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PubMed2025

S-GMAS: Genome-Wide Mediation Analysis With Brain Subcortical Shape Mediators.

Mediation analysis is widely utilized in neuroscience to investigate the role of brain image phenotypes in the neurological pathways from genetic exposures to clinical outcomes. However, it is still difficult to conduct mediation analyses with whole genome-wide exposures and brain subcortical shape mediators due to several challenges including (i) large-scale genetic exposures, that is, millions of single-nucleotide polymorphisms (SNPs); (ii) nonlinear Hilbert space for shape mediators; and (iii) statistical inference on the direct and indirect effects. To tackle these challenges, this paper proposes a genome-wide mediation analysis framework with brain subcortical shape mediators. First, to address the issue caused by the high dimensionality in genetic exposures, a fast genome-wide association analysis is conducted to discover potential genetic variants with significant genetic effects on the clinical outcome. Second, the square-root velocity function representations are extracted from the brain subcortical shapes, which fall in an unconstrained linear Hilbert subspace. Third, to identify the underlying causal pathways from the detected SNPs to the clinical outcome implicitly through the shape mediators, we utilize a shape mediation analysis framework consisting of a shape-on-scalar model and a scalar-on-shape model. Furthermore, the bootstrap resampling approach is adopted to investigate both global and spatial significant mediation effects. Finally, our framework is applied to the corpus callosum shape data from the Alzheimer's Disease Neuroimaging Initiative.

Humans↗
PubMed2003

A mammalian mediator subunit that shares properties with Saccharomyces cerevisiae mediator subunit Cse2.

The multiprotein Mediator complex is a coactivator required for activation of RNA polymerase II transcription by DNA bound transcription factors. We previously identified and partially purified a mammalian Mediator complex from rat liver nuclei (Brower, C.S., Sato, S., Tomomori-Sato, C., Kamura, T., Pause, A., Stearman, R., Klausner, R.D., Malik, S., Lane, W.S., Sorokina, I., Roeder, R.G., Conaway, J.W., and Conaway, R.C. (2002) Proc. Natl. Acad. Sci. U. S. A. 99, 10353-10358). Analysis by tandem mass spectrometry of proteins present in the most highly purified rat Mediator fractions led to the identification of a collection of new mammalian Mediator subunits, as well as several potential Mediator subunits including a previously uncharacterized protein encoded by the FLJ10193 open reading frame. In this study, we present direct biochemical evidence that the FLJ10193 protein, which we designate Med25, is a bona fide subunit of the mammalian Mediator complex. In addition, we present evidence that Med25 shares structural and functional properties with Saccharomyces cerevisiae Mediator subunit Cse2 and may be a mammalian Cse2 ortholog. Taken together, our findings identify a novel mammalian Mediator subunit and shed new light on the architecture of the mammalian Mediator complex.

Amino Acid Sequence↗
PubMed2005

NFkappaB-mediated upregulation of bcl-xl restrains TRAIL-mediated apoptosis in murine viral hepatitis.

Inhibition of NFkappaB enhances the susceptibility of cancer to TRAIL-mediated apoptosis and is suggested as a strategy for cancer therapy. Because the role of NFkappaB in TRAIL-mediated apoptosis of hepatocytes is unknown, we investigated the influence of NFkappaB-inhibition in death ligand-mediated apoptosis in hepatitis. Adenoviral hepatitis resulted in upregulation of NFkappaB-activity, which could be inhibited by expression of IkappaBalpha-superrepressor. We treated mice after the onset of adenoviral hepatitis with adenoviruses expressing FasL (AdFasL), TRAIL (AdTRAIL), or GFP (AdGFP). In contrast to apoptosis induced by AdFasL, NFkappaB inhibition strongly enhanced AdTRAIL-mediated apoptosis of hepatocytes. Expression of IkappaBalpha inhibits adenoviral infection-mediated overexpression of bcl-xl, providing a molecular mechanism for TRAIL sensitization. In agreement with this hypothesis, downregulation of bcl-xl by siRNA enhanced susceptibility of hepatocytes to TRAIL, but not to FasL-mediated apoptosis, resulting in TRAIL-mediated severe liver damage after AdTRAIL application. Our data demonstrate that inhibition of NFkappaB in adenoviral hepatitis strongly sensitizes hepatocytes to TRAIL-mediated apoptosis. Bcl-xl, in contrast to bcl-2 and c-FLIP, is strongly upregulated after viral infection and represents an essential NFkappaB-dependent survival factor against TRAIL-mediated apoptosis. In conclusion, inhibition of NFkappaB or bcl-xl during TRAIL therapy may harbor a risk of liver damage in patients with viral hepatitis.

Animals↗
PubMed2005

Uteroglobin inhibits prostaglandin F2alpha receptor-mediated expression of genes critical for the production of pro-inflammatory lipid mediators.

Prematurity is one of the leading causes of infant mortality. It may result from intrauterine infection, which mediates premature labor by stimulating the production of inflammatory lipid mediators such as prostaglandin F2alpha (PGF2alpha). The biological effects of PGF2alpha are mediated via the G protein-coupled receptor FP; however, the molecular mechanism(s) of FP signaling that mediates inflammatory lipid mediator production remains unclear. We reported previously that in the human uterus, a composite organ in which fibroblast, epithelial, and smooth muscle cells are the major constituents, an inverse relationship exists between the levels of PGF2alpha and a steroid-inducible anti-inflammatory protein, uteroglobin. Here we report that, in NIH 3T3 fibroblasts and human uterine smooth muscle cells, FP signaling is mediated via multi-kinase pathways in a cell type-specific manner to activate NF-kappaB, thus stimulating the expression of cyclooxygenase-2. Cyclooxygenase-2 is a critical enzyme for the production of prostaglandins from arachidonic acid, which is released from membrane phospholipids by phospholipase A2, the expression of which is also stimulated by PGF2alpha. Most importantly, uteroglobin inhibits FP-mediated NF-kappaB activation and cyclooxygenase-2 gene expression by binding and most likely by sequestering PGF2alpha into its central hydrophobic cavity, thereby preventing FP-PGF2alpha interaction and suppressing the production of inflammatory lipid mediators. We propose that uteroglobin plays important roles in maintaining homeostasis in organs that are vulnerable to inadvertent stimulation of FP-mediated inflammatory response.

Animals↗
PubMed1983

Clonal analysis of the lymphoid cells mediating skin allograft rejection. Mediation of graft rejection in vivo by cloned Lyt-1+2- proliferative, noncytotoxic long-term cell lines.

Studies were undertaken to elucidate the cellular basis of skin allograft rejection mediated by long-term cultured cell lines and clones. The adoptive transfer, in vivo, of in-vitro-sensitized cells, from B6AF1 anti B10.BR or from C57BL/6 anti DBA/2 cultures, and expanded eight-fold to ten-fold for one week in lectin-free interleukin 2 (LF-IL-2) were able to mediate specific skin allograft rejection. These same cells lost the ability to mediate accelerated skin graft rejection when they were expanded more than 100-fold during three weeks of culture in LF-IL-2 even though these cultures mediated high levels of specific in vitro cytotoxicity for the appropriate allosensitizing cells. When Lyt-2+ cells were depleted using monoclonal antibodies and complement prior to in vitro sensitization and expansion in LF-IL-2, these cells lines retained the ability to mediate skin allograft rejection in vivo when expanded more than 100-fold for three culture generations in vitro. These latter lines were greatly enriched for Lyt-1+2- cells and had little or no cytolytic activity, but they retained specific in vitro proliferative responses to the sensitizing alloantigen. Several Lyt-1-2+ cloned long-term lymphoid cell lines with high levels of specific cytolytic activity against the sensitizing alloantigen were derived and none was capable of mediating the accelerated rejection of skin grafts in vivo. However, cloned lymphoid cell lines that were phenotypically Lyt-1+2- and were capable of proliferating when in contact with specific alloantigen, but were not cytolytic, were capable of mediating the accelerated rejection of skin grafts in vivo both in irradiated mice and in nude mice. These studies demonstrate that skin allograft rejection can be mediated by Lyt-1+2- cell lines with specific in vitro proliferative activity to alloantigen although Lyt-1-2+ cell lines with cytolytic but not proliferative activity to alloantigen in vitro are ineffective in mediating graft rejection in vivo. Specific proliferative activity and no cytolysis appears to be a good in vitro correlate of the in vivo activity of long-term cultured cell lines.

Animals↗
PubMed1993

Response of nonreentrant catecholamine-mediated ventricular tachycardia to endogenous adenosine and acetylcholine. Evidence for myocardial receptor-mediated effects.

BACKGROUND: Reentrant ventricular tachycardia (VT) is known to be insensitive to the nucleoside adenosine. However, we have previously identified a form of nonreentrant, catecholamine-mediated VT that can be initiated with rapid pacing, demonstrates cycle length dependence, and is sensitive to exogenous adenosine as well as to the Valsalva maneuver. The mechanism of this tachycardia is thought to be due to a catecholamine-induced, cAMP-mediated increase in intracellular calcium, resulting in delayed afterdepolarizations and triggered activity. The antiarrhythmic effects of exogenous adenosine and Valsalva on this form of VT may be due to receptor-mediated inhibition of adenylate cyclase or to noncardiac receptor-mediated effects, i.e., exogenous adenosine may modulate VT through alterations in autonomic tone by activation of arterial chemoreceptors, and Valsalva has been shown to decrease venous return, resulting in a reduction in cardiac dimensions and myocardial stretch. To clarify this issue and circumvent both autonomic and noncardiac receptor effects, the response of nonreentrant catecholamine-mediated VT to endogenous adenosine and acetylcholine was evaluated. METHODS AND RESULTS: Group 1 (n = 8): Dipyridamole (0.56 mg/kg i.v.), a nucleoside transport blocker that potentiates the effects of endogenous adenosine, reproducibly abolished sustained nonreentrant, nonautomatic, catecholamine-mediated VT in the five patients in whom it was evaluated. VT recurred with the addition of aminophylline, a competitive adenosine A1-receptor antagonist. Edrophonium (10 mg i.v.), a cholinesterase inhibitor that potentiates the effects of acetylcholine at the muscarinic cholinergic receptor, terminated VT in four of four patients, an effect that was reversed by atropine. Group 2 (n = 6): In patients with reentrant VT, dipyridamole and edrophonium had no effect on VT cycle length or duration. Group 3 (n = 4): Adenosine and vagal maneuvers had no effect on catecholamine-mediated VT caused by automaticity in three of four patients tested. In one patient, adenosine transiently suppressed VT (< 5 seconds), after which it spontaneously resumed. CONCLUSIONS: The results of this study further delineate the mechanism of a newly recognized form of clinical VT. It can be identified by termination of the tachycardia in response to activation of the adenosine A1 or muscarinic cholinergic receptor, which results in inhibition of adenylate cyclase. These receptor-mediated effects appear to be specific for identifying nonreentrant, nonautomatic, catecholamine-mediated VT.

Acetylcholine↗
PubMed1982

Chemical mediator or mediators of insulin action: response to insulin and mode of action.

Studies with a subcellular system demonstrated that the interaction of insulin with the adipocyte plasma membrane resulted in the generation from the plasma membrane of a mediator that activated mitochondrial pyruvate dehydrogenase (EC 1.2.4.1). The insulin-sensitive chemical mediator from the plasma membrane has been partially characterized. It has a molecular weight of 1000-1500. The chemical mediator has been extracted from skeletal muscle, adipocytes, hepatoma cells, and IM-9 lymphocytes. Insulin increased the amount or activity of the mediator in the first three cell types, whereas insulin decreased the activity or amount of the mediator in IM-9 lymphocytes. These insulin-induced variations were consistent with the biological responses of these cells to insulin treatment. The activities of insulin-sensitive enzymes, including pyruvate dehydrogenase, adipocyte low Km 3':5'-cyclic-AMP phosphodiesterase (EC 3.1.4.17), and adipocyte plasma membrane [Ca2+ + Mg2+]-ATPase were shown to be altered by the chemical mediator. The mediator may act by altering various protein kinases and phosphoprotein phosphatases that modulate the state of phosphorylation and activity of these enzyme systems. The existence of two mediators is proposed. The first may mediate dephosphorylation of various substrates, and the second may influence phosphorylation.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed2002

A coregulatory role for the TRAP-mediator complex in androgen receptor-mediated gene expression.

The human thyroid hormone receptor-associated protein (TRAP)-Mediator complex was originally identified as a large multimeric complex that copurifies with the thyroid hormone receptor (TR) from HeLa cells and markedly enhances TR-mediated transcription in vitro. More recent studies have implicated TRAP-Mediator as a coactivator for a broad range of nuclear hormone receptors as well as other classes of transcriptional activators. Here we present evidence that TRAP-Mediator plays a functional role in androgen receptor (AR)-mediated transcription. We show that several subunits of the complex ligand-dependently coimmunoprecipitate with AR from both prostate cancer LNCaP cells and from HeLa cells stably transfected with AR. The 220-kDa subunit of the complex (TRAP220) can contact the ligand-binding domain of AR in vitro, possibly implicating TRAP220 involvement in targeting AR to the holocomplex. Consistent with a TRAP-Mediator coactivator role, transient overexpression of the TRAP220, TRAP170, and TRAP100 subunits enhanced ligand-dependent transcription by AR in cultured cells. Finally, chromatin immunoprecipitation assays show that TRAP220 is recruited to the androgen-responsive prostate-specific antigen gene promoter in vivo in ligand-stimulated LNCaP cells. Collectively, these data suggest that TRAP-Mediator may play an important coregulatory role in AR-mediated gene expression.

Antigens, Neoplasm↗
PubMed2007

Estimation of the mediation effect with a binary mediator.

A mediator acts as a third variable in the causal pathway between a risk factor and an outcome. In this paper, we consider the estimation of the mediation effect when the mediator is a binary variable. We give a precise definition of the mediation effect and examine asymptotic properties of five different estimators of the mediation effect. Our theoretical developments, which are supported by a Monte Carlo study, show that the estimators that account for the binary nature of the mediator are consistent for the mediation effect defined in this paper while other estimators are inconsistent. We use these estimators to study the mediation effect of chronic cerebral infarction in the causal relationship between the apolipoprotein E epsilon4 allele and cognitive function among 233 deceased participants from the Religious Orders Study, a longitudinal, clinical-pathologic study of aging and Alzheimer's disease.

Aging↗
PubMed1991

Regional differences of beta 1- and beta 2-adrenoceptor-mediated functions in feline heart. A beta 2-adrenoceptor-mediated positive inotropic effect possibly unrelated to cyclic AMP.

The effects of (-)-adrenaline and (-)-noradrenaline were studied on isolated preparations of kitten heart. To define the contribution of beta 1-adrenoceptors (beta 1AR) and beta 2-adrenoceptors (beta 2AR) we used as tools the highly beta 1AR-selective antagonist CGP 20,712 A and non-linear analysis of antagonism. The beta 2AR-mediated responses to the catecholamines, disclosed by CGP 20,712 A, were verified by blockade with the beta 2AR-selective ICI 118,551. The relative density and contribution of beta 1AR and beta 2AR to (-)-adrenaline- and (-)-noradrenaline-induced adenylyl cyclase stimulation was also estimated in right ventricular membranes. 1. In the sinoatrial pacemaker (-)-adrenaline caused positive chronotropic effects through both beta 1AR and beta 2AR while (-)-noradrenaline does so predominantly through beta 1AR. During beta 1AR blockade (-)-adrenaline did cause the same maximum effects through beta 2AR as (-)-noradrenaline did through beta 1AR. 2. In left atria (-)-adrenaline caused positive inotropic effects predominantly through beta 1AR. CGP 20,712 A also uncovered a beta 2AR component at high (-)-adrenaline concentrations comprising one third of the maximum beta 1AR-mediated response. 3. Receptor binding assays revealed that 80% of right ventricular beta AR were beta 1AR and 20% beta 2AR. Consistent with this finding, around 80% of the adenylyl cyclase stimulation by both (-)-noradrenaline and (-)-adrenaline was mediated through beta 1AR, around 20% through beta 2AR. The positive inotropic effects of (-)-noradrenaline appeared to be nearly exclusively mediated through beta 1AR in right ventricular papillary muscles. 4. The positive inotropic effects of (-)-adrenaline were quite variable with regard to beta 1AR and beta 2AR in right ventricular papillary muscles. Although beta 1AR-mediated effects are predominant in many muscles with only a small contribution of beta 2AR, in some muscles beta 2AR mediated around 50% of the maximum effect elicited through beta 1AR. In 3 out of 17 muscles beta 2AR mediated the same maximum effect of (-)-adrenaline as beta 1AR. 5. On occasion, we found marked beta AR heterogeneity amongst two muscles from the same right ventricle. One muscle only exhibited beta 1AR-mediated effects of (-)-adrenaline whereas in the other muscle maximal effects could be elicited through beta 2AR.(ABSTRACT TRUNCATED AT 400 WORDS)

Adrenergic beta-Antagonists↗
PubMed1997

Gbetagamma subunits mediate Src-dependent phosphorylation of the epidermal growth factor receptor. A scaffold for G protein-coupled receptor-mediated Ras activation.

In many cells, stimulation of mitogen-activated protein kinases by both receptor tyrosine kinases and receptors that couple to pertussis toxin-sensitive heterotrimeric G proteins proceed via convergent signaling pathways. Both signals are sensitive to inhibitors of tyrosine protein kinases and require Ras activation via phosphotyrosine-dependent recruitment of Ras guanine nucleotide exchange factors. Receptor tyrosine kinase stimulation mediates ligand-induced receptor autophosphorylation, which creates the initial binding sites for SH2 domain-containing docking proteins. However, the mechanism whereby G protein-coupled receptors mediate the phosphotyrosine-dependent assembly of a mitogenic signaling complex is poorly understood. We have studied the role of Src family nonreceptor tyrosine kinases in G protein-coupled receptor-mediated tyrosine phosphorylation in a transiently transfected COS-7 cell system. Stimulation of Gi-coupled lysophosphatidic acid and alpha2A adrenergic receptors or overexpression of Gbeta1gamma2 subunits leads to tyrosine phosphorylation of the Shc adapter protein, which then associates with tyrosine phosphoproteins of approximately 130 and 180 kDa, as well as Grb2. The 180-kDa Shc-associated tyrosine phosphoprotein band contains both epidermal growth factor (EGF) receptor and p185(neu). 3-5-fold increases in EGF receptor but not p185(neu) tyrosine phosphorylation occur following Gi-coupled receptor stimulation. Inhibition of endogenous Src family kinase activity by cellular expression of a dominant negative kinase-inactive mutant of c-Src inhibits Gbeta1gamma2 subunit-mediated and Gi-coupled receptor-mediated phosphorylation of both EGF receptor and Shc. Expression of Csk, which inactivates Src family kinases by phosphorylating the regulatory carboxyl-terminal tyrosine residue, has the same effect. The Gi-coupled receptor-mediated increase in EGF receptor phosphorylation does not reflect increased EGF receptor autophosphorylation, assayed using an autophosphorylation-specific EGF receptor monoclonal antibody. Lysophosphatidic acid stimulates binding of EGF receptor to a GST fusion protein containing the c-Src SH2 domain, and this too is blocked by Csk expression. These data suggest that Gbetagamma subunit-mediated activation of Src family nonreceptor tyrosine kinases can account for the Gi-coupled receptor-mediated tyrosine phosphorylation events that direct recruitment of the Shc and Grb2 adapter proteins to the membrane.

Animals↗
PubMed2001

Unsaturated fatty acid-mediated decreases in sterol regulatory element-mediated gene transcription are linked to cellular sphingolipid metabolism.

A major physiological feedback mechanism of cholesterol in transcription of a number of lipid metabolism-related genes is mediated by sterol regulatory elements (SREs) and their binding proteins (SREBPs). Polyunsaturated free fatty acids alone, as well as synergistically with sterols, decrease SRE-mediated gene expression up to 80% in a dose-dependent manner by decreasing levels of the active transcription factor SREBP. We investigated potential mechanisms for this effect. We hypothesized that free fatty acids reduce SREBP-mediated gene transcription by increasing intracellular cholesterol content through the hydrolysis of cellular sphingomyelin, which has a high affinity for free cholesterol. We also questioned whether the lipid second messenger ceramide, a product of sphingomyelin hydrolysis, can decrease SRE-mediated gene transcription. First we investigated the effect of fatty acids on sphingomyelin hydrolysis. Incubation of [(3)H]choline-labeled cells with unsaturated (but not saturated) fatty acids induced hydrolysis of [(3)H]choline-labeled sphingomyelin. Also, incubation of cell extracts from fatty acid-treated cells with [(3)H]sphingomyelin increased generation of [(3)H]ceramide compared with control cells in vitro. We found that addition of ceramide analogs alone and additively with fatty acids decreased SRE expression and that ceramide analogs reduced levels of the transcriptionally active forms of SREBP-1 and SREBP-2. Increasing intracellular ceramide levels by exogenous sphingomyelinase or inhibition of ceramidase decreased SRE-mediated gene expression. None of the above conditions induced apoptosis. Incubation with U18666A, a compound that inhibits intracellular cholesterol movement, increased SRE-mediated gene transcription. C(2)-ceramide abrogated the effect of U18666A on SRE-mediated gene transcription, suggesting cholesterol-independent regulation of SREBP. We provide evidence that sphingomyelin hydrolysis and intermediates of sphingomyelin metabolism (in addition to cholesterol and fatty acids) contribute to regulation of SRE-mediated gene transcription.

Animals↗
PubMedDate not supplied

Interaction between beta 2-adrenoceptor-mediated vasodilation and alpha 1-adrenoceptor-mediated vasoconstriction in the pithed normotensive rat.

With pithed normotensive rats we studied the interaction between beta 2-adrenoceptor-mediated vasodilation and alpha 1-adrenoceptor-mediated vasoconstriction. The selective beta 2-adrenoceptor agonist salbutamol was used to elicit vasodilatation. To induce alpha 1-adrenoceptor-mediated vasoconstriction, the selective alpha 1-adrenoceptor agonists cirazoline, St 587, and methoxamine were used. Furthermore, the alpha 1-adrenoceptor-mediated vasopressor effects of intravenously administered (--)-norepinephrine, and (--)-norepinephrine released from neurons by the nicotinic agonist 1,1-dimethyl-4-phenylpiperazine iodide (DMPP), the muscarinic ganglionic stimulant McN-A-343, electrical stimulation of the spinal cord, and the indirect sympathomimetic agent tyramine, were studied. By using the selective beta 2-adrenoceptor antagonist ICI 118,551, the interaction between the alpha 1-adrenoceptor-mediated vasoconstriction of (--)-epinephrine and alpha-methylnorepinephrine with their intrinsic beta 2-adrenoceptor agonistic effects was investigated. Two types of interaction between alpha 1-and beta 2-adrenoceptor-mediated vascular effects were found. Cirazoline and McN-A-343 activated alpha 1-adrenoceptors, inducing a vasoconstriction not affected by beta 2-adrenoceptor-mediated vasodilation. However, methoxamine at low doses, St 487, DMPP, electrical stimulation, intravenously administered (--)-norepinephrine, (--)-epinephrine, and alpha-methylnorepinephrine activated alpha 1-adrenoceptors, and their effect was attenuated by vasodilation. At low doses, tyramine stimulated alpha 1-adrenoceptors that were not sensitive to beta 2-adrenoceptor-mediated vasodilation, in contrast to the population of alpha 1-adrenoceptors activated at high doses of tyramine. It is hypothesized that there exist two different populations of alpha 1-adrenoceptors.

Adrenergic alpha-Agonists↗
PubMed1997

Comparison between copper-mediated and hypochlorite-mediated modifications of human low density lipoproteins evaluated by protein carbonyl formation.

The purpose of this study was to evaluate the mechanisms of apolipoprotein B (apoB) modification during oxidation of human low density lipoproteins (LDL) mediated either by copper or by hypochlorite (HOCl). The kinetics of protein carbonyl formation, the relationship of apoB carbonyl formation to lipid peroxidation, and the loss of apoB lysine residues were determined. During copper-mediated LDL oxidation, apoB carbonyls appeared to increase slowly, displayed saturation kinetics in response to increasing copper concentrations, and correlated with lipid peroxidation. During HOCl-mediated LDL oxidation, apoB carbonyls increased with increasing HOCl concentrations reaching plateau with time; however, lipid peroxidation was not observed. During copper-mediated but not during HOCl-mediated LDL oxidation, LDL vitamin E was depleted. ApoB carbonyls formed more efficiently during copper-mediated LDL oxidation at low (< 5 microM) copper concentrations compared with higher copper concentrations or during HOCl-mediated LDL oxidation. The differences in oxidation kinetics between copper- and HOCl-mediated LDL oxidation support the concept that the binding of copper to LDL is a site specific process, and suggest that HOCl modifies apoB amino acids randomly.

Apolipoproteins B↗
PubMed2004

Statistical assessment of mediational effects for logistic mediational models.

The concept of mediation has broad applications in medical health studies. Although the statistical assessment of a mediational effect under the normal assumption has been well established in linear structural equation models (SEM), it has not been extended to the general case where normality is not a usual assumption. In this paper, we propose to extend the definition of mediational effects through causal inference. The new definition is consistent with that in linear SEM and does not rely on the assumption of normality. Here, we focus our attention on the logistic mediation model, where all variables involved are binary. Three approaches to the estimation of mediational effects-Delta method, bootstrap, and Bayesian modelling via Monte Carlo simulation are investigated. Simulation studies are used to examine the behaviour of the three approaches. Measured by 95 per cent confidence interval (CI) coverage rate and root mean square error (RMSE) criteria, it was found that the Bayesian method using a non-informative prior outperformed both bootstrap and the Delta methods, particularly for small sample sizes. Case studies are presented to demonstrate the application of the proposed method to public health research using a nationally representative database. Extending the proposed method to other types of mediational model and to multiple mediators are also discussed.

Adolescent↗
PubMed2005

Role of clathrin- and caveolae-mediated endocytosis in gene transfer mediated by lipo- and polyplexes.

We investigated the effects of inhibitors of clathrin-mediated endocytosis (chlorpromazine and K(+) depletion) and of caveolae-mediated uptake (filipin and genistein) on internalization of FITC-poly-l-lysine-labeled DOTAP/DNA lipoplexes and PEI/DNA polyplexes by A549 pneumocytes and HeLa cells and on the transfection efficiencies of these complexes with the luciferase gene. Uptake of the complexes was assayed by fluorescence-activated cell sorting. Lipoplex internalization was inhibited by chlorpromazine and K(+) depletion but unaffected by filipin and genistein. In contrast, polyplex internalization was inhibited by all four inhibitors. We conclude that lipoplex uptake proceeds only by clathrin-mediated endocytosis, while polyplexes are taken up by two mechanisms, one involving caveolae and the other clathrin-coated pits. Transfection by lipoplexes was entirely abolished by blocking clathrin-mediated endocytosis, whereas inhibition of the caveolae pathway had no effect. By contrast, transfection mediated by polyplexes was completely blocked by genistein and filipin but was unaffected by inhibitors of clathrin-mediated endocytosis. Fluorescence colocalization studies with a lysosomal marker, AlexaFluor-dextran, revealed that polyplexes taken up by clathrin-mediated endocytosis are targeted to the lysosomal compartment for degradation, while the polyplexes internalized via caveolae escape this compartment, permitting efficient transfection.

Caveolae↗
PubMed2004

Alcohol expectancy mediation of biopsychosocial risk: complex patterns of mediation.

Alcohol expectancies' mediation of many known antecedents' influence on drinking has been well established, but most models have omitted theoretically relevant mediational pathways through both common (overlapping) and unique (nonshared) expectancy variance. The authors of this study evaluated expectancy mediation using a hierarchical expectancy model that specified both common and unique expectancy factors. The hierarchical model mediated a significantly (p <.05) larger percentage of the antecedent influences than a model specifying only common expectancy variance, revealed previously unseen mediational paths, and showed, to the authors' knowledge for the first time, expectancy mediation of protective as well as risk-related antecedents. It also allows the modeling of specific pathways for particular antecedents, rather than treating all expectancy mediation as though it flows through a general expectancy factor.

Adaptation, Psychological↗
PubMed2001

Piceatannol is an effective inhibitor of IgE-mediated secretion from human basophils but is neither selective for this receptor nor acts on syk kinase at concentrations where mediator release inhibition occurs.

BACKGROUND: Syk kinase is probably an early necessary tyrosine kinase involved in IgE-mediated secretion from human basophils. Causal testing of the role of syk kinase in the secretion requires a selective pharmacological agent. Piceatannol has previously been used to demonstrate the causal role of syk in secretion but its selectively has recently come into question. OBJECTIVE: To determine whether piceatannol inhibits IgE-mediated signalling events in a manner consistent with its putative inhibitory effects on syk kinase and at concentrations relevant to its inhibition of mediator release. METHODS: Human basophils were examined for the effects of piceatannol on mediator release or various signalling steps. RESULTS: We show that while piceatannol has an IC50 for inhibition of IgE-mediated histamine release of 3-5 microm, these same concentrations inhibit secretion of phorbol 12-myristate 13-acetate (PMA)-induced histamine release (as previously shown) and leukotriene C (LTC)4 release induced by fMLP. Concentrations of piceatannol up to 100 microm also did not inhibit IgE-mediated phosphorylation of shc, a immediate downstream target of syk kinase. Similar concentrations also did not inhibit IgE-mediated cytosolic calcium elevations, another downstream signal thought to be dependent on syk kinase. In contrast, piceatannol did modify the cytosolic calcium response that follows stimulation with formyl methionyl-leucyl-phenylalanine (fMLP). CONCLUSION: Taken together with published studies using other cell types, we conclude that piceatannol does not inhibit secretion from human basophils by inhibiting the activity of syk kinase.

Antibodies, Anti-Idiotypic↗