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The Role of TIMP3 in Models of Inflammation and ImmunitySmookler, David 01 September 2010 (has links)
The inter-relation between inflammation, the immune system and leukocytes is multifaceted, with communication between stroma and immune cells mediated by cytokines, growth factors, chemokines, integrins and other molecules. Proteolysis plays an important role in regulating these molecules. Proteolytic cleavage can not only destroy some molecules but can activate or shed others, converting local juxtacrine signalling proteins into effectors that act at a distance. Shedding can also convert membrane-bound receptors into soluble ligand-binding inhibitors. Finally, cleavage can convert agonist molecules into antagonists. As a wide-ranging inhibitor of metalloproteinases, tissue inhibitor of metalloproteinase 3 (TIMP3) has the potential to down-regulate many of these activities. We explore the role of TIMP3 in the regulation of inflammation, revealing that loss of TIMP3 leads to a more rapid increase of soluble TNF, higher levels of soluble TNF receptors and ultimately to increased TNF signalling in systemic inflammation. We also demonstrate TIMP3 loss impacts local inflammation. In addition we investigate the importance of TIMP3 in the expansion of hematopoietic cells.
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Soluble receptors for advanced glycation end products as predictors of restenosis following percutaneous coronary interventionMcNair, Erick Donnell 11 September 2009
The principal cause of non-ST-segment myocardial infarction (NSTEMI), a subclass of acute coronary syndrome (ACS), is thrombosis and the underlying cause is atherosclerosis. Percutaneous coronary intervention (PCI) is one of the treatments to attenuate the ischemic effects of severe coronary artery stenosis. However, restenosis following PCI (post-PCI) is a major problem for the long-term success of the procedure. Recently, the interaction of advanced glycation end products (AGE) with the receptor for advanced glycation end products (RAGE) has been implicated in the development of atherosclerosis in animal models. Interaction of AGE with RAGE results in activation of nuclear factor kappa-B (NF-êB), release of cytokines including tumor necrosis factor-alpha (TNF-á), the expression of adhesion molecules including soluble vascular adhesion molecule-1 (sVCAM-1) and induction of oxidative stress all of which have been implicated in the development of atherosclerosis. The soluble receptor for advanced glycation end products (sRAGE) acts as a decoy for RAGE ligands (AGEs) and this occurs by competing with RAGE. In animal models, balloon inflation and de-endothelialization of the carotid artery increase the concentration of AGE and RAGE in the arterial wall and induces neointimal hyperplasia and stenosis. Treatment with sRAGE in animal models reduces neointimal growth and decreases smooth muscle cell migration and proliferation and expression of extracellular matrix.<p>
It is hypothesized that NSTEMI and post-PCI restenosis may be due to low levels of serum sRAGE resulting in increased AGE and RAGE interactions. Low levels of sRAGE would also increase the levels of serum TNF-á and sVCAM-1.<p>
The objectives of this study were to determine whether: (1) the levels of serum sRAGE are lower and the levels of AGE, TNF-á and sVCAM-1 are higher in NSTEMI patients compared to control; (2) the levels of serum sRAGE are lower and the levels of AGE, TNF-á and sVCAM-1 are higher in NSTEMI patients with restenosis compared to those without restenosis; and (3) sRAGE or AGE/sRAGE ratio may serve as a biomarker/ predictor of NSTEMI and post-PCI restenosis.<p>
The study objectives include 46 consecutive NSTEMI patients undergoing elective PCI and 28 healthy age-matched male controls. Pre-PCI and 6 month post-PCI angiography were performed in all NSTEMI patients. Blood samples were collected at designated intervals for the measurement of sRAGE, AGE, TNF-á, and sVCAM-1 using commercially available enzyme-linked immunosorbent assay (ELISA) kits.<p>
The levels of serum sRAGE were lower and those of TNF-á, sVCAM-1, AGE and AGE/sRAGE were higher in NSTEMI patients compared to control subjects. (sRAGE, 884.55 ± 50 vs. 1287 ± 41.5 pg/mL{p<0.001}; TNF-á, 23.1 ± 2.3 vs. 10.3 ± 0.8 pg/mL {p<0.002}; sVCAM-1, 1059.62 ± 70.8 vs. 651 ± 35.5 ng/mL {p<0.0003}, AGE, 1192.50 ± 82.6 vs. 669.40 ± 47.9 ng/mL {p<0.001}; and AGE/sRAGE, 1.75 ± 0.17 vs. 0.52 ± 0.06 {p<0.001}).<p>
The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and accuracy of the sRAGE biomarker test were 59%, 100%, 100%, 100% and 74% respectively, while those of the AGE/sRAGE test were 85%, 91%, 97%, 67% and 86%, respectively, in the diagnosis of patients with NSTEMI acute coronary syndrome.
The pre-PCI levels of serum sRAGE in patients with restenosis were significantly lower (p<0.001) than in patients without restenosis (610.6 ± 24.1 vs. 1143.8 ± 52.5 pg/mL). The post-PCI levels of serum sRAGE were significantly lower (p<0.0001) in patients with restenosis compared to those without restenosis (477 ± 18.6 vs. 1106.7 ± 41.9 pg/mL). The pre-PCI levels of serum TNF-á and sVCAM-1 were significantly higher (p<0.001 and p<0.001) in patients with restenosis compared to those without restenosis (TNF-á, 37.9 ± 2.5 vs. 11.6 ± 0.41 pg/mL; sVCAM-1, 1381.8 ± 63.5 vs. 811.37 ± 26.5 ng/mL). The post PCI levels of serum TNF-á and sVCAM-1 were also significantly higher (p<0.0001 and p<0.0001) in patients with restenosis than in patients without restenosis (TNF-á, 48.4 ± 1.4 vs. 12.5 ± 0.44 pg/mL; sVCAM-1, 1381.8 ± 63.5 vs. 762.2 ± 26.4 ng/mL). The post-PCI levels sRAGE were lower while those of TNF-á and sVCAM-1 were higher compared to the pre-PCI levels in patients with restenosis. However, the pre- and post-PCI levels of serum sRAGE, TNF-á and sVCAM-1 were similar in patients without restenosis. The pre-PCI levels of serum AGE and AGE/sRAGE were significantly higher (p<0.001 and p<0.001) in patients with restenosis compared to those without restenosis (AGE, 1512.1 ± 84.53 vs. 891.7 ± 92.4 ng/mL; 2.39 ± 0.20 vs. 1.03 ± 0.17).<p>
The sensitivity, specificity, PPV, NPV, and accuracy of the pre-PCI sRAGE tests were 73%, 100%, 100%, 80%, and 87%, respectively while those of the AGE/sRAGE tests were 81%, 94%, 93%, 84% and 88%, respectively in identifying patients with post-PCI restenosis.<p>
In conclusion the results suggest that: (1) the levels of serum sRAGE are lower while those of TNF-á, sVCAM-1, AGE and AGE/sRAGE are higher in NSTEMI patients compared to control subjects; (2) serum levels of sRAGE are negatively correlated with the number of diseased vessels; (3) Both low sRAGE and high AGE/sRAGE may serve as a biomarker/predictor of NSTEMI, but AGE/sRAGE has a greater sensitivity compared to sRAGE; (4) the pre-PCI levels of serum sRAGE are lower while those of AGE/sRAGE are higher in patients with restenosis compared to those without restenosis; and (5) both low sRAGE and high AGE/sRAGE may serve as a predictor/ biomarker of post-PCI restenosis; however, AGE/sRAGE has a greater sensitivity than sRAGE.
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The Role of TIMP3 in Models of Inflammation and ImmunitySmookler, David 01 September 2010 (has links)
The inter-relation between inflammation, the immune system and leukocytes is multifaceted, with communication between stroma and immune cells mediated by cytokines, growth factors, chemokines, integrins and other molecules. Proteolysis plays an important role in regulating these molecules. Proteolytic cleavage can not only destroy some molecules but can activate or shed others, converting local juxtacrine signalling proteins into effectors that act at a distance. Shedding can also convert membrane-bound receptors into soluble ligand-binding inhibitors. Finally, cleavage can convert agonist molecules into antagonists. As a wide-ranging inhibitor of metalloproteinases, tissue inhibitor of metalloproteinase 3 (TIMP3) has the potential to down-regulate many of these activities. We explore the role of TIMP3 in the regulation of inflammation, revealing that loss of TIMP3 leads to a more rapid increase of soluble TNF, higher levels of soluble TNF receptors and ultimately to increased TNF signalling in systemic inflammation. We also demonstrate TIMP3 loss impacts local inflammation. In addition we investigate the importance of TIMP3 in the expansion of hematopoietic cells.
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Molecular Insights into the Distinct Mechanisms Regulating the TLR4 Mediated Activation, Shut Down, and Endotoxin Tolerance of the IL1B and TNF GenesAdamik, Juraj 11 October 2013 (has links)
The first wave of the inducible gene network up-regulated by pathogen-stimulated mononuclear cells encodes a variety of effector proteins with pleitropic biological activities. This class of primary immediate early (IE) genes codes for potent pro-inflammatory cytokines and chemokines that play a prominent role during the manifestation of inflammatory response. In an attempt to better understand induction mechanisms for such genes, I have focused on those coding for human interleukin-1β (IL1B) and tumor necrosis factor α (TNF), which exhibit both transient IE induction as well as cell-type restriction. Employing a combined approach using cell lines and primary cells, reporter transient transfection, chromatin conformational capture and immunoprecipitation, evaluation of transcript integrity, ectopic expression in a non-competent cell type, and comparison to mouse orthologs, I have determined that a complex array of mechanisms interplay in order to distinctly regulate the Toll-like receptor (TLR) signaling-dependent induction of these two important pro-inflammatory genes whose deregulation provides the etiology for numerous diseases. Prior to induction, TNF exhibited pre-bound TATA Binding Protein (TBP) and paused RNA Polymerase II (Pol II), which are the hallmarks of poised IE genes. In contrast, IL1B is stringently regulated by long-distance chromosome gyrations, multistep activation through a unique doubly-paused Pol II which, in association with the monocyte lineage factor Spi1/PU.1 (Spi1), maintains a low TBP and Pol II occupancy prior to activation. Activation and DNA binding of the transcription factors C/EBPβ and NF-κB resulted in de novo recruitment of TBP and Pol II to IL1B in concert with a permissive state for elongation mediated by the recruitment of the positive elongation factor b (P-TEFb). This Spi1-dependent mechanism for IL1B transcription, which is unique for a rapidly-induced/poised IE gene, was more dependent upon P-TEFb than was the case for the TNF gene. Nucleosome occupancy and chromatin modification analyses of the IL1B and TNF promoters, revealed activation-specific changes in chromatin marks that are supportive for nucleosome clearance and formation of nucleosome free regions (NFR). Furthermore, ectopic expression of Spi1, along with a TLR surrogate (over-expressed TNF receptor associated factor 6, TRAF6), in a cell line incompetent for IL1B transcription, is observed to prime the cell's endogenous genome for IL1B induction by appropriately phasing promoter nucleosomes and recruiting paused Pol II in a manner reminiscent of that observed in competent monocytes. Here I report a novel connection between the metabolic state of cells and HIF-1α in regulating murine Il1b gene expression. With regard to the lipopolysaccharide (LPS) unresponsive state known as endotoxin tolerance, my data revealed that following transient induction, IL1B and TNF remained marked with paused Pol II complexes for up to 24 hours post-stimulation. Upon subsequent LPS exposure, tolerized TNF remained in an unresponsive paused state, while IL1B resumed transcription due to recruitment of positive elongation kinase P-TEFb. Emerging evidence suggests that inflammatory responses of LPS/TLR4 activated macrophages are interconnected with metabolic pathways, resulting in the shift of energy utilization by the cells. Here I report that inhibition of either phosphoinositide 3-kinase (PI3K) or glucose metabolism had a greater affect on the transcriptional response of Il1b than of Tnf. The differences between these two genes, especially for endotoxin tolerance, suggest that il1b may play a distinct role from tnf in chronic inflammation. / Bayer School of Natural and Environmental Sciences; / Biological Sciences / PhD; / Dissertation;
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Hepatic Steatosis and TNF-α SignalingModi, Nita January 2007 (has links)
The overall objective of this research was to investigate the status of tumor necrosis factor-α (TNF-α), and molecules associated with its signaling, in the pathological state of hepatic steatosis. The effect of NSAID piroxicam, a cancer preventive agent also known to affect TNF-α signaling on hepatic steatosis, was also investigated. The biological state of the tissue was assessed by examining the expression of TNF-α signaling molecule in whole tissue, as well as in hepatic lipid raft. Lipid rafts are dynamic assemblies of cholesterol and sphingolipids, microdomains that form in the exoplasmic leaflet of the biological membranes shown to play a role in compartmentalization, modulation and integration of the cell signaling.
In the present research, Zucker obese rats were used as a model of human obesity and insulin resistant state. These rats exhibit hepatic steatosis in adulthood similar to those noted in obese individuals. Female Zucker obese and lean rats (5 weeks old) were fed a semisynthetic diet with or without piroxicam (150 ppm). Zucker lean counterparts served as control. After 8 weeks of feeding, rats were euthanized and liver from each animal was collected. Liver tissue from each animal was processed for histology and biochemical analysis which included lipids and proteins (COX-1 and 2, TNF-α, TNF-RI and RII, IKK-β, IκB-α and NF-κB). Liver histology and the level of total lipids confirmed that Zucker obese rats had hepatic steatosis, which was further augmented by piroxicam treatment. Whole tissue protein expression, using western blot, showed that the steatotic liver differed from non-steatotic livers by having lower levels of TNF-RII. TNF-RII showed a trend which was inversely proportional to the pathological state of the tissue. The obese-piroxicam liver had the lowest level of TNF-RII and lean livers had the highest (p<0.05). The total NF-κB level was higher in the obese and obese-piroxicam groups compared to the lean or lean-piroxicam groups (p<0.05). Piroxicam treatment lowered the level of NF-κB in obese and lean livers. IκB-α was higher in obese livers than in lean livers. The nuclear level of NF-κB by western blot analysis showed the same pattern as noted in the whole tissue homogenate. However, the difference in the level between obese and lean was marked. The obese nuclei contained two to three fold higher levels of NF-κB protein than the lean liver nuclei. IκB-α level was significantly higher in the obese liver tissues and nuclei than their lean counterparts. While transcriptionally active NF-κB was higher (p<0.05) in the obese livers than in the lean livers, the difference between obese and lean groups was not as significant as that noted for the level of NF-κB assessed by western blot. This suggests that the proportion of active NF-κB present in the nuclear fraction is much higher in the lean than in the obese nuclei.
Lipid raft was extracted and identified successfully from obese and lean livers. The total caveolin and flotillin levels were significantly higher in the liver lipid rafts of the obese-piroxicam than that of the other groups. This is the group that also exhibited higher steatosis. Piroxicam treatment significantly decreased the level of caveolin in the lean liver and significantly increased the level of flotillin in the obese liver. While COX-1 was not detectable, however, the level of COX-2 and TNF-RII in lipid raft was opposite to the level noted in the whole tissue homogenate. TNFRII was highest in the obese-piroxicam lipid raft and lowest in the lean-piroxicam lipid raft. TNF-RII, COX-2, IκB-α and NF-κB proteins were the molecules profoundly affected by the pathological state of the tissue and piroxicam treatment. This research is the first to report the presence of IκB-α in the nuclear compartment with a higher level in the nuclei and whole tissue in the obese liver than in the lean liver. This research demonstrates that TNF-α to NF-κB axis is altered in steatotic liver, and analysis of lipid rafts in steatotic and non-steatotic liver demonstrates that lipid rafts play a distinct role in modifying the biological availability of key proteins in the pathological state of liver steatosis.
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Soluble receptors for advanced glycation end products as predictors of restenosis following percutaneous coronary interventionMcNair, Erick Donnell 11 September 2009 (has links)
The principal cause of non-ST-segment myocardial infarction (NSTEMI), a subclass of acute coronary syndrome (ACS), is thrombosis and the underlying cause is atherosclerosis. Percutaneous coronary intervention (PCI) is one of the treatments to attenuate the ischemic effects of severe coronary artery stenosis. However, restenosis following PCI (post-PCI) is a major problem for the long-term success of the procedure. Recently, the interaction of advanced glycation end products (AGE) with the receptor for advanced glycation end products (RAGE) has been implicated in the development of atherosclerosis in animal models. Interaction of AGE with RAGE results in activation of nuclear factor kappa-B (NF-êB), release of cytokines including tumor necrosis factor-alpha (TNF-á), the expression of adhesion molecules including soluble vascular adhesion molecule-1 (sVCAM-1) and induction of oxidative stress all of which have been implicated in the development of atherosclerosis. The soluble receptor for advanced glycation end products (sRAGE) acts as a decoy for RAGE ligands (AGEs) and this occurs by competing with RAGE. In animal models, balloon inflation and de-endothelialization of the carotid artery increase the concentration of AGE and RAGE in the arterial wall and induces neointimal hyperplasia and stenosis. Treatment with sRAGE in animal models reduces neointimal growth and decreases smooth muscle cell migration and proliferation and expression of extracellular matrix.<p>
It is hypothesized that NSTEMI and post-PCI restenosis may be due to low levels of serum sRAGE resulting in increased AGE and RAGE interactions. Low levels of sRAGE would also increase the levels of serum TNF-á and sVCAM-1.<p>
The objectives of this study were to determine whether: (1) the levels of serum sRAGE are lower and the levels of AGE, TNF-á and sVCAM-1 are higher in NSTEMI patients compared to control; (2) the levels of serum sRAGE are lower and the levels of AGE, TNF-á and sVCAM-1 are higher in NSTEMI patients with restenosis compared to those without restenosis; and (3) sRAGE or AGE/sRAGE ratio may serve as a biomarker/ predictor of NSTEMI and post-PCI restenosis.<p>
The study objectives include 46 consecutive NSTEMI patients undergoing elective PCI and 28 healthy age-matched male controls. Pre-PCI and 6 month post-PCI angiography were performed in all NSTEMI patients. Blood samples were collected at designated intervals for the measurement of sRAGE, AGE, TNF-á, and sVCAM-1 using commercially available enzyme-linked immunosorbent assay (ELISA) kits.<p>
The levels of serum sRAGE were lower and those of TNF-á, sVCAM-1, AGE and AGE/sRAGE were higher in NSTEMI patients compared to control subjects. (sRAGE, 884.55 ± 50 vs. 1287 ± 41.5 pg/mL{p<0.001}; TNF-á, 23.1 ± 2.3 vs. 10.3 ± 0.8 pg/mL {p<0.002}; sVCAM-1, 1059.62 ± 70.8 vs. 651 ± 35.5 ng/mL {p<0.0003}, AGE, 1192.50 ± 82.6 vs. 669.40 ± 47.9 ng/mL {p<0.001}; and AGE/sRAGE, 1.75 ± 0.17 vs. 0.52 ± 0.06 {p<0.001}).<p>
The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and accuracy of the sRAGE biomarker test were 59%, 100%, 100%, 100% and 74% respectively, while those of the AGE/sRAGE test were 85%, 91%, 97%, 67% and 86%, respectively, in the diagnosis of patients with NSTEMI acute coronary syndrome.
The pre-PCI levels of serum sRAGE in patients with restenosis were significantly lower (p<0.001) than in patients without restenosis (610.6 ± 24.1 vs. 1143.8 ± 52.5 pg/mL). The post-PCI levels of serum sRAGE were significantly lower (p<0.0001) in patients with restenosis compared to those without restenosis (477 ± 18.6 vs. 1106.7 ± 41.9 pg/mL). The pre-PCI levels of serum TNF-á and sVCAM-1 were significantly higher (p<0.001 and p<0.001) in patients with restenosis compared to those without restenosis (TNF-á, 37.9 ± 2.5 vs. 11.6 ± 0.41 pg/mL; sVCAM-1, 1381.8 ± 63.5 vs. 811.37 ± 26.5 ng/mL). The post PCI levels of serum TNF-á and sVCAM-1 were also significantly higher (p<0.0001 and p<0.0001) in patients with restenosis than in patients without restenosis (TNF-á, 48.4 ± 1.4 vs. 12.5 ± 0.44 pg/mL; sVCAM-1, 1381.8 ± 63.5 vs. 762.2 ± 26.4 ng/mL). The post-PCI levels sRAGE were lower while those of TNF-á and sVCAM-1 were higher compared to the pre-PCI levels in patients with restenosis. However, the pre- and post-PCI levels of serum sRAGE, TNF-á and sVCAM-1 were similar in patients without restenosis. The pre-PCI levels of serum AGE and AGE/sRAGE were significantly higher (p<0.001 and p<0.001) in patients with restenosis compared to those without restenosis (AGE, 1512.1 ± 84.53 vs. 891.7 ± 92.4 ng/mL; 2.39 ± 0.20 vs. 1.03 ± 0.17).<p>
The sensitivity, specificity, PPV, NPV, and accuracy of the pre-PCI sRAGE tests were 73%, 100%, 100%, 80%, and 87%, respectively while those of the AGE/sRAGE tests were 81%, 94%, 93%, 84% and 88%, respectively in identifying patients with post-PCI restenosis.<p>
In conclusion the results suggest that: (1) the levels of serum sRAGE are lower while those of TNF-á, sVCAM-1, AGE and AGE/sRAGE are higher in NSTEMI patients compared to control subjects; (2) serum levels of sRAGE are negatively correlated with the number of diseased vessels; (3) Both low sRAGE and high AGE/sRAGE may serve as a biomarker/predictor of NSTEMI, but AGE/sRAGE has a greater sensitivity compared to sRAGE; (4) the pre-PCI levels of serum sRAGE are lower while those of AGE/sRAGE are higher in patients with restenosis compared to those without restenosis; and (5) both low sRAGE and high AGE/sRAGE may serve as a predictor/ biomarker of post-PCI restenosis; however, AGE/sRAGE has a greater sensitivity than sRAGE.
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C10 semi-peptoid beta-turn peptidomimetics: syntheses, characterization and biological studiesNnanabu, Ernest 02 June 2009 (has links)
Over the years, the Burgess group has been focusing on the preparation and
testing of small molecules that mimic protein secondary structures for protein-protein
interactions. The most successful compounds made are C10 peptide macrocycles that
effectively mimic β-turns and have given promising results from biological testing. These
peptide macrocycles have also been dimerized to give even more effective ligands for
protein-protein interaction.
The successes of the peptide macrocycles have enabled us to look into increasing
the chemical diversity of our libraries. This we believe will not only improve our ability
to obtain high affinity ligands for the receptors of interest, but will also allow us to
investigate other receptors. To achieve this, peptoids were incorporated into the C10
system to replace the peptides in the i+1 and i+2 positions. With the help of Microwave
irradiation, semi-peptoid macrocycles were synthesized with a total reaction time of less
than 2 h. These compounds were characterized and found to mimic β-turn, and show
promising biological activity towards the Insulin-like growth factor 1 receptor (IGF-IR).
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Association of Tumor Necrosis Factor a, Tumor Necrosis Factor Receptors and Interleukin-1b Genetic Polymorphisms in Palindromic RheumatismKeng, Hsiu-man 07 June 2006 (has links)
ABSTRACT
Background: Palindromic rheumatism (PR) is the rare disease that generally occurs with multiple recurring attacks of painful inflammation affecting joints and adjacent tissues. The thesis attempts to characterize the association in 10 instances of the single nucleotide polymorphisms (SNPs) of tumor necrosis factor genes (TNF-£\, TNFRSF1A and TNFRSF1B), Interleukin-1£] genes and Palindromic rheumatism (PR). Methods: The genetic polymorphisms of TNF-£\, TNFR£L, TNFR£S and IL-1£] genes cluster were investigated among 56 PR patients identified from the Kaohsiung Veterans General Hospital (VGHKS, Kaohsiung, Taiwan) and compared with one hundred healthy subjects. The genotypes for ten SNPs in the TNF-£\, TNFRSF1A, TNFRSF1B and IL-1£] genes among these 156 individuals were examined.
Results: Experiments indicate significant count of the TNFRSF1A+36 AG genotype in PR patients (OR=4.8, 95%CI=1.8-13.0, p=0.002) and TNFRSF1A+36G allele (OR=3.94, 95%CI=1.59-9.79, p=0.003).The results also have remarkable correlation with TNFRSF1B haplotype +676/+1663 T/A (OR=2.12, CI=1.2-3.8, p=0.010). However, on significant differences were found for all the TNF-£\and IL-1£]polymorphisms.
Conclusions: Genetic polymorphisms in TNF-£\ receptors are associated with susceptibility and severity of the inflammatory response in the PR patients.
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Modulation of the immune response following myocardial infarction utilizing biomaterial-based therapeutic delivery strategiesSomasuntharam, Inthirai 21 September 2015 (has links)
In 2015, American Heart Association (AHA) reported that 1 in 9 deaths are attributed to Heart failure (HF), the number one killer in the world. While advancements in interventional cardiology in conjunction with pharmacotherapies have significantly reduced the rate of mortality following MI, there has been a corresponding rise in chronic heart failure (CHF) in surviving patients, largely attributed to the limited regenerative capacity of the heart and the inadequate healing response. Myocardial ischemic injury triggers an exuberant local and systemic inflammation, and the extent and quality of the cardiac wound healing process is intricately tied to the delicate equilibrium of this inflammatory response. While cardiac regeneration is an important goal, it is imperative in the meantime to explore therapeutic strategies that target these inflammatory mediators of early cardiac repair. These interventions to influence and improve cardiac wound healing can represent a new therapeutic window to halt the progression of heart failure between the few hours that may be used to limit infarct size by reperfusion and an irreversible non-contractile cardiac scar. This dissertation examines three therapeutic delivery strategies aimed at modulating the immune response to enhance cardiac repair in rodent models MI: 1) Polyketal nanoparticles as siRNA delivery vehicles for antioxidant therapy; 2) Spherical nucleic acid particles for anti-inflammatory therapy and; 3) Bioactive PEG (polyethyleneglycol)-based hydrogel for immunomodulation. The work presented here applies novel nucleic acid delivery strategies for cardiac gene silencing and has contributed to new knowledge with regard to modulating the immune response following MI.
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MODULATION OF ENDOTHELIAL ACTIVATION AND CEREBRAL ANGIOGENESIS BY TNF FAMILY LIGANDS AND RESVERATROL: AN IN VITRO STUDYChen, Pei-Lin 10 December 2010 (has links)
Vascular endothelial cell activation and apoptosis (programmed cell death) are critical in inflammation and angiogenesis (the formation of new blood vessels). Tumor necrosis factor (TNF) is a pro-inflammatory cytokine known for its ability to induce endothelial cell activation and apoptosis. However, the ability of two death ligands in the TNF superfamily: TRAIL (TNF-Related Apoptosis-Inducing Ligand) and Fas ligand (FasL), to activate vascular endothelium is less well defined, and forms the basis of this work. We find that in the human endothelial cell line EA.hy926, TRAIL induces endothelial cell activation (activation of the transcription factor NF-?B with increased expression of the adhesion protein ICAM-1 and adhesion of human neutrophils) when it concurrently induces apoptosis. In addition, angiogenesis is implicated in diseases of the central nervous system, and its modulation represents an attractive therapeutic strategy. We investigated the modulatory potential of the two endogenous molecules TRAIL and FasL as well as an exogenous molecule resveratrol, a phytochemical present in red wine, in angiogenesis. We modeled cerebral angiogenesis with the human brain endothelial cell line hCMEC/D3 and primary bovine brain endothelial cells. Resveratrol inhibited several parameters of angiogenesis (proliferation, migration and tube formation) in human umbilical vein endothelial cells, however, neither TRAIL nor FasL had an effect on this model. By contrast, in hCMEC/D3 cells both resveratrol and TRAIL inhibited all parameters while FasL had minimal effects. Resveratrol did not induce apoptosis in hCMEC/D3 but arrested cell cycle progression to G2/M phase and inhibited phosphorylation of Akt/PKB, a key cell survival protein kinase. This leads to a reduction in cell growth, endothelial migration and tube formation, hence, inhibition of in vitro angiogenesis. TRAIL induced anti-angiogenic effects in hCMEC/D3 due to apoptosis. The data suggests that TRAIL primarily influences angiogenesis through induction of vascular endothelial apoptosis while resveratrol induces cell cycle arrest, both of which may lead to vessel regression. These are the first studies to report the modulation of different aspects of endothelial cell activation by TRAIL and resveratrol in several endothelial cell culture models, with a particular focus on the central nervous system.
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