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Journal of Lipid Research Feb 1999Apolipoprotein[a] phenotyping is a critically important method to explore the role of kringle-4 repeat number as a modulator of lipoprotein[a]-associated cardiovascular...
Apolipoprotein[a] phenotyping is a critically important method to explore the role of kringle-4 repeat number as a modulator of lipoprotein[a]-associated cardiovascular risk. The availability of a kringle-4 number-based reference standard is therefore necessary for a reliable and generally accepted classification of apo[a] phenotypes. We propose here a battery of recombinant apo[a] isoforms that may be used as the reference standard in various gel systems. Five plasmids encoding for r-apo[a] containing a known number (n = 9, 13, 17, 25, 33) of plasminogen-like kringle-4 copies were constructed, and transfected into the human embryonic kidney cell line 293. The electrophoretic mobility of the recombinant apo[a] isoforms expressed by these cells in a hollow-fiber bioreactor was determined after reduction by SDS-gel (agarose, acrylamide or a mixture of both) electrophoresis and immunoblotting using an antibody specific for human apo[a]. The equation of the linear relationship between log r-apo[a] kringle number and relative migration was used to determine the isoform size of apo[a] in normal human plasma. A very good correlation (r = 0.97) was found with the genotype (pulsed-field gel eletrophoresis of kpnI-digested restriction fragments of genomic DNA) and among electrophoretic methods. The proposed recombinant standard offers the possibility to identify apo[a] isoforms within a large range of molecular sizes, 9 to 33 kringle-4 copies, using simple electrophoretic techniques and a nomenclature based on its molecular structure, i.e., the number of kringle-4 repeats.-Anglés-Cano, E., S. Loyau, G. Cardoso-Saldaña, R. Couderc, and P. Gillery. A novel kringle-4 number-based recombinant apo[a] standard for human apo[a] phenotyping.
Topics: Apolipoproteins A; Cell Line; Electrophoresis, Polyacrylamide Gel; Genotype; Humans; Kidney; Kringles; Linear Models; Phenotype; Protein Isoforms; Recombinant Proteins; Reference Standards; Repetitive Sequences, Nucleic Acid
PubMed: 9925666
DOI: No ID Found -
Journal of Lipid Research May 1991The kinetics of apolipoprotein A-IV associated with high density lipoproteins (HDL) of plasma from fasting human subjects was followed for 15 days in five healthy... (Comparative Study)
Comparative Study
The kinetics of apolipoprotein A-IV associated with high density lipoproteins (HDL) of plasma from fasting human subjects was followed for 15 days in five healthy normolipidemic volunteers. Purified apoA-IV and apoA-I were radioiodinated, respectively, with 125I and 131I, incubated in vitro with normal HDL, isolated at density 1.250 g/ml, and finally reinjected intravenously as HDL-125I-labeled apoA-IV and HDL-131I-labeled apoA-I. Blood samples were withdrawn at regular intervals for 15 days, and 24-h urine samples were collected. More than 93% (93.5 +/- 0.9%) of apoA-IV was recovered in apoA-I-containing lipoprotein particles after affinity chromatography on an anti-apoA-I column and 69.7 +/- 4.8% was bound to apoA-II in apoA-I:A-II particles separated on an anti-apoA-II column. 125I-labeled apoA-IV showed a much faster decay than 131I-labeled apoA-I for the first 5 days and thereafter the curves became parallel. Urinary/plasma ratios (U/P) for the 125I-labeled parallel. Urinary/plasma ratios (U/P) for the 125I-labeled apoA-IV were much higher than those for 131I-labeled apoA-I for the first days, but the U/P curves became parallel for the last 7 days, suggesting heterogeneity of apoA-IV metabolism. A heterogeneous multicompartmental model was constructed to describe the metabolism of lipoprotein particles containing apoA-IV and apoA-I and to calculate the kinetic parameters, fitting simultaneously all plasma and urine data for both tracers.(ABSTRACT TRUNCATED AT 250 WORDS)
Topics: Adult; Apolipoprotein A-I; Apolipoproteins A; Female; Humans; Iodine Radioisotopes; Lipoproteins, HDL; Male; Models, Biological
PubMed: 1906522
DOI: No ID Found -
Journal of Lipid Research Sep 2000Viscoelastic behavior of proteins at interfaces is a critical determinant of their ability to stabilize emulsions. We therefore used air bubble surfactometry and drop...
Viscoelastic behavior of proteins at interfaces is a critical determinant of their ability to stabilize emulsions. We therefore used air bubble surfactometry and drop volume tensiometry to examine the dynamic interfacial properties of two plasma apolipoproteins involved in chylomicron assembly: apolipoprotein A-IV and apolipoprotein B-17, a recombinant, truncated apolipoprotein B. At the air/water interface apolipoproteins A-IV and B-17 displayed wide area - tension loops with positive phase angles indicative of viscoelastic behavior, and suggesting that they undergo rate-dependent changes in surface conformation in response to changes in interfacial area. At the triolein/water interface apolipoprotein A-IV displayed maximal surface activity only at long interface ages, with an adsorption rate constant of 1.0 3 10(-)(3) sec(-)(1), whereas apolipoprotein B-17 lowered interfacial tension even at the shortest interface ages, with an adsorption rate constant of 9.3 3 10(-)(3) sec(-)(1). Apolipoprotein A-IV displayed an expanded conformation at the air/water interface and a biphasic compression isotherm, suggesting that its hydrophilic amphipathic helices move in and out of the interface in response to changes in surface pressure. We conclude that apolipoproteins A-IV and B-17 display a combination of interfacial activity and elasticity particularly suited to stabilizing the surface of expanding triglyceride-rich particles.
Topics: Air; Apolipoprotein A-I; Apolipoproteins A; Apolipoproteins B; Elasticity; Humans; Oils; Pressure; Protein Conformation; Recombinant Proteins; Surface Properties; Surface Tension; Viscosity; Water
PubMed: 10974049
DOI: No ID Found -
Journal of Lipid Research Jan 1994This study illustrates that genetic strain and feeding status can markedly influence tissue lipid concentrations and mRNA levels of apolipoprotein genes. C57BL/6 and... (Comparative Study)
Comparative Study
This study illustrates that genetic strain and feeding status can markedly influence tissue lipid concentrations and mRNA levels of apolipoprotein genes. C57BL/6 and BALB/c mice were maintained for 2 weeks on four test diets differing in amount of cholesterol and type of fat, and fasted for 4 h or 16 h prior to collection of tissues. For both strains, the primary effect of fasting from 4 h to 16 h was to paradoxically elevate triglyceride levels in plasma and liver, and to elevate hepatic apoA-IV mRNA levels. Triglyceride secretion rates, estimated after the injection of Triton WR-1339, suggested that elevations in plasma triglyceride levels were due to reduced clearance of very low density lipoproteins. Although plasma glucose levels decreased with fasting time for both strains, insulin levels decreased for BALB/c but not C57BL/6 mice regardless of diet. This suggests that factors thought to be mediated by insulin, (e.g., plasma free fatty acid concentrations; hepatic apoA-IV mRNA levels) may be influenced by local changes in insulin sensitivity, which are controlled genetically and are not reflected by plasma insulin levels. In summary, nutritional status influences a constellation of factors involved in lipid transport that also show strong genetic components and may influence subsequent analyses of gene expression in the mouse system.
Topics: Animals; Apolipoprotein A-I; Apolipoprotein A-II; Apolipoproteins A; Blood Glucose; Fatty Acids, Nonesterified; Female; Insulin; Lipid Metabolism; Lipids; Lipoproteins, VLDL; Liver; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Nutritional Status; RNA, Messenger; Species Specificity; Triglycerides
PubMed: 8138713
DOI: No ID Found -
International Journal of Molecular... Apr 2021Hepatocyte growth factor (HGF) is an endogenously induced bioactive molecule that has strong anti-apoptotic and tissue repair activities. In this research, we identified...
BACKGROUND
Hepatocyte growth factor (HGF) is an endogenously induced bioactive molecule that has strong anti-apoptotic and tissue repair activities. In this research, we identified APOA4 as a novel pharmacodynamic (PD) marker of the recombinant human HGF (rh-HGF), E3112.
METHODS
rh-HGF was administered to mice, and their livers were investigated for the PD marker. Candidates were identified from soluble proteins and validated by using human hepatocytes in vitro and an animal disease model in vivo, in which its c-Met dependency was also ensured.
RESULTS
Among the genes induced or highly enhanced after rh-HGF exposure in vivo, a soluble apolipoprotein, , was found to be induced by rh-HGF in the murine liver. By using primary cultured human hepatocytes, the significant induction of human APOA4 was observed at the mRNA and protein levels, and it was inhibited in the presence of a c-Met inhibitor. Although mice constitutively expressed mRNA in the small intestine and the liver, the liver was the primary organ affected by administered rh-HGF to strongly induce APOA4 in a dose- and c-Met-dependent manner. Serum APOA4 levels were increased after rh-HGF administration, not only in normal mice but also in anti-Fas-induced murine acute liver failure (ALF), which confirmed the pharmacodynamic nature of APOA4.
CONCLUSIONS
APOA4 was identified as a soluble PD marker of rh-HGF with c-Met dependency. It should be worthwhile to clinically validate its utility through clinical trials with healthy subjects and ALF patients.
Topics: Animals; Apolipoproteins A; Biomarkers, Pharmacological; Cells, Cultured; Gene Expression Regulation; Hepatocyte Growth Factor; Hepatocytes; Humans; Liver; Liver Failure, Acute; Male; Mice, Inbred BALB C; Proto-Oncogene Proteins c-met; Recombinant Proteins; Mice
PubMed: 33925510
DOI: 10.3390/ijms22094578 -
Journal of the Royal Society of Medicine Feb 1989Plasma and lipoprotein cholesterol and triglycerides, and plasma apolipoproteins AI, AII and B were compared in patients with chronic airflow limitation, and normal...
Plasma and lipoprotein cholesterol and triglycerides, and plasma apolipoproteins AI, AII and B were compared in patients with chronic airflow limitation, and normal controls matched for body mass index. The controls were non-smokers, and free from respiratory disease. High-density lipoprotein (HDL) cholesterol concentration was significantly elevated in the patients, due mainly to a raised HDL2 cholesterol level. HDL triglyceride was significantly lower in the patients. All other lipids were not different from normal. Apolipoprotein AI levels were significantly raised in the patients but other apolipoproteins were unchanged. The changes found may account in part for the fact that patients with chronic airflow limitation have a lower incidence of atherosclerotic heart disease.
Topics: Adult; Aged; Apolipoprotein A-I; Apolipoprotein A-II; Apolipoproteins; Apolipoproteins A; Apolipoproteins B; Cholesterol, HDL; Humans; Lipoproteins; Lipoproteins, HDL; Lung Diseases, Obstructive; Male; Middle Aged; Triglycerides
PubMed: 2494326
DOI: 10.1177/014107688908200212 -
Journal of Lipid Research Dec 1996Lp[a] concentrations in nmol/L and apo[a] size isoforms, expressed in terms of the relative number of apo[a] kringle 4 (K4) repeats, were determined in 3959 whites and...
Lp[a] concentrations in nmol/L and apo[a] size isoforms, expressed in terms of the relative number of apo[a] kringle 4 (K4) repeats, were determined in 3959 whites and blacks from four U.S. communities. Plasma Lp[a] analyses were performed by an ELISA method insensitive to apo[a] size heterogeneity and apo[a] size isoforms were determined by high resolution agarose gel electrophoresis. Allele frequencies were estimated by maximum likelihood methods in order to account for the presence of null alleles and coalescence of hands on gels. The apo[a] allele frequencies and phenotype distributions differed significantly between blacks and whites (P < 0.0001). Blacks had a higher relative frequency of the intermediate alleles K4(22) through K4(28) whereas whites had a higher relative frequency of the small alleles K4(17) through K4(24) and large alleles K4(29) through K4(33). The estimated frequency of the null allele was low in both blacks (1.0%) and whites (6.7%). The Lp[a] distribution was less skewed and Lp[a] concentrations were higher in blacks than whites (mean 94 nmol/L and 48 nmol/L, median 74 nmol/L and 20 nmol/L for blacks and whites, respectively). The relationship between apo[a] size and Lp[a] concentration also differed significantly between these two racial groups. For the large polymorphs (> 31 K4 repeats) both blacks and whites exhibited uniformly low Lp[a] values. For the intermediate isoforms K4(20) through K4(30), a considerable range of Lp[a] values was evident in blacks; the median Lp[a] for each isoform increased nearly linearly as the apo[a] size decreased. In contrast in whites there was little change in median Lp[a] concentrations for isoforms K4(20) through K4(30). For the small apo[a] size (< 20 K4) both blacks and whites exhibited high median Lp[a] levels and a wide variation of Lp[a] levels. The major difference in Lp[a] levels between the two racial groups occurred in the intermediate size isoform range of K4(20) through K4(25). In conclusion, whites and blacks differ significantly in Lp[a] concentrations, allele and phenotype frequencies, and in the relationship between apo[a] size isoform and Lp[a] concentration.
Topics: Adolescent; Adult; Alleles; Apolipoproteins A; Black People; Female; Gene Frequency; Humans; Lipoprotein(a); Male; Polymorphism, Genetic; United States; White People
PubMed: 9017509
DOI: No ID Found -
Journal of the American College of... May 2017
Topics: Aged; Aged, 80 and over; Amyloidosis; Apolipoproteins A; Biomarkers; Cardiomyopathies; Female; Humans; Male; Middle Aged
PubMed: 28449784
DOI: 10.1016/j.jacc.2017.02.047 -
Current Opinion in Lipidology Aug 2008We have examined the evidence from recent human studies examining the role of apolipoprotein A-V in triglyceride-rich lipoprotein metabolism and cardiovascular disease... (Review)
Review
PURPOSE OF REVIEW
We have examined the evidence from recent human studies examining the role of apolipoprotein A-V in triglyceride-rich lipoprotein metabolism and cardiovascular disease risk. Special emphasis was placed on the evidence emerging from the association between genetic variability at the apolipoprotein A5 locus, lipid phenotypes and disease outcomes. Moreover, we address recent reports evaluating apolipoprotein A5 gene-environment interactions in relation to cardiovascular disease and its common risk factors.
RECENT FINDINGS
Several genetic association studies have continued to strengthen the position of APOA5 as a major gene that is involved in triglyceride metabolism and modulated by dietary factors and pharmacological therapies. Moreover, genetic variants at this locus have been significantly associated with both coronary disease and stroke risks.
SUMMARY
Apolipoprotein A-V has an important role in lipid metabolism, specifically for triglyceride-rich lipoproteins. However, its mechanism of action is still poorly understood. Clinical significance at present comes largely from genetic studies showing a consistent association with plasma triglyceride concentrations. Moreover, the effects of common genetic variants on triglyceride concentrations and disease risk are further modulated by other factors such as diet, pharmacological interventions and BMI. Therefore, these genetic variants could be potentially used to predict cardiovascular disease risk and individualize therapeutic options to decrease cardiovascular disease risk.
Topics: Apolipoprotein A-V; Apolipoproteins A; Diabetes Complications; Endpoint Determination; Humans; Metabolic Syndrome; Polymorphism, Single Nucleotide; Triglycerides
PubMed: 18607181
DOI: 10.1097/MOL.0b013e328304b681 -
Journal of Lipid Research Dec 2015Elevated lipoprotein (a) [Lp(a)] levels are a causal risk factor for coronary heart disease. Accumulating evidence suggests that Lp(a) can stimulate cellular...
Elevated lipoprotein (a) [Lp(a)] levels are a causal risk factor for coronary heart disease. Accumulating evidence suggests that Lp(a) can stimulate cellular inflammatory responses through the kringle-containing apolipoprotein (a) [apo(a)] component. Here, we report that recombinant apo(a) containing 17 kringle (17K) IV domains elicits a dose-dependent increase in interleukin (IL)-8 mRNA and protein expression in THP-1 and U937 macrophages. This effect was blunted by mutation of the lysine binding site in apo(a) kringle IV type 10, which resulted in the loss of oxidized phospholipid (oxPL) on apo(a). Trypsin-digested 17K had the same stimulatory effect on IL-8 expression as intact apo(a), while enzymatic removal of oxPL from apo(a) significantly blunted this effect. Using siRNA to assess candidate receptors, we found that CD36 and TLR2 may play roles in apo(a)-mediated IL-8 stimulation. Downstream of these receptors, inhibitors of MAPKs, Jun N-terminal kinase and ERK1/2, abolished the effect of apo(a) on IL-8 gene expression. To assess the roles of downstream transcription factors, luciferase reporter gene experiments were conducted using an IL-8 promoter fragment. The apo(a)-induced expression of this reporter construct was eliminated by mutation of IL-8 promoter binding sites for either NF-κB or AP-1. Our results provide a mechanistic link between oxPL modification of apo(a) and stimulation of proinflammatory intracellular signaling pathways.
Topics: Apolipoproteins; Apolipoproteins A; Binding Sites; Cell Line; Humans; Interleukin-8; NF-kappa B; Phospholipids; RNA, Small Interfering; Transcription Factor AP-1
PubMed: 26474593
DOI: 10.1194/jlr.M060210