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Biomedical subjects

Robert F Jacob

Publications and source records attributed to Robert F Jacob.

15 recordsLinked to original sources
PubMed2007

Regulation of the gating of BKCa channel by lipid bilayer thickness.

Transmembrane segments of ion channels tend to match the hydrophobic thickness of lipid bilayers to minimize mismatch energy and to maintain their proper organization and function. To probe how ion channels respond to mismatch with lipid bilayers of different thicknesses, we examined the single channel activities of BK(Ca) (hSlo alpha-subunit) channels in planar bilayers of binary mixtures of DOPE (1,2-dioleoyl-sn-glycero-3-phosphoethanolamine) with phosphatidylcholines (PCs) of varying chain lengths, including PC 14:1, PC 18:1, PC 22:1, PC 24:1, and with porcine brain sphingomyelin. Bilayer thickness and structure was measured with small angle x-ray diffraction and atomic force microscopy. The open probability (P(o)) of the BK(Ca) channel was finely tuned by bilayer thickness, first decreasing with increases in bilayer thickness from PC 14:1 to PC 22:1 and then increasing from PC 22:1 to PC 24:1 and to porcine brain sphingomyelin. Single channel kinetic analyses revealed that the mean open time of the channel increased monotonically with bilayer thickness and, therefore, could not account for the biphasic changes in P(o). The mean closed time increased with bilayer thickness from PC 14:1 up to PC 22:1 and then decreased with further increases in bilayer thickness to PC 24:1 and sphingomyelin, correlating with changes in P(o). This is consistent with the proposition that bilayer thickness affects channel activity mainly through altering the stability of the closed state. We suggest a simple mechanical model that combines forces of lateral stress within the lipid bilayer with local hydrophobic mismatch between lipids and the protein to account for the biphasic modulation of BK(Ca) gating.

Calcium↗
PubMed2006

Differential effects of carotenoids on lipid peroxidation due to membrane interactions: X-ray diffraction analysis.

The biological benefits of certain carotenoids may be due to their potent antioxidant properties attributed to specific physico-chemical interactions with membranes. To test this hypothesis, we measured the effects of various carotenoids on rates of lipid peroxidation and correlated these findings with their membrane interactions, as determined by small angle X-ray diffraction approaches. The effects of the homochiral carotenoids (astaxanthin, zeaxanthin, lutein, beta-carotene, lycopene) on lipid hydroperoxide (LOOH) generation were evaluated in membranes enriched with polyunsaturated fatty acids. Apolar carotenoids, such as lycopene and beta-carotene, disordered the membrane bilayer and showed a potent pro-oxidant effect (>85% increase in LOOH levels) while astaxanthin preserved membrane structure and exhibited significant antioxidant activity (40% decrease in LOOH levels). These findings indicate distinct effects of carotenoids on lipid peroxidation due to membrane structure changes. These contrasting effects of carotenoids on lipid peroxidation may explain differences in their biological activity.

Antioxidants↗
PubMed2006

Active metabolite of atorvastatin inhibits membrane cholesterol domain formation by an antioxidant mechanism.

The advanced atherosclerotic lesion is characterized by the formation of microscopic cholesterol crystals that contribute to mechanisms of inflammation and apoptotic cell death. These crystals develop from membrane cholesterol domains, a process that is accelerated under conditions of hyperlipidemia and oxidative stress. In this study, the comparative effects of hydroxymethylglutaryl-CoA (HMG-CoA) reductase inhibitors (statins) on oxidative stress-induced cholesterol domain formation were tested in model membranes containing physiologic levels of cholesterol using small angle x-ray diffraction approaches. In the absence of HMG-CoA reductase, only the atorvastatin active o-hydroxy metabolite (ATM) blocked membrane cholesterol domain formation as a function of oxidative stress. This effect of ATM is attributed to electron donation and proton stabilization mechanisms associated with its phenoxy group located in the membrane hydrocarbon core. ATM inhibited lipid peroxidation in human low density lipoprotein and phospholipid vesicles in a dose-dependent manner, unlike its parent and other statins (pravastatin, rosuvastatin, simvastatin). These findings indicate an atheroprotective effect of ATM on membrane lipid organization through a potent antioxidant mechanism.

Antioxidants↗
PubMed2006

Rofecoxib increases susceptibility of human LDL and membrane lipids to oxidative damage: a mechanism of cardiotoxicity.

Clinical investigations have demonstrated a relationship between the extended use of rofecoxib and the increased risk for atherothrombotic events. This has led to the removal of rofecoxib from the market and concern over the cardiovascular safety of other cyclooxygenase (COX)-2 selective agents. Experimental findings from independent laboratories now indicate that the cardiotoxicity of rofecoxib may not be a class effect but because of its intrinsic chemical properties. Specifically, rofecoxib has been shown to increase the susceptibility of human low-density lipoprotein and cellular membrane lipids to oxidative modification, a contributing factor to plaque instability and thrombus formation. Independently of COX-2 inhibition, rofecoxib also promoted the nonenzymatic formation of isoprostanes and reactive aldehydes from biologic lipids. The basis for these observations is that rofecoxib alters lipid structure and readily forms a reactive maleic anhydride in the presence of oxygen. By contrast, other selective (celecoxib, valdecoxib) and nonselective (naproxen, diclofenac) inhibitors did not influence rates of low-density lipoprotein and membrane lipid oxidation. We have now further confirmed these findings by demonstrating that the prooxidant activity of rofecoxib can be blocked by the potent antioxidant astaxanthin in homochiral form (all-trans 3S, 3'S). These findings provide a mechanistic rationale for differences in cardiovascular risk among COX-selective inhibitors because of their intrinsic physicochemical properties.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed2006

Effect of nebivolol on endothelial nitric oxide and peroxynitrite release in hypertensive animals: Role of antioxidant activity.

We tested the activity of nebivolol, a beta1-selective blocker with respect to nitric oxide (NO) and peroxynitrite (ONOO) generation in the endothelium of normotensive Wistar Kyoto (WKY rats) and spontaneously hypertensive rats (SHR). The endothelial effects of nebivolol and its 2 optical enantiomers were correlated with its antioxidant activity and compared to another beta-blocker, atenolol, and 2 agonists of nitric oxide synthase (eNOS), calcium ionophore (CI) and acetylcholine (ACh). The effects of nebivolol on the bioavailability of NO and ONOO, indicators of endothelial function and dysfunction, respectively, were measured in vitro using nanosensors placed in mesenteric arteries. Compared with WKY rats, treatment of SHR vessels either with ACh (1 micromol/L) or CI (1 micromol/L) showed marked deficiencies (>40%, P < 0.01) in bioavailable NO concomitant with increased ONOO levels (>50%, P < 0.01). The [NO]/[ONOO] ratio measured after stimulation with CI was 2.77 +/- 0.05 in WKY rats and much lower (1.14 +/- 0.11) in SHR indicating significant eNOS uncoupling and endothelial dysfunction in hypertensive animals. Treatment with nebivolol (10 micromol/L) inhibited eNOS uncoupling and reduced endothelial dysfunction in SHR, as evidenced by an increase in the [NO]/[ONOO] ratio to 3.09 +/- 0.04. The basis for nebivolol activity is attributed to its unique membrane interactions as determined by small-angle x-ray diffraction, as well as its antioxidant activity at nanomolar to micromolar levels. The antioxidant effects of nebivolol and its enantiomers were not reproduced by atenolol. These results demonstrate that nebivolol inhibits endothelial dysfunction through a potent antioxidant mechanism attributed to its physicochemical interactions with the membrane, independent of beta1-blockade activity.

Acetylcholine↗
PubMed2005

Nebivolol reduces nitroxidative stress and restores nitric oxide bioavailability in endothelium of black Americans.

BACKGROUND: Alterations in endothelial function may contribute to increased susceptibility of black Americans to cardiovascular disease. The ability to pharmacologically reverse endothelial dysfunction in blacks was tested with nebivolol, a beta1-selective agent with vasodilating and antioxidant properties. METHODS AND RESULTS: The effects of nebivolol on endothelial nitric oxide (NO), superoxide (O2-), and peroxynitrite concentration (ONOO-) release were studied in human umbilical vein endothelial cells and iliac artery endothelial cells isolated from age-matched black and white donors. Kinetics and concentrations of NO/O2-/ONOO- were measured simultaneously with nanosensors from single cells and shown to have significant interracial differences. The rate of NO release was &5 times slower in blacks than in whites (94 versus 505 nmol . L(-1).s(-1)), whereas the rates of release were faster by &2 times for O2- and &4 times for ONOO- (22.1 versus 9.4 nmol.L(-1).s(-1) for O2- and 810 versus 209 nmol.L(-1).s(-1) for ONOO-). Pretreatment with 1.0 to 5.0 micromol/L nebivolol restored NO bioavailability in endothelial cells from black donors with concurrent reductions in O2- and ONOO- release, similar to levels in the endothelium of whites. The effects of nebivolol were dose-dependent and not observed with atenolol; similar effects were observed with apocynin, an NAD(P)H oxidase inhibitor. CONCLUSIONS: Reduced endothelial NO bioavailability in American blacks is mainly due to excessive O2- and ONOO- generation by NAD(P)H and uncoupled endothelial NO synthase. Nebivolol decreased O2- and ONOO- concentrations and restored NO bioavailability in blacks to the level recorded in cells from whites, independently of beta1-selective blockade.

Adult↗
PubMed2005

Lipid peroxidation induces cholesterol domain formation in model membranes.

Numerous reports have established that lipid peroxidation contributes to cell injury by altering the basic physical properties and structural organization of membrane components. Oxidative modification of polyunsaturated phospholipids has been shown, in particular, to alter the intermolecular packing, thermodynamic, and phase parameters of the membrane bilayer. In this study, the effects of oxidative stress on membrane phospholipid and sterol organization were measured using small angle x-ray diffraction approaches. Model membranes enriched in dilinoleoylphosphatidylcholine were prepared at various concentrations of cholesterol and subjected to lipid peroxidation at physiologic conditions. At cholesterol-to-phospholipid mole ratios (C/P) as low as 0.4, lipid peroxidation induced the formation of discrete, membrane-restricted cholesterol domains having a unit cell periodicity or d-space value of 34 A. The formation of cholesterol domains correlated directly with lipid hydroperoxide levels and was inhibited by treatment with vitamin E. In the absence of oxidative stress, similar cholesterol domains were observed only at C/P ratios of 1.0 or higher. In addition to changes in sterol organization, lipid peroxidation also caused reproducible changes in overall membrane structure, including a 10 A reduction in the width of the surrounding, sterol-poor membrane bilayer. These data provided direct evidence that lipid peroxidation alters the essential organization and structure of membrane lipids in a manner that may contribute to changes in membrane function during aging and oxidative stress-related disorders.

Cholesterol↗
PubMed2005

Intermolecular differences of 3-hydroxy-3-methylglutaryl coenzyme a reductase inhibitors contribute to distinct pharmacologic and pleiotropic actions.

Statin drugs inhibit 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase and share the common mechanism of lowering circulating levels of low-density lipoprotein (LDL) cholesterol, a powerful indicator of risk for cardiovascular disease. Large clinical trials have documented the benefit of hypolipidemic therapy for both primary and secondary prevention of coronary artery disease and stroke. Recent clinical findings, including direct comparator studies, now indicate that certain statins may slow progression of disease at a rate and to an extent that cannot be solely attributed to LDL reduction. The proposed mechanisms for such pleiotropic actions include enhancement of endothelial-dependent nitric oxide bioavailability, anti-inflammatory activity, and inhibition of oxidative stress. To understand the biochemical basis for such differences among statins, this article reviews their physicochemical properties and pharmacology at the molecular level.

Cholesterol, LDL↗
PubMed2004

Serum levels of thiobarbituric acid reactive substances predict cardiovascular events in patients with stable coronary artery disease: a longitudinal analysis of the PREVENT study.

OBJECTIVES: The objective of this study was to test the predictive value of an oxidative stress biomarker in 634 patients from the Prospective Randomized Evaluation of the Vascular Effects of Norvasc Trial (PREVENT). BACKGROUND: Oxidative stress contributes to mechanisms of atherosclerosis and plaque instability. Biomarkers of oxidation, such as malondialdehyde (MDA), may represent independent indicators of risk for patients with stable coronary artery disease (CAD). METHODS: Serum MDA levels were measured as thiobarbituric acid reactive substances (TBARS) in 634 patients with documented CAD using reverse-phase high-performance liquid chromatography and spectrophotometric approaches. RESULTS: During the three-year study, there were 51 major vascular events such as fatal/nonfatal myocardial infarction, 149 hospitalizations for nonfatal vascular events, and 139 patients underwent a major vascular procedure. At baseline, patients with TBARS levels in the highest quartile had a relative risk (RR) of 3.30 (95% confidence interval [CI] 1.47 to 7.42; p = 0.038) for major vascular events, RR of 4.10 (95% CI 2.55 to 6.60; p < 0.0001) for nonfatal vascular events, and RR of 3.84 (95% CI 2.56 to 5.76; p < 0.0001) for major vascular procedures. The effect of TBARS on events and procedures was also seen in a multivariate model adjusted for inflammatory markers (C-reactive protein, soluble intercellular adhesion molecule-1, interleukin-6), and other risk factors (age, low-density lipoprotein, high-density lipoprotein, total cholesterol, triglycerides, body mass index, and blood pressure). This analysis showed an independent effect of TBARS on major vascular events (p = 0.0149), nonfatal vascular events (p < 0.0001), major vascular procedures (p < 0.001), and all vascular events and procedures (p < 0.0001). CONCLUSIONS: Serum levels of TBARS were strongly predictive of cardiovascular events in patients with stable CAD, independently of traditional risk factors and inflammatory markers.

Adult↗
PubMed2004

Effects of HMG-CoA reductase inhibitors on endothelial function: role of microdomains and oxidative stress.

Certain pleiotropic activities reported for 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) are related to reductions in cellular cholesterol biosynthesis and isoprenoid levels. In endothelial cells, these metabolic changes contribute to favorable effects on nitric oxide (NO) bioavailability. Given the essential role of NO in preserving vascular structure and function, this effect of statins is of considerable therapeutic importance. Statins have been demonstrated to restore endothelial NO production by several mechanisms, including upregulating endothelial NO synthase (eNOS) protein expression and blocking formation of reactive oxygen species. In this article, we will discuss additional ways in which statins restore endothelial NO production and improve endothelial function. (1) Statins modulate membrane microdomain formation, resulting in reduced expression of proteins that specifically inhibit eNOS activation. (2) Statins reduce sterol biosynthesis, thus interfering with the formation of pathologic microdomains, including cholesterol crystalline structures. This observation has important implications for plaque stabilization, as these microdomains contribute to cholesterol crystal formation and endothelial apoptosis. Finally, (3) statins improve endothelial function by interfering with oxidative stress pathways through both enzymatic and nonenzymatic mechanisms. The relationships between membrane microdomains, cholesterol biosynthesis, and endothelial function will be discussed.

Animals↗
PubMed2004

Sulfone COX-2 inhibitors increase susceptibility of human LDL and plasma to oxidative modification: comparison to sulfonamide COX-2 inhibitors and NSAIDs.

Clinical investigations have demonstrated a link between use of the sulfone cyclooxygenase-2 (COX-2) inhibitor, rofecoxib, and increased risk for atherothrombotic events. This increased risk was not observed for a sulfonamide COX-2 inhibitor (celecoxib), indicating a potential non-enzymatic mechanism for rofexocib. To test this hypothesis, we compared the independent effects of COX-2 inhibitors on human LDL oxidation, an important contributor to atherosclerotic cardiovascular disease. The results showed that rofecoxib (100 nM) significantly decreased (>40%, p<0.001) the lag time for LDL conjugated diene formation and increased levels of thiobarbituric-acid-reactive-substances (TBARS) in vitro. The pro-oxidant activity of rofecoxib was dose-dependent and attenuated by 70% (p<0.001) with the antioxidant, Trolox. Rofecoxib and etoricoxib (100 nM) also caused a marked increase (>35%, p<0.001) in non-enzymatic generation of isoprostanes, as measured by mass spectroscopy. Addition of rofecoxib to fresh human plasma reduced the oxygen radical antioxidant capacity (ORAC) by 34% (p<0.0001). By contrast, other selective (celecoxib, valdecoxib, meloxicam) and non-selective COX inhibitors (ibuprofen, naproxen, diclofenac) had no significant effect on LDL oxidation rates or plasma ORAC values, even at suprapharmacologic levels. X-ray diffraction analysis showed that sulfone COX-2 inhibitors interact differently with membrane phospholipids, suggesting a physico-chemical basis for the pro-oxidant activity. These results demonstrate that sulfone COX-2 inhibitors increase the susceptibility of biological lipids to oxidative modification through a non-enzymatic process. These findings may provide mechanistic insight into reported differences in cardiovascular risk for COX-2 inhibitors.

Animals↗
PubMed2004

The antioxidant effects of statins.

Oxidative stress contributes to the initiation and the development of atherosclerotic plaques and adversely influences myocardial integrity. Statins interfere with oxidation in several ways that may contribute to reducing the atherogenic process. In addition to direct antioxidant effects, statins reduce circulating oxidized low-density lipoproteins (oxLDL) and inhibit their uptake by macrophages. They also reduce circulating markers of oxidation such as F2-isoprostane and nitrotyrosine. Statins inhibit oxidant enzymes activity such as that of reduced nicotinamide adenine dinucleotide phosphate (NAD[P]H) oxidase and myeloperoxidase and up-regulate the activity of antioxidant enzymes such as catalase and paraoxonase. They reduce endothelial dysfunction mainly by their ability to enhance endothelial nitric oxide bioavailability, which is achieved by several mechanisms. The antioxidant properties of statins extend to organ protection especially the myocardium and the lungs.

Animals↗
PubMed2003

Direct evidence for cholesterol crystalline domains in biological membranes: role in human pathobiology.

This review will discuss the use of small-angle X-ray diffraction approaches to study the organization of lipids in plasma membranes derived from two distinct mammalian cell types: arterial smooth muscle cells and ocular lens fiber cells. These studies indicate that cholesterol at an elevated concentration can self-associate and form immiscible domains in the plasma membrane, a phenomenon that contributes to both physiologic and pathologic cellular processes, depending on tissue source. In plasma membrane samples isolated from atherosclerotic smooth muscle cells, the formation of sterol-rich domains is associated with loss of normal cell function, including ion transport activity and control of cell replication. Analysis of meridional diffraction patterns from intact and reconstituted plasma membrane samples indicates the presence of an immiscible cholesterol domain with a unit cell periodicity of 34 A, consistent with a cholesterol monohydrate tail-to-tail bilayer, under disease conditions. These cholesterol domains were observed in smooth muscle cells enriched with cholesterol in vitro as well as from cells obtained ex vivo from an animal model of atherosclerosis. By contrast, well-defined cholesterol domains appear to be essential to the normal physiology of fiber cell plasma membranes of the human ocular lens. The organization of cholesterol into separate domains underlies the role of lens fiber cell plasma membranes in maintaining lens transparency. These domains may also interfere with cataractogenic aggregation of soluble lens proteins at the membrane surface. Taken together, these analyses provide examples of both physiologic and pathologic roles that sterol-rich domains may have in mammalian plasma membranes. These findings support a model of the membrane in which cholesterol aggregates into structurally distinct regions that regulate the function of the cell membrane.

Arteries↗