Flowchart: Preparation: Scap1
 


        

Text Box: CYP4F2

                   

                         

 


Text Box: Scap

Anthrax

Hypocholesterolemia

Myocardial

 

 


                              

Text Box: Srebp

                              

 

7/23/2007/28

J Biol Chem. 2007 Feb 23;282(8):5225-36. Epub 2006 Dec 1.Click here to read Links

Regulation of human cytochrome P450 4F2 expression by sterol regulatory element-binding protein and lovastatin.

Hsu MH, Savas U, Griffin KJ, Johnson EF.

Division of Biochemistry, Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, California 92037, USA.

This report provides the first evidence that human P450 4F2 (CYP4F2) is induced by statins, which are widely used to treat hypercholesterolemia. Real time PCR and immunoblots indicate that lovastatin treatment increases expression of the endogenous CYP4F2 gene in human primary hepatocytes and HepG2 cells. The effects of lovastatin on gene expression are often mediated through sterol regulatory element-binding proteins (SREBPs). Immunoblots indicate that lovastatin-treated human hepatocytes display increased proteolytic processing of SREBP-2. In HepG2 cells, co-administration of a potent suppressor of SREBP-2 activation, 25-hydroxycholesterol, inhibits CYP4F2 mRNA induction by lovastatin. HepG2 cells transfected with an expression vector for the active nuclear form of SREBP-1a (nSREBP-1a) also display elevated endogenous CYP4F2 expression. Luciferase reporters containing the CYP4F2 proximal promoter are transactivated by nSREBPs (-1a, -1c, and -2) or a dominant positive form of the SREBP cleavage-activating protein (SCAP), which facilitates activation of endogenous SREBPs. Lovastatin-induced reporter expression is inhibited by overexpressed Insig-1, which prevents proteolytic activation of endogenous SREBPs. Electrophoretic mobility shift assays with in vitro translated nSREBP-1a identified two SREBP binding sites at -169/-152 and -109/-92, relative to the CYP4F2 transcription start site. Mutations in each site abolish SREBP binding. Chromatin immunoprecipitation experiments indicate that more SREBP-1 is associated with the CYP4F2 promoter after overexpression of nSREBP-1a. Transfection studies and mutagenesis indicate that the -109/-92 region is the primary site responsible for the effects of statins. Collectively, these results demonstrate that SREBPs transactivate CYP4F2 transcription and that CYP4F2 induction by statins is mediated by SREBP-2.

PMID: 17142457 [PubMed - indexed for MEDLINE]

Atherosclerosis. 2007 Mar 23; [Epub ahead of print]Click here to read Links

SREBP-2 and SCAP isoforms and risk of early onset myocardial infarction.

Friedlander Y, Schwartz SM, Durst R, Meiner V, Robertson AS, Erez G, Leitersdorf E, Siscovick DS.

Unit of Epidemiology, Hebrew University-Hadassah School of Public Health, Jerusalem, Israel.

BACKGROUND: Cholesterol metabolism is mediated, in part, by the sterol-regulatory element binding proteins (SREBPs) that are activated by a SREBP cleavage-activating protein (SCAP). We examined whether coding variations in the interacting domains of both genes, are related to early-onset MI risk in a population-based case-control study from western Washington State. METHODS: Cases were 257 women, aged 18-59 years, and 320 men, aged 18-49 years, with first acute non-fatal MI; controls were 353 women and 311 men, similar in age, identified from the community who had no history of clinical CHD or stroke. Genotyping of the SREBF-2 G1784C polymorphism (SREBP-2-595A/G isoforms), and the SCAP A2386G polymorphism (SCAP-796I/V isoforms), were performed. RESULTS: After adjustment for age and race, the SREBP-2-595A isoform was associated with increased MI risk among men (OR=1.63, 95% CI=1.26-2.12). In contrast, there was little evidence for an association among women in a multiplicative model. However, compared to SREBP-2-595G homozygotes, homozygote women for the SREBP-2-595A isoform were at nearly two-fold increased risk (OR=1.95, 95% CI=1.07-3.54). Overall, SCAP genotypes were neither associated with MI in men nor in women. However, in men, SCAP genotypes were found to modify the association between SREBF-2 and MI (p-value for interaction=0.01). CONCLUSION: The SREBP-2-595A isoform was associated with an increased risk of early-onset MI in U.S. men. The SCAP polymorphism appeared to modify the associations of SREBF-2 genotype with MI risk among men. These novel findings require confirmation in other populations.

PMID: 17383658 [PubMed - as supplied by publisher]

Atherosclerosis. 2007 Mar 23; [Epub ahead of print]Click here to read Links

SREBP-2 and SCAP isoforms and risk of early onset myocardial infarction.

Friedlander Y, Schwartz SM, Durst R, Meiner V, Robertson AS, Erez G, Leitersdorf E, Siscovick DS.

Unit of Epidemiology, Hebrew University-Hadassah School of Public Health, Jerusalem, Israel.

BACKGROUND: Cholesterol metabolism is mediated, in part, by the sterol-regulatory element binding proteins (SREBPs) that are activated by a SREBP cleavage-activating protein (SCAP). We examined whether coding variations in the interacting domains of both genes, are related to early-onset MI risk in a population-based case-control study from western Washington State. METHODS: Cases were 257 women, aged 18-59 years, and 320 men, aged 18-49 years, with first acute non-fatal MI; controls were 353 women and 311 men, similar in age, identified from the community who had no history of clinical CHD or stroke. Genotyping of the SREBF-2 G1784C polymorphism (SREBP-2-595A/G isoforms), and the SCAP A2386G polymorphism (SCAP-796I/V isoforms), were performed. RESULTS: After adjustment for age and race, the SREBP-2-595A isoform was associated with increased MI risk among men (OR=1.63, 95% CI=1.26-2.12). In contrast, there was little evidence for an association among women in a multiplicative model. However, compared to SREBP-2-595G homozygotes, homozygote women for the SREBP-2-595A isoform were at nearly two-fold increased risk (OR=1.95, 95% CI=1.07-3.54). Overall, SCAP genotypes were neither associated with MI in men nor in women. However, in men, SCAP genotypes were found to modify the association between SREBF-2 and MI (p-value for interaction=0.01). CONCLUSION: The SREBP-2-595A isoform was associated with an increased risk of early-onset MI in U.S. men. The SCAP polymorphism appeared to modify the associations of SREBF-2 genotype with MI risk among men. These novel findings require confirmation in other populations.

PMID: 17383658 [PubMed - as supplied by publisher]

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Biochim Biophys Acta. 2007 Jul 6; [Epub ahead of print]

Molecular docking and spatial coarse graining simulations as tools to investigate substrate recognition, enhancer binding and conformational transitions in indoleamine-2,3-dioxygenase (IDO).

Macchiarulo A, Nuti R, Bellocchi D, Camaioni E, Pellicciari R.

Dipartimento di Chimica e Tecnologia del Farmaco, Universitą di Perugia, Via del Liceo 1, 06123 Perugia, Italy.

Indoleamine 2,3-dioxygenase (IDO) is an heme-containing enzyme involved in the regulation of important immunological responses and neurological processes. The enzyme catalyzes the oxidative cleavage of the pyrrole ring of the indole nucleus of tryptophan (Trp) to yield N-formylkynurenine, that is the initial and rate limiting step of the kynurenine pathway. Some indole derivatives have been reported to act as effectors of the enzyme by enhancing its catalytic activity. On the basis of the recent availability of the crystal structure of IDO, in this work we investigate substrate recognition and enhancer binding to IDO using molecular docking experiments. In addition, conformational transitions of IDO in response to substrate and enhancer binding are studied using coarse graining simulations with the program FIRST. The results enable us to identify (i) the binding site of enhancer modulators; (ii) the motion of an electrostatic gate that regulates the access of the substrate to the catalytic site of the enzyme; (iii) the movement of the anchoring region of a hairpin loop that may assist the shuttle of substrates/products to/from the catalytic site of IDO. These data, combined with available site-directed mutagenesis experiments, reveal that conformational transitions of IDO in response to substrate and enhancer binding are controlled by distinct combination of two conformational states (open and close) of the above structural motifs. On this basis, a molecular mechanism regarding substrate recognition and activity enhancement by indole derivatives is proposed.

PMID: 17644054 [PubMed - as supplied by publisher]

J Virol. 2007 Jul 11; [Epub ahead of print]Click here to read Links

Astrocyte indoleamine 2, 3 dioxygenase (IDO) is induced by the TLR3 ligand poly IC: mechanism of induction and role in anti-viral response.

Suh HS, Zhao ML, Rivieccio M, Choi S, Connolly E, Zhao Y, Takikawa O, Brosnan CF, Lee SC.

Department of Pathology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx NY 10461 and National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, Obu, Japan.

Indoleamine 2, 3-dioxygenase (IDO) is the first and rate-limiting enzyme in the kynurenine pathway of tryptophan catabolism and has been implicated in neurotoxicity and suppression of the antiviral T cell response in HIV encephalitis (HIVE). Here we show that the toll-like receptor 3 (TLR3) ligand polyinosinic-polycytidylic acid (PIC) induces the expression of IDO in human astrocytes. PIC was less potent than IFNgamma but more potent than IFNbeta in inducing IDO. PIC induction of IDO was in part mediated by IFNbeta but not IFNgamma, and both NF-kappaB and IRF3 were required. PIC also upregulated TLR3 thereby augmenting the primary (IFNbeta) and secondary (IDO and viperin) response genes upon subsequent stimulation with PIC. In HIVE, the transcripts for TLR3, IFNbeta, IDO and viperin were increased and IDO immunoreactivity was detected in reactive astrocytes as well as macrophages and microglia. PIC caused suppression of intracellular replication of VSVg-env HIV and human cytomegalovirus in a manner dependent on IRF3 and IDO. The involvement of IDO was demonstrated by partial but significant reversal of the PIC-mediated antiviral effect by IDO RNAi and/or tryptophan supplementation. Importantly, the cytokine IL-1 abolished IFNgamma-induced IDO enzyme activity in a nitric oxide-dependent manner without suppressing protein expression. Our results demonstrate that IDO is an innate antiviral protein induced by dsRNA and suggest a therapeutic utility for PIC in human viral infections. They also show that IDO activity can be dissociated from protein expression, indicating that the local CNS cytokine and nitric oxide environment determines IDO function.

PMID: 17626075 [PubMed - as supplied by publisher]

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Int Immunopharmacol. 2006 Jun;6(6):1034-8. Epub 2006 Feb 3.

Related Articles, Links

Click here to read 
IL-2 increased RANTES production and CD25 expression in cultured PBMCs only from antiretroviral treated HIV-1+ patients with detectable viral loads.

Lozano JM, Kindelan JM, Cabello A, Gonzalez R, Solana R, Pena J.

Department of Immunology, Hospital "Reina Sofia", University of Cordoba, Spain; Department of Virology, Pasteur Institute of Paris, France.

In order to better understand the possible beneficial effects of intermittent IL-2 treatment as complement of antiretroviral therapy in HIV-1+ patients, we have measured the levels of RANTES in the supernatants and the CD25 expression in cultured PBMCs obtained from HIV-1+ individuals in presence of IL-2. The results showed a significant increases in RANTES production and in the expression of CD25+ in the cultures with IL-2 of PBMC obtained from HIV-1+ patients with a detectable viral load in comparison with both, HIV-1+ patients with no detectable viral loads and with healthy individuals. These results suggest that therapeutic IL-2 administered in addition to highly active anti-retroviral therapy (HAART) may contribute to increase the effect of this therapy by rising both RANTES production and CD25 expression only in HIV-1+ patients with detectable viral loads.

PMID: 16644491 [PubMed - in process]

 

 

 

 

 

Science. 2005 Jun 3;308(5727):1477-80.Click here to read Links

The structure of interleukin-2 complexed with its alpha receptor.

Rickert M, Wang X, Boulanger MJ, Goriatcheva N, Garcia KC.

Departments of Microbiology and Immunology, and Structural Biology, Stanford University School of Medicine, 299 Campus Drive, Fairchild D319, Stanford, CA 94305-5124, USA.

Interleukin-2 (IL-2) is an immunoregulatory cytokine that binds sequentially to the alpha (IL-2Ralpha), beta (IL-2Rbeta), and common gamma chain (gammac) receptor subunits. Here we present the 2.8 angstrom crystal structure of a complex between human IL-2 and IL-2Ralpha, which interact in a docking mode distinct from that of other cytokine receptor complexes. IL-2Ralpha is composed of strand-swapped "sushi-like" domains, unlike the classical cytokine receptor fold. As a result of this domain swap, IL-2Ralpha uses a composite surface to dock into a groove on IL-2 that also serves as a binding site for antagonist drugs. With this complex, we now have representative structures for each class of hematopoietic cytokine receptor-docking modules.

PMID: 15933202 [PubMed - indexed for MEDLINE]

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

The effect of therapeutic hypothermia on the expression of inflammatory response genes following moderate traumatic brain injury in the rat.

Truettner JS, Suzuki T, Dietrich WD.

Department of Neurological Surgery, The Neurotrauma Research Center, The Miami Project to Cure Paralysis, University of Miami School of Medicine, Miami, FL 33136, USA.

Traumatic brain injury (TBI) initiates a cascade of cellular and molecular responses including both pro- and anti-inflammatory. Although post-traumatic hypothermia has been shown to improve outcome in various models of brain injury, the underlying mechanisms responsible for these effects have not been clarified. In this study, inflammation cDNA arrays and semi-quantitative RT-PCR were used to detect genes that are differentially regulated after TBI. In addition, the effect of post-traumatic hypothermia on the expression of selective genes was also studied. Rats (n = 6-8 per group) underwent moderate fluid-percussion (F-P) brain injury with and without hypothermic treatment (33 degrees C/3 h). RNA from 3-h or 24-h survival was analyzed for the expression of IL1-beta, IL2, IL6, TGF-beta2, growth-regulated oncogene (GRO), migration inhibitory factor (MIF), and MCP (a transcription factor). The interleukins IL-1beta, IL-2, and IL-6 and TGF-beta and GRO were strongly upregulated early and transiently from 2- to 30-fold over sham at 3 h, with normalization by 24 h. In contrast, the expressions of MIF and MCP were both reduced by TBI compared to sham. Post-traumatic hypothermia had no significant effect on the acute expression of the majority of genes investigated. However, the expression of TGF-beta2 at 24 h was significantly reduced by temperature manipulation. The mechanism by which post-traumatic hypothermia is protective may not involve a general genetic response of the inflammatory genes. However, specific genes, including TGF-beta2, may be altered and effect cell death mechanisms after TBI. Hypothermia differentially regulates certain

genes and may target more delayed responses underlying the secondary damage following TBI.

PMID: 15922484 [PubMed - in process]

 

 

 

 

 

 

 

 



 erase chain reaction in unstimulated PBMC and in PBMC incubated for 4h in the presence of concanavalin A (ConA) or phorbol 12-myristate 13-acetate plus ionomycin (PMA/I). Compared to unstimulated cells, stimulation with ConA and PMA/I increased the IL2 mRNA transcription in average by 300- and 20-fold, respectively. Nevertheless, no significant differences in IL2 transcription between the genotypes could be detected. These findings were confirmed by band shift studies using different oligonucleotides containing variations of the potential binding motif, which showed no differences in the gel mobility after incubation with nuclear extract containing GATA-3. The obtained results argue against an impact of this polymorphism on the IL2 transcription and the genetic disease resistance in sheep.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

ICAM-1 gene expression in endothelial cells: Effects on the inhibition of STAT3 phosphorylation.
Resveratrol suppresses IL-6-induced
Wung BS, Hsu MC, Wu CC, Hsieh CW.

Department of Applied Microbiology, National Chiayi University, Chiayi, Taiwan.

Resveratrol, a polyphenolic phytoaxelin present in red wine, has been suggested to protect against atherosclerosis and cardiovascular disease because of its antioxidant effects. Intercellular adhesion molecule (ICAM-1), induced by cytokines, has been hypothesized to play a role in the early events during atherosclerosis. In this study we tested the effects of resveratrol upon both IL-6-induced ICAM-1 gene expression and its underlying signaling pathways in endothelial cells (ECs). Resveratrol was found to inhibit both TNFalpha- and IL-6-induced ICAM-1 gene expression at the promoter, transcriptional and protein levels. Resveratrol also abrogates the tyr705 phosphorylation of STAT3 in IL-6-treated ECs, in a dose- and time-dependent manner. Although quercetin had similar effects, resveratrol showed higher inhibitory properties following 2-4 h pretreatments. Resveratrol has been shown to induce the activity of endothelial nitric oxide synthase (eNOS) and increase NO production. Consistent with this, the treatment of ECs with a NO donor (SNAP) reduces IL-6-induced STAT3 phosphorylation. Conversely, exposure of ECs to a NOS inhibitor reversed the effects of resveratrol upon IL-6-induced STAT3 phosphorylation. Furthermore, ECs transfected with constitutively active Rac1 (RacV12) showed increases in ICAM-1 promoter activity, intracellular reactive oxygen species (ROS) levels and STAT3 phosphorylation, and these increases were attenuated by resveratrol treatment. In summary, we demonstrate for the first time that resveratrol inhibits IL-6-induced ICAM-1 gene expression, in part, by interfering with Rac-mediated pathways via the attenuation of STAT3 phosphorylation. This study therefore provides important new insights that may contribute to the proposed beneficial effects of resveratrol in endothelial responses to cytokines during inflammation.

PMID: 16150460 [PubMed - as supplied by publisher]

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Bradykinin mimics ischemic preconditioning by generating reactive oxygen species (ROS). To identify intermediate steps leading to ROS generation, rabbit cardiomyocytes were incubated in reduced MitoTracker Red that becomes fluorescent after exposure to ROS. Fluorescence intensity in treated cells was expressed as % of that in paired untreated cells. Bradykinin (500nM) caused 51+/-16% increase in ROS generation (p<0.001). Co-incubation with either bradykinin B2 receptor blocker HOE140 (5 micro M) or free radical scavenger N-(2-mercaptopropionyl) glycine (1mM) prevented this increase confirming the response was receptor-mediated and ROS were actually being measured. Closing mitochondrial KATP (mKATP)channels with 5-hydroxydecanoate (5HD, 1mM) prevented increased ROS generation. Bradykinin-induced ROS generation was blocked by L-NAME (200 micro M) implicating nitric oxide as an intermediate. Blockade of guanylyl cyclase with ODQ (10 micro M) aborted bradykinin-induced ROS generation, but not that from diazoxide, a direct opener of mKATP channels. The PKG blocker 8-Br-cGMPS (25 micro M) eliminated bradykinin's effect. Conversely, direct activation of PKG with 8-pCPT-cGMP (100 micro M) increased ROS generation (39+/-15%, p<0.004) similar to bradykinin. This increase was blocked by 5HD. Finally, the nitric oxide donor SNAP (1 micro M)increased ROS by 34+/-6%. This increase was also blocked by 5HD. In intact rabbit hearts bradykinin (400nM) decreased infarction from 30.5+/-3.0% of the risk zone in control hearts to 11.9+/-1.4% (p<0.01). This protection was aborted by either L-NAME (200 micro M) or ODQ (2 micro M)(35.4+/-5.7% and 30.4+/-3.0% infarction, resp., pNS vs control). Hence, bradykinin preconditions through receptor-mediated production of nitric oxide which activates guanylyl cyclase. The resulting cGMP activates PKG that opens mKATP. Subsequent release of ROS triggers cardioprotection.