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Functionality of C-Reactive Protein for AtheroprotectionSingh, Sanjay K., Agrawal, Alok 01 January 2019 (has links)
C-reactive protein (CRP) is a pentameric molecule made up of identical monomers. CRP can be seen in three different forms: native pentameric CRP (native CRP), non-native pentameric CRP (nonnative CRP), and monomeric CRP (mCRP). Both native and nonnative CRP execute ligand-recognition functions for host defense. The fate of any pentameric CRP after binding to a ligand is dissociation into ligand-bound mCRP. If ligand-bound mCRP is proinflammatory, like free mCRP has been shown to be in vitro, then mCRP along with the bound ligand must be cleared from the site of inflammation. Once pentameric CRP is bound to atherogenic low-density lipoprotein (LDL), it reduces both formation of foam cells and proinflammatory effects of atherogenic LDL. A CRP mutant, that is non-native CRP, which readily binds to atherogenic LDL, has been found to be atheroprotective in a murine model of atherosclerosis. Thus, unlike statins, a drug that can lower only cholesterol levels but not CRP levels should be developed. Since non-native CRP has been shown to bind to all kinds of malformed proteins in general, it is possible that non-native CRP would be protective against all inflammatory states in which host proteins become pathogenic. If it is proven through experimentation employing transgenic mice that non-native CRP is beneficial for the host, then using a small-molecule compound to target CRP with the goal of changing the conformation of endogenous native CRP would be preferred over using recombinant non-native CRP as a biologic to treat diseases caused by pathogenic proteins such as oxidized LDL.
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Conformationally Altered C-Reactive Protein Capable of Binding to Atherogenic Lipoproteins Reduces AtherosclerosisPathak, Asmita, Singh, Sanjay K., Thewke, Douglas P., Agrawal, Alok 11 August 2020 (has links)
The aim of this study was to test the hypothesis that C-reactive protein (CRP) protects against the development of atherosclerosis and that a conformational alteration of wild-type CRP is necessary for CRP to do so. Atherosclerosis is an inflammatory cardiovascular disease and CRP is a plasma protein produced by the liver in inflammatory states. The co-localization of CRP and low-density lipoproteins (LDL) at atherosclerotic lesions suggests a possible role of CRP in atherosclerosis. CRP binds to phosphocholine-containing molecules but does not interact with LDL unless the phosphocholine groups in LDL are exposed. However, CRP can bind to LDL, without the exposure of phosphocholine groups, if the native conformation of CRP is altered. Previously, we reported a CRP mutant, F66A/T76Y/E81A, generated by site-directed mutagenesis, that did not bind to phosphocholine. Unexpectedly, this mutant CRP, without any more conformational alteration, was found to bind to atherogenic LDL. We hypothesized that this CRP mutant, unlike wild-type CRP, could be anti-atherosclerotic and, accordingly, the effects of mutant CRP on atherosclerosis in atherosclerosis-prone LDL receptor-deficient mice were evaluated. Administration of mutant CRP into mice every other day for a few weeks slowed the progression of atherosclerosis. The size of atherosclerotic lesions in the aorta of mice treated with mutant CRP for 9 weeks was ~40% smaller than the lesions in the aorta of untreated mice. Thus, mutant CRP conferred protection against atherosclerosis, providing a proof of concept that a local inflammation-induced structural change in wild-type CRP is a prerequisite for CRP to control the development of atherosclerosis.
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Influence of Stress and Blood Type on Toxicity‐preventing Activity and Other Cardiac Risk FactorsNeumann, Joseph K., Arbogast, Loretta Y., Dubberley, F. Aaron 01 January 1994 (has links)
ABO blood type has been shown to be associated with both cardiovascular risk and toxicity‐preventing activity (TxPA) stress response in elderly males. Twenty middle‐aged, healthy males, 14 blood type A and six blood type O, were involved in this project. Volunteers completed a battery of psychological assessments, then gave blood and had several psychophysiological measures taken prior to, during and after two stressors. The stressors consisted of mental arithmetic tasks plus audiotapes of combat sounds and a baby crying. The anger‐out and hard‐driving scores of blood type O subjects were significantly higher than the blood type A means. TxPA decreased significantly as a function of stress and some suggestive blood type effects of TxPA were found. Plasma protein, microhematocrit, plasma cortisol, finger temperature, skin conductance, blood pressure and two facial electromyograph (EMG) variables were also significantly affected by stressors but not by the blood type factor. No significant differences of any kind were found for total cholesterol, high‐density lipoprotein or pulse variables. The importance of age and other individual subject characteristics was discussed.
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Lipid Hydroperoxides Inhibit Nitric Oxide Production in RAW264.7 MacrophagesHuang, Annong, Li, Chuanfu, Kao, Race L., Stone, William L. 01 March 1999 (has links)
The effects of oxidatively modified low density lipoprotein (oxLDL) on atherogenesis may be partly mediated by alterations in the production of nitric oxide (NO) by vascular cells. Lipid hydroperoxides (LOOH) and lysophosphatidylcholine (lysoPC) are the major primary products of LDL oxidation. The purpose of this study was to characterize the effects of oxLDL, LOOH and lysoPC on NO production and the expression of inducible nitric oxide synthase (iNOS) gene in lipopolysaccharide (LPS) stimulated macrophages. LDL was oxidized using an azo-initiator 2,2'-azobis (2- amidinopropane) HCl (ABAP) and octadecadienoic acid was oxidized by lipoxygenase to generate 13-hydroperoxyl octadecadienoic acid (13-HPODE). Our study showed that oxLDL markedly decreased the production of NO, the levels of iNOS protein and iNOS mRNA in LPS stimulated macrophages. The inhibition potential of oxLDL on NO production and iNOS gene expression depended on the levels of LOOH formed in oxLDL and was not due to oxLDL cytotoxicity. Furthermore, 13-HPODE markedly reduced NO production and iNOS protein levels, whereas lysoPC showed only slight reduction. The effects of 13-HPODE and lysoPC did not require an acetylated LDL carrier. Our results suggest that 13-HPODE is a much more potent inhibitor of NO production and iNOS gene expression than lysoPC in LPS stimulated RAW264.7 macrophages.
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Phosphoethanolamine-Complexed C-Reactive Protein: A Pharmacological-Like Macromolecule That Binds to Native Low-Density Lipoprotein in Human SerumSingh, Sanjay, Suresh, Madathilparambil V., Prayther, Deborah C., Moorman, Jonathan P., Rusiñol, Antonio E., Agrawal, Alok 01 August 2008 (has links)
Background: C-reactive protein (CRP) is an acute phase plasma protein. An important binding specificity of CRP is for the modified forms of low-density lipoprotein (LDL) in which the phosphocholine-binding sites of CRP participate. CRP, however, does not bind to native LDL. Methods: We investigated the interaction of CRP with native LDL using sucrose density gradient ultracentrifugation. Results: We found that the blocking of the phosphocholine-binding sites of CRP with phosphoethanolamine (PEt) converted CRP into a potent molecule for binding to native LDL. In the presence of PEt, CRP acquired the ability to bind to fluid-phase purified native LDL. Because purified native LDL may undergo subtle modifications, we also used whole human serum as the source of native LDL. In the presence of PEt, CRP bound to native LDL in serum also. The effect of PEt on CRP was selective for LDL because PEt-complexed CRP did not bind to high-density lipoprotein in the serum. Conclusions: The pharmacologic intervention of endogenous CRP by PEt-based compounds, or the use of exogenously prepared CRP-PEt complexes, may turn out to be an effective approach to capture native LDL cholesterol in vivo to prevent the development of atherosclerosis.
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Assessing the efficacy of artemisinin-based combination therapies (ACTs) against Plasmodium malariae and Plasmodium ovale infections with low parasite densities: overcoming challenges during molecular analysesBroumou, Ioanna January 2020 (has links)
Background: Malaria is a major public health issue. Artemisinin-based combination therapies are the WHO recommended treatment for uncomplicated malaria. Plasmodium malariae and Plasmodium ovale infections are considered underestimated and the effectiveness of artemisinin-based combination treatments against them is poorly documented. The aim of this study was to evaluate the efficacy of dihydroartemisinin-piperaquine against low parasite density Plasmodium malariae and ovale infections. Methods: DNA was extracted from dried blood spots on filter papers with Chelex®-100 or a column-based extraction method. Species detection and determination was conducted by SYBR Green quantitative PCR targeting the cytochrome b gene (cytb-qPCR) followed by restriction fragment length polymorphism analyses. In total, 241 samples from 53 patients enrolled in a clinical trial were analysed. The obtained molecular data were compared with the microscopy data of the study. Results: Only 69 out of 143 microscopy-positive samples were confirmed as positive by cytb-qPCR. Ninety-three samples were identified as parasite negative by both microscopy and PCR. None of the 36 microscopy-defined coinfections were detected in the molecular analysis. The cytb-qPCR success rate was 72.9% (CI95% 61.4-82.6), 75.0% (CI95% 34.9-96.8) and 14.8% (CI95% 6.9-26.2) for parasite densities above 1000 parasites/ μL, between 600-1000 parasites/ μL and below 600 parasites/ μL, respectively. The observed poor qPCR success rate is most likely due to sample degradation under poor storage conditions. Conclusions: This study highlights the impact on the preservation and quality of Plasmodium genomic DNA on dried blood spots, when filter papers are stored for more than 3 years in tropical conditions.
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Injury of Arterial Endothelial Cells in Diabetic, Sucrose-Fed and Aged RatsArbogast, Bradley W., Berry, Dianne L., Newell, Chris L. 01 January 1984 (has links)
The toxicity of elevated levels of very low density lipoproteins (VLDL, d < 1.006 g/ml) was investigated using porcine aortic endothelial cells in vitro. VLDL isolated from normal rat serum and added at elevated levels was as toxic as VLDL isolated from streptozotocin-induced diabetic rat serum. Injury was detected by scanning electron microscopy in 4-day-old primary cultures of endothelial cells after a 1 2 exposure to diabetic rat serum. Bleb formation and contraction was seen first in isolated cells ( 1 2 h), followed by cells at the periphery of the monolayer (1 h) and finally in cells throughout the monolayer (4 h). By 10 h few cells remained attached to the dish. A similar sequence of events occurred in 1-day-old cultures after a 3-h lag period. Serum from sucrose-fed as well as aged rats was also found to be toxic to endothelial cells in vitro. Elevated levels of VLDL were responsible for the toxicities of these sera. Scanning electron microscopy of the aortas from diabetic and sucrose-fed rats revealed endothelial desquamation, platelet and leukocyte attachment, fibrin deposition and the presence of microthrombi. The common occurrence of both micro- and macrovascular disease in diabetic, sucrose-fed, and aged rats and the toxicity of their serum in vitro suggest that elevated levels of VLDL may initiate vascular disease in these models.
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Degradation of Microplastic Residuals in Water by Visible Light PhotocatalysisTofa, Tajkia Syeed January 2018 (has links)
Microplastic (MP) pollution has recently been recognized as a threat to the biosphere including humans due to its widespread distribution, persistent nature and infinitesimal size. This study focused on the solid phase degradation of microplastic residues (particularly low density polyethylene, LDPE) in water through heterogeneous photocatalysis process by designed photocatalysts of zinc oxide nanorods (ZnO NRs) and platinum nanoparticles deposited on zincoxide nanorods (Pt NPs-ZnO NRs) under visible light irradiation. These photocatalysts were assessed following standard protocol (ISP 10678: 2010), and characterized using SEM, EDX andoptical spectroscopies (UV-VIS and PL). Deposition of Pt-NPs on ZnO NRs for certain minutes has been found optimum that enhanced the photodegradation process about 38% under UV irradiation and 16.5% under visible light irradiation by improving of both electrons-holes pair separation process and visible light absorption. Photocatalytic degradation of LDPE films was confirmed by FTIR spectroscopy, dynamic mechanical analyzer (DMA), optical and electron microscopes. When LDPE film irradiated in presence of Pt-ZnO, degradation was found quicker than ZnO alone of similar concentration which exhibited formation of a large number of wrinkles, cracks and cavities on the film surface. Dynamic mechanical analyzer (DMA) test indicated stiffness and embrittlement of exposed LDPE films in presence of photocatalysts. Thus, the present work provides a new insight about modified catalysts for the degradation of microplastics in water using visible light.
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Differential Expression Of Proteins Involved In VLDL Trafficking Causes Reduced VLDL Secretion In Male Ames Dwarf MiceAhmed Moinuddin, Faisal 01 January 2015 (has links)
Cardiovascular diseases (CVDs) have been recorded as the number one cause of death worldwide, accounting for 32% of total deaths annually. More than two-thirds of all CVD cases are associated with atherosclerosis, which is the accumulation of fats and other substances causing plaque formation in the interior walls of major arteries. This leads to narrowing of the lumen and hardening of the arteries, ultimately resulting in angina, heart attack and/or stroke. Studies have shown that the pathogenesis of atherosclerosis and associated CVDs is strongly linked to elevated secretion of liver-specific lipoproteins called very-low-density-lipoprotein (VLDL). VLDLs are crucial lipoproteins responsible for transportation of triacylglycerides (TAGs), chemically inert particles that are physiologically significant for their energy storing capacity, from the liver to peripheral tissues. These VLDL particles are synthesized in the lumen of the endoplasmic reticulum (ER) of hepatocytes, transported from the ER to the cis-Golgi in special transport vesicles called VLDL-transport-vesicles (VTVs) and secreted into plasma through a highly regulated secretory pathway. Previous studies from our laboratory have shown that VTV-mediated ER-to-Golgi VLDL trafficking is the rate-limiting step in overall VLDL secretion from hepatocytes into plasma. In this project, we investigated intracellular VLDL trafficking and VLDL secretion in Ames dwarf (Prop1df, df/df) mice, a mutant mouse model homozygous for a recessive mutation at Prop1 gene locus (Prop1df) having deficiency of growth hormone (GH), thyroid stimulating hormone (TSH) and prolactin (PRL). This model is characteristic of prolonged longevity (~50% longer) and improved insulin sensitivity in comparison to their wild-type (N) counterparts. Ames dwarf (df/df) mice have recently been shown to have highly reduced plasma TAG levels, associating them with reduced susceptibility to atherosclerosis and associated CVDs. The underlying mechanism responsible for reduced VLDL secretion in Ames dwarf mice is yet to be characterized. We hypothesize that VTV-mediated trafficking of VLDL is reduced in Ames dwarf mice because of reduced expression of proteins regulating VLDL and VTV formation. To test our hypothesis, we first performed VTV-budding assay using cellular fractions isolated separately from Ames dwarf (df/df) and wild-type (N) mice livers. Our results show a significant (45%) reduction in VTV-budding process in Ames dwarf (df/df) mice compared to wild-type (N). Next we performed 2-dimensional differential gel electrophoresis (2-DIGE) on VTV and whole cell lysate (WCL) samples in order to examine the differences in protein expression and to have highly specific protein separation. ExPASy database was used to analyze protein spots that allowed us in identifying proteins specifically expressed in each of the mouse groups. Employing western blotting, samples (ER, cytosol, VTV and WCL) from both sets of mice were tested for expression levels of VLDL and VTV associated proteins (ApoB100, Sec22b, CideB, MTP, Apo-A1 and Apo-AIV) with ?-actin as the loading control. Significant differences in expression level of these proteins were observed which strongly suggest that the formation of VTV from ER in male Ames dwarf (df/df) mice is reduced compared to wild-type (N). Overall, we conclude that the differential expression of proteins required for VLDL transport causes reduced VLDL secretion in male Ames dwarf (df/df) mice.
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Simulation Study Of A Gpram System: Error Control Coding And ConnectionismSchultz, Steven E 01 January 2012 (has links)
A new computing platform, the General Purpose Reprsentation and Association Machine is studied and simulated. GPRAM machines use vague measurements to do a quick and rough assessment on a task; then use approximated message-passing algorithms to improve assessment; and finally selects ways closer to a solution, eventually solving it. We illustrate concepts and structures using simple examples.
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