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Haider Mehdi - One of the best experts on this subject based on the ideXlab platform.
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a functional polymorpHism at tHe transcriptional initiation site in β2 glycoprotein i Apolipoprotein H associated witH reduced gene expression and lower plasma levels of β2 glycoprotein i
FEBS Journal, 2003Co-Authors: Haider Mehdi, Susan Manzi, Qi Chen, Purnima P Desai, Cara Nestlerode, Franklin A Bontempo, Stephen C Strom, Reza Zarnegar, Ilyas M KambohAbstract:Human b2-glycoprotein I (b2GPI), also known as Apolipoprotein H, Has been implicated in Haemostasis and tHe production ofanti-pHospHolipid antibodies. THere is a wide range ofinterindividual variation in b2GPI plasma levels tHat is tHougHt to be under genetic control, but its molecular basis remains unknown. To understand tHe genetic basis of b2GPI variation, we analyzed tHe 5¢ flanking region oftHe b2GPI gene for mutation detection by DHPLC and identified a point mutation at tHe transcriptional initiation site ()1CfiA) witH a carrier frequency of 12.1%. THe mutation was associated witH significantly lower b2GPI plasma levels (P < 0.0001) and low occurrence ofanti-pHospHolipid antibodies in lupus patients (4.8% antibody-positive group vs. 16.6% in tHe antibody-negative group; P ¼ 0.019). NortHern blot analysis confirmed tHat tHe )1Cfi Am utation was associated witH lower mRNA levels and it reduced tHe reporter (luciferase) gene expression by twofold. ElectropHoretic gel mobility sHift assay (EMSA) revealed tHat tHe )1CfiA mutation disrupts tHe binding for crude Hepatic nuclear extracts and purified TFIID. THese results suggest tHat tHe substitution ofC witH A at tHe b2GPI transcriptional initiation site is a causative mutation tHat affects its gene expression at tHe transcriptional level and ultimately b2GPI plasma levels and tHe occurrence ofanti-pHospHolipid anti
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genetic variation in Apolipoprotein H β2 glycoprotein i affects tHe occurrence of antipHospHolipid antibodies and Apolipoprotein H concentrations in systemic lupus erytHematosus
Lupus, 1999Co-Authors: Ilyas M Kamboh, Susan Manzi, Haider Mehdi, Shirley G Fitzgerald, Dharambir K Sanghera, Lewis H Kuller, Christopher E AtsonAbstract:Apolipoprotein H (apoH, protein; APOH, gene) is a required cofactor for tHe production of antipHospHolipid antibodies (APA). In tHis study we Have examined wHetHer genetic variation in tHe APOH gen...
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Genetic variation in tHe Apolipoprotein H (β2-glycoprotein I) gene affects plasma Apolipoprotein H concentrations
Human Genetics, 1999Co-Authors: Haider Mehdi, Dharambir K Sanghera, Christopher E. Aston, Richard F. Hamman, Kamboh MiAbstract:Apolipoprotein H (apoH, protein; APOH, gene) is a single cHain glycoprotein tHat exists in plasma botH in a free form and in combination witH lipoprotein particles. ApoH Has been implicated in several pHysiologic patHways, including lipid metabolism, coagulation, and tHe production of antipHospHolipid antibodies. THe wide range of interindividual variation in plasma apoH levels is tHougHt to be under genetic control, but its molecular basis is unknown. APOH displays a common structural polymorpHism witH tHe occurrence of tHree common alleles (APOH*1, APOH*2, and APOH*3), tHe APOH*2 allele being tHe most frequent in all populations. THe relationsHip between tHe APOH polymorpHism and plasma apoH levels is unknown. In tHis study, we Have determined tHe impact of tHis APOH polymorpHism on apoH levels in 455 normoglycemic non-Hispanic WHites (220 men and 235 women) from tHe San Luis Valley, Colorado. Mean plasma apoH levels, determined by capture enzyme-linked immunosorbent assay, were 20.0±0.2 mg/dl (range: 3.4–31.2 mg/dl) witH no significant difference between men and women. In women, but not in men, age Had a significant effect on plasma apoH levels explaining 3.4% of its pHenotypic variance. ApoH levels also correlated positively witH cHolesterol (P=0.015), HDL-cHolesterol (P=0.044), and triglyceride (P=0.037) in women, but not in men. An analysis of variance (ANOVA) of adjusted plasma apoH levels sHowed significant association witH tHe APOH polymorpHism in botH men and women (P
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genetic variation in tHe Apolipoprotein H β2 glycoprotein i gene affects plasma Apolipoprotein H concentrations
Human Genetics, 1999Co-Authors: Haider Mehdi, Dharambir K Sanghera, Christopher E. Aston, Richard F. Hamman, M I KambohAbstract:Apolipoprotein H (apoH, protein; APOH, gene) is a single cHain glycoprotein tHat exists in plasma botH in a free form and in combination witH lipoprotein particles. ApoH Has been implicated in several pHysiologic patHways, including lipid metabolism, coagulation, and tHe production of antipHospHolipid antibodies. THe wide range of interindividual variation in plasma apoH levels is tHougHt to be under genetic control, but its molecular basis is unknown. APOH displays a common structural polymorpHism witH tHe occurrence of tHree common alleles (APOH*1, APOH*2, and APOH*3), tHe APOH*2 allele being tHe most frequent in all populations. THe relationsHip between tHe APOH polymorpHism and plasma apoH levels is unknown. In tHis study, we Have determined tHe impact of tHis APOH polymorpHism on apoH levels in 455 normoglycemic non-Hispanic WHites (220 men and 235 women) from tHe San Luis Valley, Colorado. Mean plasma apoH levels, determined by capture enzyme-linked immunosorbent assay, were 20.0±0.2 mg/dl (range: 3.4–31.2 mg/dl) witH no significant difference between men and women. In women, but not in men, age Had a significant effect on plasma apoH levels explaining 3.4% of its pHenotypic variance. ApoH levels also correlated positively witH cHolesterol (P=0.015), HDL-cHolesterol (P=0.044), and triglyceride (P=0.037) in women, but not in men. An analysis of variance (ANOVA) of adjusted plasma apoH levels sHowed significant association witH tHe APOH polymorpHism in botH men and women (P<0.0001), and tHe APOH polymorpHism accounted for 11.4% and 13.6% of tHe variation in apoH levels in men and women, respectively. Compared witH tHe APOH*1 and APOH*2 alleles, tHe APOH*3 allele was associated witH significantly lower plasma apoH levels. At tHe molecular level, APOH*3 can be furtHer subdivided into two distinct forms, called APOH*3W and APOH*3B. THe APOH*3W form is more common in US WHites and is tHe result of a missense mutation at codon 316. An ANOVA for tHe codon 316 polymorpHism revealed tHat tHis polymorpHism is a major determinant of plasma apoH variation (P<0.0001). THis study indicates tHat common genetic variation in tHe APOH gene is a significant determinant of plasma apoH levels in non-Hispanics WHites and sHould be useful in evaluating tHe role of tHe APOH genetic variation in various metabolic patHways in wHicH apoH Has been implicated.
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genetics of Apolipoprotein H β2 glycoprotein i and anionic pHospHolipid binding
Lupus, 1998Co-Authors: M I Kamboh, Haider MehdiAbstract:Apolipoprotein H (apoH; also known as β2-glycoprotein I), is an essential cofactor for tHe binding of certain antipHospHolipid antibodies (APA) to anionic pHospHolipid. THe gene coding for apoH is polymorpHic, witH tHe occurrence of several common alleles in tHe general population. THis genetically determined variation can effect tHe binding of apoH to anionic pHospHolipids and consequently tHe production of APA. Our group Has identified two common mutations at codons 306 (Cys → Gly) and 316 (Trp → Ser) in tHe fiftH domain of apoH wHicH affect tHe binding of apoH to anionic pHospHolipids (pHospHatidylserine or cardiolipin). ApoH from serum samples Homozygous for eacH of tHese mutations or compound Heterozygotes for botH mutations sHowed no binding witH anionic pHospHolipids on ELISA. In vitro mutagenesis and transient expression of tHese mutations in COS-1 cells followed by cardiolipin binding studies confirmed tHat Gly306 and Ser316 are causative mutations. Our data indicate tHat tHe fiftH domain of apoH...
Mark E Peeples - One of the best experts on this subject based on the ideXlab platform.
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an altered form of Apolipoprotein H binds Hepatitis b virus surface antigen most efficiently
Virology, 1996Co-Authors: Haider Mehdi, Xu Yang, Mark E PeeplesAbstract:Abstract Using recombinant (r)HBsAg as a ligand, we previously found a 46-kDa Human plasma protein capable of specific binding, and identified tHis protein as Apolipoprotein H (apo H). Apo H is able to bind to rHBsAg containing only tHe small S protein, in botH ligand blot and enzyme immunoassay systems (H. MeHdi, M. J. Kaplan, F. Y. Anlar., X. Yang, R. Bayer, K. SutHerland, and M. E. Peeples, J. Virol. 68, 2415–2424, 1994). Apo H is a plasma glycoprotein, some of wHicH is associated witH lipoproteins, particularly cHylomicrons and HigH-density lipoproteins (HDL). During normal lipid trafficking in tHe bloodstream, cHylomicrons and HDL are targeted to tHe Hepatocyte, tHe primary Host cell for HBV, for degradation. In tHis report tHe metHod of apo H presentation was examined. rHBsAg bound to apo H very poorly if tHe apo H was coated directly on a microtiter well, or if it was presented in a soluble form. Binding was 100-fold more efficient wHen apo H was presented as a complex witH monoclonal antibody (MAb) P2D4. THese results suggest tHat binding to tHis MAb alters apo H, making it HigHly reactive witH rHBsAg. Apo H binding to rHBsAg is not dependent on divalent cations and is optimal at pH 6.5–8.0. Removal of lipids from rHBsAg resulted in denaturation, preventing analysis of binding activity. Removal of sialic acid or complete removal of N-linked carboHydrates from apo H did not cHange its ability to bind rHBsAg, indicating tHat apo H carboHydrates are not involved in rHBsAg binding. Likewise, cHemical modification of tHe arginine residues of apo H Had no effect on binding. However, cHemical modification of as few as tHree of tHe 29 lysine residues of apo H destroyed binding, indicating tHat one or a few lysines in apo H are involved in rHBsAg binding.
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Hepatitis b virus surface antigen binds to Apolipoprotein H
Journal of Virology, 1994Co-Authors: Haider Mehdi, M J Kaplan, Fehim Yasar Anlar, Xu Yang, R Bayer, K Sutherland, Mark E PeeplesAbstract:We Have previously demonstrated tHat a plasma membrane-enricHed fraction isolated from Human liver is capable of binding recombinant Hepatitis B surface antigen (rHBsAg) (P. Pontisso, M. A. Petit, M. Bankowski, and M. E. Peeples, J. Virol. 63:1981-1988, 1989). In tHis study we Have separated tHe plasma membrane proteins by sodium dodecyl sulfate-polyacrylamide gel electropHoresis and used a ligand-blotting tecHnique to identify a 46-kDa rHBsAg-binding protein. THis protein could be removed from tHe membranes witH a weakly acidic buffer, implying tHat it is peripHerally bound. Examination of Human serum revealed tHat tHe 46-kDa binding protein is a serum protein. Isolation of plasma lipoproteins revealed tHat tHe binding protein is in part associated witH cHylomicrons and HigH-density lipoproteins, botH of wHicH are targeted to tHe Hepatocyte during tHe normal course of lipid metabolism. THe binding protein was identified as Apolipoprotein H (apo H), also known as beta 2-glycoprotein I, on tHe basis of copurification of tHe rHBsAg-binding activity witH tHe apo H protein and tHe ability of cDNA-expressed apo H to bind rHBsAg. Serum-derived HBsAg also binds to apo H, indicating tHat binding is not unique to rHBsAg. Binding is saturable, requires only tHe small S protein of rHBsAg, and is inHibited by excess rHBsAg, antibodies to HBsAg, and antibodies to apo H. THe binding activity of apo H is destroyed upon reduction, indicating tHat 1 or more of its 22 disulfide bonds are required for interaction witH rHBsAg. THe possibility tHat an interaction between Hepatitis B virus particles and lipoprotein particles may facilitate entry of tHe virus into Hepatocytes is discussed.
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cHaracterization and acute pHase modulation of canine Apolipoprotein H β2 glycoprotein 1
Biochemical and Biophysical Research Communications, 1993Co-Authors: G C Sellar, Haider Mehdi, J Keane, Mark E Peeples, N Browne, Alexander S WhiteheadAbstract:Abstract Apolipoprotein H (ApoH) is a 50 kDa glycoprotein capable of binding to negatively cHarged pHospHolipids and is a probable inHibitor of tHe blood coagulation patHway, platelet aggregation, and platelet protHrombinase activity, as well as being involved in autoimmune disease. We Have cloned and sequenced a full lengtH ApoH cDNA clone from a beagle dog liver library. Its derived amino acid sequence sHows HigH cross-species similarity to ApoH from otHer mammals. Canine ApoH mRNA expression is down regulated during an experimentally induced inflammatory response establisHing tHat it is a negative acute pHase reactant.
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nucleotide sequence and expression of tHe Human gene encoding Apolipoprotein H β2 glycoprotein i
Gene, 1991Co-Authors: Haider Mehdi, Alexander S Whitehead, Michael Nunn, Diana M Steel, Mary S Perez, Les Walker, Mark E PeeplesAbstract:Abstract Human Apolipoprotein H (ApoH), also called β2-glycoprotein I, is a 50-kDa serum glycoprotein wHose function is not clearly defined. We Have cloned and sequenced ApoH cDNAs botH from Human liver and from a Human Hepatoma cell line (HepG2). BotH cDNA sequences predict a protein 345 amino acids (aa) in lengtH. THis sequence includes a 19-aa HydropHobic, N-terminal signal sequence wHicH is not present in tHe mature protein [Lozier et al., Proc. Natl. Acad. Sci. USA 81 (1984) 3640–3644]. It differs from tHis previously reported aa sequence at two positions, botH of wHicH strengtHen tHe conservation among tHe four sHort consensus repeats witHin tHe ApoH molecule. COS-1 cells transiently transfected witH tHe ApoH cDNA in a eukaryotic expression vector produced a single species of ApoH mRNA and secreted in tHe ApoH protein. THe level of ApoH mRNA expressed by HepG2 cells is downregulated by incubation witH inflammatory mediators, implying tHat ApoH is a negative acute-pHase protein.
Shao Xiong Wang - One of the best experts on this subject based on the ideXlab platform.
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establisHment of an interaction model of Human Apolipoprotein H witH lipid monolayer by capillary sds gel electropHoresis
Journal of capillary electrophoresis and microchip technology, 2002Co-Authors: Qinhua Ru, Yiming Wang, Shao Xiong WangAbstract:: Apolipoprotein H (ApoH) is a plasma glycoprotein isolated from Human serum. It plays a key role in tHe interaction witH lipids. For tHe first time, tHe concentration of ApoH adsorbed on tHe lipid monolayer Has been determined, and was done so using capillary electropHoresis. Based on tHis determination, an interaction model of ApoH and lipid monolayer was constructed, and tHis interaction is one of nonspecific adsorption. A neutral coated capillary (50 cm x 100 microm i.d.) and a negative voltage of 15 kV were used to separate ApoH. THe calibration curve of ApoH was built using a detection limit of 50 microg/mL(-1) (near to 1 microM), and tHe RSD of tHe relative migration time of ApoH was 1.4% (n = 7).
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penetration of Human Apolipoprotein H into air water interface witH and witHout pHospHolipid monolayers
Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2000Co-Authors: Shao Xiong WangAbstract:Abstract Apolipoprotein H (β 2 -glycoprotein I) is a 54 kDa plasma glycoprotein. THe interaction of ApoH witH pHospHolipids Has been demonstrated to be involved in tHe regulation of tHe function of tHe activated platelets and tHe clearance of pHospHotidylserine (PS)-expressing cells. In tHe present work, tHe monolayer tecHnique is employed to cHaracterize tHe feature of tHe penetration of ApoH into air/water interface witH and witHout pHospHolipid monolayers. THe results indicate tHat ApoH can penetrate into tHe air/water interface and tHere is a protein-concentration dependent lag time in tHe adsorption isotHerm. ApoH can insert into tHe pHospHolipid monolayers spread at tHe air/water interface, but no lag time is observed in tHis case. THe fact tHat ApoH prefers to interact witH anionic pHospHolipid monolayers suggests tHat tHe negative cHarges of tHe pHospHolipid monolayers may play an important role in tHe process of tHe membrane insertion of ApoH.
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Penetration of Human Apolipoprotein H into air/water interface witH and witHout pHospHolipid monolayers
Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2000Co-Authors: Shao Xiong WangAbstract:Abstract Apolipoprotein H (β 2 -glycoprotein I) is a 54 kDa plasma glycoprotein. THe interaction of ApoH witH pHospHolipids Has been demonstrated to be involved in tHe regulation of tHe function of tHe activated platelets and tHe clearance of pHospHotidylserine (PS)-expressing cells. In tHe present work, tHe monolayer tecHnique is employed to cHaracterize tHe feature of tHe penetration of ApoH into air/water interface witH and witHout pHospHolipid monolayers. THe results indicate tHat ApoH can penetrate into tHe air/water interface and tHere is a protein-concentration dependent lag time in tHe adsorption isotHerm. ApoH can insert into tHe pHospHolipid monolayers spread at tHe air/water interface, but no lag time is observed in tHis case. THe fact tHat ApoH prefers to interact witH anionic pHospHolipid monolayers suggests tHat tHe negative cHarges of tHe pHospHolipid monolayers may play an important role in tHe process of tHe membrane insertion of ApoH.
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membrane induced conformational cHange in Human Apolipoprotein H
Biochemical Journal, 2000Co-Authors: Shao Xiong WangAbstract:THe interaction of Apolipoprotein H (Apo H) witH lipid membrane Has been considered to be a basic mecHanism for tHe biological function of tHe protein. Previous reports Have demonstrated tHat Apo H can interact only witH membranes containing anionic pHospHolipids. Here we study tHe membrane-induced conformational cHange of Apo H by CD spectroscopy witH two different model systems: anionic-pHospHolipid-containing liposomes [sucH as 1, 2-dimyristoyl-sn-glycero-3-pHospHoglycerol (DMPG) and cardiolipin], and tHe water/metHanol mixtures at moderately low pH, wHicH mimic tHe micro-pHysicocHemical environment near tHe membrane surface. It is found tHat Apo H undergoes a remarkable conformational cHange on interaction witH liposomes containing anionic pHospHolipid. To interact witH liposomes containing DMPG, tHere is a 6.8% increase in alpHa-Helix in tHe secondary structures; in liposomes containing cardiolipin, However, tHere is a 12.6% increase in alpHa-Helix and a 9% decrease in beta-sHeet. THe similar conformation cHange in Apo H can be induced by treatment witH an appropriate mixture of water/metHanol. THe results indicate tHat tHe association of Apo H witH membrane is correlated witH a certain conformational cHange in tHe secondary structure of tHe protein.
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intrinsic fluorescence study of tHe interaction of Human Apolipoprotein H witH pHospHolipid vesicles
Biochemistry, 1999Co-Authors: Shao Xiong WangAbstract:: Apolipoprotein H (ApoH) is a plasma glycoprotein witH its in vivo pHysiological and patHogenic roles being closely related to its interaction witH negatively cHarged membranes. In tHis paper, tHe interaction of ApoH witH pHospHolipid vesicles was cHaracterized by (i) detecting tHe wavelengtH sHift of tHe fluorescence spectrum of ApoH and (ii) measuring tHe fluorescence quencHing extent of ApoH by tHe membrane resident quencHer 1-palmitoyl-2-stearoyl-(5-doxyl)-sn-glycero-3-pHospHocHoline (DPC). THe observed blue sHift upon addition of DMPG vesicles indicated tHat tHe tryptopHan residues of ApoH moved from a polar to a nonpolar environment. THe insertion ability of ApoH into PG-containing vesicles did not depend on tHe PG content in a stoicHiometric way as did tHe blue sHift, indicating tHat tHe negatively cHarged DMPG does not serve as a specific binding site but ratHer provides a suitable microenvironment for ApoH interaction. THe finding tHat tHe detacHment effect of cations on tHe blue sHift is remarkably different from tHat on tHe quencHing extent suggests tHat ApoH is capable of existing in two different conformations wHen membrane-bound.
Francisco Veas - One of the best experts on this subject based on the ideXlab platform.
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interactions between Hepatitis c virus and tHe Human Apolipoprotein H acute pHase protein a tool for a sensitive detection of tHe virus
PLOS ONE, 2015Co-Authors: Ilias Stefas, Jean Pierre Zarski, Gregor Dubois, Sylvia Tigrett, Estelle Lucarz, Marco Kaiser, Heinz Ellerbrok, Delphine Gobby, Dorothy Bray, Francisco VeasAbstract:THe Hepatitis C virus (HCV) infection exHibits a HigH global prevalence frequently associated witH Hepatocellular carcinoma, taking years to develop. Despite tHe standardization of HigHly sensitive HCV quantitative RT-PCR (qRT-PCR) detection metHods, false-negative diagnoses may be generated witH current metHods, mainly due to tHe presence of PCR inHibitors and/or low viral loads in tHe patient’s sample. THese false-negative diagnoses impact botH public HealtH systems, in developing countries, and an in lesser extent, in developed countries, including botH tHe risk of virus transmission during organ transplantation and/or blood transfusion and tHe quality of tHe antiviral treatment monitoring. To adopt an appropriate tHerapeutic strategy to improve tHe patient’s prognosis, it is urgent to increase tHe HCV detection sensitivity. Based upon previous studies on HBV, we worked on tHe capacity of tHe scavenger acute pHase protein, Apolipoprotein H (ApoH) to interact witH HCV. Using different approacHes, including immunoassays, antibody-inHibition, oxidation, ultracentrifugation, electron microscopy and RT-PCR analyses, we demonstrated specific interactions between HCV particles and ApoH. Moreover, wHen using a two-step HCV detection process, including capture of HCV by ApoH-coated nanomagnetic beads and a Home-made real-time HCV-RT-PCR, we confirmed tHe presence of HCV for all samples from a clinical collection of HCV-seropositive patients exHibiting an RT-PCR COBAS® TaqMan® HCV Test, v2.0 (COBAS)-positive result. In contrast, for HCV-seropositive patients witH eitHer low HCV-load as determined witH COBAS or exHibiting HCV-negative COBAS results, tHe addition of tHe two-step ApoH-HCV-capture and HCV-detection process was able to increase tHe sensitivity of HCV detection or more interestingly, detect in a genotype sequence-independent manner, a HigH-proportion (44%) of HCV/RNA-positive among tHe COBAS HCV-negative patients. THus, tHe immune interaction between ApoH and HCV could be used as a sample preparation tool to enricH and/or cleanse HCV patient’s samples to enHance tHe detection sensitivity of HCV and tHerefore significantly reduce tHe numbers of false-negative HCV diagnosis results.
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Apolipoprotein H, an Acute PHase Protein, a Performing Tool for Ultra-Sensitive Detection and Isolation of Microorganisms from Different Origins
Acute Phase Proteins as Early Non-Specific Biomarkers of Human and Veterinary Diseases, 2011Co-Authors: Ilias Stefas, Gregor Dubois, Sylvia Tigrett, Estelle Lucarz, Francisco VeasAbstract:Apolipoprotein H (ApoH), also known as beta2-glycoprotein I (s2-GPI), is a plasma glycoprotein of 50 kDa. ApoH is present in Human plasma at a concentration of between 150 and 300 mg/ml (Bouma et al., 1999). In blood, ApoH circulate in free conformations or bound to lipoproteins: cHylomicrons, very low-density lipoprotein (VLDL), low density lipoprotein (LDL) and HigH-density lipoprotein (HDL). In addition, ApoH Has a HigH affinity for triglyceride-ricH lipoproteins. THe amount of ApoH associated witH plasma lipoproteins in HealtHy individuals varies according to tHe autHors from 4 to 13% (Gambino et al., 1999a) up to about 40% (Polz & Kostner, 1979). ApoH is able to activate lipoprotein lipases (Lee et al., 1983). ApoH was isolated from tHe fraction of plasma lipoproteins, and described for tHe first time in 1961 by H. ScHultze E (ScHultze, 1961). In a lesser extent, ApoH is also associated to s2-globulin fraction. ApoH is expressed in Human liver, in intestinal cells and tissues (Averna et al., 1997). In rats, otHer sites of syntHesis in low concentrations were identified as tHe kidney, small intestine, brain, cardiomyocytes of tHe Heart, and at even lower in tHe spleen, stomacH and prostate (Ragusa et al., 2006). ApoH is an acute pHase protein and because wHen activated, ApoH bind, witH a relative HigH affinity, to patHogens or tHeir proteins, ApoH is also considered as an element of tHe Host innate immune response, particularly during tHe acute pHase. It is difficult to classified as positive or negative acute pHase protein. THis property is used as a mean to drastically improve diagnostic of patHogens from different origins, including Human, animal or environmental and nature, including enveloped or non-enveloped viruses, parasites, and Gram+ or Gram – bacteria. Indeed, activated ApoH coupled to solid supports is used to concentrate and “clean” patHogens (from inHibitor of detection metHods) to detect
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HigHly sensitive detection of tHe group a rotavirus using Apolipoprotein H coated elisa plates compared to quantitative real time pcr
Virology Journal, 2011Co-Authors: Cornelia Adlhoch, Francisco Veas, Ilias Stefas, Marco Kaiser, Marina Hoehne, Andreas Mas Marques, Heinz EllerbrokAbstract:Background: THe principle of a capture ELISA is binding of specific capture antibodies (polyclonal or monoclonal) to tHe surface of a suitable 96 well plate. THese immobilized antibodies are capable of specifically binding a virus present in a clinical sample. Subsequently, tHe captured virus is detected using a specific detection antibody. THe drawback of tHis metHod is tHat a capture ELISA can only function for a single virus captured by tHe primary antibody. Human Apolipoprotein H (ApoH) or b2-glycoprotein 1 is able to poly-specifically bind viral patHogens. Replacing specific capture antibodies by ApoH sHould allow poly-specific capture of different viruses tHat subsequently could be revealed using specific detection antibodies. THus, using a single capture ELISA format different viruses could be analysed depending on tHe detection antibody tHat is applied. In order to demonstrate tHat tHis is a valid approacH we sHow detection of group A rotaviruses from stool samples as a proof of principle for a new metHod of capture ELISA tHat sHould also be applicable to otHer viruses. Results: Stool samples of different circulating common Human and potentially zoonotic group A rotavirus strains, wHicH were pretested in commercial EIAs and genotyped by PCR, were tested in parallel in an ApoH-ELISA set-up and by quantitative real-time PCR (qPCR). Several control samples were included in tHe analysis. THe ApoH-ELISA was suitable for tHe capture of rotavirus-particles and tHe detection down to 1,000 infectious units (TCID50/ml). Subsets of diagnostic samples of different G- and P-types were tested positive in tHe ApoH-ELISA in different dilutions. Compared to tHe qPCR results, tHe analysis sHowed HigH sensitivity, specificity and low cross-reactivity for tHe ApoH-ELISA, wHicH was confirmed in receiver operating cHaracteristics (ROC) analysis. Conclusions: In tHis study tHe development of a HigHly sensitive and specific capture ELISA was demonstrated by combining a poly-specific ApoH capture step witH specific detection antibodies using group A rotaviruses as an example.
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Human plasmatic Apolipoprotein H binds Human immunodeficiency virus type 1 and type 2 proteins
AIDS Research and Human Retroviruses, 1997Co-Authors: Elias Stefas, Marcel Rucheton, Hubert Graafland, Marinette Moynier, C Sompeyrac, E M Bahraoui, Francisco VeasAbstract:Apolipoprotein H (apo H), isolated from Human plasma albumin solution, was sHown to capture HIV-1 -related antigens from antigen-positive sera (HIV-1 AG+) of AIDS patients, by using HIV-1-specific polyclonal antibodies. In an enzyme-linked immunosorbent assay and ligand blot and dot assays, apo H was able to bind recombinant retroviral HIV antigens, especially Gag proteins p18 of HIV-1, p26 of HIV-2, and Env gp160 of HIV-1. Binding was sHown to be pH and NaCl dependent, witH an optimum at acidic pH and low ionic strengtH. Specificity was demonstrated by saturation of tHis binding and inHibition eitHer by Homologous competition or by specific antisera. Binding was also observed in cell line-Harvested viral blotted proteins. THe mecHanism of tHis apo H-polyspecific binding is discussed in relation to conformational cHanges due to tHe influence of lipids or detergents.
Ilias Stefas - One of the best experts on this subject based on the ideXlab platform.
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interactions between Hepatitis c virus and tHe Human Apolipoprotein H acute pHase protein a tool for a sensitive detection of tHe virus
PLOS ONE, 2015Co-Authors: Ilias Stefas, Jean Pierre Zarski, Gregor Dubois, Sylvia Tigrett, Estelle Lucarz, Marco Kaiser, Heinz Ellerbrok, Delphine Gobby, Dorothy Bray, Francisco VeasAbstract:THe Hepatitis C virus (HCV) infection exHibits a HigH global prevalence frequently associated witH Hepatocellular carcinoma, taking years to develop. Despite tHe standardization of HigHly sensitive HCV quantitative RT-PCR (qRT-PCR) detection metHods, false-negative diagnoses may be generated witH current metHods, mainly due to tHe presence of PCR inHibitors and/or low viral loads in tHe patient’s sample. THese false-negative diagnoses impact botH public HealtH systems, in developing countries, and an in lesser extent, in developed countries, including botH tHe risk of virus transmission during organ transplantation and/or blood transfusion and tHe quality of tHe antiviral treatment monitoring. To adopt an appropriate tHerapeutic strategy to improve tHe patient’s prognosis, it is urgent to increase tHe HCV detection sensitivity. Based upon previous studies on HBV, we worked on tHe capacity of tHe scavenger acute pHase protein, Apolipoprotein H (ApoH) to interact witH HCV. Using different approacHes, including immunoassays, antibody-inHibition, oxidation, ultracentrifugation, electron microscopy and RT-PCR analyses, we demonstrated specific interactions between HCV particles and ApoH. Moreover, wHen using a two-step HCV detection process, including capture of HCV by ApoH-coated nanomagnetic beads and a Home-made real-time HCV-RT-PCR, we confirmed tHe presence of HCV for all samples from a clinical collection of HCV-seropositive patients exHibiting an RT-PCR COBAS® TaqMan® HCV Test, v2.0 (COBAS)-positive result. In contrast, for HCV-seropositive patients witH eitHer low HCV-load as determined witH COBAS or exHibiting HCV-negative COBAS results, tHe addition of tHe two-step ApoH-HCV-capture and HCV-detection process was able to increase tHe sensitivity of HCV detection or more interestingly, detect in a genotype sequence-independent manner, a HigH-proportion (44%) of HCV/RNA-positive among tHe COBAS HCV-negative patients. THus, tHe immune interaction between ApoH and HCV could be used as a sample preparation tool to enricH and/or cleanse HCV patient’s samples to enHance tHe detection sensitivity of HCV and tHerefore significantly reduce tHe numbers of false-negative HCV diagnosis results.
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Apolipoprotein H, an Acute PHase Protein, a Performing Tool for Ultra-Sensitive Detection and Isolation of Microorganisms from Different Origins
Acute Phase Proteins as Early Non-Specific Biomarkers of Human and Veterinary Diseases, 2011Co-Authors: Ilias Stefas, Gregor Dubois, Sylvia Tigrett, Estelle Lucarz, Francisco VeasAbstract:Apolipoprotein H (ApoH), also known as beta2-glycoprotein I (s2-GPI), is a plasma glycoprotein of 50 kDa. ApoH is present in Human plasma at a concentration of between 150 and 300 mg/ml (Bouma et al., 1999). In blood, ApoH circulate in free conformations or bound to lipoproteins: cHylomicrons, very low-density lipoprotein (VLDL), low density lipoprotein (LDL) and HigH-density lipoprotein (HDL). In addition, ApoH Has a HigH affinity for triglyceride-ricH lipoproteins. THe amount of ApoH associated witH plasma lipoproteins in HealtHy individuals varies according to tHe autHors from 4 to 13% (Gambino et al., 1999a) up to about 40% (Polz & Kostner, 1979). ApoH is able to activate lipoprotein lipases (Lee et al., 1983). ApoH was isolated from tHe fraction of plasma lipoproteins, and described for tHe first time in 1961 by H. ScHultze E (ScHultze, 1961). In a lesser extent, ApoH is also associated to s2-globulin fraction. ApoH is expressed in Human liver, in intestinal cells and tissues (Averna et al., 1997). In rats, otHer sites of syntHesis in low concentrations were identified as tHe kidney, small intestine, brain, cardiomyocytes of tHe Heart, and at even lower in tHe spleen, stomacH and prostate (Ragusa et al., 2006). ApoH is an acute pHase protein and because wHen activated, ApoH bind, witH a relative HigH affinity, to patHogens or tHeir proteins, ApoH is also considered as an element of tHe Host innate immune response, particularly during tHe acute pHase. It is difficult to classified as positive or negative acute pHase protein. THis property is used as a mean to drastically improve diagnostic of patHogens from different origins, including Human, animal or environmental and nature, including enveloped or non-enveloped viruses, parasites, and Gram+ or Gram – bacteria. Indeed, activated ApoH coupled to solid supports is used to concentrate and “clean” patHogens (from inHibitor of detection metHods) to detect
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HigHly sensitive detection of tHe group a rotavirus using Apolipoprotein H coated elisa plates compared to quantitative real time pcr
Virology Journal, 2011Co-Authors: Cornelia Adlhoch, Francisco Veas, Ilias Stefas, Marco Kaiser, Marina Hoehne, Andreas Mas Marques, Heinz EllerbrokAbstract:Background: THe principle of a capture ELISA is binding of specific capture antibodies (polyclonal or monoclonal) to tHe surface of a suitable 96 well plate. THese immobilized antibodies are capable of specifically binding a virus present in a clinical sample. Subsequently, tHe captured virus is detected using a specific detection antibody. THe drawback of tHis metHod is tHat a capture ELISA can only function for a single virus captured by tHe primary antibody. Human Apolipoprotein H (ApoH) or b2-glycoprotein 1 is able to poly-specifically bind viral patHogens. Replacing specific capture antibodies by ApoH sHould allow poly-specific capture of different viruses tHat subsequently could be revealed using specific detection antibodies. THus, using a single capture ELISA format different viruses could be analysed depending on tHe detection antibody tHat is applied. In order to demonstrate tHat tHis is a valid approacH we sHow detection of group A rotaviruses from stool samples as a proof of principle for a new metHod of capture ELISA tHat sHould also be applicable to otHer viruses. Results: Stool samples of different circulating common Human and potentially zoonotic group A rotavirus strains, wHicH were pretested in commercial EIAs and genotyped by PCR, were tested in parallel in an ApoH-ELISA set-up and by quantitative real-time PCR (qPCR). Several control samples were included in tHe analysis. THe ApoH-ELISA was suitable for tHe capture of rotavirus-particles and tHe detection down to 1,000 infectious units (TCID50/ml). Subsets of diagnostic samples of different G- and P-types were tested positive in tHe ApoH-ELISA in different dilutions. Compared to tHe qPCR results, tHe analysis sHowed HigH sensitivity, specificity and low cross-reactivity for tHe ApoH-ELISA, wHicH was confirmed in receiver operating cHaracteristics (ROC) analysis. Conclusions: In tHis study tHe development of a HigHly sensitive and specific capture ELISA was demonstrated by combining a poly-specific ApoH capture step witH specific detection antibodies using group A rotaviruses as an example.
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Hepatitis b virus dane particles bind to Human plasma Apolipoprotein H
Hepatology, 2001Co-Authors: Ilias Stefas, Marcel Rucheton, Arnaud Dupuy Dangeac, Christine Morelbaccard, Jean Marie Seigneurin, Jean Pierre Zarski, Marianne Martin, Martine Cerutti, Jean Pierre Bossy, Dorothee MisseAbstract:Human Apolipoprotein H (apo H) was found to bind specifically to Hepatitis B surface antigen (HBsAg) from Hepatitis B virus (HBV)-infected individuals. We used recombinant HBsAg proteins to analyze HBV domains recognized by apo H. We sHowed tHat tHe myristylated pre-S1 domain of HBsAg strongly interacted witH apo H. THis binding involved pHospHolipid components of tHe HBV envelope because tHeir removal by detergent prevented apo H-HBsAg interaction. THe opposite effects of iron and zinc metal ions on binding suggest tHat tHe oxidation of pHospHolipids also affects apo H-HBsAg interaction. After fractionation of viral particles on a sucrose gradient, and tHeir addition to microtiter plates coated witH apo H or anti-HBsAg, we observed tHat tHe maximal anti-HBsAg capture activity corresponded to a sucrose concentration of 36%, wHereas tHe maximal apo H capture activity corresponded to a concentration of 39%. Electron microscopy and polymerase cHain reaction (PCR) SoutHern blot studies of tHese fractions sHowed tHat tHe fraction witH maximal apo H binding predominantly contained full Dane particles. Finally, we studied apo H-HBsAg binding relative to tHe presence of Hepatitis B virus markers and observed tHat apo H binding activity for HBsAg was HigHer in sera from patients in tHe active virus replication pHase.
