and interferon regulatory factor 3 activation induced by LPS or poly[I:C], and the LPS-induced phosphorylation of interferon regulatory factor 3 as well as interferon-inducible genes such as interferon inducible protein-10. These results suggest that parthenolide can modulate TRIF-dependent signaling pathways of TLRs, and may be the basis of effective therapeutics for chronic inflammatory diseases.

Twelve potential fibrosis markers to differentiate mild liver fibrosis from cirrhosis in patients infected with chronic hepatitis C genotype 1

Information about the stage of liver fibrosis is important for managing patients with chronic hepatitis C (CHC). The aim of this study was to evaluate 12 plasma markers for differentiating no/mild liver fibrosis from cirrhosis among patients with CHC genotype 1. Transient elastography was used to assess the stage of fibrosis for the patients included in the study. Forty patients were included (21 cirrhotic). Plasma levels of tumor necrosis factor-α (TNF-α), interleukin 8 (IL-8), interferon-γ inducible protein-10 (IP-10), monocyte chemotactic protein-1 (MCP-1), soluble urokinase-type plasminogen activator (suPAR), monokine induced by γ-interferon (MIG), human hepatocyte growth factor (HGF), insulin, interleukin 6 (IL-6), interleukin 1-β (IL-1β), leptin, and nerve growth factor (NGF) were analyzed. Concentrations of TNF-α (median 15.0 vs. 25.1 pg/ml, area under the receiver operating characteristic curve [AUC] 0.91), IL-8 (48.7 vs. 103.3 pg/ml, AUC 0.85), IP-10 (176 vs. 566 pg/ml, AUC 0.83), MCP-1 (449 vs. 735 pg/ml, AUC 0.78), suPAR (3.5 vs. 5.2 ng/ml, AUC 0.78), MIG (100 vs. 152 pg/ml, AUC 0.75), and HGF (3.69 vs. 5.58 ng/ml, AUC 0.71) were significantly higher in patients with cirrhosis. In conclusion, several of the investigated markers showed promise for differentiating cirrhosis from no/mild fibrosis among patients with CHC genotype 1.

Quercetin attenuates inflammation in human macrophages and adipocytes exposed to macrophage-conditioned media

Obesity is linked to chronic inflammation in white adipose tissue, which is exacerbated by infiltrating macrophages (MΦs). We recently demonstrated that an extract from grape powder (GPE), which is abundant in quercetin (QUE), reduced inflammation in human MΦs and prevented MΦ-mediated inflammation and insulin resistance in human adipocytes. However, we did not know how QUE individually affected these outcomes. We examined the extent to which QUE prevents inflammation in human MΦs (that is, differentiated U937 cell line) and cross-talk with human adipocytes (that is, primary cultures of newly differentiated human adipocytes). Treatment of MΦs with QUE attenuated the basal expression of inflammatory genes, such as tumor necrosis factor-α, interleukin (IL)-6, IL-8, IL-1β and interferon-γ inducible protein-10, and cyclooxygenase-2, a marker of prostaglandin production. QUE also attenuated the abundance of phosphorylated c-Jun N-terminal kinase (JNK) and c-Jun, and IκBα degradation in MΦs. Furthermore, conditioned media (CM) obtained from MΦs treated with QUE decreased the capacity of this CM to inflame adipocytes and cause insulin resistance as evidenced by decreased: (1) inflammatory gene expression, (2) phosphorylation of JNK and c-Jun, (3) serine residue 307 phosphorylation of insulin receptor substrate (IRS)-1, 4) protein tyrosine phosphatase-1B gene expression and 5) suppression of insulin-stimulated glucose uptake. Taken together, these data suggest that QUE is one of the bioactive components of GPE that prevents inflammation in MΦs and MΦ-mediated insulin resistance in adipocytes.

Potential role and mechanism of IFN-gamma inducible protein-10 on receptor activator of nuclear factor kappa-B ligand (RANKL) expression in rheumatoid arthritis

IntroductionIFN-gamma inducible protein-10 (CXCL10), a member of the CXC chemokine family, and its receptor CXCR3 contribute to the recruitment of T cells from the blood stream into the inflamed joints and have a crucial role in perpetuating inflammation in rheumatoid arthritis (RA) synovial joints. Recently we showed the role of CXCL10 on receptor activator of nuclear factor kappa-B ligand (RANKL) expression in an animal model of RA and suggested the contribution to osteoclastogenesis. We tested the effects of CXCL10 on the expression of RANKL in RA synoviocytes and T cells, and we investigated which subunit of CXCR3 contributes to RANKL expression by CXCL10.MethodsSynoviocytes derived from RA patients were kept in culture for 24 hours in the presence or absence of TNF-α. CXCL10 expression was measured by reverse transcriptase polymerase chain reaction (RT-PCR) of cultured synoviocytes. Expression of RANKL was measured by RT-PCR and western blot in cultured synoviocytes with or without CXCL10 and also measured in Jurkat/Hut 78 T cells and CD4+ T cells in the presence of CXCL10 or dexamethasone. CXCL10 induced RANKL expression in Jurkat T cells was tested upon the pertussis toxin (PTX), an inhibitor of Gi subunit of G protein coupled receptor (GPCR). The synthetic siRNA for Gαi2 was used to knock down gene expression of respective proteins.ResultsCXCL10 expression in RA synoviocytes was increased by TNF-α. CXCL10 slightly increased RANKL expression in RA synoviocytes, but markedly increased RANKL expression in Jurkat/Hut 78 T cell or CD4+ T cell. CXCL10 augmented the expression of RANKL by 62.6%, and PTX inhibited both basal level of RANKL (from 37.4 ± 16.0 to 18.9 ± 13.0%) and CXCL10-induced RANKL expression in Jurkat T cells (from 100% to 48.6 ± 27.3%). Knock down of Gαi2 by siRNA transfection, which suppressed the basal level of RANKL (from 61.8 ± 17.9% to 31.1 ± 15.9%) and CXCL10-induced RANKL expression (from 100% to 53.1 ± 27.1%) in Jurkat T cells, is consistent with PTX, which inhibited RANKL expression.ConclusionsCXCL10 increased RANKL expression in CD4+ T cells and it was mediated by Gαi subunits of CXCR3. These results indicate that CXCL10 may have a potential role in osteoclastogenesis of RA synovial tissue and subsequent joint erosion.

Role of Cytotoxic Protease Granzyme‐b in Neuronal Degeneration During Human Stroke

Infiltration of leukocytes into post-ischemic cerebrum is a well-described phenomenon in stroke injury. Because CD-8(+) T-lymphocytes secrete cytotoxic proteases, including granzyme-b (Gra-b) that exacerbates post-ischemic brain damage, we investigated roles of Gra-b in human stroke. To study the role of Gra-b in stroke, ischemic and non-ischemic tissues (from post-mortem stroke patients) were analyzed using immunoblotting, co-immunoprecipitation, terminal deoxy uridine nick end labeling (TUNEL) and Annexin-V immunostaining, and in vitro neuron survival assays. Activated CG-SH cells and supernatants were used to model leukocyte-dependent injury. Non-ischemic brain tissues were used as non-pathological controls. Non-activated CG-SH cells and supernatants were used as controls for in vitro experiments. Human stroke (ischemic) samples contained significantly higher levels of Gra-b and interferon-gamma inducible protein-10 (IP-10/CXCL10) than non-ischemic controls. In stroke, poly (ADP-ribose) polymerase-1 and heat shock protein-70 were cleaved to canonical proteolytic "signature" fragments by Gra-b. Gra-b was also found to bind to Bid and caspase-3. Gra-b also co-localized with Annexin-V(+) /TUNEL(+) in degenerating neurons. Importantly, Gra-b inhibition protected both normal and ischemia-reperfused neurons against in vitro neurotoxicity mediated by activated CG-SH cells and supernatants. These results suggest that increased leukocyte infiltration and elevated Gra-b levels in the post-stroke brain can induce contact-dependent and independent post-ischemic neuronal death to aggravate stroke injury.

Response of periodontitis and healthy patients in a Porphyromonas gingivalis-stimulated whole-blood model.

To investigate the inflammatory responses of periodontitis patients and healthy patients in a whole-blood model stimulated with Porphyromonas gingivalis. Whole blood collected from 17 periodontitis patients and six healthy patients was stimulated with Porphyromonas gingivalis cells. The secretion of cytokines and matrix metalloproteinases was quantified by enzyme-linked immunosorbent assay. An analysis of covariance with the ancova model was used to evaluate the significance of differences in secreted host molecules by whole blood from the periodontitis and healthy groups. Porphyromonas gingivalis induced the secretion of interleukin-1β, interleukin-6, interleukin-8, tumor necrosis factor-α, monocyte chemoattractant protein-1, interferon inducible protein-10 by whole blood from patients in the periodontitis and healthy groups. Matrix metalloproteinase-8 and -9 levels secreted by whole blood also increased following stimulation. No significant differences (P < 0.05) in the amounts of secreted host molecules were observed between periodontitis and healthy patients. This study suggests that Porphyromonas gingivalis can provoke an inflammatory response and promote the progression of periodontitis by inducing the secretion of high levels of cytokines and matrix metalloproteinases by a mixed leukocyte population. However, the whole-blood model did not reveal any significant differences in the inflammatory response between periodontitis patients (n = 17) and periodontally-healthy patients (n = 6).

181 Interferon-γ, Interferon-γ-Inducible Protein-10 and Tuberculin Skin Test Responses of Hiv-Positive and Hiv Negative Children in Northeast Brazil

Background and aims: Detection of latent tuberculosis (TB) infection (LTBI) and provision of chemoprophylaxis is an important strategy for TB control, particularly for HIV-infected children. Assessing LTBI in this population is difficult because the tuberculin skin test (TST) is unreliable in immunosuppressed individuals. Interferon-γ release assays (QTF) have been recently used as complementary tools for the diagnosis of LTBI and Interferon-γ inducible protein-10 (IP-10) has been evaluated as a potential marker for detection of Mycobacterium tuberculosis infection. Our aim was to describe Interferon-γ, IP-10 and TST responses in HIV-positive and negative children in Northeast Brazil. Methods: TST, Interferon-γ and IP-10 responses were evaluated in HIV-positive and HIV-negative children with known contact with adults with TB, a previous or current diagnosis of active TB and controls with no history of TB contact or disease. Results: 41 HIV-positive and 56 HIV-negative children were enrolled. The concordance of TST and QTF was high amongst HIV-positive and HIV-negative children (k 0.85 and k 0.82 respectively). IP-10 concentrations were higher for both HIV-positive and negative children with positive QTF/TST agreement. HIV-positive children had lower IP-10 concentrations when compared with HIV-negative children. Interferon-γ and IP-10 concentrations did not significantly differ amongst HIV-positive children with active TB, TB Contact or controls. Among HIV-positive children, higher IP-10 concentrations were observed in children with higher CD4 counts. Conclusions: IP-10 concentrations had good correlation with positive QTF/TST agreement, reinforcing its potential role for the diagnosis of LTBI. Interferon-γ and IP-10 responses in HIV-positive children need to be further investigated.

C1q Deficiency Leads to the Defective Suppression of IFN-α in Response to Nucleoprotein Containing Immune Complexes

Almost all humans with homozygous deficiency of C1q develop systemic lupus erythematosus (SLE). The precise cellular mechanism(s) by which C1q prevents the development of SLE remains unclear. In this study, we tested the role of C1q in the regulation of IFN-α induced by immune complexes (ICs) in vitro, as well as the consequences of lack of C1q in vivo. Our experiments revealed that C1q preferentially promotes the binding of SLE ICs to monocytes rather than plasmacytoid dendritic cells, but this inhibition was not due to the induction of inhibitory soluble factors. The presence of C1q also altered the trafficking of ICs within monocytes such that ICs persisted in early endosomes. In patients with C1q deficiency, serum and cerebrospinal fluid levels of IFN-α and IFN-γ-inducible protein-10 levels were elevated and strongly correlated with Ro autoantibodies, demonstrating the clinical significance of these observations. These studies therefore associate C1q deficiency with defective regulation of IFN-α and provide a better understanding of the cellular mechanisms by which C1q prevents the development of IC-stimulated autoimmunity.